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Sample records for egfr tyrosine kinase

  1. Design, Synthesis and Evaluation of Ribose-modified Anilinopyrimidine Derivatives as EGFR Tyrosine Kinase Inhibitors

    Science.gov (United States)

    Hu, Xiuqin; Wang, Disha; Tong, Yi; Tong, Linjiang; Wang, Xia; Zhu, Lili; Xie, Hua; Li, Shiliang; Yang, You; Xu, Yufang

    2017-11-01

    The synthesis of a series of ribose-modified anilinopyrimidine derivatives was efficiently achieved by utilizing DBU or tBuOLi-promoted coupling of ribosyl alcohols with 2,4,5-trichloropyrimidine as key step. Preliminary biological evaluation of this type of compounds as new EGFR tyrosine kinase inhibitors for combating EGFR L858R/T790M mutant associated with drug resistance in the treatment of non-small cell lung cancer revealed that 3-N-acryloyl-5-O-anilinopyrimidine ribose derivative 1a possessed potent and specific inhibitory activity against EGFR L858R/T790M over WT EGFR. Based upon molecular docking studies of the binding mode between compound 1a and EGFR, the distance between the Michael receptor and the pyrimidine scaffold is considered as an important factor for the inhibitory potency and future design of selective EGFR tyrosine kinase inhibitors against EGFR L858R/T790M mutants.

  2. Erkitinib, a novel EGFR tyrosine kinase inhibitor screened using a ProteoChip system from a phytochemical library

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    Kim, Eung-Yoon; Choi, Young-Jin [Biochip Research Center, Hoseo University, Asan 336-795 (Korea, Republic of); Innopharmascreen, Inc., Asan 336-795 (Korea, Republic of); Park, Chan-Won [Biochip Research Center, Hoseo University, Asan 336-795 (Korea, Republic of); Dept. of Biological Science, Hoseo University, Asan 336-795 (Korea, Republic of); Kang, In-Cheol, E-mail: ickang@hoseo.edu [Biochip Research Center, Hoseo University, Asan 336-795 (Korea, Republic of); Dept. of Biological Science, Hoseo University, Asan 336-795 (Korea, Republic of); Innopharmascreen, Inc., Asan 336-795 (Korea, Republic of)

    2009-11-20

    Receptor tyrosine kinases (PTKs) play key roles in the pathogenesis of numerous human diseases, including cancer. Therefore PTK inhibitors are currently under intensive investigation as potential drug candidates. Herein, we report on a ProteoChip-based screening of an epidermal growth factor receptor (EGFR) tyrosine kinase (TK) inhibitor, Erkitinibs, from phytochemical libraries. PLC-{gamma}-1 was used as a substrate immobilized on a ProteoChip and incubated with an EGFR kinase to phosphorylate tyrosine residues of the substrate, followed by a fluorescence detection of the substrate recognized by a phospho-specific monoclonal antibody. Erkitinibs inhibited HeLa cell proliferation in a dose-dependent manner. In conclusion, these data suggest that Erkitinibs can be a specific inhibitor of an EGFR kinase and can be further developed as a potent anti-tumor agent.

  3. Erkitinib, a novel EGFR tyrosine kinase inhibitor screened using a ProteoChip system from a phytochemical library

    International Nuclear Information System (INIS)

    Kim, Eung-Yoon; Choi, Young-Jin; Park, Chan-Won; Kang, In-Cheol

    2009-01-01

    Receptor tyrosine kinases (PTKs) play key roles in the pathogenesis of numerous human diseases, including cancer. Therefore PTK inhibitors are currently under intensive investigation as potential drug candidates. Herein, we report on a ProteoChip-based screening of an epidermal growth factor receptor (EGFR) tyrosine kinase (TK) inhibitor, Erkitinibs, from phytochemical libraries. PLC-γ-1 was used as a substrate immobilized on a ProteoChip and incubated with an EGFR kinase to phosphorylate tyrosine residues of the substrate, followed by a fluorescence detection of the substrate recognized by a phospho-specific monoclonal antibody. Erkitinibs inhibited HeLa cell proliferation in a dose-dependent manner. In conclusion, these data suggest that Erkitinibs can be a specific inhibitor of an EGFR kinase and can be further developed as a potent anti-tumor agent.

  4. Quantitative Tyrosine Phosphoproteomics of Epidermal Growth Factor Receptor (EGFR) Tyrosine Kinase Inhibitor-treated Lung Adenocarcinoma Cells Reveals Potential Novel Biomarkers of Therapeutic Response.

    Science.gov (United States)

    Zhang, Xu; Maity, Tapan; Kashyap, Manoj K; Bansal, Mukesh; Venugopalan, Abhilash; Singh, Sahib; Awasthi, Shivangi; Marimuthu, Arivusudar; Charles Jacob, Harrys Kishore; Belkina, Natalya; Pitts, Stephanie; Cultraro, Constance M; Gao, Shaojian; Kirkali, Guldal; Biswas, Romi; Chaerkady, Raghothama; Califano, Andrea; Pandey, Akhilesh; Guha, Udayan

    2017-05-01

    Mutations in the Epidermal growth factor receptor (EGFR) kinase domain, such as the L858R missense mutation and deletions spanning the conserved sequence 747 LREA 750 , are sensitive to tyrosine kinase inhibitors (TKIs). The gatekeeper site residue mutation, T790M accounts for around 60% of acquired resistance to EGFR TKIs. The first generation EGFR TKIs, erlotinib and gefitinib, and the second generation inhibitor, afatinib are FDA approved for initial treatment of EGFR mutated lung adenocarcinoma. The predominant biomarker of EGFR TKI responsiveness is the presence of EGFR TKI-sensitizing mutations. However, 30-40% of patients with EGFR mutations exhibit primary resistance to these TKIs, underscoring the unmet need of identifying additional biomarkers of treatment response. Here, we sought to characterize the dynamics of tyrosine phosphorylation upon EGFR TKI treatment of mutant EGFR-driven human lung adenocarcinoma cell lines with varying sensitivity to EGFR TKIs, erlotinib and afatinib. We employed stable isotope labeling with amino acids in cell culture (SILAC)-based quantitative mass spectrometry to identify and quantify tyrosine phosphorylated peptides. The proportion of tyrosine phosphorylated sites that had reduced phosphorylation upon erlotinib or afatinib treatment correlated with the degree of TKI-sensitivity. Afatinib, an irreversible EGFR TKI, more effectively inhibited tyrosine phosphorylation of a majority of the substrates. The phosphosites with phosphorylation SILAC ratios that correlated with the TKI-sensitivity of the cell lines include sites on kinases, such as EGFR-Y1197 and MAPK7-Y221, and adaptor proteins, such as SHC1-Y349/350, ERRFI1-Y394, GAB1-Y689, STAT5A-Y694, DLG3-Y705, and DAPP1-Y139, suggesting these are potential biomarkers of TKI sensitivity. DAPP1, is a novel target of mutant EGFR signaling and Y-139 is the major site of DAPP1 tyrosine phosphorylation. We also uncovered several off-target effects of these TKIs, such as MST1R-Y1238

  5. Discovery of novel EGFR tyrosine kinase inhibitors by structure-based virtual screening.

    Science.gov (United States)

    Li, Siyuan; Sun, Xianqiang; Zhao, Hongli; Tang, Yun; Lan, Minbo

    2012-06-15

    By using of structure-based virtual screening, 13 novel epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors were discovered from 197,116 compounds in the SPECS database here. Among them, 8 compounds significantly inhibited EGFR kinase activity with IC(50) values lower than 10 μM. 3-{[1-(3-Chloro-4-fluorophenyl)-3,5-dioxo-4-pyrazolidinylidene]methyl}phenyl 2-thiophenecarboxylate (13), particularly, was the most potent inhibitor possessing the IC(50) value of 3.5 μM. The docking studies also provide some useful information that the docking models of the 13 compounds are beneficial to find a new path for designing novel EGFR inhibitors. Copyright © 2012 Elsevier Ltd. All rights reserved.

  6. The Use of EGFR Tyrosine Kinase Inhibitors in EGFR Wild-Type Non-Small-Cell Lung Cancer.

    Science.gov (United States)

    Stinchcombe, Thomas E

    2016-04-01

    The objective response rate and progression-free survival observed with epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs) in patients with metastatic epidermal growth factor receptor (EGFR) wild-type non-small cell lung cancer (NSCLC) are modest. The adverse events associated with EGFR TKIs are manageable but they must be considered in the context of the limited efficacy. The development of anti-PD-1 immunotherapy as second-line therapy has reduced the role of EGFR TKIs in EGFR wild-type NSCLC. Recently, there has been increased recognition of the benefit of the earlier integration of palliative care and symptom management, and this is reasonable alternative to treatment with an EGFR TKI for many patients. My practice pattern for patients with EGFR wild-type NSCLC is platinum-based chemotherapy as first-line therapy, immunotherapy as second-line therapy, and single-agent chemotherapy as third-line therapy for patients with preserved performance status who want to pursue further therapy. Only a small proportion of patients are eligible for fourth-line therapy, and I prefer to enroll them in clinical trials rather than use EGFR TKIs. I suspect that the use of EGFR TKIs in clinical use and as a comparator arm for clinical trials will continue to decline over the next several years.

  7. Activity of EGFR-tyrosine kinase and ALK inhibitors for EML4–ALK-rearranged non–small–cell lung cancer harbored coexisting EGFR mutation

    International Nuclear Information System (INIS)

    Miyanaga, Akihiko; Kawamoto, Masashi; Tsuchiya, Shinichi; Hagiwara, Koichi; Soda, Manabu; Takeuchi, Kengo; Yamamoto, Nobuyuki; Mano, Hiroyuki; Ishikawa, Yuichi; Gemma, Akihiko; Shimizu, Kumi; Noro, Rintaro; Seike, Masahiro; Kitamura, Kazuhiro; Kosaihira, Seiji; Minegishi, Yuji; Shukuya, Takehito; Yoshimura, Akinobu

    2013-01-01

    The EML4–ALK (echinoderm microtubule-associated protein-like 4 gene and the anaplastic lymphoma kinase gene) fusion oncogene represents a novel molecular target in a small subset of non–small–cell lung cancers (NSCLCs). The EML4–ALK fusion gene occurs generally in NSCLC without mutations in epidermal growth factor receptor (EGFR) and KRAS. We report that a case of EML4–ALK-positive NSCLC with EGFR mutation had a response of stable disease to both an EGFR tyrosine kinase inhibitor (EGFR-TKI) and ALK inhibitor. We described the first clinical report of a patient with EML4–ALK-positive NSCLC with EGFR mutation that had a response of stable disease to both single-agent EGFR-TKI and ALK inhibitor. EML4–ALK translocation may be associated with resistance to EGFR-TKI, and EGFR signaling may contribute to resistance to ALK inhibitor in EML4–ALK-positive NSCLC

  8. Development of a PET tracer for imaging EGFR tyrosine kinase: evaluation of the suitability of PKI166

    International Nuclear Information System (INIS)

    Kernchen, R.; Brust, P.; Krause, M.; Baumann, M.

    2002-01-01

    The suitability of PKI166 for the development of a PET tracer for imaging EGFR tyrosine kinase was investigated. Binding studies using EGFR positive tumour tissue and tritiated PKI166 as the radioligand indicated a low binding affinity of PKI166 to the target tissue. PKI166 is therefore not recommended for PET tracer development. (orig.)

  9. Small tyrosine kinase inhibitors interrupt EGFR signaling by interacting with erbB3 and erbB4 in glioblastoma cell lines

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    Carrasco-Garcia, Estefania; Saceda, Miguel [Instituto de Biologia Molecular y Celular, Universidad Miguel Hernandez, 03202 Elche (Alicante) (Spain); Unidad de Investigacion, Hospital General Universitario de Elche, 03203 Elche (Alicante) (Spain); Grasso, Silvina; Rocamora-Reverte, Lourdes; Conde, Mariano; Gomez-Martinez, Angeles [Instituto de Biologia Molecular y Celular, Universidad Miguel Hernandez, 03202 Elche (Alicante) (Spain); Garcia-Morales, Pilar [Instituto de Biologia Molecular y Celular, Universidad Miguel Hernandez, 03202 Elche (Alicante) (Spain); Unidad de Investigacion, Hospital General Universitario de Elche, 03203 Elche (Alicante) (Spain); Ferragut, Jose A. [Instituto de Biologia Molecular y Celular, Universidad Miguel Hernandez, 03202 Elche (Alicante) (Spain); Martinez-Lacaci, Isabel, E-mail: imlacaci@umh.es [Instituto de Biologia Molecular y Celular, Universidad Miguel Hernandez, 03202 Elche (Alicante) (Spain); Unidad AECC de Investigacion Traslacional en Cancer, Hospital Universitario Virgen de la Arrixaca, 30120 Murcia (Spain)

    2011-06-10

    Signaling through the epidermal growth factor receptor (EGFR) is relevant in glioblastoma. We have determined the effects of the EGFR inhibitor AG1478 in glioblastoma cell lines and found that U87 and LN-229 cells were very sensitive to this drug, since their proliferation diminished and underwent a marked G{sub 1} arrest. T98 cells were a little more refractory to growth inhibition and A172 cells did not undergo a G{sub 1} arrest. This G{sub 1} arrest was associated with up-regulation of p27{sup kip1}, whose protein turnover was stabilized. EGFR autophosphorylation was blocked with AG1478 to the same extent in all the cell lines. Other small-molecule EGFR tyrosine kinase inhibitors employed in the clinic, such as gefitinib, erlotinib and lapatinib, were able to abrogate proliferation of glioblastoma cell lines, which underwent a G{sub 1} arrest. However, the EGFR monoclonal antibody, cetuximab had no effect on cell proliferation and consistently, had no effect on cell cycle either. Similarly, cetuximab did not inhibit proliferation of U87 {Delta}EGFR cells or primary glioblastoma cell cultures, whereas small-molecule EGFR inhibitors did. Activity of downstream signaling molecules of EGFR such as Akt and especially ERK1/2 was interrupted with EGFR tyrosine kinase inhibitors, whereas cetuximab treatment could not sustain this blockade over time. Small-molecule EGFR inhibitors were able to prevent phosphorylation of erbB3 and erbB4, whereas cetuximab only hindered EGFR phosphorylation, suggesting that EGFR tyrosine kinase inhibitors may mediate their anti-proliferative effects through other erbB family members. We can conclude that small-molecule EGFR inhibitors may be a therapeutic approach for the treatment of glioblastoma patients.

  10. Anti-tumor activity of high-dose EGFR tyrosine kinase inhibitor and sequential docetaxel in wild type EGFR non-small cell lung cancer cell nude mouse xenografts

    OpenAIRE

    Tang, Ning; Zhang, Qianqian; Fang, Shu; Han, Xiao; Wang, Zhehai

    2016-01-01

    Treatment of non-small-cell lung cancer (NSCLC) with wild-type epidermal growth factor receptor (EGFR) is still a challenge. This study explored antitumor activity of high-dose icotinib (an EGFR tyrosine kinase inhibitor) plus sequential docetaxel against wild-type EGFR NSCLC cells-generated nude mouse xenografts. Nude mice were subcutaneously injected with wild-type EGFR NSCLC A549 cells and divided into different groups for 3-week treatment. Tumor xenograft volumes were monitored and record...

  11. Icotinib (BPI-2009H), a novel EGFR tyrosine kinase inhibitor, displays potent efficacy in preclinical studies.

    Science.gov (United States)

    Tan, Fenlai; Shen, Xiaoyan; Wang, Dechang; Xie, Guojian; Zhang, Xiaodong; Ding, Lieming; Hu, Yunyan; He, Wei; Wang, Yanping; Wang, Yinxiang

    2012-05-01

    Icotinib, one of the leading compounds selected from our compound library, was found to be a potent and specific epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) with an IC(50) of 5 nM. When profiled with 88 kinases, Icotinib only showed meaningful inhibitory activity to EGFR and its mutants. Icotinib blocked EGFR-mediated intracellular tyrosine phosphorylation (IC(50)=45 nM) in the human epidermoid carcinoma A431 cell line and inhibits tumor cell proliferation. In vivo studies demonstrated that Icotinib exhibited potent dose-dependent antitumor effects in nude mice carrying a variety of human tumor-derived xenografts. The drug was well tolerated at doses up to 120 mg/kg/day in mice without mortality or significant body weight loss during the treatment. A head to head randomized, double blind phase III trial using Gefitinib as an active control for patients with advanced non-small cell lung cancer (NSCLC) was finished recently (Trial registration ID: NCT01040780). The data shows that Icotinib was non-inferior to Gefitinib in terms of median progression free survival (PFS) and safety superior favor to Icotinib compared to Gefitinib. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  12. Antibacterial and EGFR-Tyrosine Kinase Inhibitory Activities of Polyhydroxylated Xanthones from Garcinia succifolia

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    Susawat Duangsrisai

    2014-11-01

    Full Text Available Chemical investigation of the methanol extract of the wood of Garcinia succifolia Kurz (Clusiaceae led to the isolation of 1,5-dihydroxyxanthone (1, 1,7-dihydroxyxanthone (2, 1,3,7-trihydroxyxanthone (3, 1,5,6-trihydroxyxanthone (4, 1,6,7-trihydroxyxanthone (5, and 1,3,6,7-tetrahydroxyxanthone (6. All of the isolated xanthones were evaluated for their antibacterial activity against bacterial reference strains, two Gram-positive (Staphylococcus aureus ATTC 25923, Bacillus subtillis ATCC 6633 and two Gram-negative (Escherichia coli ATCC 25922 and Pseudomonas aeruginosa ATCC 27853, and environmental drug-resistant isolates (S. aureus B1, Enteroccoccus faecalis W1, and E. coli G1, as well as for their epidermal growth factor receptor (EGFR of tyrosine kinase inhibitory activity. Only 1,5,6-trihydroxy-(4, 1,6,7-trihydroxy-(5, and 1,3,6,7-tetrahydroxyxanthones (6 exhibited antibacterial activity against Gram-positive bacteria, however none was active against vancomycin-resistant E. faecalis. Additionally, 1,7-dihydroxyxanthone (2 showed synergism with oxacillin, but not with ampicillin. On the other hand, only 1,5-dihydroxyxanthone (1 and 1,7-dihydroxyxanthone (2 were found to exhibit the EGFR-tyrosine kinase inhibitory activity, with IC50 values of 90.34 and 223 nM, respectively.

  13. Sequential treatment of tyrosine kinase inhibitors and chemotherapy for EGFR-mutated non-small cell lung cancer: a meta-analysis of Phase III trials

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    Zhang Y

    2013-11-01

    Full Text Available Yiliang Zhang,1,* Yihua Sun,1,* Lei Wang,1 Ting Ye,1 Yunjian Pan,1 Haichuan Hu,1 Yongfu Yu,2 Naiqing Zhao,2 Yanyan Song,3 David Garfield,4 Haiquan Chen1 1Department of Thoracic Surgery, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, 2Department of Biostatistics, School of Public Health, Fudan University, 3Department of Pharmacology and Biostatistics, Institute of Medical Science, Shanghai Jiaotong University School of Medicine, 4ProMed Cancer Centers, Shanghai, People’s Republic of China *These authors contributed equally to this work Background: This aim of this study was to compare the efficacy of first-line tyrosine kinase inhibitor therapy followed, upon progression, by chemotherapy with the reverse sequence in patients with EGFR-mutated non-small cell lung cancer (NSCLC in terms of overall survival. Methods: We performed a meta-analysis of studies that met the following criteria: Phase III clinical trial comparing the sequencing of epidermal growth factor receptor (EGFR tyrosine kinase inhibitors with chemotherapy in the treatment of advanced EGFR-mutated NSCLC; activating mutations reported; and availability of hazard ratio estimates with 95% confidence intervals (CIs for overall survival. Results: Six clinical trials were included in this study. The pooled hazard ratio for overall survival of the EGFR-mutated population that completed sequential treatment was 1.03 (95% CI 0.86–1.22, P=0.776. There was no statistically significant heterogeneity between the studies (tau2 =0; I2=0, 95% CI 0–0.37, P=0.548. Evidence of marked publication bias for the two treatment sequences was insufficient (P=0.145. Conclusion: In patients with advanced NSCLC and activating EGFR mutations, first-line chemotherapy followed upon progression by a tyrosine kinase inhibitor was not inferior in terms of overall survival compared with the inverse sequence. This may serve as an indication that

  14. EGFR tyrosine kinase inhibitory peptide attenuates Helicobacter pylori-mediated hyper-proliferation in AGS enteric epithelial cells

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    Himaya, S.W.A. [Marine Bio-Process Research Center, Pukyong National University, Nam-Gu, Busan, 608-737 (Korea, Republic of); Dewapriya, Pradeep [Department of Chemistry, Pukyong National University, Nam-Gu, Busan, 608-737 (Korea, Republic of); Kim, Se-Kwon, E-mail: sknkim@pknu.ac.kr [Marine Bio-Process Research Center, Pukyong National University, Nam-Gu, Busan, 608-737 (Korea, Republic of); Department of Chemistry, Pukyong National University, Nam-Gu, Busan, 608-737 (Korea, Republic of)

    2013-06-15

    Helicobacter pylori infection is one of the most critical causes of stomach cancer. The current study was conducted to explore the protective effects of an isolated active peptide H-P-6 (Pro-Gln-Pro-Lys-Val-Leu-Asp-Ser) from microbial hydrolysates of Chlamydomonas sp. against H. pylori-induced carcinogenesis. The peptide H-P-6 has effectively suppressed H. pylori-induced hyper-proliferation and migration of gastric epithelial cells (AGS). However, the peptide did not inhibit the viability of the bacteria or invasion into AGS cells. Therefore, the effect of the peptide on regulating H. pylori-induced molecular signaling was investigated. The results indicated that H. pylori activates the EGFR tyrosine kinase signaling and nuclear translocation of the β-catenin. The EGFR activation has led to the up-regulation of PI3K/Akt signaling pathway. Moreover, the nuclear translocation levels of β-catenin were significantly increased as a result of Akt mediated down-regulation of GSK3/β protein levels in the cytoplasm. Both of these consequences have resulted in increased expression of cell survival and migration related genes such as c-Myc, cyclin-D, MMP-2 and matrilysin. Interestingly, the isolated peptide potently inhibited H. pylori-mediated EGFR activation and thereby down-regulated the subsequent P13K/Akt signaling leading to β-catenin nuclear translocation. The effect of the peptide was confirmed with the use of EGFR tyrosine kinase inhibitor AG1487 and molecular docking studies. Collectively this study identifies a potent peptide which regulates the H. pylori-induced hyper-proliferation and migration of AGS cells at molecular level. - Highlights: • Chlamydomonas sp. derived peptide H-P-6 inhibits H. pylori-induced pathogenesis. • H-P-6 suppresses H. pylori-induced hyper-proliferation and migration of AGS cells. • The peptide inhibits H. pylori-induced EGFR activation.

  15. Loss of activating EGFR mutant gene contributes to acquired resistance to EGFR tyrosine kinase inhibitors in lung cancer cells.

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    Keisuke Tabara

    Full Text Available Non-small-cell lung cancer harboring epidermal growth factor receptor (EGFR mutations attains a meaningful response to EGFR-tyrosine kinase inhibitors (TKIs. However, acquired resistance to EGFR-TKIs could affect long-term outcome in almost all patients. To identify the potential mechanisms of resistance, we established cell lines resistant to EGFR-TKIs from the human lung cancer cell lines PC9 and11-18, which harbored activating EGFR mutations. One erlotinib-resistant cell line from PC9 and two erlotinib-resistant cell lines and two gefitinib-resistant cell lines from 11-18 were independently established. Almost complete loss of mutant delE746-A750 EGFR gene was observed in the erlotinib-resistant cells isolated from PC9, and partial loss of the mutant L858R EGFR gene copy was specifically observed in the erlotinib- and gefitinib-resistant cells from 11-18. However, constitutive activation of EGFR downstream signaling, PI3K/Akt, was observed even after loss of the mutated EGFR gene in all resistant cell lines even in the presence of the drug. In the erlotinib-resistant cells from PC9, constitutive PI3K/Akt activation was effectively inhibited by lapatinib (a dual TKI of EGFR and HER2 or BIBW2992 (pan-TKI of EGFR family proteins. Furthermore, erlotinib with either HER2 or HER3 knockdown by their cognate siRNAs also inhibited PI3K/Akt activation. Transfection of activating mutant EGFR complementary DNA restored drug sensitivity in the erlotinib-resistant cell line. Our study indicates that loss of addiction to mutant EGFR resulted in gain of addiction to both HER2/HER3 and PI3K/Akt signaling to acquire EGFR-TKI resistance.

  16. Radiotherapy of non-small-cell lung cancer in the era of EGFR gene mutations and EGF receptor tyrosine kinase inhibitors.

    Science.gov (United States)

    Moschini, Ilaria; Dell'Anna, Cristina; Losardo, Pier Luigi; Bordi, Paola; D'Abbiero, Nunziata; Tiseo, Marcello

    2015-01-01

    Non-small-cell lung cancer (NSCLC) occurs, approximately, in 80-85% of all cases of lung cancer. The majority of patients present locally advanced or metastatic disease when diagnosed, with poor prognosis. The discovery of activating mutations in the EGFR gene has started a new era of personalized treatment for NSCLC patients. To improve the treatment outcome in patients with unresectable NSCLC and, in particular, EGFR mutated, a combined strategy of radiotherapy and medical treatment can be undertaken. In this review we will discuss preclinical data regarding EGF receptor (EGFR) tyrosine kinase inhibitors (TKIs) and radiotherapy, available clinical trials investigating efficacy and toxicity of combined treatment (thoracic or whole brain radiotherapy and EGFR-TKIs) and, also, the role of local radiation in mutated EGFR patients who developed EGFR-TKI resistance.

  17. Epidermal growth factor receptor tyrosine kinase (EGFR-TK) mutation testing in adults with locally advanced or metastatic non-small cell lung cancer: A systematic review and cost-effectiveness analysis

    NARCIS (Netherlands)

    M. Westwood (Marie); M.A. Joore (Manuela); P. Whiting (Penny); T. van Asselt (Thea); B.L.T. Ramaekers (Bram); N. Armstrong (Nigel); K. Misso (Kate); J.L. Severens (Hans); J. Kleijnen (Jos)

    2014-01-01

    markdownabstract__Abstract__ Background: Non-small cell lung cancer (NSCLC) is the most common form of lung cancer. Some epidermal growth factor receptor tyrosine kinase (EGFR-TK) mutations make tumours responsive to treatment with EGFR-TK inhibitors (EGFR-TKIs) but less responsive to treatment

  18. Epidermal growth factor receptor tyrosine kinase (EGFR-TK) mutation testing in adults with locally advanced or metastatic non-small cell lung cancer : a systematic review and cost-effectiveness analysis

    NARCIS (Netherlands)

    Westwood, Marie; Joore, Manuela; Whiting, Penny; van Asselt, Thea; Ramaekers, Bram; Armstrong, Nigel; Misso, Kate; Severens, Johan; Kleijnen, Jos

    BACKGROUND: Non-small cell lung cancer (NSCLC) is the most common form of lung cancer. Some epidermal growth factor receptor tyrosine kinase (EGFR-TK) mutations make tumours responsive to treatment with EGFR-TK inhibitors (EGFR-TKIs) but less responsive to treatment with standard chemotherapy.

  19. Mechanisms of acquired resistance to EGFR-tyrosine kinase inhibitor in Korean patients with lung cancer

    International Nuclear Information System (INIS)

    Ji, Wonjun; Lee, Dae Ho; Lee, Jae Cheol; Choi, Chang-Min; Rho, Jin Kyung; Jang, Se Jin; Park, Young Soo; Chun, Sung-Min; Kim, Woo Sung; Lee, Jung-Shin; Kim, Sang-We

    2013-01-01

    Despite an initial good response to epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor (TKI), resistance to treatment eventually develops. Although several resistance mechanisms have been discovered, little data exist regarding Asian patient populations. Among patients at a tertiary referral hospital in Korea who initially responded well to gefitinib and later acquired resistance to treatment, we selected those with enough tissues obtained before EGFR-TKI treatment and after the onset of resistance to examine mutations by mass spectrometric genotyping technology (Asan-Panel), MET amplification by fluorescence in situ hybridization (FISH), and analysis of AXL status, epithelial-to-mesenchymal transition (EMT) and neuroendocrine markers by immunohistochemistry. Twenty-six patients were enrolled, all of whom were diagnosed with adenocarcinoma with EGFR mutations (19del: 16, L858R: 10) except one (squamous cell carcinoma with 19del). Secondary T790M mutation was detected in 11 subjects (42.3%) and four of these patients had other co-existing resistance mechanisms; increased AXL expression was observed in 5/26 patients (19.2%), MET gene amplification was noted in 3/26 (11.5%), and one patient acquired a mutation in the phosphatidylinositol-4, 5-bisphosphate 3-kinase catalytic subunit alpha isoform (PIK3CA) gene. None of the patients exhibited EMT; however, increased CD56 expression suggesting neuroendocrine differentiation was observed in two patients. Interestingly, conversion from L858R-mutant to wild-type EGFR occurred in one patient. Seven patients (26.9%) did not exhibit any known resistance mechanisms. Patients with a T790M mutation showed a more favorable prognosis. The mechanisms and frequency of acquired EGFR-TKI resistance in Koreans are comparable to those observed in Western populations; however, more data regarding the mechanisms that drive EGFR-TKI resistance are necessary

  20. Convergent Akt activation drives acquired EGFR inhibitor resistance in lung cancer

    DEFF Research Database (Denmark)

    Jacobsen, Kirstine; Bertran-Alamillo, Jordi; Molina, Miguel Angel

    2017-01-01

    Non-small-cell lung cancer patients with activating epidermal growth factor receptor (EGFR) mutations typically benefit from EGFR tyrosine kinase inhibitor treatment. However, virtually all patients succumb to acquired EGFR tyrosine kinase inhibitor resistance that occurs via diverse mechanisms....... The diversity and unpredictability of EGFR tyrosine kinase inhibitor resistance mechanisms presents a challenge for developing new treatments to overcome EGFR tyrosine kinase inhibitor resistance. Here, we show that Akt activation is a convergent feature of acquired EGFR tyrosine kinase inhibitor resistance......, across a spectrum of diverse, established upstream resistance mechanisms. Combined treatment with an EGFR tyrosine kinase inhibitor and Akt inhibitor causes apoptosis and synergistic growth inhibition in multiple EGFR tyrosine kinase inhibitor-resistant non-small-cell lung cancer models. Moreover...

  1. Survey of tyrosine kinase signaling reveals ROS kinase fusions in human cholangiocarcinoma.

    Directory of Open Access Journals (Sweden)

    Ting-Lei Gu

    Full Text Available Cholangiocarcinoma, also known as bile duct cancer, is the second most common primary hepatic carcinoma with a median survival of less than 2 years. The molecular mechanisms underlying the development of this disease are not clear. To survey activated tyrosine kinases signaling in cholangiocarcinoma, we employed immunoaffinity profiling coupled to mass spectrometry and identified DDR1, EPHA2, EGFR, and ROS tyrosine kinases, along with over 1,000 tyrosine phosphorylation sites from about 750 different proteins in primary cholangiocarcinoma patients. Furthermore, we confirmed the presence of ROS kinase fusions in 8.7% (2 out of 23 of cholangiocarcinoma patients. Expression of the ROS fusions in 3T3 cells confers transforming ability both in vitro and in vivo, and is responsive to its kinase inhibitor. Our data demonstrate that ROS kinase is a promising candidate for a therapeutic target and for a diagnostic molecular marker in cholangiocarcinoma. The identification of ROS tyrosine kinase fusions in cholangiocarcinoma, along with the presence of other ROS kinase fusions in lung cancer and glioblastoma, suggests that a more broadly based screen for activated ROS kinase in cancer is warranted.

  2. Combined effects of EGFR tyrosine kinase inhibitors and vATPase inhibitors in NSCLC cells

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    Jin, Hyeon-Ok [KIRAMS Radiation Biobank, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Hong, Sung-Eun [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Kim, Chang Soon [Department of Microbiological Engineering, Kon-Kuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul, 143–701 (Korea, Republic of); Park, Jin-Ah; Kim, Jin-Hee; Kim, Ji-Young; Kim, Bora [KIRAMS Radiation Biobank, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Chang, Yoon Hwan; Hong, Seok-Il; Hong, Young Jun [Department of Laboratory Medicine, Korea Cancer Center Hospital, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Park, In-Chul, E-mail: parkic@kirams.re.kr [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Lee, Jin Kyung, E-mail: jklee@kirams.re.kr [KIRAMS Radiation Biobank, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of); Department of Laboratory Medicine, Korea Cancer Center Hospital, Korea Institute of Radiological and Medical Sciences, 75 Nowon-ro, Nowon-gu, Seoul, 139–706 (Korea, Republic of)

    2015-08-15

    Despite excellent initial clinical responses of non-small cell lung cancer (NSCLC) patients to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs), many patients eventually develop resistance. According to a recent report, vacuolar H + ATPase (vATPase) is overexpressed and is associated with chemotherapy drug resistance in NSCLC. We investigated the combined effects of EGFR TKIs and vATPase inhibitors and their underlying mechanisms in the regulation of NSCLC cell death. We found that combined treatment with EGFR TKIs (erlotinib, gefitinib, or lapatinib) and vATPase inhibitors (bafilomycin A1 or concanamycin A) enhanced synergistic cell death compared to treatments with each drug alone. Treatment with bafilomycin A1 or concanamycin A led to the induction of Bnip3 expression in an Hif-1α dependent manner. Knock-down of Hif-1α or Bnip3 by siRNA further enhanced cell death induced by bafilomycin A1, suggesting that Hif-1α/Bnip3 induction promoted resistance to cell death induced by the vATPase inhibitors. EGFR TKIs suppressed Hif-1α and Bnip3 expression induced by the vATPase inhibitors, suggesting that they enhanced the sensitivity of the cells to these inhibitors by decreasing Hif-1α/Bnip3 expression. Taken together, we conclude that EGFR TKIs enhance the sensitivity of NSCLC cells to vATPase inhibitors by decreasing Hif-1α/Bnip3 expression. We suggest that combined treatment with EGFR TKIs and vATPase inhibitors is potentially effective for the treatment of NSCLC. - Highlights: • Co-treatment with EGFR TKIs and vATPase inhibitors induces synergistic cell death • EGFR TKIs enhance cell sensitivity to vATPase inhibitors via Hif-1α downregulation • Co-treatment of these inhibitors is potentially effective for the treatment of NSCLC.

  3. Anti-tumor activity of high-dose EGFR tyrosine kinase inhibitor and sequential docetaxel in wild type EGFR non-small cell lung cancer cell nude mouse xenografts

    Science.gov (United States)

    Tang, Ning; Zhang, Qianqian; Fang, Shu; Han, Xiao; Wang, Zhehai

    2017-01-01

    Treatment of non-small-cell lung cancer (NSCLC) with wild-type epidermal growth factor receptor (EGFR) is still a challenge. This study explored antitumor activity of high-dose icotinib (an EGFR tyrosine kinase inhibitor) plus sequential docetaxel against wild-type EGFR NSCLC cells-generated nude mouse xenografts. Nude mice were subcutaneously injected with wild-type EGFR NSCLC A549 cells and divided into different groups for 3-week treatment. Tumor xenograft volumes were monitored and recorded, and at the end of experiments, tumor xenografts were removed for Western blot and immunohistochemical analyses. Compared to control groups (negative control, regular-dose icotinib [IcoR], high-dose icotinib [IcoH], and docetaxel [DTX]) and regular icotinib dose (60 mg/kg) with docetaxel, treatment of mice with a high-dose (1200 mg/kg) of icotinib plus sequential docetaxel for 3 weeks (IcoH-DTX) had an additive effect on suppression of tumor xenograft size and volume (P Icotinib-containing treatments markedly reduced phosphorylation of EGFR, mitogen activated protein kinase (MAPK), and protein kinase B (Akt), but only the high-dose icotinib-containing treatments showed an additive effect on CD34 inhibition (P icotinib plus docetaxel had a similar effect on mouse weight loss (a common way to measure adverse reactions in mice), compared to the other treatment combinations. The study indicate that the high dose of icotinib plus sequential docetaxel (IcoH-DTX) have an additive effect on suppressing the growth of wild-type EGFR NSCLC cell nude mouse xenografts, possibly through microvessel density reduction. Future clinical trials are needed to confirm the findings of this study. PMID:27852073

  4. Effects of icotinib, a novel epidermal growth factor receptor tyrosine kinase inhibitor, in EGFR-mutated non-small cell lung cancer.

    Science.gov (United States)

    Yang, Guangdie; Yao, Yinan; Zhou, Jianya; Zhao, Qiong

    2012-06-01

    Epidermal growth factor receptor (EGFR) is one of the most promising targets for non-small cell lung cancer (NSCLC). Our study demonstrated the antitumor effects of icotinib hydrochloride, a highly selective epidermal growth factor receptor tyrosine kinase inhibitor (EGFR TKI), in two EGFR-mutated lung cancer cell lines compared to A549, a cell line without EGFR mutations. We incubated PC-9 and HCC827 human lung cancer cell lines both with (E746-A750) mutations with various concentrations of icotinib and gefitinib for 48 h. Cell proliferation and migration were determined using a real-time cell invasion and migration assay and cytotoxicity assay. Apoptosis was assessed by measuring Annexin V staining using flow cytometry. The antitumor effects of icotinib compared to gefitinib were similar and were most effective in reducing the proliferation of EGFR-mutated cells compared to non-mutated controls. Our results suggest the possibility of icotinib as a new therapeutic agent of EGFR-mutated cancer cells, which has the potential to be used in the first-line treatment of EGFR-mutated NSCLC.

  5. Epidermal growth factor receptor tyrosine kinase (EGFR-TK) mutation testing in adults with locally advanced or metastatic non-small cell lung cancer: A systematic review and cost-effectiveness analysis

    OpenAIRE

    Westwood, Marie; Joore, Manuela; Whiting, Penny; Asselt, Thea; Ramaekers, Bram; Armstrong, Nigel; Misso, Kate; Severens, Hans; Kleijnen, Jos

    2014-01-01

    markdownabstract__Abstract__ Background: Non-small cell lung cancer (NSCLC) is the most common form of lung cancer. Some epidermal growth factor receptor tyrosine kinase (EGFR-TK) mutations make tumours responsive to treatment with EGFR-TK inhibitors (EGFR-TKIs) but less responsive to treatment with standard chemotherapy. Patients with NSCLC are therefore tested for EGFR-TK tumour gene mutations to inform treatment decisions. There are a variety of tests available to detect these mutations. T...

  6. Tyrosine dephosphorylation enhances the therapeutic target activity of epidermal growth factor receptor (EGFR) by disrupting its interaction with estrogen receptor (ER).

    Science.gov (United States)

    Ma, Shao; Yin, Ning; Qi, Xiaomei; Pfister, Sandra L; Zhang, Mei-Jie; Ma, Rong; Chen, Guan

    2015-05-30

    Protein-protein interactions can increase or decrease its therapeutic target activity and the determining factors involved, however, are largely unknown. Here, we report that tyrosine-dephosphorylation of epidermal growth factor receptor (EGFR) increases its therapeutic target activity by disrupting its interaction with estrogen receptor (ER). Protein tyrosine phosphatase H1 (PTPH1) dephosphorylates the tyrosine kinase EGFR, disrupts its interaction with the nuclear receptor ER, and increases breast cancer sensitivity to small molecule tyrosine kinase inhibitors (TKIs). These effects require PTPH1 catalytic activity and its interaction with EGFR, suggesting that the phosphatase may increase the sensitivity by dephosphorylating EGFR leading to its dissociation with ER. Consistent with this notion, a nuclear-localization defective ER has a higher EGFR-binding activity and confers the resistance to TKI-induced growth inhibition. Additional analysis show that PTPH1 stabilizes EGFR, stimulates the membranous EGFR accumulation, and enhances the growth-inhibitory activity of a combination therapy of TKIs with an anti-estrogen. Since EGFR and ER both are substrates for PTPH1 in vitro and in intact cells, these results indicate that an inhibitory EGFR-ER protein complex can be switched off through a competitive enzyme-substrate binding. Our results would have important implications for the treatment of breast cancer with targeted therapeutics.

  7. Phase I study of icotinib, an EGFR tyrosine kinase inhibitor combined with IMRT in nasopharyngeal carcinoma.

    Science.gov (United States)

    Hu, Wei; Wang, Wei; Yang, Peinong; Zhou, Chao; Yang, Weifang; Wu, Bo; Lu, Hongsheng; Yang, Haihua

    2015-01-01

    Epidermal growth factor receptor (EGFR) is a new target for nasopharyngeal carcinoma (NPC) therapy. This prospective phase I study sought to determine the safety and recommended phase II dose of icotinib, a novel highly selective oral EGFR tyrosine kinase inhibitor, in combination with intensity-modulated radiotherapy (IMRT) in patients with NPC. Eligible patients with NPC received escalating doses of icotinib during IMRT. We treated six patients at a particular dose level until the maximum tolerated dose (MTD) was determined. The starting dose was 125 mg, once-daily and the dose was escalated to another level 125 mg, twice- and thrice- daily, until dose-limiting toxicity (DLT) occurred in two or more patients at a dose level. Expression and mutation analysis of EGFR were performed in all cases. A total of twelve patients were enrolled. Three patients experienced DLT (250 mg/day cohort) and MTD was 125 mg/day. Mucositis toxicity appears to be the major DLT. While EGFR expression in tumor tissue was detected in 75% (9/12) patients, EGFR mutation was detected in 16.67% (1/6) patients in 125 mg/day cohort, and 50% (3/6) in 250 mg/day cohort. The combination of icotinib (125 mg/day) and IMRT in patients with locally NPC had an acceptable safety profile and was well tolerated.

  8. MLH1 V384D polymorphism associates with poor response to EGFR tyrosine kinase inhibitors in patients with EGFR L858R-positive lung adenocarcinoma.

    Science.gov (United States)

    Chiu, Chao-Hua; Ho, Hsiang-Ling; Doong, Howard; Yeh, Yi-Chen; Chen, Mei-Yu; Chou, Teh-Ying; Tsai, Chun-Ming

    2015-04-10

    A significant fraction of patients with lung adenocarcinomas harboring activating epidermal growth factor receptor (EGFR) mutations do not experience clinical benefits from EGFR tyrosine kinase inhibitor (TKI) therapy. Using next-generation sequencing, we screened 739 mutation hotspots in 46 cancer-related genes in EGFR L858R-mutant lung adenocarcinomas from 29 patients who received EGFR-TKI therapy; 13 had short ( 1 year) progression-free survival (PFS). We discovered MLH1 V384D as a genetic variant enriched in the group of patients with short PFS. Next, we investigated this genetic variation in 158 lung adenocarcinomas with the EGFR L858R mutation and found 14 (8.9%) patients had MLH1 V384D; available blood or non-tumor tissues from patients were also tested positive for MLH1 V384D. Patients with MLH1 V384D had a significantly shorter median PFS than those without (5.1 vs. 10.6 months; P= 0.001). Multivariate analysis showed that MLH1 V384D polymorphism was an independent predictor for a reduced PFS time (hazard ratio, 3.5; 95% confidence interval, 1.7 to 7.2; P= 0.001). In conclusion, MLH1 V384D polymorphism is associated with primary resistance to EGFR-TKIs in patients with EGFR L858R-positive lung adenocarcinoma and may potentially be a novel biomarker to guide treatment decisions.

  9. Dialkoxyquinazolines: Screening Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitors for Potential Tumor Imaging Probes

    International Nuclear Information System (INIS)

    VanBrocklin, Henry F.; Lim, John K.; Coffing, Stephanie L.; Hom, Darren L.; Negash, Kitaw; Ono, Michele Y.; Hanrahan, Stephen M.; Taylor, Scott E.; Vanderpoel, Jennifer L.; Slavik, Sarah M.; Morris, Andrew B.; Riese II, David J.

    2005-01-01

    The epidermal growth factor receptor (EGFR), a long-standing drug development target, is also a desirable target for imaging. Sixteen dialkoxyquinazoline analogs, suitable for labeling with positron-emitting isotopes, have been synthesized and evaluated in a battery of in vitro assays to ascertain their chemical and biological properties. These characteristics provided the basis for the adoption of a selection schema to identify lead molecules for labeling and in vivo evaluation. A newEGFR tyrosine kinase radiometric binding assay revealed that all of the compounds possessed suitable affinity (IC50 = 0.4 - 51 nM) for the EGFR tyrosine kinase. All of the analogs inhibited ligand-induced EGFR tyrosine phosphorylation (IC50 = 0.8 - 20 nM). The HPLC-estimated octanol/water partition coefficients ranged from 2.0-5.5. Four compounds,4-(2'-fluoroanilino)- and 4-(3'-fluoroanilino)-6,7-diethoxyquinazoline as well as 4-(3'-chloroanilino)- and4-(3'-bromoanilino)-6,7-dimethoxyquinazoline, possess the best combination of characteristics that warrant radioisotope labeling and further evaluation in tumor-bearing mice

  10. Second-generation inhibitors of Bruton tyrosine kinase

    Directory of Open Access Journals (Sweden)

    Jingjing Wu

    2016-09-01

    Full Text Available Abstract Bruton tyrosine kinase (BTK is a critical effector molecule for B cell development and plays a major role in lymphoma genesis. Ibrutinib is the first-generation BTK inhibitor. Ibrutinib has off-target effects on EGFR, ITK, and Tec family kinases, which explains the untoward effects of ibrutinib. Resistance to ibrutinib was also reported. The C481S mutation in the BTK kinase domain was reported to be a major mechanism of resistance to ibrutinib. This review summarizes the clinical development of novel BTK inhibitors, ACP-196 (acalabrutinib, ONO/GS-4059, and BGB-3111.

  11. Tyrosine kinase domain mutations of EGFR gene in head and neck squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Vatte C

    2017-03-01

    Full Text Available Chittibabu Vatte,1 Ali M Al Amri,2 Cyril Cyrus,1 Shahanas Chathoth,1 Sadananda Acharya,3 Tariq Mohammad Hashim,4 Zhara Al Ali,2 Saleh Tawfeeq Alshreadah,2 Ahmed Alsayyah,4 Amein K Al-Ali5 1Department of Genetic Research, Institute for Research and Medical Consultation, University of Dammam, Dammam, 2Department of Internal Medicine, King Fahd Hospital of the University, University of Dammam, Al-Khobar, 3Department of Stemcell Research, Institute for Research and Medical Consultation, 4Department of Pathology, King Fahd Hospital of the University, University of Dammam, Al-Khobar, 5Department of Biochemistry, College of Medicine, University of Dammam, Dammam, Kingdom of Saudi Arabia Background: Epidermal growth factor receptor (EGFR is a commonly altered gene that is identified in various cancers, including head and neck squamous cell carcinoma (HNSCC. Therefore, EGFR is a promising molecular marker targeted by monoclonal antibodies and small molecule inhibitors targeting the tyrosine kinase (TK domain. Objective: The objective of this study was to investigate the spectrum of mutations in exons 18, 19, 20, and 21 of the EGFR gene in HNSCC patients. Materials and methods: This retrospective study included 47 confirmed HNSCC cases. Mutations in the TK domain, exons 18, 19, 20, and 21 of the EGFR gene, were detected by Scorpion® chemistry and ARMS® technologies on Rotor-Gene Q real-time polymerase chain reaction.Results: The tumors exhibited EGFR-TK domain mutations in 57% of cases. Four cases of T790M mutations were reported for the first time among HNSCC patients. Out of the total mutations, L861Q (exon 21, exon 20 insertions and deletions of exon 19 accounted for the majority of mutations (21%, 19%, and 17%, respectively. EGFR mutation status was correlated with the higher grade (P=0.026 and advanced stage (P=0.034 of HNSCC tumors.Conclusion: Higher frequency of EGFR-TK domain mutations together with the presence of the T790M mutation suggests

  12. Associations of mRNA:microRNA for the shared downstream molecules of EGFR and alternative tyrosine kinase receptors in Non-Small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Fengfeng Wang

    2016-10-01

    Full Text Available Lung cancer is the top cancer killer worldwide with high mortality rate. Majority belong to non-small cell lung cancers (NSCLCs. The epidermal growth factor receptor (EGFR has been broadly explored as a drug target for therapy. However, the drug responses are not durable due to the acquired resistance. MicroRNAs (miRNAs are small noncoding and endogenous molecules that can inhibit mRNA translation initiation and degrade mRNAs. We wonder if some downstream molecules shared by EGFR and the other tyrosine kinase receptors (TKRs further transduce the signals alternatively, and some miRNAs play the key roles in affecting the expression of these downstream molecules. In this study, we investigated the mRNA:miRNA associations for the direct EGFR downstream molecules in the EGFR signaling pathway shared with the other TKRs, including c-MET (hepatocyte growth factor receptor, Ron (a protein tyrosine kinase related to c-MET, PDGFR (platelet-derived growth factor receptor, and IGF-1R (insulin-like growth factor receptor-1. The multiple linear regression and support vector regression (SVR models were used to discover the statistically significant and the best weighted miRNAs regulating the mRNAs of these downstream molecules. These two models revealed the similar mRNA:miRNA associations. It was found that the miRNAs significantly affecting the mRNA expressions in the multiple regression model were also those with the largest weights in the SVR model. To conclude, we effectively identified a list of meaningful mRNA:miRNA associations: phospholipase C, gamma 1 (PLCG1 with miR-34a, phosphoinositide-3-kinase, regulatory subunit 2 (PIK3R2 with miR-30a-5p, growth factor receptor-bound protein 2 (GRB2 with miR-27a, and Janus kinase 1 (JAK1 with miR-302b and miR-520e. These associations could make great contributions to explore new mechanism in NSCLCs. These candidate miRNAs may be regarded as the potential drug targets for treating NSCLCs with acquired drug

  13. Mutational profiling of non-small-cell lung cancer patients resistant to first-generation EGFR tyrosine kinase inhibitors using next generation sequencing

    Science.gov (United States)

    Jin, Ying; Shao, Yang; Shi, Xun; Lou, Guangyuan; Zhang, Yiping; Wu, Xue; Tong, Xiaoling; Yu, Xinmin

    2016-01-01

    Patients with advanced non-small-cell lung cancer (NSCLC) harboring sensitive epithelial growth factor receptor (EGFR) mutations invariably develop acquired resistance to EGFR tyrosine kinase inhibitors (TKIs). Identification of actionable genetic alterations conferring drug-resistance can be helpful for guiding the subsequent treatment decision. One of the major resistant mechanisms is secondary EGFR-T790M mutation. Other mechanisms, such as HER2 and MET amplifications, and PIK3CA mutations, were also reported. However, the mechanisms in the remaining patients are still unknown. In this study, we performed mutational profiling in a cohort of 83 NSCLC patients with TKI-sensitizing EGFR mutations at diagnosis and acquired resistance to three different first-generation EGFR TKIs using targeted next generation sequencing (NGS) of 416 cancer-related genes. In total, we identified 322 genetic alterations with a median of 3 mutations per patient. 61% of patients still exhibit TKI-sensitizing EGFR mutations, and 36% of patients acquired EGFR-T790M. Besides other known resistance mechanisms, we identified TET2 mutations in 12% of patients. Interestingly, we also observed SOX2 amplification in EGFR-T790M negative patients, which are restricted to Icotinib treatment resistance, a drug widely used in Chinese NSCLC patients. Our study uncovered mutational profiles of NSCLC patients with first-generation EGFR TKIs resistance with potential therapeutic implications. PMID:27528220

  14. Antibody-induced dimerization activates the epidermal growth factor receptor tyrosine kinase

    NARCIS (Netherlands)

    Spaargaren, M.; Defize, L. H.; Boonstra, J.; de Laat, S. W.

    1991-01-01

    The relationship between epidermal growth factor receptor (EGF-R) protein tyrosine kinase activation and ligand-induced receptor dimerization was investigated using several bivalent anti-EGF-R antibodies directed against various receptor epitopes. In A431 membrane preparations and permeabilized

  15. Tyrosine kinase, aurora kinase and leucine aminopeptidase as attractive drug targets in anticancer therapy - characterisation of their inhibitors.

    Science.gov (United States)

    Ziemska, Joanna; Solecka, Jolanta

    Cancers are the leading cause of deaths all over the world. Available anticancer agents used in clinics exhibit low therapeutic index and usually high toxicity. Wide spreading drug resistance of cancer cells induce a demanding need to search for new drug targets. Currently, many on-going studies on novel compounds with potent anticancer activity, high selectivity as well as new modes of action are conducted. In this work, we describe in details three enzyme groups, which are at present of extensive interest to medical researchers and pharmaceutical companies. These include receptor tyrosine kinases (e.g. EGFR enzymes) and non-receptor tyrosine kinases (Src enzymes), type A, B and C Aurora kinases and aminopeptidases, especially leucine aminopeptidase. We discuss classification of these enzymes, biochemistry as well as their role in the cell cycle under normal conditions and during cancerogenesis. Further on, the work describes enzyme inhibitors that are under in vitro, preclinical, clinical studies as well as drugs available on the market. Both, chemical structures of discovered inhibitors and the role of chemical moieties in novel drug design are discussed. Described enzymes play essential role in cell cycle, especially in mitosis (Aurora kinases), cell differentiation, growth and apoptosis (tyrosine kinases) as well as G1/S transition (leucine aminopeptidase). In cancer cells, they are overexpressed and only their inhibition may stop tumor progression. This review presents the clinical outcomes of selected inhibitors and argues the safety of drug usage in human volunteers. Clinical studies of EGFR and Src kinase inhibitors in different tumors clearly show the need for molecular selection of patients (to those with mutations in genes coding EGFR and Src) to achieve positive clinical response. Current data indicates the great necessity for new anticancer treatment and actions to limit off-target activity.

  16. Targeted sequencing identifies genetic alterations that confer primary resistance to EGFR tyrosine kinase inhibitor (Korean Lung Cancer Consortium).

    Science.gov (United States)

    Lim, Sun Min; Kim, Hye Ryun; Cho, Eun Kyung; Min, Young Joo; Ahn, Jin Seok; Ahn, Myung-Ju; Park, Keunchil; Cho, Byoung Chul; Lee, Ji-Hyun; Jeong, Hye Cheol; Kim, Eun Kyung; Kim, Joo-Hang

    2016-06-14

    Non-small-cell lung cancer (NSCLC) patients with activating epidermal growth factor receptor (EGFR) mutations may exhibit primary resistance to EGFR tyrosine kinase inhibitor (TKI). We aimed to examine genomic alterations associated with de novo resistance to gefitinib in a prospective study of NSCLC patients. One-hundred and fifty two patients with activating EGFR mutations were included in this study and 136 patients' tumor sample were available for targeted sequencing of genomic alterations in 22 genes using the Colon and Lung Cancer panel (Ampliseq, Life Technologies). All 132 patients with EGFR mutation were treated with gefitinib for their treatment of advanced NSCLC. Twenty patients showed primary resistance to EGFR TKI, and were classified as non-responders. A total of 543 somatic single-nucleotide variants (498 missense, 13 nonsense) and 32 frameshift insertions/deletions, with a median of 3 mutations per sample. TP53 was most commonly mutated (47%) and mutations in SMAD4 was also common (19%), as well as DDR2 (16%), PIK3CA (15%), STK11 (14%), and BRAF (7%). Genomic mutations in the PI3K/Akt/mTOR pathway were commonly found in non-responders (45%) compared to responders (27%), and they had significantly shorter progression-free survival and overall survival compared to patients without mutations (2.1 vs. 12.8 months, P=0.04, 15.7 vs. not reached, PAkt/mTOR pathway were commonly identified in non-responders and may confer resistance to EGFR TKI. Screening lung adenocarcinoma patients with clinical cancer gene test may aid in selecting out those who show primary resistance to EGFR TKI (NCT01697163).

  17. Convergent Akt activation drives acquired EGFR inhibitor resistance in lung cancer

    DEFF Research Database (Denmark)

    Jacobsen, Kirstine; Bertran-Alamillo, Jordi; Molina, Miguel Angel

    2017-01-01

    Non-small-cell lung cancer patients with activating epidermal growth factor receptor (EGFR) mutations typically benefit from EGFR tyrosine kinase inhibitor treatment. However, virtually all patients succumb to acquired EGFR tyrosine kinase inhibitor resistance that occurs via diverse mechanisms....

  18. Icotinib, a selective EGF receptor tyrosine kinase inhibitor, for the treatment of non-small-cell lung cancer.

    Science.gov (United States)

    Tan, Fenlai; Shi, Yuankai; Wang, Yinxiang; Ding, Lieming; Yuan, Xiaobin; Sun, Yan

    2015-01-01

    Advanced non-small-cell lung cancer (NSCLC) is the main cause for cancer-related mortality. Treatments for advanced NSCLC are largely palliative and a benefit plateau appears to have reached with the platinum-based chemotherapy regimens. EGF receptor (EGFR) tyrosine kinase inhibitors gefitinib, erlotinib and afatinib came up with prolonged progression-free survival and improved quality of life, especially in EGFR-mutated patients. Icotinib is an oral selective EGFR tyrosine kinase, which was approved by China Food and Drug administration in June 2011 for treating advanced NSCLC. Its approval was based on the registered Phase III trial (ICOGEN), which showed icotinib is noninferior to gefitinib. This review will discuss the role of icotinib in NSCLC, and its potential application and ongoing investigations.

  19. EGFR mutation is a better predictor of response to tyrosine kinase inhibitors in non-small cell lung carcinoma than FISH, CISH, and immunohistochemistry.

    Science.gov (United States)

    Sholl, Lynette M; Xiao, Yun; Joshi, Victoria; Yeap, Beow Y; Cioffredi, Leigh-Anne; Jackman, David M; Lee, Charles; Jänne, Pasi A; Lindeman, Neal I

    2010-06-01

    About 10% of patients with non-small cell lung carcinoma (NSCLC) respond to epidermal growth factor receptor (EGFR)-targeted tyrosine kinase inhibitors (TKIs). More than 75% of "responders" have activating mutations in EGFR. However, mutation analysis is not widely available, and proposed alternatives (in situ hybridization and immunohistochemical analysis) have shown inconsistent associations with outcome. Fluorescence in situ hybridization (FISH), chromogenic in situ hybridization (CISH), immunohistochemical analysis, and DNA sequencing were compared in this study of 40 NSCLC samples from TKI-treated patients. Response rates were 12 of 19 in EGFR-mutant vs 1 of 20 EGFR wild-type tumors (P = .0001), 7 of 19 FISH+ vs 4 of 17 FISH- tumors (not significant [NS]), 5 of 16 CISH+ vs 6 of 21 CISH- tumors (NS), and 3 of 9 immunohistochemically positive vs 7 of 22 immunohistochemically negative tumors (NS). EGFR mutation was associated with improved progression-free survival (P = .0004). Increased copy number (FISH or CISH) and protein expression (immunohistochemical) did not independently predict outcome. Thus, EGFR sequence analysis was the only method useful for predicting response and progression-free survival following TKI therapy in NSCLC.

  20. The combi-targeting concept: a novel 3,3-disubstituted nitrosourea with EGFR tyrosine kinase inhibitory properties.

    Science.gov (United States)

    Qiu, Qiyu; Dudouit, Fabienne; Matheson, Stephanie L; Brahimi, Fouad; Banerjee, Ranjita; McNamee, James P; Jean-Claude, Bertrand J

    2003-01-01

    To study the dual mechanism of action of FD137, a 3,3-disubstituted nitrosourea designed to block signaling mediated by the epidermal growth factor receptor (EGFR) on its own and to be hydrolyzed to an inhibitor of EGFR plus a DNA-damaging species. HPLC was used to determine the half-life (t(1/2)) of FD137 and to characterize its derived metabolite FD110. The dual mechanisms of DNA damaging and EGFR tyrosine kinase (TK) targeting were ascertained by the comet assay for DNA damage and by inmunodetection of phosphotyrosine in an ELISA and a whole-cell assay for EGFR-mediated signaling. The antiproliferative effects of the different drugs and their combinations were determined by the sulforhodamine B (SRB) assay. In contrast to BCNU, FD137 significantly blocked EGF-induced EGFR autophosphorylation (IC(50) 4 micro M) in the human solid tumor cell line A431. DNA damage induced by FD137 could only be observed after 24 h exposure, but the level of DNA damage remained 3.6-fold lower than that induced by BCNU. This difference was rationalized by the 160-fold greater stability of FD137 when compared with BCNU in serum-containing medium. Further, degradation of FD137 was accompanied by the slow release of FD110, an extremely potent inhibitor of EGFR TK [IC(50) (EGFR autophosphorylation) <0.3 micro M]. The complex properties of FD137 translated into a 55-fold greater antiproliferative activity than BCNU against the EGFR-overexpressing A431 cells that coexpresses the O(6)-alkylguanine transferase (AGT). Depletion of AGT in these cells by the use of O(6)-benzylguanine (O(6)-BG) enhanced their sensitivity to BCNU by 8-fold, but only by 3-fold to FD137. The results overall suggest that the superior antiproliferative activity of FD137 when compared with BCNU may be associated with its ability to behave as a combination of many species with different mechanisms of action. However, the enhancement of its potency by O(6)-BG suggests that its antiproliferative effect was at least

  1. Tyrosine kinase receptor status in endometrial stromal sarcoma: an immunohistochemical and genetic-molecular analysis.

    Science.gov (United States)

    Cossu-Rocca, Paolo; Contini, Marcella; Uras, Maria Gabriela; Muroni, Maria Rosaria; Pili, Francesca; Carru, Ciriaco; Bosincu, Luisanna; Massarelli, Giovannino; Nogales, Francisco F; De Miglio, Maria Rosaria

    2012-11-01

    Endometrial stromal sarcomas (ESS) are rare uterine malignant mesenchymal neoplasms, which are currently treated by surgery, as effective adjuvant therapies have not yet been established. Tyrosine kinase inhibitors have rarely been applied in ESS therapy, with few reports describing imatinib responsivity. The aim of this study was to analyze the status of different tyrosine kinase receptors in an ESS series, in order to evaluate their potential role as molecular targets. Immunohistochemistry was performed for EGFR, c-KIT, PDGFR-α, PDGFR-β, and ABL on 28 ESS. EGFR, PDGFR-α, and PDGFR-β gene expression was investigated by real-time polymerase chain reaction (qRT-PCR) on selected cases. "Hot-spot" mutations were screened for on EGFR, c-KIT, PDGFR-α, and PDGFR-β genes, by sequencing. All analysis was executed from formalin-fixed, paraffin-embedded specimens. Immunohistochemical overexpression of 2 or more tyrosine kinase receptors was observed in 18 of 28 tumors (64%), whereas only 5 tumors were consistently negative. Gene expression profiles were concordant with immunohistochemical overexpression in only 1 tumor, which displayed both high mRNA levels and specific immunoreactivity for PDGFR-α, and PDGFR-β. No activating mutations were found on the tumors included in the study. This study confirms that TKRs expression is frequently observed in ESS. Considering that the responsiveness to tyrosine kinase inhibitors is known to be related to the presence of specific activating mutations or gene over-expression, which are not detectable in ESS, TKRs immunohistochemical over-expression alone should not be considered as a reliable marker for targeted therapies in ESS. Specific post-translational abnormalities, responsible for activation of TKRs, should be further investigated.

  2. Melanoma-associated antigen expression and the efficacy of tyrosine kinase inhibitors in head and neck cancer

    DEFF Research Database (Denmark)

    Hartmann, Stefan; Brands, Roman C; Küchler, Nora

    2015-01-01

    receptor (EGFR). The efficacy of tyrosine kinase inhibitors (TKI) in the context of melanoma-associated antigens is discussed in the present study. Five human squamous cell carcinoma cell lines were treated with the EGFR TKIs, erlotinib and gefitinib. The efficacy of these agents was measured using...

  3. Interactions between Type III receptor tyrosine phosphatases and growth factor receptor tyrosine kinases regulate tracheal tube formation in Drosophila

    Directory of Open Access Journals (Sweden)

    Mili Jeon

    2012-04-01

    The respiratory (tracheal system of the Drosophila melanogaster larva is an intricate branched network of air-filled tubes. Its developmental logic is similar in some ways to that of the vertebrate vascular system. We previously described a unique embryonic tracheal tubulogenesis phenotype caused by loss of both of the Type III receptor tyrosine phosphatases (RPTPs, Ptp4E and Ptp10D. In Ptp4E Ptp10D double mutants, the linear tubes in unicellular and terminal tracheal branches are converted into bubble-like cysts that incorporate apical cell surface markers. This tube geometry phenotype is modulated by changes in the activity or expression of the epidermal growth factor receptor (Egfr tyrosine kinase (TK. Ptp10D physically interacts with Egfr. Here we demonstrate that the Ptp4E Ptp10D phenotype is the consequence of the loss of negative regulation by the RPTPs of three growth factor receptor TKs: Egfr, Breathless and Pvr. Reducing the activity of any of the three kinases by tracheal expression of dominant-negative mutants suppresses cyst formation. By competing dominant-negative and constitutively active kinase mutants against each other, we show that the three RTKs have partially interchangeable activities, so that increasing the activity of one kinase can compensate for the effects of reducing the activity of another. This implies that SH2-domain downstream effectors that are required for the phenotype are likely to be able to interact with phosphotyrosine sites on all three receptor TKs. We also show that the phenotype involves increases in signaling through the MAP kinase and Rho GTPase pathways.

  4. Gefitinib, an EGFR Tyrosine Kinase inhibitor, Prevents Smoke-Mediated Ciliated Airway Epithelial Cell Loss and Promotes Their Recovery.

    Directory of Open Access Journals (Sweden)

    Monica Valencia-Gattas

    Full Text Available Cigarette smoke exposure is a major health hazard. Ciliated cells in the epithelium of the airway play a critical role in protection against the noxious effects of inhaled cigarette smoke. Ciliated cell numbers are reduced in smokers which weakens host defense and leads to disease. The mechanisms for the loss of ciliated cells are not well understood. The effects of whole cigarette smoke exposure on human airway ciliated ciliated cells were examined using in vitro cultures of normal human bronchial epithelial cells and a Vitrocell® VC 10® Smoking Robot. These experiments showed that whole cigarette smoke causes the loss of differentiated ciliated cells and inhibits differentiation of ciliated cells from undifferentiated basal cells. Furthermore, treatment with the epidermal growth factor receptor (EGFR tyrosine kinase inhibitor, Gefitinib, during smoke exposure prevents ciliated cell loss and promotes ciliated cell differentiation from basal cells. Finally, restoration of ciliated cells was inhibited after smoke exposure was ceased but was enhanced by Gefitinib treatment. These data suggest that inhibition of EGFR activity may provide therapeutic benefit for treating smoke related diseases.

  5. Mechanism of c-Met and EGFR tyrosine kinase inhibitor resistance through epithelial mesenchymal transition in non-small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Rastogi, Ichwaku; Rajanna, Supriya; Webb, Andrew; Chhabra, Gagan; Foster, Brad [Department of Biomedical Sciences, University of Illinois College of Medicine at Rockford, Illinois (United States); Webb, Brian [Thermo Fisher Scientific, Rockford, Illinois (United States); Puri, Neelu, E-mail: neelupur@uic.edu [Department of Biomedical Sciences, University of Illinois College of Medicine at Rockford, Illinois (United States)

    2016-09-02

    According to currently available estimates from Cancer Research UK, 14.1 million new lung cancer cases were diagnosed and a staggering 8.2 million people worldwide died from lung cancer in 2012. EGFR and c-Met are two tyrosine kinase receptors most commonly overexpressed or mutated in Non-small Cell Lung Cancer (NSCLC) resulting in increased proliferation and survival of lung cancer cells. Tyrosine kinase inhibitors (TKIs), such as erlotinib, approved by the FDA as first/second line therapy for NSCLC patients have limited clinical efficacy due to acquired resistance. In this manuscript, we investigate and discuss the role of epithelial mesenchymal transition (EMT) in the development of resistance against EGFR and c-Met TKIs in NSCLC. Our findings show that Zeb-1, a transcriptional repressor of E-Cadherin, is upregulated in TKI-resistant cells causing EMT. We observed that TKI-resistant cells have increased gene and protein expression of EMT related proteins such as Vimentin, N-Cadherin, β-Catenin and Zeb-1, while expression of E-Cadherin, an important cell adhesion molecule, was suppressed. We also confirmed that TKI-resistant cells display mesenchymal cell type morphology, and have upregulation of β-Catenin which may regulate expression of Zeb-1, a transcriptional repressor of E-Cadherin in TKI-resistant NSCLC cells. Finally, we show that down-regulating Zeb-1 by inducing miR-200a or β-Catenin siRNA can increase drug sensitivity of TKI-resistant cells. - Highlights: • Resistance to TKIs in NSCLC cells is mediated via modulation in EMT related proteins. • EMT may induce c-Met mediated TKI resistance, similar to EGFR TKI resistance. • Role of β-catenin and cadherins in TKI resistance was validated by FACS and qPCR. • Knockdown of β-catenin or Zeb-1 can increase TKI sensitivity in TKI-resistant cells. • Targeting key EMT related proteins may overcome TKI resistance in NSCLC.

  6. Negative Regulation of Receptor Tyrosine Kinase (RTK Signaling: A Developing Field

    Directory of Open Access Journals (Sweden)

    Fernanda Ledda

    2007-01-01

    Full Text Available ophic factors control cellular physiology by activating specific receptor tyrosine kinases (RTKs. While the over activation of RTK signaling pathways is associated with cell growth and cancer, recent findings support the concept that impaired down-regulation or deactivation of RTKs may also be a mechanism involved in tumor formation. Under this perspective, the molecular determinants of RTK signaling inhibition may act as tumor-suppressor genes and have a potential role as tumor markers to monitor and predict disease progression. Here, we review the current understanding of the physiological mechanisms that attenuate RTK signaling and discuss evidence that implicates deregulation of these events in cancer.Abbreviations: BDP1: Brain-derived phosphatase 1; Cbl: Casitas B-lineage lymphoma; CIN-85: Cbl-interacting protein of 85 kDa; DER: Drosophila EGFR; EGFR: Epidermal growth factor receptor; ERK 1/2: Extracellular signal-regulated kinase 1/2; Grb2: Growth factor receptor-bound protein 2; HER2: Human epidermal growth factor receptor 2; LRIG: Leucine-rich repeats and immunoglobulin-like domain 1; MAPK: Mitogen-activated protein kinase; Mig 6: Mitogen-inducible gene 6; PTEN: Phosphatase and tensin homologue; RET: Rearranged in transformation; RTK: Receptor tyrosine kinase. SH2 domain: Src-homology 2 domain; SH3 domain: Src-homology 3 domain; Spry: Sprouty.

  7. Protein Kinase G facilitates EGFR-mediated cell death in MDA-MB-468 cells

    Energy Technology Data Exchange (ETDEWEB)

    Jackson, Nicole M.; Ceresa, Brian P., E-mail: brian.ceresa@louisville.edu

    2016-08-15

    The Epidermal Growth Factor Receptor (EGFR) is a transmembrane receptor tyrosine kinase with critical implications in cell proliferation, migration, wound healing and the regulation of apoptosis. However, the EGFR has been shown to be hyper-expressed in a number of human malignancies. The MDA-MB-468 metastatic breast cell line is one example of this. This particular cell line hyper-expresses the EGFR and undergoes EGFR-mediated apoptosis in response to EGF ligand. The goal of this study was to identify the kinases that could be potential intermediates for the EGFR-mediated induction of apoptosis intracellularly. After identifying Cyclic GMP-dependent Protein Kinase G (PKG) as a plausible intermediate, we wanted to determine the temporal relationship of these two proteins in the induction of apoptosis. We observed a dose-dependent decrease in MDA-MB-468 cell viability, which was co-incident with increased PKG activity as measured by VASPSer239 phosphorylation. In addition, we observed a dose dependent decrease in cell viability, as well as an increase in apoptosis, in response to two different PKG agonists, 8-Bromo-cGMP and 8-pCPT-cGMP. MDA-MB-468 cells with reduced PKG activity had attenuated EGFR-mediated apoptosis. These findings indicate that PKG does not induce cell death via transphosphorylation of the EGFR. Instead, PKG activity occurs following EGFR activation. Together, these data indicate PKG as an intermediary in EGFR-mediated cell death, likely via apoptotic pathway.

  8. Ibrutinib Inhibits ERBB Receptor Tyrosine Kinases and HER2-Amplified Breast Cancer Cell Growth.

    Science.gov (United States)

    Chen, Jun; Kinoshita, Taisei; Sukbuntherng, Juthamas; Chang, Betty Y; Elias, Laurence

    2016-12-01

    Ibrutinib is a potent, small-molecule Bruton tyrosine kinase (BTK) inhibitor developed for the treatment of B-cell malignancies. Ibrutinib covalently binds to Cys481 in the ATP-binding domain of BTK. This cysteine residue is conserved among 9 other tyrosine kinases, including HER2 and EGFR, which can be targeted. Screening large panels of cell lines demonstrated that ibrutinib was growth inhibitory against some solid tumor cells, including those inhibited by other HER2/EGFR inhibitors. Among sensitive cell lines, breast cancer lines with HER2 overexpression were most potently inhibited by ibrutinib (ibrutinib coincided with downregulation of phosphorylation on HER2 and EGFR and their downstream targets, AKT and ERK. Irreversible inhibition of HER2 and EGFR in breast cancer cells was established after 30-minute incubation above 100 nmol/L or following 2-hour incubation at lower concentrations. Furthermore, ibrutinib inhibited recombinant HER2 and EGFR activity that was resistant to dialysis and rapid dilution, suggesting an irreversible interaction. The dual activity toward TEC family (BTK and ITK) and ERBB family kinases was unique to ibrutinib, as ERBB inhibitors do not inhibit or covalently bind BTK or ITK. Xenograft studies with HER2 + MDA-MB-453 and BT-474 cells in mice in conjunction with determination of pharmacokinetics demonstrated significant exposure-dependent inhibition of growth and key signaling molecules at levels that are clinically achievable. Ibrutinib's unique dual spectrum of activity against both TEC family and ERBB kinases suggests broader applications of ibrutinib in oncology. Mol Cancer Ther; 15(12); 2835-44. ©2016 AACR. ©2016 American Association for Cancer Research.

  9. CBL is frequently altered in lung cancers: its relationship to mutations in MET and EGFR tyrosine kinases.

    Directory of Open Access Journals (Sweden)

    Yi-Hung Carol Tan

    2010-01-01

    Full Text Available Non-small cell lung cancer (NSCLC is a heterogeneous group of disorders with a number of genetic and proteomic alterations. c-CBL is an E3 ubiquitin ligase and adaptor molecule important in normal homeostasis and cancer. We determined the genetic variations of c-CBL, relationship to receptor tyrosine kinases (EGFR and MET, and functionality in NSCLC.Using archival formalin-fixed paraffin embedded (FFPE extracted genomic DNA, we show that c-CBL mutations occur in somatic fashion for lung cancers. c-CBL mutations were not mutually exclusive of MET or EGFR mutations; however they were independent of p53 and KRAS mutations. In normal/tumor pairwise analysis, there was significant loss of heterozygosity (LOH for the c-CBL locus (22%, n = 8/37 and none of these samples revealed any mutation in the remaining copy of c-CBL. The c-CBL LOH also positively correlated with EGFR and MET mutations observed in the same samples. Using select c-CBL somatic mutations such as S80N/H94Y, Q249E and W802* (obtained from Caucasian, Taiwanese and African-American samples, respectively transfected in NSCLC cell lines, there was increased cell viability and cell motility.Taking the overall mutation rate of c-CBL to be a combination as somatic missense mutation and LOH, it is clear that c-CBL is highly mutated in lung cancers and may play an essential role in lung tumorigenesis and metastasis.

  10. Intracellular signaling of the Ufo/Axl receptor tyrosine kinase is mediated mainly by a multi-substrate docking-site.

    Science.gov (United States)

    Braunger, J; Schleithoff, L; Schulz, A S; Kessler, H; Lammers, R; Ullrich, A; Bartram, C R; Janssen, J W

    1997-06-05

    Ufo/Axl belongs to a new family of receptor tyrosine kinases with an extracellular structure similar to that of neural cell adhesion molecules. In order to elucidate intracellular signaling, the cytoplasmic moiety of Ufo/Axl was used to screen an expression library according to the CORT (cloning of receptor targets) method. Three putative Ufo substrates were identified: phospholipase Cgamma1 (PLCgamma), as well as p85alpha and p85beta subunits of phosphatidylinositol 3'-kinase (PI3-kinase). Subsequently, chimeric EGFR/Ufo receptors consisting of the extracellular domains of the epidermal growth factor receptor (EGFR) and the transmembrane and intracellular moiety of Ufo were engineered. Using different far-Western blot analyses and coimmunoprecipitation assays, receptor binding of PLCgamma and p85 proteins as well as GRB2, c-src and lck was examined in vitro and in vivo. Competitive inhibition of substrate binding and mutagenesis experiments with EGFR/Ufo constructs revealed C-terminal tyrosine 821 (EILpYVNMDEG) as a docking site for multiple effectors, namely PLCgamma, p85 proteins, GRB2, c-src and lck. Tyrosine 779 (DGLpYALMSRC) demonstrated an additional, but lower binding affinity for the p85 proteins in vitro. In addition, binding of PLCgamma occurred through tyrosine 866 (AGRpYVLCPST). Moreover, our in vivo data indicate that further direct or indirect binding sites for PLCgamma, GRB2, c-src and lck on the human Ufo receptor may exist.

  11. HER2 mutated breast cancer responds to treatment with single agent neratinib, a second generation HER2/EGFR tyrosine kinase inhibitor

    Science.gov (United States)

    Ben–Baruch, Noa Efrat; Bose, Ron; Kavuri, Shyam M.; Ma, Cynthia X.; Ellis, Matthew J.

    2015-01-01

    Activating mutations in the HER2 tyrosine kinase have been identified in human breast cancers that lack HER2 gene amplification. These patients are not candidates for HER2 targeted drugs under current standards of care, but preclinical data strongly suggest that these patients will benefit from anti-HER2 drugs. In this case report, we describe a young woman with metastatic breast cancer whose tumor was found to carry a HER2 L755S mutation, which is in the kinase domain of HER2. Treatment with the second generation HER2/EGFR tyrosine kinase inhibitor, neratinib, resulted in partial response and dramatic improvement in the patient’s function status. This partial response lasted 11 months and when the patient’s cancer progressed, she was treated with neratinib plus capecitabine and her cancer again responded. This second response parallels the benefit seen with continuing trastuzumab in HER2 amplified breast cancer after disease progression. This case is the first report, to our knowledge, of successful single agent treatment of HER2 mutated breast cancer. Two clinical trials of neratinib for HER2 mutated, metastatic breast cancer are currently enrolling patients. Further, data from The Cancer Genome Atlas project have identified HER2 mutations in a wide range of solid tumors, including bladder, colorectal, and non-small cell lung cancer, suggesting that clinical trials of neratinib or neratinib-based combinations for HER2 mutated solid tumors is warranted. PMID:26358790

  12. HER2-Mutated Breast Cancer Responds to Treatment With Single-Agent Neratinib, a Second-Generation HER2/EGFR Tyrosine Kinase Inhibitor.

    Science.gov (United States)

    Ben-Baruch, Noa Efrat; Bose, Ron; Kavuri, Shyam M; Ma, Cynthia X; Ellis, Matthew J

    2015-09-01

    Activating mutations in the HER2 tyrosine kinase have been identified in human breast cancers that lack HER2 gene amplification. These patients are not candidates for HER2-targeted drugs under current standards of care, but preclinical data strongly suggest that these patients will benefit from anti-HER2 drugs. This case report describes a young woman with metastatic breast cancer whose tumor was found to carry a HER2 L755S mutation, which is in the kinase domain of HER2. Treatment with the second-generation HER2/EGFR tyrosine kinase inhibitor neratinib resulted in partial response and dramatic improvement in the patient's functional status. This partial response lasted 11 months, and when the patient's cancer progressed, she was treated with neratinib plus capecitabine and her cancer again responded. This second response parallels the benefit seen with continuing trastuzumab in HER2-amplified breast cancer after disease progression. This case represents the first report, to our knowledge, of successful single-agent treatment of HER2-mutated breast cancer. Two clinical trials of neratinib for HER2-mutated metastatic breast cancer are currently enrolling patients. Further, data from The Cancer Genome Atlas project have identified HER2 mutations in a wide range of solid tumors, including bladder, colorectal, and non-small cell lung cancers, suggesting that clinical trials of neratinib or neratinib-based combinations for HER2-mutated solid tumors is warranted. Copyright © 2015 by the National Comprehensive Cancer Network.

  13. Bacterial Protein-Tyrosine Kinases

    DEFF Research Database (Denmark)

    Shi, Lei; Kobir, Ahasanul; Jers, Carsten

    2010-01-01

    in exopolysaccharide production, virulence, DNA metabolism, stress response and other key functions of the bacterial cell. BY-kinases act through autophosphorylation (mainly in exopolysaccharide production) and phosphorylation of other proteins, which have in most cases been shown to be activated by tyrosine......Bacteria and Eukarya share essentially the same family of protein-serine/threonine kinases, also known as the Hanks-type kinases. However, when it comes to protein-tyrosine phosphorylation, bacteria seem to have gone their own way. Bacterial protein-tyrosine kinases (BY-kinases) are bacterial...... and highlighted their importance in bacterial physiology. Having no orthologues in Eukarya, BY-kinases are receiving a growing attention from the biomedical field, since they represent a particularly promising target for anti-bacterial drug design....

  14. Receptor Tyrosine Kinases in Drosophila Development

    Science.gov (United States)

    Sopko, Richelle; Perrimon, Norbert

    2013-01-01

    Tyrosine phosphorylation plays a significant role in a wide range of cellular processes. The Drosophila genome encodes more than 20 receptor tyrosine kinases and extensive studies in the past 20 years have illustrated their diverse roles and complex signaling mechanisms. Although some receptor tyrosine kinases have highly specific functions, others strikingly are used in rather ubiquitous manners. Receptor tyrosine kinases regulate a broad expanse of processes, ranging from cell survival and proliferation to differentiation and patterning. Remarkably, different receptor tyrosine kinases share many of the same effectors and their hierarchical organization is retained in disparate biological contexts. In this comprehensive review, we summarize what is known regarding each receptor tyrosine kinase during Drosophila development. Astonishingly, very little is known for approximately half of all Drosophila receptor tyrosine kinases. PMID:23732470

  15. Adaptation to TKI Treatment Reactivates ERK Signaling in Tyrosine Kinase-Driven Leukemias and Other Malignancies.

    Science.gov (United States)

    Bruner, J Kyle; Ma, Hayley S; Li, Li; Qin, Alice Can Ran; Rudek, Michelle A; Jones, Richard J; Levis, Mark J; Pratz, Keith W; Pratilas, Christine A; Small, Donald

    2017-10-15

    FMS-like tyrosine kinase-3 (FLT3) tyrosine kinase inhibitors (TKI) have been tested extensively to limited benefit in acute myeloid leukemia (AML). We hypothesized that FLT3/internal tandem duplication (ITD) leukemia cells exhibit mechanisms of intrinsic signaling adaptation to TKI treatment that are associated with an incomplete response. Here, we identified reactivation of ERK signaling within hours following treatment of FLT3/ITD AML cells with selective inhibitors of FLT3. When these cells were treated with inhibitors of both FLT3 and MEK in combination, ERK reactivation was abrogated and anti-leukemia effects were more pronounced compared with either drug alone. ERK reactivation was also observed following inhibition of other tyrosine kinase-driven cancer cells, including EGFR-mutant lung cancer, HER2-amplified breast cancer, and BCR-ABL leukemia. These studies reveal an adaptive feedback mechanism in tyrosine kinase-driven cancers associated with reactivation of ERK signaling in response to targeted inhibition. Cancer Res; 77(20); 5554-63. ©2017 AACR . ©2017 American Association for Cancer Research.

  16. Tyrosine kinases in rheumatoid arthritis

    Directory of Open Access Journals (Sweden)

    Kobayashi Akiko

    2011-08-01

    Full Text Available Abstract Rheumatoid arthritis (RA is an inflammatory, polyarticular joint disease. A number of cellular responses are involved in the pathogenesis of rheumatoid arthritis, including activation of inflammatory cells and cytokine expression. The cellular responses involved in each of these processes depends on the specific signaling pathways that are activated; many of which include protein tyrosine kinases. These pathways include the mitogen-activated protein kinase pathway, Janus kinases/signal transducers and activators transcription pathway, spleen tyrosine kinase signaling, and the nuclear factor κ-light-chain-enhancer of activated B cells pathway. Many drugs are in development to target tyrosine kinases for the treatment of RA. Based on the number of recently published studies, this manuscript reviews the role of tyrosine kinases in the pathogenesis of RA and the potential role of kinase inhibitors as new therapeutic strategies of RA.

  17. Raman Microspectroscopic Evidence for the Metabolism of a Tyrosine Kinase Inhibitor, Neratinib, in Cancer Cells.

    Science.gov (United States)

    Aljakouch, Karim; Lechtonen, Tatjana; Yosef, Hesham K; Hammoud, Mohamad K; Alsaidi, Wissam; Kötting, Carsten; Mügge, Carolin; Kourist, Robert; El-Mashtoly, Samir F; Gerwert, Klaus

    2018-06-11

    Tyrosine kinase receptors are one of the main targets in cancer therapy. They play an essential role in the modulation of growth factor signaling and thereby inducing cell proliferation and growth. Tyrosine kinase inhibitors such as neratinib bind to EGFR and HER2 receptors and exhibit antitumor activity. However, little is known about their detailed cellular uptake and metabolism. Here, we report for the first time the intracellular spatial distribution and metabolism of neratinib in different cancer cells using label-free Raman imaging. Two new neratinib metabolites were detected and fluorescence imaging of the same cells indicate that neratinib accumulates in lysosomes. The results also suggest that both EGFR and HER2 follow the classical endosome lysosomal pathway for degradation. A combination of Raman microscopy, DFT calculations, and LC-MS was used to identify the chemical structure of neratinib metabolites. These results show the potential of Raman microscopy to study drug pharmacokinetics. © 2018 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

  18. Sensitivity to epidermal growth factor receptor tyrosine kinase inhibitors in males, smokers, and non-adenocarcinoma lung cancer in patients with EGFR mutations.

    Science.gov (United States)

    Zeng, Zhu; Chen, Hua-Jun; Yan, Hong-Hong; Yang, Jin-Ji; Zhang, Xu-Chao; Wu, Yi-Long

    2013-09-27

    The demographical/clinical characteristics of being Asian, having an adenocarcinoma, being female, and being a "never-smoker" are regarded as favorable predictors for epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) efficacy in non-small cell lung cancer (NSCLC) with unknown EGFR gene status. In this study, we examined the effects of the supposedly unfavorable clinical variables in EGFR-mutant patients. In total, 159 EGFR-mutant NSCLC patients' clinical features were correlated with progression-free survival (PFS), response rate (RR), and overall survival (OS). Multivariate analysis of clinical characteristics was performed using the Cox and logistic regression methods. There were 90 females (56.6%), 112 never-smokers (70.4%), and 153 patients with adenocarcinomas (96.2%). All patients were treated with EGFR-TKI, and 52.8% received TKI in a first-line setting. The median PFS of patients receiving first-line TKI was similar, regardless of gender (males vs females: 9.1 vs 9.7 months, p=0.793), smoking status (never-smokers vs smokers: 9.9 vs 9.1 months, p=0.570), or histology (adenocarcinoma vs non-adenocarcinoma: 9.7 vs 9.2 months, p=0.644). OS curves of first-line TKI-treated patients were also not associated with gender (p=0.722), smoking status (p=0.579), or histology (p=0.480). Similar results of PFS and OS were obtained for patients who received TKI beyond first-line. Multivariate analysis indicated that none of these clinical factors was an independent predictor of survival. The supposedly 'favorable' clinical factors of female gender, non-smoking status, and adenocarcinoma were not independent predictive factors for PFS or OS in this population of EGFR-mutant NSCLC patients.

  19. Clinical responses to EGFR-tyrosine kinase inhibitor retreatment in non-small cell lung cancer patients who benefited from prior effective gefitinib therapy: a retrospective analysis

    International Nuclear Information System (INIS)

    Watanabe, Satoshi; Miyabayashi, Takao; Narita, Ichiei; Yoshizawa, Hirohisa; Tanaka, Junta; Ota, Takeshi; Kondo, Rie; Tanaka, Hiroshi; Kagamu, Hiroshi; Ichikawa, Kosuke; Koshio, Jun; Baba, Junko

    2011-01-01

    Gefitinib was the first epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) approved for the treatment of advanced non-small cell lung cancer (NSCLC). Few treatment options are available for NSCLC patients who have responded to gefitinib treatment and demonstrated tumor progression. The present study was conducted to evaluate the efficacy and toxicity of the 2 nd EGFR-TKI administration. We retrospectively analyzed 11 patients who had obtained a partial response (PR) or stable disease (SD) with gefitinib treatment and were re-treated with EGFR-TKI after failure of the initial gefitinib treatment. Three patients (27%) were treated with gefitinib as the 2 nd EGFR-TKI, and 8 patients (73%) received erlotinib. Only one patient (9%) showed PR, 7 (64%) achieved SD, and 3 (27%) had progressive disease. The disease control rate was 73% (95% CI, 43% - 91%) and the median progression-free survival was 3.4 months (95% CI, 2 - 5.2). The median overall survival from the beginning of the 2 nd EGFR-TKI and from diagnosis were 7.3 months (95% CI, 2.7 - 13) and 36.7 months (95% CI, 23.6 - 43.9), respectively. No statistical differences in PFS or OS were observed between gefitinib and erlotinib as the 2 nd EGFR-TKI (PFS, P = 0.23 and OS, P = 0.052). The toxicities associated with the 2 nd EGFR-TKI were generally acceptable and comparable to those observed for the initial gefitinib therapy. Our results indicate that a 2 nd EGFR-TKI treatment can be an effective treatment option for gefitinib responders

  20. Antibody-induced activation of the epidermal growth factor receptor tyrosine kinase requires the presence of detergent

    NARCIS (Netherlands)

    Spaargaren, M.; Defize, L. H.; de Laat, S. W.; Boonstra, J.

    1990-01-01

    Activation of the epidermal growth factor receptor (EGF-R) tyrosine kinase was investigated in membrane preparations as well as intact A431 cells, using anti-EGF-R antibodies directed against extra- and intracellular receptor domains. In vitro assay conditions were mimicked on whole cells by a mild

  1. A novel mutation in the tyrosine kinase domain of ERBB2 in hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Bekaii-Saab, Tanios; Williams, Nita; Plass, Christoph; Calero, Miguel Villalona; Eng, Charis

    2006-01-01

    Several studies showed that gain-of-function somatic mutations affecting the catalytic domain of EGFR in non-small cell lung carcinomas were associated with response to gefitinib and erlotinib, both EGFR-tyrosine kinase inhibitors. In addition, 4% of non-small cell lung carcinomas were shown to have ERBB2 mutations in the kinase domain. In our study, we sought to determine if similar respective gain-of-function EGFR and ERBB2 mutations were present in hepatoma and/or biliary cancers. We extracted genomic DNA from 40 hepatoma (18) and biliary cancers (22) samples, and 44 adenocarcinomas of the lung, this latter as a positive control for mutation detection. We subjected those samples to PCR-based semi-automated double stranded nucleotide sequencing targeting exons 18–21 of EGFR and ERBB2. All samples were tested against matched normal DNA. We found 11% of hepatoma, but no biliary cancers, harbored a novel ERBB2 H878Y mutation in the activating domain. These newly described mutations may play a role in predicting response to EGFR-targeted therapy in hepatoma and their role should be explored in prospective studies

  2. Three generations of epidermal growth factor receptor tyrosine kinase inhibitors developed to revolutionize the therapy of lung cancer

    Directory of Open Access Journals (Sweden)

    Zhang H

    2016-11-01

    Full Text Available Haijun Zhang Department of Oncology, Zhongda Hospital, Medical School, Southeast University, Nanjing, People’s Republic of China Abstract: Lung cancer, ~80%–85% of which is non-small-cell lung cancer (NSCLC, is the leading cause of cancer-related mortality worldwide. Sensitizing mutations in epidermal growth factor receptor (EGFR gene (EGFRm+, such as exon 19 deletions and exon 21 L858R point mutations, are the most important drivers in NSCLC patients. In this respect, small-molecule EGFR tyrosine kinase inhibitors (TKIs have been designed and developed, which launched the era of targeted, personalized and precise medicine for lung cancer. Patients with EGFRm+ could achieve good responses to the treatment with the first-generation EGFR TKIs, such as erlotinib and gefitinib. However, most patients develop acquired drug resistance mostly driven by the T790M mutation occurring within exon 20. Although the second-generation EGFR TKIs, such as afatinib, dacomitinib and neratinib, demonstrated promising activity against T790M in preclinical models, they have failed to overcome resistance in patients due to dose-limiting toxicity. Recently, the third-generation EGFR TKIs have shown to be effective against cell lines and murine models harboring T790M mutations while sparing wild-type EGFR, which represents a promising breakthrough approach in overcoming T790M-mediated resistance in NSCLC patients. This article provides a comprehensive review of the therapy revolution for NSCLC with three generations of EGFR TKIs. Keywords: lung cancer, epidermal growth factor receptor, tyrosine kinase inhibitors, T790M mutation

  3. Patients harboring EGFR mutation after primary resistance to crizotinib and response to EGFR-tyrosine kinase inhibitor

    Directory of Open Access Journals (Sweden)

    Wang WX

    2016-01-01

    Full Text Available Wenxian Wang,1 Xiaowen Jiang,1 Zhengbo Song,1,2 Yiping Zhang1,2 1Department of Chemotherapy, Zhejiang Cancer Hospital, 2Key Laboratory Diagnosis and Treatment Technology on Thoracic Oncology, Hangzhou, Zhejiang, People’s Republic of China Abstract: Anaplastic lymphoma kinase (ALK rearrangement lung cancer responds to ALK tyrosine kinase inhibitors. It is known that many cases ultimately acquired resistance to crizotinib. However, a case of primary resistance is rare. We present a case of harboring exon 19 deletion in epidermal growth factor receptor in ALK rearranged lung adenocarcinoma, who experienced a partial tumor response to icotinib after failure with crizotinib therapy and chemotherapy. Considering the partial response, we conclude that it is important to find the cause of resistance to crizotinib. We detected gene mutations with plasma by the next-generation sequencing; the next-generation sequencing demonstrates an attractive system to identify mutations improving the outcome of patients with a deadly disease. Keywords: non-small cell lung cancer, anaplastic lymphoma kinase, crizotinib, epidermal growth factor receptor

  4. Icotinib in Patients with Pretreated Advanced Esophageal Squamous Cell Carcinoma with EGFR Overexpression or EGFR Gene Amplification: A Single-Arm, Multicenter Phase 2 Study.

    NARCIS (Netherlands)

    Huang, J.; Fan, Q.; Lu, P.; Ying, J.; Ma, C.; Liu, W.; Liu, Y.; Tan, F.; Sun, Y

    2016-01-01

    INTRODUCTION: Epidermal growth factor receptor (EGFR) has been reported to be overexpressed and amplified in a high percentage of patients with esophageal squamous cell carcinoma (ESCC). The activity of icotinib, an EGFR tyrosine kinase inhibitor, was assessed in previously treated ESCC with EGFR

  5. EGFR kinase-dependent and kinase-independent roles in clear cell renal cell carcinoma.

    Science.gov (United States)

    Cossu-Rocca, Paolo; Muroni, Maria R; Sanges, Francesca; Sotgiu, Giovanni; Asunis, Anna; Tanca, Luciana; Onnis, Daniela; Pira, Giovanna; Manca, Alessandra; Dore, Simone; Uras, Maria G; Ena, Sara; De Miglio, Maria R

    2016-01-01

    Epidermal growth factor receptor (EGFR) is associated with progression of many epithelial malignancies and represents a significant therapeutic target. Although clear cell renal cell carcinoma (CCRCC) has been widely investigated for EGFR molecular alterations, genetic evidences of EGFR gene activating mutations and/or gene amplification have been rarely confirmed in the literature. Therefore, until now EGFR-targeted therapies in clinical trials have been demonstrated unsuccessful. New evidence has been given about the interactions between EGFR and the sodium glucose co-transporter-1 (SGLT1) in maintaining the glucose basal intracellular level to favour cancer cell growth and survival; thus a new functional role may be attributed to EGFR, regardless of its kinase activity. To define the role of EGFR in CCRCC an extensive investigation of genetic changes and functional kinase activities was performed in a series of tumors by analyzing the EGFR mutational status and expression profile, together with the protein expression of downstream signaling pathways members. Furthermore, we investigated the co-expression of EGFR and SGLT1 proteins and their relationships with clinic-pathological features in CCRCC. EGFR protein expression was identified in 98.4% of CCRCC. Furthermore, it was described for the first time that SGLT1 is overexpressed in CCRCC (80.9%), and that co-expression with EGFR is appreciable in 79.4% of the tumours. Moreover, the activation of downstream EGFR pathways was found in about 79.4% of SGLT1-positive CCRCCs. The mutational status analysis of EGFR failed to demonstrate mutations on exons 18 to 24 and the presence of EGFR-variantIII (EGFRvIII) in all CCRCCs analyzed. FISH analysis revealed absence of EGFR amplification, and high polysomy of chromosome 7. Finally, the EGFR gene expression profile showed gene overexpression in 38.2% of CCRCCs. Our study contributes to define the complexity of EGFR role in CCRCC, identifying its bivalent kinase

  6. Epidermal growth factor receptor (EGFR) and EGFR mutations, function and possible role in clinical trials

    DEFF Research Database (Denmark)

    Voldborg, B R; Damstrup, L; Spang-Thomsen, M

    1997-01-01

    The epidermal growth factor receptor (EGFR) is a growth factor receptor that induces cell differentiation and proliferation upon activation through the binding of one of its ligands. The receptor is located at the cell surface, where the binding of a ligand activates a tyrosine kinase in the intr...... aspects of therapeutic targeting of EGFR....

  7. Ibrutinib targets mutant-EGFR kinase with a distinct binding conformation.

    Science.gov (United States)

    Wang, Aoli; Yan, Xiao-E; Wu, Hong; Wang, Wenchao; Hu, Chen; Chen, Cheng; Zhao, Zheng; Zhao, Peng; Li, Xixiang; Wang, Li; Wang, Beilei; Ye, Zi; Wang, Jinhua; Wang, Chu; Zhang, Wei; Gray, Nathanael S; Weisberg, Ellen L; Chen, Liang; Liu, Jing; Yun, Cai-Hong; Liu, Qingsong

    2016-10-25

    Ibrutinib, a clinically approved irreversible BTK kinase inhibitor for Mantle Cell Lymphoma (MCL) and Chronic Lymphocytic Leukemia (CLL) etc, has been reported to be potent against EGFR mutant kinase and currently being evaluated in clinic for Non Small Cell Lung Cancer (NSCLC). Through EGFR wt/mutant engineered isogenic BaF3 cell lines we confirmed the irreversible binding mode of Ibrutinib with EGFR wt/mutant kinase via Cys797. However, comparing to typical irreversible EGFR inhibitor, such as WZ4002, the washing-out experiments revealed a much less efficient covalent binding for Ibrutinib. The biochemical binding affinity examination in the EGFR L858R/T790M kinase revealed that, comparing to more efficient irreversible inhibitor WZ4002 (Kd: 0.074 μM), Ibrutinib exhibited less efficient binding (Kd: 0.18 μM). An X-ray crystal structure of EGFR (T790M) in complex with Ibrutinib exhibited a unique DFG-in/c-Helix-out inactive binding conformation, which partially explained the less efficiency of covalent binding and provided insight for further development of highly efficient irreversible binding inhibitor for the EGFR mutant kinase. These results also imply that, unlike the canonical irreversible inhibitor, sustained effective concentration might be required for Ibrutinib in order to achieve the maximal efficacy in the clinic application against EGFR driven NSCLC.

  8. Lead acetate induces EGFR activation upstream of SFK and PKCα linkage to the Ras/Raf-1/ERK signaling

    International Nuclear Information System (INIS)

    Wang, C.-Y.; Wang, Y.-T.; Tzeng, D.-W.; Yang, J.-L.

    2009-01-01

    Lead acetate (Pb), a probable human carcinogen, can activate protein kinase C (PKC) upstream of extracellular signal-regulated kinase 1 and 2 (ERK1/2). Yet, it remains unclear whether Pb activation of PKC → ERK1/2 involves receptor/non-receptor tyrosine kinases and the Ras signaling transducer. Here we demonstrate a novel mechanism elicited by Pb for transmitting ERK1/2 signaling in CL3 human non-small-cell lung adenocarcinoma cells. Pb induction of higher steady-state levels of Ras-GTP was essential for increasing phospho-Raf-1 S338 and phospho-ERK1/2. Pre-treatment of the cells with a conventional PKC inhibitor Goe6976 or depleting PKCα using specific small interfering RNA blocked Pb induction of Ras-GTP. Pb also activated cellular tyrosine kinases. Specific pharmacological inhibitors, PD153035 for epidermal growth factor receptor (EGFR) and SU6656 for Src family tyrosine kinases (SFK), but not AG1296 for platelet-derived growth factor receptor, could suppress the Pb-induced tyrosine kinases, PKCα, Ras-GTP, phospho-Raf-1 S338 and phospho-ERK1/2. Furthermore, phosphorylation of tyrosines on the EGFR multiple autophosphorylation sites and the conserved SFK autophosphorylation site occurred during exposure of cells to Pb for 1-5 min and 5-30 min, respectively. Intriguingly, Pb activation of EGFR required the intrinsic kinase activity but not dimerization of the receptor. Inhibition of SFK or PKCα activities did not affect EGFR phosphorylation, while knockdown of EGFR blocked SFK phosphorylation and PKCα activation following Pb. Together, these results indicate that immediate activation of EGFR in response to Pb is obligatory for activation of SFK and PKCα and subsequent the Ras-Raf-1-MKK1/2-ERK1/2 signaling cascade

  9. Her4 and Her2/neu tyrosine kinase domains dimerize and activate in a reconstituted in vitro system.

    Science.gov (United States)

    Monsey, John; Shen, Wei; Schlesinger, Paul; Bose, Ron

    2010-03-05

    Her4 (ErbB-4) and Her2/neu (ErbB-2) are receptor-tyrosine kinases belonging to the epidermal growth factor receptor (EGFR) family. Crystal structures of EGFR and Her4 kinase domains demonstrate kinase dimerization and activation through an allosteric mechanism. The kinase domains form an asymmetric dimer, where the C-lobe surface of one monomer contacts the N-lobe of the other monomer. EGFR kinase dimerization and activation in vitro was previously reported using a nickel-chelating lipid-liposome system, and we now apply this system to all other members of the EGFR family. Polyhistidine-tagged Her4, Her2/neu, and Her3 kinase domains are bound to these nickel-liposomes and are brought to high local concentration, mimicking what happens to full-length receptors in vivo following ligand binding. Addition of nickel-liposomes to Her4 kinase domain results in 40-fold activation in kinase activity and marked enhancement of C-terminal tail autophosphorylation. Activation of Her4 shows a sigmoidal dependence on kinase concentration, consistent with a cooperative process requiring kinase dimerization. Her2/neu kinase activity is also activated by nickel-liposomes, and is increased further by heterodimerization with Her3 or Her4. The ability of Her3 and Her4 to heterodimerize and activate other family members is studied in vitro. Her3 kinase domain readily activates Her2/neu but is a poor activator of Her4, which differs from the prediction made by the asymmetric dimer model. Mutation of Her3 residues (952)ENI(954) to the corresponding sequence in Her4 enhanced the ability of Her3 to activate Her4, demonstrating that sequence differences on the C-lobe surface influence the heterodimerization and activation of ErbB kinase domains.

  10. A case of Trousseau syndrome caused by pulmonary adenocarcinoma that was controlled for one year and 10 months with thrombosis treatment using an EGFR tyrosine kinase inhibitor and chemotherapy

    Directory of Open Access Journals (Sweden)

    Hiroaki Masubuchi

    2015-01-01

    Trousseau syndrome carries a poor prognosis and controlling thrombosis is difficult. In this case, the administration of anticancer therapy allowed use to control the patient's thrombosis. Therefore, this case highlights the importance of treating cancer in patients with Trousseau syndrome. In addition, the FDP and D-dimer levels changed in parallel with changes in the CEA level, which suggests that the activity of cancer is related to an internal thrombotic tendency. Hence, changes in the FDP and D-dimer values are associated with the efficacy of treatment with EGFR tyrosine kinase inhibitors and chemotherapy and may function as markers of recurrence.

  11. A targeted enzyme approach to sensitization of tyrosine kinase inhibitor-resistant breast cancer cells.

    Science.gov (United States)

    Giordano, Courtney R; Mueller, Kelly L; Terlecky, Laura J; Krentz, Kendra A; Bollig-Fischer, Aliccia; Terlecky, Stanley R; Boerner, Julie L

    2012-10-01

    Gefitinib is an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) of potential use in patients with breast cancer. Unfortunately, in clinical studies, gefitinib is often ineffective indicating that resistance to EGFR inhibitors may be a common occurrence in cancer of the breast. EGFR has been shown to be overexpressed in breast cancer, and in particular remains hyperphosphorylated in cell lines such as MDA-MB-468 that are resistant to EGFR inhibitors. Here, we investigate the cause of this sustained phosphorylation and the molecular basis for the ineffectiveness of gefitinib. We show that reactive oxygen species (ROS), known to damage cellular macromolecules and to modulate signaling cascades in a variety of human diseases including cancers, appear to play a critical role in mediating EGFR TKI-resistance. Furthermore, elimination of these ROS through use of a cell-penetrating catalase derivative sensitizes the cells to gefitinib. These results suggest a new approach for the treatment of TKI-resistant breast cancer patients specifically, the targeting of ROS and attendant downstream oxidative stress and their effects on signaling cascades. Copyright © 2012. Published by Elsevier Inc.

  12. Small Molecule Tyrosine Kinase Inhibitors of ErbB2/HER2/Neu in the Treatment of Aggressive Breast Cancer

    Directory of Open Access Journals (Sweden)

    Richard L. Schroeder

    2014-09-01

    Full Text Available The human epidermal growth factor receptor 2 (HER2 is a member of the erbB class of tyrosine kinase receptors. These proteins are normally expressed at the surface of healthy cells and play critical roles in the signal transduction cascade in a myriad of biochemical pathways responsible for cell growth and differentiation. However, it is widely known that amplification and subsequent overexpression of the HER2 encoding oncogene results in unregulated cell proliferation in an aggressive form of breast cancer known as HER2-positive breast cancer. Existing therapies such as trastuzumab (Herceptin® and lapatinib (Tyverb/Tykerb®, a monoclonal antibody inhibitor and a dual EGFR/HER2 kinase inhibitor, respectively, are currently used in the treatment of HER2-positive cancers, although issues with high recurrence and acquired resistance still remain. Small molecule tyrosine kinase inhibitors provide attractive therapeutic targets, as they are able to block cell signaling associated with many of the proposed mechanisms for HER2 resistance. In this regard we aim to present a review on the available HER2 tyrosine kinase inhibitors, as well as those currently in development. The use of tyrosine kinase inhibitors as sequential or combinatorial therapeutic strategies with other HER family inhibitors is also discussed.

  13. Administration of Menadione, Vitamin K3, Ameliorates Off-Target Effects on Corneal Epithelial Wound Healing Due to Receptor Tyrosine Kinase Inhibition.

    Science.gov (United States)

    Rush, Jamie S; Bingaman, David P; Chaney, Paul G; Wax, Martin B; Ceresa, Brian P

    2016-11-01

    The antiangiogenic receptor tyrosine kinase inhibitor (RTKi), 3-[(4-bromo-2,6-difluorophenyl)methoxy]-5-[[[[4-(1-pyrrolidinyl) butyl] amino] carbonyl]amino]-4-isothiazolecarboxamide hydrochloride, targets VEGFR2 (half maximal inhibitory concentration [IC50] = 11 nM); however, off-target inhibition of epidermal growth factor receptor (EGFR) occurs at higher concentrations. (IC50 = 5.8 μM). This study was designed to determine the effect of topical RTKi treatment on EGF-mediated corneal epithelial wound healing and to develop new strategies to minimize off-target EGFR inhibition. In vitro corneal epithelial wound healing was measured in response to EGF using a transformed human cell line (hTCEpi cells). In vivo corneal wound healing was assessed using a murine model. In these complementary assays, wound healing was measured in the presence of varying RTKi concentrations. Immunoblot analysis was used to examine EGFR and VEGFR2 phosphorylation and the kinetics of EGFR degradation. An Alamar Blue assay measured VEGFR2-mediated cell biology. Receptor tyrosine kinase inhibitor exposure caused dose-dependent inhibition of EGFR-mediated corneal epithelial wound healing in vitro and in vivo. Nanomolar concentrations of menadione, a vitamin K3 analog, when coadministered with the RTKi, slowed EGFR degradation and ameliorated the inhibitory effects on epithelial wound healing both in vitro and in vivo. Menadione did not alter the RTKi's IC50 against VEGFR2 phosphorylation or its inhibition of VEGF-induced retinal endothelial cell proliferation. An antiangiogenic RTKi exhibited off-target effects on the corneal epithelium that can be minimized by menadione without deleteriously affecting its on-target VEGFR2 blockade. These data indicate that menadione has potential as a topical supplement for individuals suffering from perturbations in corneal epithelial homeostasis, especially as an untoward side effect of kinase inhibitors.

  14. Effects of the EGFR Inhibitor Erlotinib on Magnesium Handling

    NARCIS (Netherlands)

    Dimke, Henrik; van der Wijst, Jenny; Alexander, Todd R.; Meijer, Inez M. J.; Mulder, Gemma M.; van Goor, Harry; Tejpar, Sabine; Hoenderop, Joost G.; Bindels, Rene J.

    A mutation in pro-EGF causes isolated hypomagnesemia, and monoclonal antibodies targeting the extracellular domain of the EGF receptor (EGFR) affect epithelial Mg2+ transport. The effect of the EGFR tyrosine kinase inhibitor erlotinib on Mg2+ homeostasis, however, remains unknown. Here, we injected

  15. Tyrosine kinase inhibitors: Multi-targeted or single-targeted?

    Science.gov (United States)

    Broekman, Fleur; Giovannetti, Elisa; Peters, Godefridus J

    2011-02-10

    Since in most tumors multiple signaling pathways are involved, many of the inhibitors in clinical development are designed to affect a wide range of targeted kinases. The most important tyrosine kinase families in the development of tyrosine kinase inhibitors are the ABL, SCR, platelet derived growth factor, vascular endothelial growth factor receptor and epidermal growth factor receptor families. Both multi-kinase inhibitors and single-kinase inhibitors have advantages and disadvantages, which are related to potential resistance mechanisms, pharmacokinetics, selectivity and tumor environment. In different malignancies various tyrosine kinases are mutated or overexpressed and several resistance mechanisms exist. Pharmacokinetics is influenced by interindividual differences and differs for two single targeted inhibitors or between patients treated by the same tyrosine kinase inhibitor. Different tyrosine kinase inhibitors have various mechanisms to achieve selectivity, while differences in gene expression exist between tumor and stromal cells. Considering these aspects, one type of inhibitor can generally not be preferred above the other, but will depend on the specific genetic constitution of the patient and the tumor, allowing personalized therapy. The most effective way of cancer treatment by using tyrosine kinase inhibitors is to consider each patient/tumor individually and to determine the strategy that specifically targets the consequences of altered (epi)genetics of the tumor. This strategy might result in treatment by a single multi kinase inhibitor for one patient, but in treatment by a couple of single kinase inhibitors for other patients.

  16. Mig6 Puts the Brakes on Mutant EGFR-Driven Lung Cancer | Center for Cancer Research

    Science.gov (United States)

    Lung cancer is the most common cause of cancer-related death worldwide. These cancers are often induced by mutations in the epidermal growth factor receptor (EGFR), resulting in constitutive activation of the protein’s tyrosine kinase domain. Lung cancers expressing these EGFR mutants are initially sensitive to tyrosine kinase inhibitors (TKIs), such as erlotinib, but often become resistant by developing compensatory mutations in EGFR or other growth-promoting pathways. To better understand how mutant EGFR initiates and maintains tumor growth in the hopes of identifying novel targets for drug development, Udayan Guha, M.D., Ph.D., of CCR’s Thoracic and Gastrointestinal Oncology Branch, and his colleagues examined the landscape of proteins phosphorylated in EGFR wild type and mutant cells. One protein hyper-phosphorylated in mutant EGFR cells was Mig6, a putative tumor suppressor.

  17. Dialkoxyquinazolines: Screening Epidermal Growth Factor ReceptorTyrosine Kinase Inhibitors for Potential Tumor Imaging Probes

    Energy Technology Data Exchange (ETDEWEB)

    VanBrocklin, Henry F.; Lim, John K.; Coffing, Stephanie L.; Hom,Darren L.; Negash, Kitaw; Ono, Michele Y.; Hanrahan, Stephen M.; Taylor,Scott E.; Vanderpoel, Jennifer L.; Slavik, Sarah M.; Morris, Andrew B.; Riese II, David J.

    2005-09-01

    The epidermal growth factor receptor (EGFR), a long-standingdrug development target, is also a desirable target for imaging. Sixteendialkoxyquinazoline analogs, suitable for labeling with positron-emittingisotopes, have been synthesized and evaluated in a battery of in vitroassays to ascertain their chemical and biological properties. Thesecharacteristics provided the basis for the adoption of a selection schemato identify lead molecules for labeling and in vivo evaluation. A newEGFR tyrosine kinase radiometric binding assay revealed that all of thecompounds possessed suitable affinity (IC50 = 0.4 - 51 nM) for the EGFRtyrosine kinase. All of the analogs inhibited ligand-induced EGFRtyrosine phosphorylation (IC50 = 0.8 - 20 nM). The HPLC-estimatedoctanol/water partition coefficients ranged from 2.0-5.5. Four compounds,4-(2'-fluoroanilino)- and 4-(3'-fluoroanilino)-6,7-diethoxyquinazoline aswell as 4-(3'-chloroanilino)- and4-(3'-bromoanilino)-6,7-dimethoxyquinazoline, possess the bestcombination of characteristics that warrant radioisotope labeling andfurther evaluation in tumor-bearing mice.

  18. EGFR Mutation Status in Uighur Lung Adenocarcinoma Patients

    Directory of Open Access Journals (Sweden)

    Li SHAN

    2013-02-01

    Full Text Available Background and objective Epidermal growth factor receptor (EGFR, a transmembrane protein, is a member of the tyrosine kinase family. Gefitinib, an EGFR tyrosine-kinase inhibitors, has shown a high response rate in the treatment of lung cancer in patients with EGFR mutation. However, significant differences in EGFR mutations exist among different ethnic groups. The aim of this study is to investigate the prevalence of EGFR mutations in Uighur lung adenocarcinoma patients by using a rapid and sensitive detection method and to analyze EGFR mutation differences compared with Han lung adenocarcinoma patients. Methods We examined lung adenocarcinoma tissues from 138 patients, including 68 Uighur lung adenocarcinoma patients and 70 Han lung adenocarcinoma patients, for EGFR mutations in exons 18, 19, 20, and 21 by using the amplification refractory mutation system (ARMS PCR method. The mutation differences between Uighur and Han lung adenocarcinoma were compared by using the chi-square test method. Results EGFR mutations were detected in 43 (31.2% of the 138 lung adenocarcinoma patients. EGFR mutations were detected in 11 (16.2% of the 68 Uighur lung adenocarcinoma patients and in 32 (45.7% of the 70 Han lung adenocarcinoma patients. Significant differences were observed in the EGFR mutations between Uighur lung adenocarcinoma patients and Han lung adenocarcinoma patients (P<0.001. Conclusion Our results indicate that the EGFR mutation in Uighur lung adenocarcinoma patients (16.2% is significantly lower than that in Han lung adenocarcinoma patients (45.7%.

  19. Gefitinib-induced killing of NSCLC cell lines expressing mutant EGFR requires BIM and can be enhanced by BH3 mimetics.

    Directory of Open Access Journals (Sweden)

    Mark S Cragg

    2007-10-01

    Full Text Available The epidermal growth factor receptor (EGFR plays a critical role in the control of cellular proliferation, differentiation, and survival. Abnormalities in EGF-EGFR signaling, such as mutations that render the EGFR hyperactive or cause overexpression of the wild-type receptor, have been found in a broad range of cancers, including carcinomas of the lung, breast, and colon. EGFR inhibitors such as gefitinib have proven successful in the treatment of certain cancers, particularly non-small cell lung cancers (NSCLCs harboring activating mutations within the EGFR gene, but the molecular mechanisms leading to tumor regression remain unknown. Therefore, we wished to delineate these mechanisms.We performed biochemical and genetic studies to investigate the mechanisms by which inhibitors of EGFR tyrosine kinase activity, such as gefitinib, inhibit the growth of human NSCLCs. We found that gefitinib triggered intrinsic (also called "mitochondrial" apoptosis signaling, involving the activation of BAX and mitochondrial release of cytochrome c, ultimately unleashing the caspase cascade. Gefitinib caused a rapid increase in the level of the proapoptotic BH3-only protein BIM (also called BCL2-like 11 through both transcriptional and post-translational mechanisms. Experiments with pharmacological inhibitors indicated that blockade of MEK-ERK1/2 (mitogen-activated protein kinase kinase-extracellular signal-regulated protein kinase 1/2 signaling, but not blockade of PI3K (phosphatidylinositol 3-kinase, JNK (c-Jun N-terminal kinase or mitogen-activated protein kinase 8, or AKT (protein kinase B, was critical for BIM activation. Using RNA interference, we demonstrated that BIM is essential for gefitinib-induced killing of NSCLC cells. Moreover, we found that gefitinib-induced apoptosis is enhanced by addition of the BH3 mimetic ABT-737.Inhibitors of the EGFR tyrosine kinase have proven useful in the therapy of certain cancers, in particular NSCLCs possessing

  20. Normalization of TAM post-receptor signaling reveals a cell invasive signature for Axl tyrosine kinase.

    Science.gov (United States)

    Kimani, Stanley G; Kumar, Sushil; Davra, Viralkumar; Chang, Yun-Juan; Kasikara, Canan; Geng, Ke; Tsou, Wen-I; Wang, Shenyan; Hoque, Mainul; Boháč, Andrej; Lewis-Antes, Anita; De Lorenzo, Mariana S; Kotenko, Sergei V; Birge, Raymond B

    2016-09-06

    Tyro3, Axl, and Mertk (TAMs) are a family of three conserved receptor tyrosine kinases that have pleiotropic roles in innate immunity and homeostasis and when overexpressed in cancer cells can drive tumorigenesis. In the present study, we engineered EGFR/TAM chimeric receptors (EGFR/Tyro3, EGFR/Axl, and EGF/Mertk) with the goals to interrogate post-receptor functions of TAMs, and query whether TAMs have unique or overlapping post-receptor activation profiles. Stable expression of EGFR/TAMs in EGFR-deficient CHO cells afforded robust EGF inducible TAM receptor phosphorylation and activation of downstream signaling. Using a series of unbiased screening approaches, that include kinome-view analysis, phosphor-arrays, RNAseq/GSEA analysis, as well as cell biological and in vivo readouts, we provide evidence that each TAM has unique post-receptor signaling platforms and identify an intrinsic role for Axl that impinges on cell motility and invasion compared to Tyro3 and Mertk. These studies demonstrate that TAM show unique post-receptor signatures that impinge on distinct gene expression profiles and tumorigenic outcomes.

  1. Genistein and tyrphostin AG556 decrease ultra-rapidly activating delayed rectifier K+ current of human atria by inhibiting EGF receptor tyrosine kinase.

    Science.gov (United States)

    Xiao, Guo-Sheng; Zhang, Yan-Hui; Wu, Wei; Sun, Hai-Ying; Wang, Yan; Li, Gui-Rong

    2017-03-01

    The ultra-rapidly activating delayed rectifier K + current I Kur (encoded by K v 1.5 or KCNA5) plays an important role in human atrial repolarization. The present study investigates the regulation of this current by protein tyrosine kinases (PTKs). Whole-cell patch voltage clamp technique and immunoprecipitation and Western blotting analysis were used to investigate whether the PTK inhibitors genistein, tyrphostin AG556 (AG556) and PP2 regulate human atrial I Kur and hKv1.5 channels stably expressed in HEK 293 cells. Human atrial I Kur was decreased by genistein (a broad-spectrum PTK inhibitor) and AG556 (a highly selective EGFR TK inhibitor) in a concentration-dependent manner. Inhibition of I Kur induced by 30 μM genistein or 10 μM AG556 was significantly reversed by 1 mM orthovanadate (a protein tyrosine phosphatase inhibitor). Similar results were observed in HEK 293 cells stably expressing hK v 1.5 channels. On the other hand, the Src family kinase inhibitor PP2 (1 μM) slightly enhanced I Kur and hK v 1.5 current, and the current increase was also reversed by orthovanadate. Immunoprecipitation and Western blotting analysis showed that genistein, AG556, and PP2 decreased tyrosine phosphorylation of hK v 1.5 channels and that the decrease was countered by orthovanadate. The PTK inhibitors genistein and AG556 decrease human atrial I Kur and cloned hK v 1.5 channels by inhibiting EGFR TK, whereas the Src kinase inhibitor PP2 increases I Kur and hK v 1.5 current. These results imply that EGFR TK and the soluble Src kinases may have opposite effects on human atrial I Kur . © 2017 The British Pharmacological Society.

  2. Cancer Cell-derived Exosomes Induce Mitogen-activated Protein Kinase-dependent Monocyte Survival by Transport of Functional Receptor Tyrosine Kinases*

    Science.gov (United States)

    Song, Xiao; Ding, Yanping; Liu, Gang; Yang, Xiao; Zhao, Ruifang; Zhang, Yinlong; Zhao, Xiao; Anderson, Gregory J.; Nie, Guangjun

    2016-01-01

    Tumor-associated macrophages (TAM) play pivotal roles in cancer initiation and progression. Monocytes, the precursors of TAMs, normally undergo spontaneous apoptosis within 2 days, but can subsist in the inflammatory tumor microenvironment for continuous survival and generation of sufficient TAMs. The mechanisms underlying tumor-driving monocyte survival remain obscure. Here we report that cancer cell-derived exosomes were crucial mediators for monocyte survival in the inflammatory niche. Analysis of the survival-promoting molecules in monocytes revealed that cancer cell-derived exosomes activated Ras and extracellular signal-regulated kinases in the mitogen-activated protein kinase (MAPK) pathway, resulting in the prevention of caspase cleavage. Phosphorylated receptor tyrosine kinases (RTKs), such as phosphorylated epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER-2), were abundantly expressed in cancer cell-derived exosomes. Knock-out of EGFR or/and HER-2, or alternatively, inhibitors against their phosphorylation significantly disturbed the exosome-mediated activation of the MAPK pathway, inhibition of caspase cleavage, and increase in survival rate in monocytes. Moreover, the deprived survival-stimulating activity of exosomes due to null expression of EGFR and HER-2 could be restored by activation of another RTK, insulin receptor. Overall, our study uncovered a mechanism of tumor-associated monocyte survival and demonstrated that cancer cell-derived exosomes can stimulate the MAPK pathway in monocytes through transport of functional RTKs, leading to inactivation of apoptosis-related caspases. This work provides insights into the long sought question on monocyte survival prior to formation of plentiful TAMs in the tumor microenvironment. PMID:26895960

  3. Ethacrynic acid improves the antitumor effects of irreversible epidermal growth factor receptor tyrosine kinase inhibitors in breast cancer.

    Science.gov (United States)

    Liu, Bing; Huang, XinPing; Hu, YunLong; Chen, TingTing; Peng, BoYa; Gao, NingNing; Jin, ZhenChao; Jia, TieLiu; Zhang, Na; Wang, ZhuLin; Jin, GuangYi

    2016-09-06

    Prolonged treatment of breast cancer with epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) often results in acquired resistance and a narrow therapeutic index. One strategy to improve the therapeutic effects of EGFR TKIs is to combine them with drugs used for other clinical indications. Ethacrynic acid (EA) is an FDA approved drug that may have antitumor effects and may enhance the cytotoxicity of chemotherapeutic agents by binding to glutathione and inhibiting WNT signaling. While the α,β-unsaturated-keto structure of EA is similar to that of irreversible TKIs, the mechanism of action of EA when combined with irreversible EGFR TKIs in breast cancer remains unknown. We therefore investigated the combination of irreversible EGFR TKIs and EA. We found that irreversible EGFR TKIs and EA synergistically inhibit breast cancer both in vitro and in vivo. The combination of EGFR TKIs and EA induces necrosis and cell cycle arrest and represses WNT/β-catenin signaling as well as MAPK-ERK1/2 signaling. We conclude that EA synergistically enhances the antitumor effects of irreversible EGFR TKIs in breast cancer.

  4. Alteration of radiation response by two tyrosine kinase inhibitors: STI571 (Glivec) and BIBW 2992

    International Nuclear Information System (INIS)

    Huguet, F.

    2010-01-01

    Concurrent chemo-radiation is one of the main weapon in the treatment of cancer. The targeted therapies may act on the mechanisms of tumor resistance to radiation and are therefore very promising in combination with radiotherapy. The STI571 (imatinib or Gleevec) inhibits specifically the Bcr-Abl tyrosine kinase. It leads to radiosensitization in K562 chronic myeloid leukemia cell line by alterations of the cell cycle. The BIBW2992 is a selective inhibitor of EGFR and HER2. The BIBW 2992 shows cytotoxic and radiosensitizing effects on pancreatic adenocarcinoma cells BxPC3 and Capan-2, regardless of KRAS status. The mechanism underlying this radiosensitization is not unequivocal, involving both changes in the cell cycle and induction of mitotic death. Our results show that the combination of an inhibitor of tyrosine kinase with ionizing radiation may lead to a radiosensitization in vitro with mechanisms depending on the type of cell line. (author)

  5. Differential role of EGF and BFGF in human GBM-TIC proliferation: relationship to EGFR-tyrosine kinase inhibitor sensibility.

    Science.gov (United States)

    Bajetto, A; Porcile, C; Pattarozzi, A; Scotti, L; Aceto, A; Daga, A; Barbieri, F; Florio, T

    2013-01-01

    Glioblastoma multiforme (GBM) is among the most devastating human tumors being rapidly fatal despite aggressive surgery, radiation and chemotherapies. It is characterized by extensive dissemination of tumor cells within the brain that hinders complete surgical resection. GBM tumor initiating-cells (TICs) are a rare subpopulation of cells responsible for tumor development, growth, invasiveness and recurrence after chemotherapy. TICs from human GBM can be selected in vitro using the same conditions permissive for the growth of normal neural cells, of which share some features including marker expression, self-renewal capacity, long-term proliferation, and ability to differentiate into neuronal and glial cells. EGFR overexpression and its constitutive activation is one of the most important signaling alteration identified in GBM, and its pharmacological targeting represents an attractive therapeutic goal. We previously demonstrated that human GBM TICs have different sensitivity to the EGFR kinase inhibitors erlotinib and gefitinib, depending on the differential modulation of downstream signaling cascades. In this work we investigated the mechanisms of resistance to erlotinib in two human GBM TIC cultures, analyzing EGF and bFGF individual contribution to proliferation, clonogenicity, and migration. We demonstrated the presence of a small cell subpopulation whose proliferation is supported by EGF and a larger one mainly dependent on bFGF. Thus, insensitivity to EGFR kinase inhibitors as far as TIC proliferation results from a predominant FGFR activation that hides the inhibitory effects induced on EGFR signaling. Conversely, EGF and bFGF induced cell migration with similar efficacy. In addition, unlike neural stem/progenitors cells, the removal of chondroitin sulphate proteoglycans from cell surface was unable to discern EGF- and bFGF-dependent subpopulations in GBM TICs.

  6. Src mediates cigarette smoke-induced resistance to tyrosine kinase inhibitors in NSCLC cells.

    Science.gov (United States)

    Filosto, Simone; Baston, David S; Chung, Samuel; Becker, Cathleen R; Goldkorn, Tzipora

    2013-08-01

    The EGF receptor (EGFR) is a proto-oncogene commonly dysregulated in several cancers including non-small cell lung carcinoma (NSCLC) and, thus, is targeted for treatment using tyrosine kinase inhibitors (TKI) such as erlotinib. However, despite the efficacy observed in patients with NSCLC harboring oncogenic variants of the EGFR, general ineffectiveness of TKIs in patients with NSCLC who are current and former smokers necessitates identification of novel mechanisms to overcome this phenomenon. Previously, we showed that NSCLC cells harboring either wild-type (WT) EGFR or oncogenic mutant (MT) L858R EGFR become resistant to the effects of TKIs when exposed to cigarette smoke, evidenced by their autophosphorylation and prolonged downstream signaling. Here, we present Src as a target mediating cigarette smoke-induced resistance to TKIs in both WT EGFR- and L858R MT EGFR-expressing NSCLC cells. First, we show that cigarette smoke exposure of A549 cells leads to time-dependent activation of Src, which then abnormally binds to the WT EGFR causing TKI resistance, contrasting previous observations of constitutive binding between inactive Src and TKI-sensitive L858R MT EGFR. Next, we show that Src inhibition restores TKI sensitivity in cigarette smoke-exposed NSCLC cells, preventing EGFR autophosphorylation in the presence of erlotinib. Furthermore, we show that overexpression of a dominant-negative Src (Y527F/K295R) restores TKI sensitivity to A549 exposed to cigarette smoke. Importantly, the TKI resistance that emerges even in cigarette smoke-exposed L858R EGFR-expressing NSCLC cells could be eliminated with Src inhibition. Together, these findings offer new rationale for using Src inhibitors for treating TKI-resistant NSCLC commonly observed in smokers.

  7. Single-label kinase and phosphatase assays for tyrosine phosphorylation using nanosecond time-resolved fluorescence detection.

    Science.gov (United States)

    Sahoo, Harekrushna; Hennig, Andreas; Florea, Mara; Roth, Doris; Enderle, Thilo; Nau, Werner M

    2007-12-26

    The collision-induced fluorescence quenching of a 2,3-diazabicyclo[2.2.2]oct-2-ene-labeled asparagine (Dbo) by hydrogen atom abstraction from the tyrosine residue in peptide substrates was introduced as a single-labeling strategy to assay the activity of tyrosine kinases and phosphatases. The assays were tested for 12 different combinations of Dbo-labeled substrates and with the enzymes p60c-Src Src kinase, EGFR kinase, YOP protein tyrosine phosphatase, as well as acid and alkaline phosphatases, thereby demonstrating a broad application potential. The steady-state fluorescence changed by a factor of up to 7 in the course of the enzymatic reaction, which allowed for a sufficient sensitivity of continuous monitoring in steady-state experiments. The fluorescence lifetimes (and intensities) were found to be rather constant for the phosphotyrosine peptides (ca. 300 ns in aerated water), while those of the unphosphorylated peptides were as short as 40 ns (at pH 7) and 7 ns (at pH 13) as a result of intramolecular quenching. Owing to the exceptionally long fluorescence lifetime of Dbo, the assays were alternatively performed by using nanosecond time-resolved fluorescence (Nano-TRF) detection, which leads to an improved discrimination of background fluorescence and an increased sensitivity. The potential for inhibitor screening was demonstrated through the inhibition of acid and alkaline phosphatases by molybdate.

  8. Cancer Cell-derived Exosomes Induce Mitogen-activated Protein Kinase-dependent Monocyte Survival by Transport of Functional Receptor Tyrosine Kinases.

    Science.gov (United States)

    Song, Xiao; Ding, Yanping; Liu, Gang; Yang, Xiao; Zhao, Ruifang; Zhang, Yinlong; Zhao, Xiao; Anderson, Gregory J; Nie, Guangjun

    2016-04-15

    Tumor-associated macrophages (TAM) play pivotal roles in cancer initiation and progression. Monocytes, the precursors of TAMs, normally undergo spontaneous apoptosis within 2 days, but can subsist in the inflammatory tumor microenvironment for continuous survival and generation of sufficient TAMs. The mechanisms underlying tumor-driving monocyte survival remain obscure. Here we report that cancer cell-derived exosomes were crucial mediators for monocyte survival in the inflammatory niche. Analysis of the survival-promoting molecules in monocytes revealed that cancer cell-derived exosomes activated Ras and extracellular signal-regulated kinases in the mitogen-activated protein kinase (MAPK) pathway, resulting in the prevention of caspase cleavage. Phosphorylated receptor tyrosine kinases (RTKs), such as phosphorylated epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER-2), were abundantly expressed in cancer cell-derived exosomes. Knock-out of EGFR or/and HER-2, or alternatively, inhibitors against their phosphorylation significantly disturbed the exosome-mediated activation of the MAPK pathway, inhibition of caspase cleavage, and increase in survival rate in monocytes. Moreover, the deprived survival-stimulating activity of exosomes due to null expression of EGFR and HER-2 could be restored by activation of another RTK, insulin receptor. Overall, our study uncovered a mechanism of tumor-associated monocyte survival and demonstrated that cancer cell-derived exosomes can stimulate the MAPK pathway in monocytes through transport of functional RTKs, leading to inactivation of apoptosis-related caspases. This work provides insights into the long sought question on monocyte survival prior to formation of plentiful TAMs in the tumor microenvironment. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. Non small-cell lung cancer and treatment options after tyrosine kinase inhibitors failure in the first line

    International Nuclear Information System (INIS)

    Chowaniecova, G.

    2014-01-01

    Introduction: Advanced non-small cell lung cancer with present epidermal growth factor receptor (EGFR) sensitising mutation is standardly treated with tyrosine kinase inhibitors (TKI). During treatment a resistance to TKI develops, disease progresses. We differ primary and secondary resistance. The most effective treatment after TKI failure is not definitively proven. Standard chemotherapy is usually introduced, eventually it is possible to use other TKI in the next lines. Case: The author presents a case of 60-year old patient with lung adenocarcinoma with EGFR sensitising mutation, where primary resistance to TKI was observed. Chemotherapy after progression was introduced. Planned therapy with afatnib was not carried out due to deterioration of patient´s condition. Conclusion: Presented case of EGFR mutation-positive patient represents an example of not very frequent primary resistance to TKI. Mechanisms of primary resistance are not well understood. Treatment after first line TKI failure in non-small cell lung cancer with EGFR mutation represents a challenge for medical research. (author)

  10. Nuclear EGFR as a molecular target in cancer

    International Nuclear Information System (INIS)

    Brand, Toni M.; Iida, Mari; Luthar, Neha; Starr, Megan M.; Huppert, Evan J.; Wheeler, Deric L.

    2013-01-01

    The epidermal growth factor receptor (EGFR) has been one of the most targeted receptors in the field of oncology. While anti-EGFR inhibitors have demonstrated clinical success in specific cancers, most patients demonstrate either intrinsic or acquired resistance within one year of treatment. Many mechanisms of resistance to EGFR inhibitors have been identified, one of these being attributed to alternatively localized EGFR from the cell membrane into the cell’s nucleus. Inside the nucleus, EGFR functions as a co-transcription factor for several genes involved in cell proliferation and angiogenesis, and as a tyrosine kinase to activate and stabilize proliferating cell nuclear antigen and DNA dependent protein kinase. Nuclear localized EGFR is highly associated with disease progression, worse overall survival in numerous cancers, and enhanced resistance to radiation, chemotherapy, and the anti-EGFR therapies gefitinib and cetuximab. In this review the current knowledge of how nuclear EGFR enhances resistance to cancer therapeutics is discussed, in addition to highlighting ways to target nuclear EGFR as an anti-cancer strategy in the future

  11. Putative tyrosine kinases expressed in K-562 human leukemia cells

    International Nuclear Information System (INIS)

    Partanen, J.; Maekelae, T.P.; Lehvaeslaiho, H.; Alitalo, K.; Alitalo, R.

    1990-01-01

    Tyrosine phosphorylation is important in the transmission of growth and differentiation signals; known tyrosine kinases include several oncoproteins and growth factor receptors. Interestingly, some differentiated cell types, such as erythrocytes and platelets contain high amounts of phosphotyrosine. The authors analyzed tyrosine kinases expressed in the K-562 chronic myelogenous leukemia cell line, which has a bipotential erythroid and megakaryoblastoid differentiation capacity. Analysis of 359 polymerase chain reaction-amplified cDNA clones led to the identification of 14 different tyrosine kinase-related sequences (JTK1-14). Two of the clones (JTK2 and JTK4) represent unusual members of the fibroblast growth factor receptor gene family, and the clones JTK5, JTK11, and JTK14 may also belong to the family of receptor tyrosine kinases but lack a close relationship to any known tyrosine kinase. Each of these different genes has its own characteristic expression pattern in K-562 cells and several other human tumor cell lines. In addition, the JTK11 and JTK14 mRNAs are induced during the megakaryoblastoid differentiation of K-562 cells. These tyrosine kinases may have a role in the differentiation of megakaryoblasts or in the physiology of platelets

  12. Epidermal Growth Factor Receptor Mutation (EGFR) Testing for Prediction of Response to EGFR-Targeting Tyrosine Kinase Inhibitor (TKI) Drugs in Patients with Advanced Non-Small-Cell Lung Cancer: An Evidence-Based Analysis.

    Science.gov (United States)

    2010-01-01

    In February 2010, the Medical Advisory Secretariat (MAS) began work on evidence-based reviews of the literature surrounding three pharmacogenomic tests. This project came about when Cancer Care Ontario (CCO) asked MAS to provide evidence-based analyses on the effectiveness and cost-effectiveness of three oncology pharmacogenomic tests currently in use in Ontario.Evidence-based analyses have been prepared for each of these technologies. These have been completed in conjunction with internal and external stakeholders, including a Provincial Expert Panel on Pharmacogenetics (PEPP). Within the PEPP, subgroup committees were developed for each disease area. For each technology, an economic analysis was also completed by the Toronto Health Economics and Technology Assessment Collaborative (THETA) and is summarized within the reports.THE FOLLOWING REPORTS CAN BE PUBLICLY ACCESSED AT THE MAS WEBSITE AT: http://www.health.gov.on.ca/mas or at www.health.gov.on.ca/english/providers/program/mas/mas_about.htmlGENE EXPRESSION PROFILING FOR GUIDING ADJUVANT CHEMOTHERAPY DECISIONS IN WOMEN WITH EARLY BREAST CANCER: An Evidence-Based AnalysisEpidermal Growth Factor Receptor Mutation (EGFR) Testing for Prediction of Response to EGFR-Targeting Tyrosine Kinase Inhibitor (TKI) Drugs in Patients with Advanced Non-Small-Cell Lung Cancer: an Evidence-Based AnalysisK-RAS testing in Treatment Decisions for Advanced Colorectal Cancer: an Evidence-Based Analysis The Medical Advisory Secretariat undertook a systematic review of the evidence on the clinical effectiveness and cost-effectiveness of epidermal growth factor receptor (EGFR) mutation testing compared with no EGFR mutation testing to predict response to tyrosine kinase inhibitors (TKIs), gefitinib (Iressa(®)) or erlotinib (Tarceva(®)) in patients with advanced non-small cell lung cancer (NSCLC). TARGET POPULATION AND CONDITION With an estimated 7,800 new cases and 7,000 deaths last year, lung cancer is the leading cause of cancer

  13. Src protein-tyrosine kinase structure and regulation

    International Nuclear Information System (INIS)

    Roskoski, Robert

    2004-01-01

    Src and Src-family protein kinases are proto-oncogenes that play key roles in cell morphology, motility, proliferation, and survival. v-Src (a viral protein) is encoded by the chicken oncogene of Rous sarcoma virus, and Src (the cellular homologue) is encoded by a physiological gene, the first of the proto-oncogenes. From the N- to C-terminus, Src contains an N-terminal 14-carbon myristoyl group, a unique segment, an SH3 domain, an SH2 domain, a protein-tyrosine kinase domain, and a C-terminal regulatory tail. The chief phosphorylation sites of Src include tyrosine 416 that results in activation from autophosphorylation and tyrosine 527 that results in inhibition from phosphorylation by C-terminal Src kinase. In the restrained state, the SH2 domain forms a salt bridge with phosphotyrosine 527, and the SH3 domain binds to the kinase domain via a polyproline type II left-handed helix. The SH2 and SH3 domains occur on the backside of the kinase domain away from the active site where they stabilize a dormant enzyme conformation. Protein-tyrosine phosphatases such as PTPα displace phosphotyrosine 527 from the Src SH2 domain and mediate its dephosphorylation leading to Src kinase activation. C-terminal Src kinase consists of an SH3, SH2, and kinase domain; it lacks an N-terminal myristoyl group and a C-terminal regulatory tail. Its X-ray structure has been determined, and the SH2 lobe occupies a position that is entirely different from that of Src. Unlike Src, the C-terminal Src kinase SH2 and SH3 domains stabilize an active enzyme conformation. Amino acid residues in the αD helix near the catalytic loop in the large lobe of C-terminal Src kinase serve as a docking site for the physiological substrate (Src) but not for an artificial substrate (polyGlu 4 Tyr)

  14. Ror receptor tyrosine kinases: orphans no more

    OpenAIRE

    Green, Jennifer L.; Kuntz, Steven G.; Sternberg, Paul W.

    2008-01-01

    Receptor tyrosine kinase-like orphan receptor (Ror) proteins are a conserved family of tyrosine kinase receptors that function in developmental processes including skeletal and neuronal development, cell movement and cell polarity. Although Ror proteins were originally named because the associated ligand and signaling pathway were unknown, recent studies in multiple species have now established that Ror proteins are Wnt receptors. Depending on the cellular context, Ror proteins can either act...

  15. Personalized Radiation Oncology: Epidermal Growth Factor Receptor and Other Receptor Tyrosine Kinase Inhibitors.

    Science.gov (United States)

    Higgins, Geoff S; Krause, Mechthild; McKenna, W Gillies; Baumann, Michael

    Molecular biomarkers are currently evaluated in preclinical and clinical studies in order to establish predictors for treatment decisions in radiation oncology. The receptor tyrosine kinases (RTK) are described in the following text. Among them, the most data are available for the epidermal growth factor receptor (EGFR) that plays a major role for prognosis of patients after radiotherapy, but seems also to be involved in mechanisms of radioresistance, specifically in repopulation of tumour cells between radiotherapy fractions. Monoclonal antibodies against the EGFR improve locoregional tumour control and survival when applied during radiotherapy, however, the effects are heterogeneous and biomarkers for patient selection are warranted. Also other RTK´s such as c-Met and IGF-1R seem to play important roles in tumour radioresistance. Beside the potential to select patients for molecular targeting approaches combined with radiotherapy, studies are also needed to evluate radiotherapy adaptation approaches for selected patients, i.e. adaptation of radiation dose, or, more sophisticated, of target volumes.

  16. Acquired MET expression confers resistance to EGFR inhibition in a mouse model of glioblastoma multiforme.

    Science.gov (United States)

    Jun, H J; Acquaviva, J; Chi, D; Lessard, J; Zhu, H; Woolfenden, S; Bronson, R T; Pfannl, R; White, F; Housman, D E; Iyer, L; Whittaker, C A; Boskovitz, A; Raval, A; Charest, A

    2012-06-21

    Glioblastoma multiforme (GBM) is an aggressive brain tumor for which there is no cure. Overexpression of wild-type epidermal growth factor receptor (EGFR) and loss of the tumor suppressor genes Ink4a/Arf and PTEN are salient features of this deadly cancer. Surprisingly, targeted inhibition of EGFR has been clinically disappointing, demonstrating an innate ability for GBM to develop resistance. Efforts at modeling GBM in mice using wild-type EGFR have proven unsuccessful to date, hampering endeavors at understanding molecular mechanisms of therapeutic resistance. Here, we describe a unique genetically engineered mouse model of EGFR-driven gliomagenesis that uses a somatic conditional overexpression and chronic activation of wild-type EGFR in cooperation with deletions in the Ink4a/Arf and PTEN genes in adult brains. Using this model, we establish that chronic activation of wild-type EGFR with a ligand is necessary for generating tumors with histopathological and molecular characteristics of GBMs. We show that these GBMs are resistant to EGFR kinase inhibition and we define this resistance molecularly. Inhibition of EGFR kinase activity using tyrosine kinase inhibitors in GBM tumor cells generates a cytostatic response characterized by a cell cycle arrest, which is accompanied by a substantial change in global gene expression levels. We demonstrate that an important component of this pattern is the transcriptional activation of the MET receptor tyrosine kinase and that pharmacological inhibition of MET overcomes the resistance to EGFR inhibition in these cells. These findings provide important new insights into mechanisms of resistance to EGFR inhibition and suggest that inhibition of multiple targets will be necessary to provide therapeutic benefit for GBM patients.

  17. Constitutive Activity in an Ancestral Form of Abl Tyrosine Kinase.

    Directory of Open Access Journals (Sweden)

    Saadat U Aleem

    Full Text Available The c-abl proto-oncogene encodes a nonreceptor tyrosine kinase that is found in all metazoans, and is ubiquitously expressed in mammalian tissues. The Abl tyrosine kinase plays important roles in the regulation of mammalian cell physiology. Abl-like kinases have been identified in the genomes of unicellular choanoflagellates, the closest relatives to the Metazoa, and in related unicellular organisms. Here, we have carried out the first characterization of a premetazoan Abl kinase, MbAbl2, from the choanoflagellate Monosiga brevicollis. The enzyme possesses SH3, SH2, and kinase domains in a similar arrangement to its mammalian counterparts, and is an active tyrosine kinase. MbAbl2 lacks the N-terminal myristoylation and cap sequences that are critical regulators of mammalian Abl kinase activity, and we show that MbAbl2 is constitutively active. When expressed in mammalian cells, MbAbl2 strongly phosphorylates cellular proteins on tyrosine, and transforms cells much more potently than mammalian Abl kinase. Thus, MbAbl2 appears to lack the autoinhibitory mechanism that tightly constrains the activity of mammalian Abl kinases, suggesting that this regulatory apparatus arose more recently in metazoan evolution.

  18. Epidermal growth factor receptor activation by diesel particles is mediated by tyrosine phosphatase inhibition

    International Nuclear Information System (INIS)

    Tal, Tamara L.; Bromberg, Philip A.; Kim, Yumee; Samet, James M.

    2008-01-01

    Exposure to particulate matter (PM) is associated with increased cardiopulmonary morbidity and mortality. Diesel exhaust particles (DEP) are a major component of ambient PM and may contribute to PM-induced pulmonary inflammation. Proinflammatory signaling is mediated by phosphorylation-dependent signaling pathways whose activation is opposed by the activity of protein tyrosine phosphatases (PTPases) which thereby function to maintain signaling quiescence. PTPases contain an invariant catalytic cysteine that is susceptible to electrophilic attack. DEP contain electrophilic oxy-organic compounds that may contribute to the oxidant effects of PM. Therefore, we hypothesized that exposure to DEP impairs PTPase activity allowing for unopposed basal kinase activity. Here we report that exposure to 30 μg/cm 2 DEP for 4 h induces differential activation of signaling in primary cultures of human airway epithelial cells (HAEC), a primary target cell in PM inhalation. In-gel kinase activity assay of HAEC exposed to DEPs of low (L-DEP), intermediate (I-DEP) or high (H-DEP) organic content showed differential activation of intracellular kinases. Exposure to these DEP also induced varying levels of phosphorylation of the receptor tyrosine kinase EGFR in a manner that requires EGFR kinase activity but does not involve receptor dimerization. We demonstrate that treatment with DEP results in an impairment of total and EGFR-directed PTPase activity in HAEC with a potency that is independent of the organic content of these particles. These data show that DEP-induced EGFR phosphorylation in HAEC is the result of a loss of PTPase activities which normally function to dephosphorylate EGFR in opposition to baseline EGFR kinase activity

  19. Importance of tyrosine phosphorylation in receptor kinase complexes.

    Science.gov (United States)

    Macho, Alberto P; Lozano-Durán, Rosa; Zipfel, Cyril

    2015-05-01

    Tyrosine phosphorylation is an important post-translational modification that is known to regulate receptor kinase (RK)-mediated signaling in animals. Plant RKs are annotated as serine/threonine kinases, but recent work has revealed that tyrosine phosphorylation is also crucial for the activation of RK-mediated signaling in plants. These initial observations have paved the way for subsequent detailed studies on the mechanism of activation of plant RKs and the biological relevance of tyrosine phosphorylation for plant growth and immunity. In this Opinion article we review recent reports on the contribution of RK tyrosine phosphorylation in plant growth and immunity; we propose that tyrosine phosphorylation plays a major regulatory role in the initiation and transduction of RK-mediated signaling in plants. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Consensus for EGFR mutation testing in non-small cell lung cancer: results from a European workshop

    DEFF Research Database (Denmark)

    Pirker, Robert; Herth, Felix J F; Kerr, Keith M

    2010-01-01

    Activating somatic mutations of the tyrosine kinase domain of epidermal growth factor receptor (EGFR) have recently been characterized in a subset of patients with advanced non-small cell lung cancer (NSCLC). Patients harboring these mutations in their tumors show excellent response to EGFR tyros...

  1. Anti-EGFR Therapy: Mechanism and Advances in Clinical Efficacy in Breast Cancer

    Directory of Open Access Journals (Sweden)

    John F. Flynn

    2009-01-01

    Full Text Available This review will focus on recent advances in the application of antiepidermal growth factor receptor (anti-EGFR for the treatment of breast cancer. The choice of EGFR, a member of the ErbB tyrosine kinase receptor family, stems from evidence pinpointing its role in various anti-EGFR therapies. Therefore, an increase in our understanding of EGFR mechanism and signaling might reveal novel targets amenable to intervention in the clinic. This knowledge base might also improve existing medical treatment options and identify research gaps in the design of new therapeutic agents. While the approved use of drugs like the dual kinase inhibitor Lapatinib represents significant advances in the clinical management of breast cancer, confirmatory studies must be considered to foster the use of anti-EGFR therapies including safety, pharmacokinetics, and clinical efficacy.

  2. PTP1B-dependent regulation of receptor tyrosine kinase signaling by the actin-binding protein Mena.

    Science.gov (United States)

    Hughes, Shannon K; Oudin, Madeleine J; Tadros, Jenny; Neil, Jason; Del Rosario, Amanda; Joughin, Brian A; Ritsma, Laila; Wyckoff, Jeff; Vasile, Eliza; Eddy, Robert; Philippar, Ulrike; Lussiez, Alisha; Condeelis, John S; van Rheenen, Jacco; White, Forest; Lauffenburger, Douglas A; Gertler, Frank B

    2015-11-01

    During breast cancer progression, alternative mRNA splicing produces functionally distinct isoforms of Mena, an actin regulator with roles in cell migration and metastasis. Aggressive tumor cell subpopulations express Mena(INV), which promotes tumor cell invasion by potentiating EGF responses. However, the mechanism by which this occurs is unknown. Here we report that Mena associates constitutively with the tyrosine phosphatase PTP1B and mediates a novel negative feedback mechanism that attenuates receptor tyrosine kinase signaling. On EGF stimulation, complexes containing Mena and PTP1B are recruited to the EGFR, causing receptor dephosphorylation and leading to decreased motility responses. Mena also interacts with the 5' inositol phosphatase SHIP2, which is important for the recruitment of the Mena-PTP1B complex to the EGFR. When Mena(INV) is expressed, PTP1B recruitment to the EGFR is impaired, providing a mechanism for growth factor sensitization to EGF, as well as HGF and IGF, and increased resistance to EGFR and Met inhibitors in signaling and motility assays. In sum, we demonstrate that Mena plays an important role in regulating growth factor-induced signaling. Disruption of this attenuation by Mena(INV) sensitizes tumor cells to low-growth factor concentrations, thereby increasing the migration and invasion responses that contribute to aggressive, malignant cell phenotypes. © 2015 Hughes, Oudin, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  3. Co-activation of STAT3 and YES-Associated Protein 1 (YAP1) Pathway in EGFR-Mutant NSCLC

    DEFF Research Database (Denmark)

    Chaib, Imane; Karachaliou, Niki; Pilotto, Sara

    2017-01-01

    Background: The efficacy of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in EGFR-mutant non-small cell lung cancer (NSCLC) is limited by adaptive activation of cell survival signals. We hypothesized that both signal transducer and activator of transcription 3 (STAT3) ...

  4. Gefitinib: a pharmacoeconomic profile of its use in patients with Non Small Cell Lung Cancer EGFR+

    Directory of Open Access Journals (Sweden)

    Viola Sacchi

    2011-06-01

    Full Text Available Lung cancer is the most common form of cancer with the highest incidence worldwide. The mortality rates are highest in males and second highest in females, after breast cancer. The genetic predisposition to Non Small Cell Lung Cancer (NSCLC is still under investigation, however, studies have shown that the Epidermal Growth Factor Receptor (EGFR, a receptor tyrosine kinase is frequently over-expressed and activated to a phosphorylated state in NSCLC. The activity of EGFR in cancer cells results in the phosphorylation of downstream proteins that promote cell proliferation, invasion, metastasis, and inhibition of apoptosis. Targeting the EGFR pathway therefore constitutes a relevant strategy for cancer therapy. Gefitinib is a selective inhibitor of the EGFR tyrosine kinase and is indicated for the treatment of adult patients with locally advanced or metastatic NSCLC with activating mutations of EGFR-TK. From the pharmacoeconomic point of view gefitinib is dominant (more effective and less expensive compared to the alternatives. In conclusion, gefitinib is a treatment option for NSCLC tumors with a high clinical and economic value in the Italian setting.

  5. Interaction between EGFR and EphA2

    DEFF Research Database (Denmark)

    Larsen, Alice Bjerregaard

    2010-01-01

    Enhanced or altered epidermal growth factor receptor (EGFR) activity has been reported in many human cancers and several molecular targeting therapies has been developed. However, despite intense research, therapies targeting EGFR have shown conflicting results in clinical studies, indicating...... the involvement of other important molecular players. Several different EGFR mutations have been reported in cancer, one of which is the cancer specific type III EGFR deletion mutant (EGFRvIII, de2-7EGFR, ΔEGFR). In a global search for EGFR and EGFRvIII regulated genes we identified the receptor tyrosine kinase...... (RTK) EphA2. EphA2 belongs to the large Eph-receptor family, which has mainly been associated with neuronal development. More recently, expression of several Eph-receptors has been detected in many different cancer types. Elevated EphA2 expression has been reported in a broad range of human cancer...

  6. Interaction between EGFR and EphA2

    DEFF Research Database (Denmark)

    Larsen, Alice Bjerregaard

    2010-01-01

    Enhanced or altered epidermal growth factor receptor (EGFR) activity has been reported in many human cancers and several molecular targeting therapies has been developed. However, despite intense research, therapies targeting EGFR have shown conflicting results in clinical studies, indicating...... the involvement of other important molecular players. Several different EGFR mutations have been reported in cancer, one of which is the cancer specific type III EGFR deletion mutant (EGFRvIII, de2-7EGFR, ¿EGFR). In a global search for EGFR and EGFRvIII regulated genes we identified the receptor tyrosine kinase...... (RTK) EphA2. EphA2 belongs to the large Eph-receptor family, which has mainly been associated with neuronal development. More recently, expression of several Eph-receptors has been detected in many different cancer types. Elevated EphA2 expression has been reported in a broad range of human cancer...

  7. Drug Resistance to EGFR Inhibitors in Lung Cancer | Office of Cancer Genomics

    Science.gov (United States)

    The discovery of mutations in epidermal growth factor receptor (EGFR) has dramatically changed the treatment of patients with non-small-cell lung cancer (NSCLC), the leading cause of cancer deaths worldwide. EGFR-targeted therapies show considerable promise, but drug resistance has become a substantial issue. We reviewed the literature to provide an overview of the drug resistance to EGFR tyrosine kinase inhibitors (TKIs) in NSCLC. The mechanisms causing primary, acquired and persistent drug resistance to TKIs vary.

  8. SH2 domains: modulators of nonreceptor tyrosine kinase activity.

    Science.gov (United States)

    Filippakopoulos, Panagis; Müller, Susanne; Knapp, Stefan

    2009-12-01

    The Src homology 2 (SH2) domain is a sequence-specific phosphotyrosine-binding module present in many signaling molecules. In cytoplasmic tyrosine kinases, the SH2 domain is located N-terminally to the catalytic kinase domain (SH1) where it mediates cellular localization, substrate recruitment, and regulation of kinase activity. Initially, structural studies established a role of the SH2 domain stabilizing the inactive state of Src family members. However, biochemical characterization showed that the presence of the SH2 domain is frequently required for catalytic activity, suggesting a crucial function stabilizing the active state of many nonreceptor tyrosine kinases. Recently, the structure of the SH2-kinase domain of Fes revealed that the SH2 domain stabilizes the active kinase conformation by direct interactions with the regulatory helix alphaC. Stabilizing interactions between the SH2 and the kinase domains have also been observed in the structures of active Csk and Abl. Interestingly, mutations in the SH2 domain found in human disease can be explained by SH2 domain destabilization or incorrect positioning of the SH2. Here we summarize our understanding of mechanisms that lead to tyrosine kinase activation by direct interactions mediated by the SH2 domain and discuss how mutations in the SH2 domain trigger kinase inactivation.

  9. Characterizing tyrosine phosphorylation signaling in lung cancer using SH2 profiling.

    Directory of Open Access Journals (Sweden)

    Kazuya Machida

    2010-10-01

    Full Text Available Tyrosine kinases drive the proliferation and survival of many human cancers. Thus profiling the global state of tyrosine phosphorylation of a tumor is likely to provide a wealth of information that can be used to classify tumors for prognosis and prediction. However, the comprehensive analysis of tyrosine phosphorylation of large numbers of human cancer specimens is technically challenging using current methods.We used a phosphoproteomic method termed SH2 profiling to characterize the global state of phosphotyrosine (pTyr signaling in human lung cancer cell lines. This method quantifies the phosphorylated binding sites for SH2 domains, which are used by cells to respond to changes in pTyr during signaling. Cells could be grouped based on SH2 binding patterns, with some clusters correlated with EGF receptor (EGFR or K-RAS mutation status. Binding of specific SH2 domains, most prominently RAS pathway activators Grb2 and ShcA, correlated with EGFR mutation and sensitivity to the EGFR inhibitor erlotinib. SH2 binding patterns also reflected MET activation and could identify cells driven by multiple kinases. The pTyr responses of cells treated with kinase inhibitors provided evidence of distinct mechanisms of inhibition.This study illustrates the potential of modular protein domains and their proteomic binding profiles as powerful molecular diagnostic tools for tumor classification and biomarker identification.

  10. Solubilization and characterization of a novel tyrosine kinase from rat adipocytes

    International Nuclear Information System (INIS)

    Yagaloff, K.A.; Czech, M.P.

    1987-01-01

    The authors report the efficient solubilization and characterization of a Triton X-100 insoluble tyrosine kinase from rat adipocytes. Plasma membranes were prepared from rat epididymal fat pads and were solubilized in 1% Triton X-100. Following centrifugation, the pellet was solubilized for 15 min at 4 0 C using both ionic and non-ionic detergents. Tyrosine kinase activity was measured in the soluble and particulate fractions using the exogenous substrate poly(glu-tyr) in a TCA precipitation assay. Reactions were performed in 50mM Hepes, 10mM MgCl 2 and 100μM gamma[ 32 P]-ATP (10Ci/mmol) at 4 0 C with or without 1mg/ml of the polyaminoacid. Incorporation rates of 100 to 1000 pmol/min/mg were obtained, while endogenous [ 32 P] incorporation was typically less than 10% of that in the presence of poly(glu-tyr). More than 75% of the tyrosine kinase activity was recovered in the soluble supernatant using this assay methodology. The solubilized tyrosine kinase was found to require Mg 2+ or Mn 2+ but preferred Mg 2+ and was inhibited by high levels of Mn 2+ . Kinase activity was strongly inhibited by Ca 2+ (>50% at 1mM), NaCl (>50% at 250mM) and NH 4 SO 4 (>50% at 50mM) but was activated by 10μM heparin and 5mM dithiothreitol. These properties distinguish the solubilized tyrosine kinase from other cellular tyrosine kinases

  11. Cationic Polyamidoamine Dendrimers as Modulators of EGFR Signaling In Vitro and In Vivo.

    Directory of Open Access Journals (Sweden)

    Saghir Akhtar

    Full Text Available Cationic polyamidoamine (PAMAM dendrimers are branch-like spherical polymers being investigated for a variety of applications in nanomedicine including nucleic acid drug delivery. Emerging evidence suggests they exhibit intrinsic biological and toxicological effects but little is known of their interactions with signal transduction pathways. We previously showed that the activated (fragmented generation (G 6 PAMAM dendrimer, Superfect (SF, stimulated epidermal growth factor receptor (EGFR tyrosine kinase signaling-an important signaling cascade that regulates cell growth, survival and apoptosis- in cultured human embryonic kidney (HEK 293 cells. Here, we firstly studied the in vitro effects of Polyfect (PF, a non-activated (intact G6 PAMAM dendrimer, on EGFR tyrosine kinase signaling via extracellular-regulated kinase 1/2 (ERK1/2 and p38 mitogen-activated protein kinase (MAPK in cultured HEK 293 cells and then compared the in vivo effects of a single administration (10mg/kg i.p of PF or SF on EGFR signaling in the kidneys of normal and diabetic male Wistar rats. Polyfect exhibited a dose- and time-dependent inhibition of EGFR, ERK1/2 and p38 MAPK phosphorylation in HEK-293 cells similar to AG1478, a selective EGFR inhibitor. Administration of dendrimers to non-diabetic or diabetic animals for 24h showed that PF inhibited whereas SF stimulated EGFR phosphorylation in the kidneys of both sets of animals. PF-mediated inhibition of EGFR phosphorylation as well as SF or PF-mediated apoptosis in HEK 293 cells could be significantly reversed by co-treatment with antioxidants such as tempol implying that both these effects involved an oxidative stress-dependent mechanism. These results show for the first time that SF and PF PAMAM dendrimers can differentially modulate the important EGFR signal transduction pathway in vivo and may represent a novel class of EGFR modulators. These findings could have important clinical implications for the use of PAMAM

  12. Involvement of the N-terminal unique domain of Chk tyrosine kinase in Chk-induced tyrosine phosphorylation in the nucleus

    International Nuclear Information System (INIS)

    Nakayama, Yuji; Kawana, Akiko; Igarashi, Asae; Yamaguchi, Naoto

    2006-01-01

    Chk tyrosine kinase phosphorylates Src-family kinases and suppresses their kinase activity. We recently showed that Chk localizes to the nucleus as well as the cytoplasm and inhibits cell proliferation. In this study, we explored the role of the N-terminal unique domain of Chk in nuclear localization and Chk-induced tyrosine phosphorylation in the nucleus. In situ binding experiments showed that the N-terminal domain of Chk was associated with the nucleus and the nuclear matrix. The presence of the N-terminal domain of Chk led to a fourfold increase in cell population exhibiting Chk-induced tyrosine phosphorylation in the nucleus. Expression of Chk but not kinase-deficient Chk induced tyrosine phosphorylation of a variety of proteins ranging from 23 kDa to ∼200 kDa, especially in Triton X-100-insoluble fraction that included chromatin and the nuclear matrix. Intriguingly, in situ subnuclear fractionations revealed that Chk induced tyrosine phosphorylation of proteins that were associated with the nuclear matrix. These results suggest that various unidentified substrates of Chk, besides Src-family kinases, may be present in the nucleus. Thus, our findings indicate that the importance of the N-terminal domain to Chk-induced tyrosine phosphorylation in the nucleus, implicating that these nuclear tyrosine-phosphorylated proteins may contribute to inhibition of cell proliferation

  13. Acquired resistance mechanisms to tyrosine kinase inhibitors in lung cancer with activating epidermal growth factor receptor mutation--diversity, ductility, and destiny.

    Science.gov (United States)

    Suda, Kenichi; Mizuuchi, Hiroshi; Maehara, Yoshihiko; Mitsudomi, Tetsuya

    2012-12-01

    Lung cancers that harbor somatic activating mutations in the gene for the epidermal growth factor receptor (EGFR) depend on mutant EGFR for their proliferation and survival; therefore, lung cancer patients with EGFR mutations often dramatically respond to orally available EGFR tyrosine kinase inhibitors (TKIs). However, emergence of acquired resistance is virtually inevitable, thus limiting improvement in patient outcomes. To elucidate and overcome this acquired resistance, multidisciplinary basic and clinical investigational approaches have been applied, using in vitro cell line models or samples obtained from lung cancer patients treated with EGFR-TKIs. These efforts have revealed several acquired resistance mechanisms and candidates, including EGFR secondary mutations (T790M and other rare mutations), MET amplification, PTEN downregulation, CRKL amplification, high-level HGF expression, FAS-NFκB pathway activation, epithelial-mesenchymal transition, and conversion to small cell lung cancer. Interestingly, cancer cells harbor potential destiny and ductility together in acquiring resistance to EGFR-TKIs, as shown in in vitro acquired resistance models. Molecular mechanisms of "reversible EGFR-TKI tolerance" that occur in early phase EGFR-TKI exposure have been identified in cell line models. Furthermore, others have reported molecular markers that can predict response to EGFR-TKIs in clinical settings. Deeper understanding of acquired resistance mechanisms to EGFR-TKIs, followed by the development of molecular target drugs that can overcome the resistance, might turn this fatal disease into a chronic disorder.

  14. Resistance to EGF receptor inhibitors in glioblastoma mediated by phosphorylation of the PTEN tumor suppressor at tyrosine 240.

    Science.gov (United States)

    Fenton, Tim R; Nathanson, David; Ponte de Albuquerque, Claudio; Kuga, Daisuke; Iwanami, Akio; Dang, Julie; Yang, Huijun; Tanaka, Kazuhiro; Oba-Shinjo, Sueli Mieko; Uno, Miyuki; Inda, Maria del Mar; Wykosky, Jill; Bachoo, Robert M; James, C David; DePinho, Ronald A; Vandenberg, Scott R; Zhou, Huilin; Marie, Suely K N; Mischel, Paul S; Cavenee, Webster K; Furnari, Frank B

    2012-08-28

    Glioblastoma multiforme (GBM) is the most aggressive of the astrocytic malignancies and the most common intracranial tumor in adults. Although the epidermal growth factor receptor (EGFR) is overexpressed and/or mutated in at least 50% of GBM cases and is required for tumor maintenance in animal models, EGFR inhibitors have thus far failed to deliver significant responses in GBM patients. One inherent resistance mechanism in GBM is the coactivation of multiple receptor tyrosine kinases, which generates redundancy in activation of phosphoinositide-3'-kinase (PI3K) signaling. Here we demonstrate that the phosphatase and tensin homolog deleted on chromosome 10 (PTEN) tumor suppressor is frequently phosphorylated at a conserved tyrosine residue, Y240, in GBM clinical samples. Phosphorylation of Y240 is associated with shortened overall survival and resistance to EGFR inhibitor therapy in GBM patients and plays an active role in mediating resistance to EGFR inhibition in vitro. Y240 phosphorylation can be mediated by both fibroblast growth factor receptors and SRC family kinases (SFKs) but does not affect the ability of PTEN to antagonize PI3K signaling. These findings show that, in addition to genetic loss and mutation of PTEN, its modulation by tyrosine phosphorylation has important implications for the development and treatment of GBM.

  15. Apoptosis-related molecular differences for response to tyrosin kinase inhibitors in drug-sensitive and drug-resistant human bladder cancer cells

    Directory of Open Access Journals (Sweden)

    Jixia Li

    2013-01-01

    Full Text Available Context: The epidermal growth factor receptor (EGFR family is reportedly overexpressed in bladder cancer, and tyrosine kinaseinhibitors (TKIs have been suggested as treatment. Gefitinib is a selective inhibitor of the EGFR and lapatinib is a dual inhibitor of both the EGFR and HER2 (human EGFR type 2 receptor. Both compounds compete with the binding of adenosine triphosphate (ATP to the tyrosine kinase domain of the respective receptors to inhibit receptor autophosphorylation causing suppression of signal transduction. Unfortunately, resistance to these inhibitors is a major clinical problem. Aims: To compare the apoptosis signaling pathway(s induced by gefitinib and lapatinib, in UM-UC-5 (drug-sensitive and UM-UC-14 (drug-resistant bladder cancer cells and to identify molecular differences that might be useful predictors of their efficacy. Materials and Methods: Cell proliferation, cell cycle and apoptosis assay were used to detect the effect of TKIs on UM-UC-5 and UM-UC-14 cells. Molecular differences for response to TKIs were examined by protein array. Results: TKIs strongly inhibited cell proliferation and induced cell cycle G1 arrest and apoptosis in UM-UC-5 cells. Most notable apoptosis molecular differences included decreased claspin, trail, and survivin by TKIs in the sensitive cells. In contrast, TKIs had no effect on resistant cells. Conclusions: Claspin, trail, and survivin might be used to determine the sensitivity of bladder cancers to TKIs.

  16. Hierarchical modeling of activation mechanisms in the ABL and EGFR kinase domains: thermodynamic and mechanistic catalysts of kinase activation by cancer mutations.

    Directory of Open Access Journals (Sweden)

    Anshuman Dixit

    2009-08-01

    Full Text Available Structural and functional studies of the ABL and EGFR kinase domains have recently suggested a common mechanism of activation by cancer-causing mutations. However, dynamics and mechanistic aspects of kinase activation by cancer mutations that stimulate conformational transitions and thermodynamic stabilization of the constitutively active kinase form remain elusive. We present a large-scale computational investigation of activation mechanisms in the ABL and EGFR kinase domains by a panel of clinically important cancer mutants ABL-T315I, ABL-L387M, EGFR-T790M, and EGFR-L858R. We have also simulated the activating effect of the gatekeeper mutation on conformational dynamics and allosteric interactions in functional states of the ABL-SH2-SH3 regulatory complexes. A comprehensive analysis was conducted using a hierarchy of computational approaches that included homology modeling, molecular dynamics simulations, protein stability analysis, targeted molecular dynamics, and molecular docking. Collectively, the results of this study have revealed thermodynamic and mechanistic catalysts of kinase activation by major cancer-causing mutations in the ABL and EGFR kinase domains. By using multiple crystallographic states of ABL and EGFR, computer simulations have allowed one to map dynamics of conformational fluctuations and transitions in the normal (wild-type and oncogenic kinase forms. A proposed multi-stage mechanistic model of activation involves a series of cooperative transitions between different conformational states, including assembly of the hydrophobic spine, the formation of the Src-like intermediate structure, and a cooperative breakage and formation of characteristic salt bridges, which signify transition to the active kinase form. We suggest that molecular mechanisms of activation by cancer mutations could mimic the activation process of the normal kinase, yet exploiting conserved structural catalysts to accelerate a conformational transition

  17. Expression Profiling of Tyrosine Kinase Genes

    National Research Council Canada - National Science Library

    Weier, Heinz

    2000-01-01

    ... of these genes parallels the progression of tumors to a more malignant phenotype. We developed a DNA micro-array based screening system to monitor the level of expression of tyrosine kinase (tk...

  18. Efficacy of EGFR-TKI therapy in patients with brain metastases from ...

    African Journals Online (AJOL)

    Purpose: This meta-analysis aimed to systematically evaluate the efficacy of tyrosine kinase inhibitors of epidermal growth factor receptor (EGFR-TKIs) for patients with brain metastases (BM) from nonsmall- cell lung cancer (NSCLC), and to compare this treatment modality to chemotherapy or radiotherapy. Methods: ...

  19. A marked response to icotinib in a patient with large cell neuroendocrine carcinoma harboring an EGFR mutation: A case report.

    Science.gov (United States)

    Wang, Yuehong; Shen, Yi Hong; Ma, Shanni; Zhou, Jianying

    2015-09-01

    The present study reports the case of an 84-year-old male with primary pulmonary large cell neuroendocrine carcinoma (LCNEC) harboring an epidermal growth factor receptor (EGFR) gene mutation that exhibited a long-lasting response to the EGFR-tyrosine kinase inhibitor (EGFR-TKI) icotinib. The patient had an extensive smoking history, a poor performance status, and presented with an irregular mass in the middle lobe of the right lung on computed tomography (CT) and an enlarged left supraclavicular lymph node on physical examination. Right middle lobe bronchial brushing during fiberoptic bronchoscopy identified poorly-differentiated cancer cells. The left supraclavicular lymph node was biopsied and a diagnosis of metastatic LCNEC was determined. Furthermore, an EGFR exon 19 deletion was identified by DNA sequencing. Following diagnosis, icotinib was administered at a dose of 125 mg three times a day. Chest CT scans were performed after 1 month of treatment, which indicated that the tumor was in partial remission. This marked response to icotinib lasted for 8 months. Thus, the present case illustrates the possibility of identifying EGFR mutations in LCNEC and indicates that EGFR-tyrosine kinase inhibitors may be an alternative treatment strategy for patients with LCNEC harboring activating EGFR mutations.

  20. Neratinib, A Novel HER2-Targeted Tyrosine Kinase Inhibitor.

    Science.gov (United States)

    Tiwari, Shruti Rakesh; Mishra, Prasun; Abraham, Jame

    2016-10-01

    HER2 gene amplification and receptor overexpression is identified in 20% to 25% of human breast cancers. Use of targeted therapy for HER2-amplified breast cancer has led to improvements in disease-free and overall survival in this subset of patients. Neratinib is an oral pan HER inhibitor, that irreversibly inhibits the tyrosine kinase activity of epidermal growth factor receptor (EGFR or HER1), HER2, and HER4, which leads to reduced phosphorylation and activation of downstream signaling pathways. Neratinib is currently being tested in a number of clinical trials for its safety and efficacy in lung cancer, and colorectal, bladder, and breast cancers. In this review we discuss the available phase I, II, and III data for use of neratinib in the metastatic, adjuvant, neoadjuvant, and extended adjuvant settings along with the ongoing clinical trials of neratinib in breast cancer. We also elaborate on the side effect profile of this relatively new drug and provide guidelines for its use in clinical practice. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. Can EGFR mutation status be reliably determined in pre-operative needle biopsies from adenocarcinomas of the lung?

    DEFF Research Database (Denmark)

    Lindahl, Kim Hein; Sørensen, Flemming Brandt; Jonstrup, Søren Peter

    2015-01-01

    The identification of EGFR mutations in non-small-cell lung cancer is important for selecting patients, who may benefit from treatment with EGFR tyrosine kinase inhibitors. The analysis is usually performed on cytological aspirates and/or histological needle biopsies, representing a small fraction....... Moreover, several inconclusive results in the diagnostic biopsies reveal that attention must be paid on the suitability of pre-operative biopsies for EGFR mutation analysis....

  2. Epidermal growth factor receptor (EGFR) mutations in lung cancer: preclinical and clinical data

    Energy Technology Data Exchange (ETDEWEB)

    Jorge, S.E.D.C.; Kobayashi, S.S.; Costa, D.B. [Harvard Medical School, Beth Israel Deaconess Medical Center, Department of Medicine, Division of Hematology/Oncology, Boston, MA (United States)

    2014-09-05

    Lung cancer leads cancer-related mortality worldwide. Non-small-cell lung cancer (NSCLC), the most prevalent subtype of this recalcitrant cancer, is usually diagnosed at advanced stages, and available systemic therapies are mostly palliative. The probing of the NSCLC kinome has identified numerous nonoverlapping driver genomic events, including epidermal growth factor receptor (EGFR) gene mutations. This review provides a synopsis of preclinical and clinical data on EGFR mutated NSCLC and EGFR tyrosine kinase inhibitors (TKIs). Classic somatic EGFR kinase domain mutations (such as L858R and exon 19 deletions) make tumors addicted to their signaling cascades and generate a therapeutic window for the use of ATP-mimetic EGFR TKIs. The latter inhibit these kinases and their downstream effectors, and induce apoptosis in preclinical models. The aforementioned EGFR mutations are stout predictors of response and augmentation of progression-free survival when gefitinib, erlotinib, and afatinib are used for patients with advanced NSCLC. The benefits associated with these EGFR TKIs are limited by the mechanisms of tumor resistance, such as the gatekeeper EGFR-T790M mutation, and bypass activation of signaling cascades. Ongoing preclinical efforts for treating resistance have started to translate into patient care (including clinical trials of the covalent EGFR-T790M TKIs AZD9291 and CO-1686) and hold promise to further boost the median survival of patients with EGFR mutated NSCLC.

  3. Epidermal growth factor receptor (EGFR) mutations in lung cancer: preclinical and clinical data

    International Nuclear Information System (INIS)

    Jorge, S.E.D.C.; Kobayashi, S.S.; Costa, D.B.

    2014-01-01

    Lung cancer leads cancer-related mortality worldwide. Non-small-cell lung cancer (NSCLC), the most prevalent subtype of this recalcitrant cancer, is usually diagnosed at advanced stages, and available systemic therapies are mostly palliative. The probing of the NSCLC kinome has identified numerous nonoverlapping driver genomic events, including epidermal growth factor receptor (EGFR) gene mutations. This review provides a synopsis of preclinical and clinical data on EGFR mutated NSCLC and EGFR tyrosine kinase inhibitors (TKIs). Classic somatic EGFR kinase domain mutations (such as L858R and exon 19 deletions) make tumors addicted to their signaling cascades and generate a therapeutic window for the use of ATP-mimetic EGFR TKIs. The latter inhibit these kinases and their downstream effectors, and induce apoptosis in preclinical models. The aforementioned EGFR mutations are stout predictors of response and augmentation of progression-free survival when gefitinib, erlotinib, and afatinib are used for patients with advanced NSCLC. The benefits associated with these EGFR TKIs are limited by the mechanisms of tumor resistance, such as the gatekeeper EGFR-T790M mutation, and bypass activation of signaling cascades. Ongoing preclinical efforts for treating resistance have started to translate into patient care (including clinical trials of the covalent EGFR-T790M TKIs AZD9291 and CO-1686) and hold promise to further boost the median survival of patients with EGFR mutated NSCLC

  4. Novel Bruton's tyrosine kinase inhibitors currently in development

    Directory of Open Access Journals (Sweden)

    D'Cruz OJ

    2013-03-01

    Full Text Available Osmond J D'Cruz,1 Fatih M Uckun1,21Children's Center for Cancer and Blood Diseases, Children's Hospital Los Angeles, Los Angeles, CA, USA; 2Department of Pediatrics, University of Southern California, Los Angeles, CA, USAAbstract: Bruton's tyrosine kinase (Btk is intimately involved in multiple signal-transduction pathways regulating survival, activation, proliferation, and differentiation of B-lineage lymphoid cells. Btk is overexpressed and constitutively active in several B-lineage lymphoid malignancies. Btk has emerged as a new antiapoptotic molecular target for treatment of B-lineage leukemias and lymphomas. Preclinical and early clinical results indicate that Btk inhibitors may be useful in the treatment of leukemias and lymphomas.Keywords: tyrosine kinase, personalized therapy, kinase inhibitors, Btk, leukemia, lymphoma

  5. Efficacy of EGFR-TKI therapy in patients with brain metastases from ...

    African Journals Online (AJOL)

    Purpose: This meta-analysis aimed to systematically evaluate the efficacy of tyrosine kinase inhibitors of epidermal growth factor receptor (EGFR-TKIs) for patients with brain metastases (BM) from non- small-cell lung cancer (NSCLC), and to compare this treatment modality to chemotherapy or radiotherapy. Methods: ...

  6. Clinical efficacy of first-generation EGFR-TKIs in patients with advanced non-small-cell lung cancer harboring EGFR exon 20 mutations

    Directory of Open Access Journals (Sweden)

    Chen D

    2016-07-01

    Full Text Available Dan Chen,1 Zhengbo Song,2 Guoping Cheng3 1Department of Cardiothoracic Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing, 2Department of Chemotherapy, 3Department of Pathology, Zhejiang Cancer Hospital, Hangzhou, People’s Republic of China Purpose: Subsets of non-small-cell lung cancer patients with epidermal growth factor receptor (EGFR mutations carry uncommon subtypes. We evaluated the efficacy of first-generation EGFR-tyrosine kinase inhibitors (TKIs; erlotinib, gefitinib, and icotinib in patients with non-small-cell lung cancer carrying insertions and T790M and S768I mutations in EGFR exon 20. Patients and methods: Patients carrying EGFR exon 20 insertion/T790M/S768I mutations and treated with EGFR-TKIs were evaluated from 2005 to 2014 in Zhejiang Cancer Hospital. The efficacy was evaluated using the Kaplan–Meier method and compared with the log-rank test. Results: Sixty-two patients with exon 20 insertion/T790M/S768I mutations were enrolled. Mutations including exon 20 insertions and T790M and S768I mutations were observed in 29, 23, and ten patients, respectively. In total, the response rate and median progression-free survival (PFS were 8.1% and 2.1 months, respectively. Patients with S768I mutation manifested the longest median PFS (2.7 months, followed by those with T790M (2.4 months and exon 20 insertions (1.9 months; P=0.022. Patients with complex mutations show a better PFS than those with single mutations (2.7 months vs 1.9 months; P=0.034. Conclusion: First-generation EGFR-TKIs are less effective in patients with exon 20 uncommon mutations than in those with common mutations. Patients with complex mutations benefited more from first-generation EGFR-TKIs than those with single mutations. Keywords: non-small cell lung cancer, epidermal growth factor receptor, EGFR mutations, exon 20, tyrosine kinase inhibitor

  7. Phase I study of icotinib hydrochloride (BPI-2009H), an oral EGFR tyrosine kinase inhibitor, in patients with advanced NSCLC and other solid tumors.

    Science.gov (United States)

    Zhao, Qiong; Shentu, Jianzhong; Xu, Nong; Zhou, Jianya; Yang, Guangdie; Yao, Yinan; Tan, Fenlai; Liu, Dongyang; Wang, Yingxiang; Zhou, Jianying

    2011-08-01

    The goal of this study was to assess the safety and tolerability of icotinib hydrochloride (BPI-2009H), a new selective epidermal growth factor receptor tyrosine kinase inhibitor (EGFR TKI), and to explore its pharmacokinetics (PK) and clinical activity in patients with advanced solid tumors, mainly those with non-small-cell lung cancer (NSCLC) after the failure of the prior platinum-based chemotherapy. Different doses of oral icotinib were administered once every 8 h (Q8H) for a 28-continuous-day cycle until disease progression and or undue toxicity was observed. PK studies of subjects' blood were performed during cycle one (day 1 through 28). Patients aged ≥18 and ≤70 years with an Eastern Cooperative Oncology Group (ECOG) performance status (PS) of 0-1 and adequate organ functions eligible for the study. Tumor responses were assessed by Response Evaluation Criteria in Solid Tumors (RECIST). K-ras and EGFR mutations in the extracted DNA of fourteen specimens were examined using PCR-based direct sequencing assay. Thirty-six patients were enrolled in the study. PK analysis demonstrated that the mean elimination half-life of icotinib was 6 h, and the T(max) was around 2 h. The steady-state concentration of icotinib administered at a dose of 125 mg once every 8 h (Q8H) was significantly higher than that achieved by a dose of 100mg Q8H. The most frequent treatment-related adverse events (TRAEs) were an acne-like (folliculitis) rash (16/36, 44.4%), diarrhea (8/36, 22.2%) and a decrease in white blood cells (4/36, 11.1%). The maximum-tolerated dose (MTD) was not reached. Among 33 patients with NSCLC, 7 patients exhibited a partial response, 7 showed stable disease at the 24 weeks. Among 14 patients undergoing DNA sequence for K-ras and EGFR mutations, 3 with K-ras mutation presented 2 stable disease (SD) and 1 partial response (PR), 5 with EGFR exon 19 or 21 mutation 2 PR and 3 SD within 4 weeks. Oral icotinib was generally well tolerated, with manageable and

  8. The Drosophila Arf GEF Steppke controls MAPK activation in EGFR signaling.

    Science.gov (United States)

    Hahn, Ines; Fuss, Bernhard; Peters, Annika; Werner, Tamara; Sieberg, Andrea; Gosejacob, Dominic; Hoch, Michael

    2013-06-01

    Guanine nucleotide exchange factors (GEFs) of the cytohesin protein family are regulators of GDP/GTP exchange for members of the ADP ribosylation factor (Arf) of small GTPases. They have been identified as modulators of various receptor tyrosine kinase signaling pathways including the insulin, the vascular epidermal growth factor (VEGF) and the epidermal growth factor (EGF) pathways. These pathways control many cellular functions, including cell proliferation and differentiation, and their misregulation is often associated with cancerogenesis. In vivo studies on cytohesins using genetic loss of function alleles are lacking, however, since knockout mouse models are not available yet. We have recently identified mutants for the single cytohesin Steppke (Step) in Drosophila and we could demonstrate an essential role of Step in the insulin signaling cascade. In the present study, we provide in vivo evidence for a role of Step in EGFR signaling during wing and eye development. By analyzing step mutants, transgenic RNA interference (RNAi) and overexpression lines for tissue specific as well as clonal analysis, we found that Step acts downstream of the EGFR and is required for the activation of mitogen-activated protein kinase (MAPK) and the induction of EGFR target genes. We further demonstrate that step transcription is induced by EGFR signaling whereas it is negatively regulated by insulin signaling. Furthermore, genetic studies and biochemical analysis show that Step interacts with the Connector Enhancer of KSR (CNK). We propose that Step may be part of a larger signaling scaffold coordinating receptor tyrosine kinase-dependent MAPK activation.

  9. Development of new mouse lung tumor models expressing EGFR T790M mutants associated with clinical resistance to kinase inhibitors.

    Science.gov (United States)

    Regales, Lucia; Balak, Marissa N; Gong, Yixuan; Politi, Katerina; Sawai, Ayana; Le, Carl; Koutcher, Jason A; Solit, David B; Rosen, Neal; Zakowski, Maureen F; Pao, William

    2007-08-29

    The EGFR T790M mutation confers acquired resistance to kinase inhibitors in human EGFR mutant lung adenocarcinoma, is occasionally detected before treatment, and may confer genetic susceptibility to lung cancer. To study further its role in lung tumorigenesis, we developed mice with inducible expression in type II pneumocytes of EGFR(T790M) alone or together with a drug-sensitive L858R mutation. Both transgenic lines develop lung adenocarcinomas that require mutant EGFR for tumor maintenance but are resistant to an EGFR kinase inhibitor. EGFR(L858R+T790M)-driven tumors are transiently targeted by hsp90 inhibition. Notably, EGFR(T790M)-expressing animals develop tumors with longer latency than EGFR(L858R+T790M)-bearing mice and in the absence of additional kinase domain mutations. These new mouse models of mutant EGFR-dependent lung adenocarcinomas provide insight into clinical observations. The models should also be useful for developing improved therapies for patients with lung cancers harboring EGFR(T790M) alone or in conjunction with drug-sensitive EGFR kinase domain mutations.

  10. Development of new mouse lung tumor models expressing EGFR T790M mutants associated with clinical resistance to kinase inhibitors.

    Directory of Open Access Journals (Sweden)

    Lucia Regales

    2007-08-01

    Full Text Available The EGFR T790M mutation confers acquired resistance to kinase inhibitors in human EGFR mutant lung adenocarcinoma, is occasionally detected before treatment, and may confer genetic susceptibility to lung cancer.To study further its role in lung tumorigenesis, we developed mice with inducible expression in type II pneumocytes of EGFR(T790M alone or together with a drug-sensitive L858R mutation. Both transgenic lines develop lung adenocarcinomas that require mutant EGFR for tumor maintenance but are resistant to an EGFR kinase inhibitor. EGFR(L858R+T790M-driven tumors are transiently targeted by hsp90 inhibition. Notably, EGFR(T790M-expressing animals develop tumors with longer latency than EGFR(L858R+T790M-bearing mice and in the absence of additional kinase domain mutations.These new mouse models of mutant EGFR-dependent lung adenocarcinomas provide insight into clinical observations. The models should also be useful for developing improved therapies for patients with lung cancers harboring EGFR(T790M alone or in conjunction with drug-sensitive EGFR kinase domain mutations.

  11. Identification and analysis of a novel protein-tyrosine kinase from bovine thymus

    International Nuclear Information System (INIS)

    Zioncheck, T.F.; Harrison, M.L.; Geahlen, R.L.

    1986-01-01

    A cytosolic protein-tyrosine kinase has been identified and purified to near homogeneity from calf thymus by using the phosphorylation of the tyrosine-containing peptide angiotensin I as an assay. Specific peptide phosphorylating activity was enhanced by carrying out the assay at high ionic strength (2M NaCl). The inclusion of NaCl at this concentration acts to stimulate endogenous protein-tyrosine kinase activity while simultaneously inhibiting other endogenous kinases. The purification procedure involved extraction of the enzyme from calf-thymus and sequential chromatography on columns of DEAE-cellulose, heparin-agarose, casein-sepharose, butylagarose, and Sephadex G-75. Analysis of the most highly purified preparations by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single Coomassie blue-stained band of 41 KDa. This molecular weight was consistent with results obtained from gel filtration, indicating that the enzyme exists as a monomer. The enzyme has also been found to catalyze an autophosphorylation reaction. Incubation of the enzyme with Mn 2+ and [γ- 32 P]ATP led to its modification on a tyrosine residue. Phosphopeptide mapping experiments indicated that the 41 KDa kinase was distinct from p56, the major membrane-associated protein-tyrosine kinase in T lymphocytes

  12. Requirement for tyrosine phosphatase during serotonergic neuromodulation by protein kinase C.

    Science.gov (United States)

    Catarsi, S; Drapeau, P

    1997-08-01

    Tyrosine kinases and phosphatases are abundant in the nervous system, where they signal cellular differentiation, mediate the responses to growth factors, and direct neurite outgrowth during development. Tyrosine phosphorylation can also alter ion channel activity, but its physiological significance remains unclear. In an identified leech mechanosensory neuron, the ubiquitous neuromodulator serotonin increases the activity of a cation channel by activating protein kinase C (PKC), resulting in membrane depolarization and modulation of the receptive field properties. We observed that the effects on isolated neurons and channels were blocked by inhibiting tyrosine phosphatases. Serotonergic stimulation of PKC thus activates a tyrosine phosphatase activity associated with the channels, which reverses their constitutive inhibition by tyrosine phosphorylation, representing a novel form of neuromodulation.

  13. Receptor tyrosine kinase signaling: a view from quantitative proteomics

    DEFF Research Database (Denmark)

    Dengjel, Joern; Kratchmarova, Irina; Blagoev, Blagoy

    2009-01-01

    Growth factor receptor signaling via receptor tyrosine kinases (RTKs) is one of the basic cellular communication principals found in all metazoans. Extracellular signals are transferred via membrane spanning receptors into the cytoplasm, reversible tyrosine phosphorylation being the hallmark of all...

  14. Coarse-grained molecular simulation of epidermal growth factor receptor protein tyrosine kinase multi-site self-phosphorylation.

    Directory of Open Access Journals (Sweden)

    John G Koland

    2014-01-01

    Full Text Available Upon the ligand-dependent dimerization of the epidermal growth factor receptor (EGFR, the intrinsic protein tyrosine kinase (PTK activity of one receptor monomer is activated, and the dimeric receptor undergoes self-phosphorylation at any of eight candidate phosphorylation sites (P-sites in either of the two C-terminal (CT domains. While the structures of the extracellular ligand binding and intracellular PTK domains are known, that of the ∼225-amino acid CT domain is not, presumably because it is disordered. Receptor phosphorylation on CT domain P-sites is critical in signaling because of the binding of specific signaling effector molecules to individual phosphorylated P-sites. To investigate how the combination of conventional substrate recognition and the unique topological factors involved in the CT domain self-phosphorylation reaction lead to selectivity in P-site phosphorylation, we performed coarse-grained molecular simulations of the P-site/catalytic site binding reactions that precede EGFR self-phosphorylation events. Our results indicate that self-phosphorylation of the dimeric EGFR, although generally believed to occur in trans, may well occur with a similar efficiency in cis, with the P-sites of both receptor monomers being phosphorylated to a similar extent. An exception was the case of the most kinase-proximal P-site-992, the catalytic site binding of which occurred exclusively in cis via an intramolecular reaction. We discovered that the in cis interaction of P-site-992 with the catalytic site was facilitated by a cleft between the N-terminal and C-terminal lobes of the PTK domain that allows the short CT domain sequence tethering P-site-992 to the PTK core to reach the catalytic site. Our work provides several new mechanistic insights into the EGFR self-phosphorylation reaction, and demonstrates the potential of coarse-grained molecular simulation approaches for investigating the complexities of self-phosphorylation in

  15. Gallic acid induces apoptosis in EGFR-mutant non-small cell lung cancers by accelerating EGFR turnover.

    Science.gov (United States)

    Nam, Boas; Rho, Jin Kyung; Shin, Dong-Myung; Son, Jaekyoung

    2016-10-01

    Gallic acid is a common botanic phenolic compound, which is present in plants and foods worldwide. Gallic acid is implicated in various biological processes such as cell growth and apoptosis. Indeed, gallic acid has been shown to induce apoptosis in many cancer types. However, the molecular mechanisms of gallic acid-induced apoptosis in cancer, particularly lung cancer, are still unclear. Here, we report that gallic acid induces apoptosis in EGFR-mutant non-small cell lung cancer (NSCLC) cells, but not in EGFR-WT NSCLC cells. Treatment with gallic acid resulted in a significant reduction in proliferation and induction of apoptosis, only in EGFR-mutant NSCLC cells. Interestingly, treatment with gallic acid led to a robust decrease in EGFR levels, which is critical for NSCLC survival. Treatment with gallic acid had no significant effect on transcription, but induced EGFR turnover. Indeed, treatment with a proteasome inhibitor dramatically reversed gallic acid-induced EGFR downregulation. Moreover, treatment with gallic acid induced EGFR turnover leading to apoptosis in EGFR-TKI (tyrosine kinase inhibitor)-resistant cell lines, which are dependent on EGFR signaling for survival. Thus, these studies suggest that gallic acid can induce apoptosis in EGFR-dependent lung cancers that are dependent on EGFR for growth and survival via acceleration of EGFR turnover. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Pervanadate induces Mammalian Ste20 Kinase 3 (MST3) tyrosine phosphorylation but not activation.

    Science.gov (United States)

    Kan, Wei-Chih; Lu, Te-Ling; Ling, Pin; Lee, Te-Hsiu; Cho, Chien-Yu; Huang, Chi-Ying F; Jeng, Wen-Yih; Weng, Yui-Ping; Chiang, Chun-Yen; Wu, Jin Bin; Lu, Te-Jung

    2016-07-01

    The yeast Ste20 (sterile) protein kinase, which is a serine/threonine kinase, responds to the stimulation of the G proteincoupled receptor (GPCR) pheromone receptor. Ste20 protein kinase serves as the critical component that links signaling from the GPCR/G proteins to the mitogen-activated protein kinase (MAPK) cascade in yeast. The yeast Ste20p functions as a MAP kinase kinase kinase kinase (MAP4K) in the pheromone response. Ste20-like kinases are structurally conserved from yeast to mammals. The mechanism by which MAP4K links GPCR to the MAPK pathway is less clearly defined in vertebrates. In addition to MAP4K, the tyrosine kinase cascade bridges G proteins and the MAPK pathway in vertebrate cells. Mammalian Ste20 Kinase 3 (MST3) has been categorized into the Ste20 family and has been reported to function in the regulation of cell polarity and migration. However, whether MST3 tyrosine phosphorylation regulates diverse signaling pathways is unknown. In this study, the tyrosine phosphatase inhibitor pervanadate was found to induce MST3 tyrosine phosphorylation in intact cells, and the activity of tyrosine-phosphorylated MST3 was measured. This tyrosine-directed phosphorylation was independent of MST3 activity. Parameters including protein conformation, Triton concentration and ionic concentration influenced the sensitivity of MST3 activity. Taken together, our data suggests that the serine/threonine kinase MST3 undergoes tyrosinedirected phosphorylation. The tyrosine-phosphorylated MST3 may create a docking site for the structurally conserved SH2/SH3 (Src Homology 2 and 3) domains within the Src oncoprotein. The unusual tyrosinephosphorylated MST3 may recruit MST3 to various signaling components. Copyright © 2016. Published by Elsevier Inc.

  17. Evidence for association of the cloned liver growth hormone receptor with a tyrosine kinase

    DEFF Research Database (Denmark)

    Wang, X; Uhler, M D; Billestrup, N

    1992-01-01

    The ability of the cloned liver growth hormone (GH) receptor, when expressed in mammalian cell lines, to copurify with tyrosine kinase activity and be tyrosyl phosphorylated was examined. 125I-human growth hormone-GH receptor complexes isolated from COS-7 cells transiently expressing high levels...... of tyrosine kinase activity with cloned liver GH receptor. The level of phosphorylation of the GH receptor was very low, as compared with the endogenous GH receptor in 3T3-F442A cells, suggesting that tyrosine kinase activity is not intrinsic to the cloned GH receptor but rather resides with a kinase present...... in a variety of cell types. The finding that the level of phosphorylation of GH receptor appears to vary with cell type is consistent with the cloned liver GH receptor being a substrate for an associated tyrosine kinase and with the amount of such a GH receptor-associated tyrosine kinase being cell type-specific....

  18. Ror receptor tyrosine kinases: orphans no more.

    Science.gov (United States)

    Green, Jennifer L; Kuntz, Steven G; Sternberg, Paul W

    2008-11-01

    Receptor tyrosine kinase-like orphan receptor (Ror) proteins are a conserved family of tyrosine kinase receptors that function in developmental processes including skeletal and neuronal development, cell movement and cell polarity. Although Ror proteins were originally named because the associated ligand and signaling pathway were unknown, recent studies in multiple species have now established that Ror proteins are Wnt receptors. Depending on the cellular context, Ror proteins can either activate or repress transcription of Wnt target genes and can modulate Wnt signaling by sequestering Wnt ligands. New evidence implicates Ror proteins in planar cell polarity, an alternative Wnt pathway. Here, we review the progress made in understanding these mysterious proteins and, in particular, we focus on their function as Wnt receptors.

  19. SOCS proteins in regulation of receptor tyrosine kinase signaling

    DEFF Research Database (Denmark)

    Kazi, Julhash U.; Kabir, Nuzhat N.; Flores Morales, Amilcar

    2014-01-01

    Receptor tyrosine kinases (RTKs) are a family of cell surface receptors that play critical roles in signal transduction from extracellular stimuli. Many in this family of kinases are overexpressed or mutated in human malignancies and thus became an attractive drug target for cancer treatment....... The signaling mediated by RTKs must be tightly regulated by interacting proteins including protein-tyrosine phosphatases and ubiquitin ligases. The suppressors of cytokine signaling (SOCS) family proteins are well-known negative regulators of cytokine receptors signaling consisting of eight structurally similar...

  20. Tyrosine Kinase Ligand-Receptor Pair Prediction by Using Support Vector Machine

    Directory of Open Access Journals (Sweden)

    Masayuki Yarimizu

    2015-01-01

    Full Text Available Receptor tyrosine kinases are essential proteins involved in cellular differentiation and proliferation in vivo and are heavily involved in allergic diseases, diabetes, and onset/proliferation of cancerous cells. Identifying the interacting partner of this protein, a growth factor ligand, will provide a deeper understanding of cellular proliferation/differentiation and other cell processes. In this study, we developed a method for predicting tyrosine kinase ligand-receptor pairs from their amino acid sequences. We collected tyrosine kinase ligand-receptor pairs from the Database of Interacting Proteins (DIP and UniProtKB, filtered them by removing sequence redundancy, and used them as a dataset for machine learning and assessment of predictive performance. Our prediction method is based on support vector machines (SVMs, and we evaluated several input features suitable for tyrosine kinase for machine learning and compared and analyzed the results. Using sequence pattern information and domain information extracted from sequences as input features, we obtained 0.996 of the area under the receiver operating characteristic curve. This accuracy is higher than that obtained from general protein-protein interaction pair predictions.

  1. EGFR T790M mutation after chemotherapy for small cell lung cancer transformation of EGFR-positive non-small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Tomoaki Sonoda

    Full Text Available In non-small cell lung cancer (NSCLC with an epidermal growth factor receptor (EGFR mutation, 50%–65% of cases acquire resistance after treatment with EGFR-tyrosine kinase inhibitors (EGFR-TKIs because of an EGFR T790M point mutation and 3%–14% of these cases transformed to small cell lung cancer (SCLC. Generally, the EGFR T790M secondary mutation develops with ongoing ATP competitive inhibition. We present a case of a 76-year-old woman with lung adenocarcinoma harboring an EGFR-L858R mutation who received first-line gefitinib and developed SCLC transformation. She was administered several chemotherapy agents, including a platinum doublet. The primary lesion that showed SCLC transformation had reconverted to adenocarcinoma with EGFR L858R and T790M mutations at the time of a second re-biopsy. Therefore, she was administered osimertinib, which resulted in clinical remission. This case suggested that serial biopsies are necessary even after SCLC transformation. Keywords: NSCLC, EGFR mutation, SCLC transformation, T790M, Osimertinib

  2. Activation of a Neospora caninum EGFR-Like Kinase Facilitates Intracellular Parasite Proliferation

    Directory of Open Access Journals (Sweden)

    Xiaoxia Jin

    2017-10-01

    Full Text Available The Apicomplexan parasite Neospora caninum, an obligate intracellular protozoan, causes serious diseases in a number of mammalian species, especially in cattle. Infection with N. caninum is associated with abortions in both dairy and beef cattle worldwide which have a major economic impact on the cattle industry. However, the mechanism by which N. caninum proliferates within host cells is poorly understood. Epidermal growth factor receptor (EGFR is a protein kinase ubiquitously expressed, present on cell surfaces in numerous species, which has been confirmed to be essential in signal transduction involved in cell growth, proliferation, survival, and many other intracellular processes. However, the presence of EGFR in N. caninum and its role in N. caninum proliferation remain unclear. In the present study, we identified a putative EGFR-like kinase in N. caninum, which could be activated in tachyzoites by infection or treatment with rNcMIC3 [containing four epidermal growth factor (EGF domains] or human EGF. Blockade of EGFR-like in tachyzoites by AG1478 significantly reduced parasite proliferation in host cells. Our data suggested that the activation of tachyzoite EGFR-like might facilitate the intracellular proliferation of N. caninum.

  3. Tyrosine kinase fusion genes in pediatric BCR-ABL1-like acute lymphoblastic leukemia

    Science.gov (United States)

    Boer, Judith M.; Steeghs, Elisabeth M.P.; Marchante, João R.M.; Boeree, Aurélie; Beaudoin, James J.; Berna Beverloo, H.; Kuiper, Roland P.; Escherich, Gabriele; van der Velden, Vincent H.J.; van der Schoot, C. Ellen; de Groot-Kruseman, Hester A.; Pieters, Rob; den Boer, Monique L.

    2017-01-01

    Approximately 15% of pediatric B cell precursor acute lymphoblastic leukemia (BCP-ALL) is characterized by gene expression similar to that of BCR-ABL1-positive disease and unfavorable prognosis. This BCR-ABL1-like subtype shows a high frequency of B-cell development gene aberrations and tyrosine kinase-activating lesions. To evaluate the clinical significance of tyrosine kinase gene fusions in children with BCP-ALL, we studied the frequency of recently identified tyrosine kinase fusions, associated genetic features, and prognosis in a representative Dutch/German cohort. We identified 14 tyrosine kinase fusions among 77 BCR-ABL1-like cases (18%) and none among 76 non-BCR-ABL1-like B-other cases. Novel exon fusions were identified for RCSD1-ABL2 and TERF2-JAK2. JAK2 mutation was mutually exclusive with tyrosine kinase fusions and only occurred in cases with high CRLF2 expression. The non/late response rate and levels of minimal residual disease in the fusion-positive BCR-ABL1-like group were higher than in the non-BCR-ABL1-like B-others (p<0.01), and also higher, albeit not statistically significant, compared with the fusion-negative BCR-ABL1-like group. The 8-year cumulative incidence of relapse in the fusion-positive BCR-ABL1-like group (35%) was comparable with that in the fusion-negative BCR-ABL1-like group (35%), and worse than in the non-BCR-ABL1-like B-other group (17%, p=0.07). IKZF1 deletions, predominantly other than the dominant-negative isoform and full deletion, co-occurred with tyrosine kinase fusions. This study shows that tyrosine kinase fusion-positive cases are a high-risk subtype of BCP-ALL, which warrants further studies with specific kinase inhibitors to improve outcome. PMID:27894077

  4. Tyrosine kinase signalling in breast cancer

    International Nuclear Information System (INIS)

    Hynes, Nancy E

    2000-01-01

    Cells are continuously exposed to diverse stimuli ranging from soluble endocrine and paracrine factors to signalling molecules on neighbouring cells. Receptors of the tyrosine kinase family play an important role in the integration and interpretation of these external stimuli, allowing a cell to respond appropriately to its environment. The activation of receptor tyrosine kinases (RTKs) is tightly controlled, allowing a normal cell to correctly integrate its external environment with internal signal transduction pathways. In contrast, due to numerous molecular alterations arising during the course of malignancy, a tumour is characterized by an abnormal response to its environment, which allows cancer cells to evade the normal mechanisms controlling cellular proliferation. Alterations in the expression of various RTKs, in their activation, and in the signalling molecules lying downstream of the receptors play important roles in the development of cancer. This topic is the major focus of the thematic review section of this issue of Breast Cancer Research

  5. PTB domain-directed substrate targeting in a tyrosine kinase from the unicellular choanoflagellate Monosiga brevicollis.

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    Victoria Prieto-Echagüe

    2011-04-01

    Full Text Available Choanoflagellates are considered to be the closest living unicellular relatives of metazoans. The genome of the choanoflagellate Monosiga brevicollis contains a surprisingly high number and diversity of tyrosine kinases, tyrosine phosphatases, and phosphotyrosine-binding domains. Many of the tyrosine kinases possess combinations of domains that have not been observed in any multicellular organism. The role of these protein interaction domains in M. brevicollis kinase signaling is not clear. Here, we have carried out a biochemical characterization of Monosiga HMTK1, a protein containing a putative PTB domain linked to a tyrosine kinase catalytic domain. We cloned, expressed, and purified HMTK1, and we demonstrated that it possesses tyrosine kinase activity. We used immobilized peptide arrays to define a preferred ligand for the third PTB domain of HMTK1. Peptide sequences containing this ligand sequence are phosphorylated efficiently by recombinant HMTK1, suggesting that the PTB domain of HMTK1 has a role in substrate recognition analogous to the SH2 and SH3 domains of mammalian Src family kinases. We suggest that the substrate recruitment function of the noncatalytic domains of tyrosine kinases arose before their roles in autoinhibition.

  6. Characterization and response of newly developed high-grade glioma cultures to the tyrosine kinase inhibitors, erlotinib, gefitinib and imatinib.

    LENUS (Irish Health Repository)

    Kinsella, Paula

    2012-03-10

    High-grade gliomas (HGG), are the most common aggressive brain tumours in adults. Inhibitors targeting growth factor signalling pathways in glioma have shown a low clinical response rate. To accurately evaluate response to targeted therapies further in vitro studies are necessary. Growth factor pathway expression using epidermal growth factor receptor (EGFR), mutant EGFR (EGFRvIII), platelet derived growth factor receptor (PDGFR), C-Kit and C-Abl together with phosphatase and tensin homolog (PTEN) expression and downstream activation of AKT and phosphorylated ribosomal protein S6 (P70S6K) was analysed in 26 primary glioma cultures treated with the tyrosine kinase inhibitors (TKIs) erlotinib, gefitinib and imatinib. Response to TKIs was assessed using 50% inhibitory concentrations (IC(50)). Response for each culture was compared with the EGFR\\/PDGFR immunocytochemical pathway profile using hierarchical cluster analysis (HCA) and principal component analysis (PCA). Erlotinib response was not strongly associated with high expression of the growth factor pathway components. PTEN expression did not correlate with response to any of the three TKIs. Increased EGFR expression was associated with gefitinib response; increased PDGFR-α expression was associated with imatinib response. The results of this in vitro study suggest gefitinib and imatinib may have therapeutic potential in HGG tumours with a corresponding growth factor receptor expression profile.

  7. Ethyl p-methoxycinnamate from Kaempferia galanga inhibits angiogenesis through tyrosine kinase

    Directory of Open Access Journals (Sweden)

    Juni Ekowati

    2015-04-01

    Full Text Available Background Many tumors express on their receptor tyrosine kinases vascular endothelial growth factor activity associated with angiogenesis. Inhibition of angiogenesis through reduction of tyrosine kinase activity is a promising strategy for cancer therapy. The present study aimed to determine the mechanism and potency of ethyl p-methoxycinnamate (EPMC isolated from Kaempferia galanga as angiogenesis inhibitor. Methods A laboratory experimental study was conducted using chorio-allantoic membranes (CAMs of nine-day old chicken eggs induced by 60ng basic fibroblast growth factor (bFGF. Ethyl p-methoxycinnamate (EPMC potency was determined at dosages of 30, 60, 90 and 120 mg and compared with celecoxib 60 mg as reference drug and one negative bFGF-induced control group. Neovascularization and endothelial cell count in CAM blood vessels were evaluated. To predict the antiangiogenic mechanism of EPMC, a docking study was performed with the Molegro Virtual Docker program on tyrosine kinase as receptor (PDB 1XKK. Results Angiogenesis stimulation by bFGF was prevented significantly (p<0.05 by EPMC at dosages of 30, 60, 90 and 120 mg and this activity was dose dependent. Molecular docking showed interaction between EPMC functional groups and tyrosine kinase amino acids at Met766, Met793, Thr854, Thr790, Gln791 and Ala743. There was an association between EPMC antiangiogenic activity and docking study results. Conclusions Ethyl p-methoxycinnamate is a potential new angiogenesis inhibitor through interaction with tyrosine kinase. EPMC could be a promising therapeutic agent for treatment of angiogenesis-related diseases.

  8. Inhibitory Activities of Epidermal Growth Factor Receptor Tyrosine Kinase-Targeted Dihydroxyisoflavone and Trihydroxydeoxybenzoin Derivatives on Sarcocystis neurona, Neospora caninum, and Cryptosporidium parvum Development

    Science.gov (United States)

    Gargala, G.; Baishanbo, A.; Favennec, L.; François, A.; Ballet, J. J.; Rossignol, J.-F.

    2005-01-01

    Several gene sequences of parasitic protozoa belonging to protein kinase gene families and epidermal growth factor (EGF)-like peptides, which act via binding to receptor tyrosine kinases of the EGF receptor (EGFR) family, appear to mediate host-protozoan interactions. As a clue to EGFR protein tyrosine kinase (PTK) mediation and a novel approach for identifying anticoccidial agents, activities against Sarcocystis neurona, Neospora caninum, and Cryptosporidium parvum grown in BM and HCT-8 cell cultures of 52 EGFR PTK inhibitor isoflavone analogs (dihydroxyisoflavone and trihydroxydeoxybenzoine derivatives) were investigated. Their cytotoxicities against host cells were either absent, mild, or moderate by a nitroblue tetrazolium test. At concentrations ranging from 5 to 10 μg/ml, 20 and 5 analogs, including RM-6427 and RM-6428, exhibited an in vitro inhibitory effect of ≥95% against at least one parasite or against all three, respectively. In immunosuppressed Cryptosporidium parvum-infected Mongolian gerbils orally treated with either 200 or 400 mg of agent RM-6427/kg of body weight/day for 8 days, fecal microscopic oocyst shedding was abolished in 6/10 animals (P of 0.05, respectively). After RM-6427 therapy (200 mg/kg/day for 8 days), the reduction in the ratio of animals with intracellular parasites was nearly significant in ileum (P = 0.067) and more marked in the biliary tract (P < 0.0013) than after nitazoxanide or paromomycin treatment (0.05 < P < 0.004). RM-6428 treatment at a regimen of 400 mg/kg/day for 12 days inhibited oocyst shedding, measured using flow cytometry from day 4 (P < 0.05) to day 12 (P < 0.02) of therapy, when 2/15 animals had no shedding (P < 0.0001) and 11/15 were free of gut and/or biliary tract parasites (P < 0.01). No mucosal alteration was microscopically observed for treated or untreated infected gerbils. To our knowledge, this report is the first to suggest that the isoflavone class of agents has the potential for

  9. A case of lung adenocarcinoma harboring EGFR mutation and EML4-ALK fusion gene

    International Nuclear Information System (INIS)

    Tanaka, Hisashi; Hayashi, Akihito; Morimoto, Takeshi; Taima, Kageaki; Tanaka, Yoshihito; Shimada, Michiko; Kurose, Akira; Takanashi, Shingo; Okumura, Ken

    2012-01-01

    Lung cancer is the leading cause of cancer-related death worldwide. Epidermal growth factor receptor (EGFR) - tyrosine kinase inhibitor (TKI) is used for the patients with EGFR-mutant lung cancer. Recently, phase III studies in the patients with EGFR-mutant demonstrated that EGFR-TKI monotherapy improved progression-free survival compared with platinum-doublet chemotherapy. The echinoderm microtubule-associated protein-like 4 (EML4) - anaplastic lymphoma kinase (ALK) fusion oncogene represents one of the newest molecular targets in non-small cell lung cancer (NSCLC). Patients who harbor EML4-ALK fusions have been associated with a lack of EGFR or KRAS mutations. We report a 39-year-old patient diagnosed as adenocarcinoma harboring EGFR mutation and EML4-ALK fusion gene. We treated this patient with erlotinib as the third line therapy, but no clinical benefit was obtained. We experienced a rare case with EGFR mutation and EML4-ALK. Any clinical benefit using EGFR-TKI was not obtained in our case. The therapeutic choice for the patients with more than one driver mutations is unclear. We needs further understanding of the lung cancer molecular biology and the biomarker infomation

  10. SH2/SH3 adaptor proteins can link tyrosine kinases to a Ste20-related protein kinase, HPK1.

    Science.gov (United States)

    Anafi, M; Kiefer, F; Gish, G D; Mbamalu, G; Iscove, N N; Pawson, T

    1997-10-31

    Ste20-related protein kinases have been implicated as regulating a range of cellular responses, including stress-activated protein kinase pathways and the control of cytoskeletal architecture. An important issue involves the identities of the upstream signals and regulators that might control the biological functions of mammalian Ste20-related protein kinases. HPK1 is a protein-serine/threonine kinase that possesses a Ste20-like kinase domain, and in transfected cells activates a protein kinase pathway leading to the stress-activated protein kinase SAPK/JNK. Here we have investigated candidate upstream regulators that might interact with HPK1. HPK1 possesses an N-terminal catalytic domain and an extended C-terminal tail with four proline-rich motifs. The SH3 domains of Grb2 bound in vitro to specific proline-rich motifs in the HPK1 tail and functioned synergistically to direct the stable binding of Grb2 to HPK1 in transfected Cos1 cells. Epidermal growth factor (EGF) stimulation did not affect the binding of Grb2 to HPK1 but induced recruitment of the Grb2.HPK1 complex to the autophosphorylated EGF receptor and to the Shc docking protein. Several activated receptor and cytoplasmic tyrosine kinases, including the EGF receptor, stimulated the tyrosine phosphorylation of the HPK1 serine/threonine kinase. These results suggest that HPK1, a mammalian Ste20-related protein-serine/threonine kinase, can potentially associate with protein-tyrosine kinases through interactions mediated by SH2/SH3 adaptors such as Grb2. Such interaction may provide a possible mechanism for cross-talk between distinct biochemical pathways following the activation of tyrosine kinases.

  11. Structural basis for the regulation mechanism of the tyrosine kinase CapB from Staphylococcus aureus.

    Directory of Open Access Journals (Sweden)

    Vanesa Olivares-Illana

    2008-06-01

    Full Text Available Bacteria were thought to be devoid of tyrosine-phosphorylating enzymes. However, several tyrosine kinases without similarity to their eukaryotic counterparts have recently been identified in bacteria. They are involved in many physiological processes, but their accurate functions remain poorly understood due to slow progress in their structural characterization. They have been best characterized as copolymerases involved in the synthesis and export of extracellular polysaccharides. These compounds play critical roles in the virulence of pathogenic bacteria, and bacterial tyrosine kinases can thus be considered as potential therapeutic targets. Here, we present the crystal structures of the phosphorylated and unphosphorylated states of the tyrosine kinase CapB from the human pathogen Staphylococcus aureus together with the activator domain of its cognate transmembrane modulator CapA. This first high-resolution structure of a bacterial tyrosine kinase reveals a 230-kDa ring-shaped octamer that dissociates upon intermolecular autophosphorylation. These observations provide a molecular basis for the regulation mechanism of the bacterial tyrosine kinases and give insights into their copolymerase function.

  12. Effects of icotinib on advanced non-small cell lung cancer with different EGFR phenotypes.

    Science.gov (United States)

    Pan, Huiyun; Liu, Rong; Li, Shengjie; Fang, Hui; Wang, Ziwei; Huang, Sheng; Zhou, Jianying

    2014-09-01

    Icotinib is the first oral epidermal growth factor receptor (EGFR) tyrosine kinase receptor inhibitor, which has been proven to exert significant inhibitory effects on non-small cell lung cancer in vitro. Clinical evidence has showed that the efficacy of Icotinib on retreating advanced non-small cell lung cancer is comparable to Gefitinib. However, different phenotypes of EGFR can affect the therapeutic outcomes of EGFR tyrosine kinase receptor inhibitor. Therefore, our study focused on efficacy and safety of Icotinib in patients with advanced non-small cell lung cancer of different EGPR phenotypes. Clinical data of patients with advanced non-small cell lung cancer who received Icotinib treatment from August, 2011 to May, 2013 were retrospectively analyzed. Kaplan-Meier analysis was used for survival analysis and comparison. 18 wild-type EGFR and 51 mutant type were found in a total of 69 patients. Objective response rate of patients with mutant type EGFR was 54.9 % and disease control rate was 86.3 %. Objective response rate of wild-type patients was 11.1 % (P = 0.0013 vs mutant type), disease control rate was 50.0 % (P = 0.0017). Median progression-free survival (PFS) of mutant type and wild-type patients were 9.7 and 2.6 months, respectively (P Icotinib included rash, diarrhea, itching skin with occurrence rates of 24.6 % (17/69), 13.0 % (9/69), and 11.6 % (8/69), respectively. Most adverse reactions were grade I-II. Icotinib has great efficacy in EGFR mutated patients, making it an optimal regimen to treat EGFR mutated patients. Furthermore, most of adverse reactions associated with Icotinib treatment were tolerable.

  13. Tyrosine Kinase Gene Expression Profiling in Prostate Cancer

    National Research Council Canada - National Science Library

    Weier, Heinz-Ulrich

    2001-01-01

    ... of these genes parallels the progression of tumors to a more malignant phenotype. We developed a DNA micro-array based screening system to monitor the level of expression of tyrosine kinase (tk...

  14. Tyrosine Kinase Gene Expression Profiling in Prostate Cancer

    National Research Council Canada - National Science Library

    Weier, Heinz-Ulrich

    2002-01-01

    ... of these genes parallels the progression of tumors to a more malignant phenotype. We developed a DNA micro-array based screening system to monitor the level of expression of tyrosine kinase (tk...

  15. Anterior Gradient 2 (AGR2) Induced Epidermal Growth Factor Receptor (EGFR) Signaling Is Essential for Murine Pancreatitis-Associated Tissue Regeneration

    Science.gov (United States)

    Wodziak, Dariusz; Dong, Aiwen; Basin, Michael F.; Lowe, Anson W.

    2016-01-01

    A recently published study identified Anterior Gradient 2 (AGR2) as a regulator of EGFR signaling by promoting receptor presentation from the endoplasmic reticulum to the cell surface. AGR2 also promotes tissue regeneration in amphibians and fish. Whether AGR2-induced EGFR signaling is essential for tissue regeneration in higher vertebrates was evaluated using a well-characterized murine model for pancreatitis. The impact of AGR2 expression and EGFR signaling on tissue regeneration was evaluated using the caerulein-induced pancreatitis mouse model. EGFR signaling and cell proliferation were examined in the context of the AGR2-/- null mouse or with the EGFR-specific tyrosine kinase inhibitor, AG1478. In addition, the Hippo signaling coactivator YAP1 was evaluated in the context of AGR2 expression during pancreatitis. Pancreatitis-induced AGR2 expression enabled EGFR translocation to the plasma membrane, the initiation of cell signaling, and cell proliferation. EGFR signaling and tissue regeneration were partially inhibited by the tyrosine kinase inhibitor AG1478, but absent in the AGR2-/- null mouse. AG1478-treated and AGR2-/- null mice with pancreatitis died whereas all wild-type controls recovered. YAP1 activation was also dependent on pancreatitis-induced AGR2 expression. AGR2-induced EGFR signaling was essential for tissue regeneration and recovery from pancreatitis. The results establish tissue regeneration as a major function of AGR2-induced EGFR signaling in adult higher vertebrates. Enhanced AGR2 expression and EGFR signaling are also universally present in human pancreatic cancer, which support a linkage between tissue injury, regeneration, and cancer pathogenesis. PMID:27764193

  16. Identifying EGFR-Expressed Cells and Detecting EGFR Multi-Mutations at Single-Cell Level by Microfluidic Chip

    Science.gov (United States)

    Li, Ren; Zhou, Mingxing; Li, Jine; Wang, Zihua; Zhang, Weikai; Yue, Chunyan; Ma, Yan; Peng, Hailin; Wei, Zewen; Hu, Zhiyuan

    2018-03-01

    EGFR mutations companion diagnostics have been proved to be crucial for the efficacy of tyrosine kinase inhibitor targeted cancer therapies. To uncover multiple mutations occurred in minority of EGFR-mutated cells, which may be covered by the noises from majority of un-mutated cells, is currently becoming an urgent clinical requirement. Here we present the validation of a microfluidic-chip-based method for detecting EGFR multi-mutations at single-cell level. By trapping and immunofluorescently imaging single cells in specifically designed silicon microwells, the EGFR-expressed cells were easily identified. By in situ lysing single cells, the cell lysates of EGFR-expressed cells were retrieved without cross-contamination. Benefited from excluding the noise from cells without EGFR expression, the simple and cost-effective Sanger's sequencing, but not the expensive deep sequencing of the whole cell population, was used to discover multi-mutations. We verified the new method with precisely discovering three most important EGFR drug-related mutations from a sample in which EGFR-mutated cells only account for a small percentage of whole cell population. The microfluidic chip is capable of discovering not only the existence of specific EGFR multi-mutations, but also other valuable single-cell-level information: on which specific cells the mutations occurred, or whether different mutations coexist on the same cells. This microfluidic chip constitutes a promising method to promote simple and cost-effective Sanger's sequencing to be a routine test before performing targeted cancer therapy.[Figure not available: see fulltext.

  17. Giant hub Src and Syk tyrosine kinase thermodynamic profiles recapitulate evolution

    Science.gov (United States)

    Phillips, J. C.

    2017-10-01

    Thermodynamic scaling theory, previously applied mainly to small proteins, here analyzes quantitative evolution of the titled functional network giant hub enzymes. The broad domain structure identified homologically is confirmed hydropathically using amino acid sequences only. The most surprising results concern the evolution of the tyrosine kinase globular surface roughness from avians to mammals, which is first order, compared to the evolution within mammals from rodents to humans, which is second order. The mystery of the unique amide terminal region of proto oncogene tyrosine protein kinase is resolved by the discovery there of a rare hydroneutral septad targeting cluster, which is paralleled by an equally rare octad catalytic cluster in tyrosine kinase in humans and a few other species (cat and dog). These results, which go far towards explaining why these proteins are among the largest giant hubs in protein interaction networks, use no adjustable parameters.

  18. p-Benzoquinone initiates non-invasive urothelial cancer through aberrant tyrosine phosphorylation of EGFR, MAP kinase activation and cell cycle deregulation: Prevention by vitamin C

    Directory of Open Access Journals (Sweden)

    Shinjini Ganguly

    Full Text Available According to WHO classification system, non-invasive urothelial carcinoma represents urothelial carcinoma in situ (CIS and dysplasia. Dysplastic urothelium often progresses to CIS that further advances to urothelial carcinoma (UC. The strongest risk factor for UC is cigarette smoking. However, the pathogenesis of cigarette smoke (CS-induced UC is poorly understood. Earlier we had shown that p-benzoquinone (p-BQ, a major toxic quinone derived from p-benzosemiquinone of CS in vivo, is a causative factor for various CS-induced diseases. Here, using a guinea pig model we showed that prolonged treatment with p-BQ led to non-invasive UC, specifically carcinoma in situ (CIS of the renal pelvis and dysplasia in the ureter and bladder. The mechanisms of carcinogenesis were p-BQ-induced oxidative damage and apoptosis that were later suppressed and followed by activation of epidermal growth factor receptor, aberrant phosphorylation of intracellular tyrosine residues, activation of MAP kinase pathway and persistent growth signaling. This was accompanied by deregulation of cell cycle as shown by marked decrease in the expression of p21waf1/cip1 and cyclin D1 proteins as well as hyperphosphorylation of pRb. UC has been characterised by histopathology and immunohistochemistry showing aberrant CK20, increased Ki-67, and marked p53 nuclear immunopositivity with uniformly negative labelling of CD44. Oral supplementation of vitamin C (30 mg/kg body weight/day prevented CIS of the renal pelvis and dysplasia in the ureter and bladder. Since majority of non-invasive UC progresses to invasive cancer with increased risk of mortality, our preclinical study might help to devise effective strategies for early intervention of the disease. Abbreviations: Bax, Bcl-2, CS, DNPH, GAPDH, IARC, p-BQ, p-BSQ, PAHs, PBS, ROS, SDS PAGE, TUNEL, WHO, UC, CIS, EGFR, MAPK, Keywords: p-Benzoquinone, Carcinoma in situ, Dysplasia, Aberrant EGFR activation, Cell cycle deregulation

  19. SH2 domains: modulators of nonreceptor tyrosine kinase activity

    OpenAIRE

    Filippakopoulos, Panagis; Müller, Susanne; Knapp, Stefan

    2009-01-01

    The Src homology 2 (SH2) domain is a sequence-specific phosphotyrosine-binding module present in many signaling molecules. In cytoplasmic tyrosine kinases, the SH2 domain is located N-terminally to the catalytic kinase domain (SH1) where it mediates cellular localization, substrate recruitment, and regulation of kinase activity. Initially, structural studies established a role of the SH2 domain stabilizing the inactive state of Src family members. However, biochemical characterization showed ...

  20. The non-small cell lung cancer EGFR extracellular domain mutation, M277E, is oncogenic and drug-sensitive

    Directory of Open Access Journals (Sweden)

    Yu S

    2017-09-01

    Full Text Available Su Yu,1,2 Yang Zhang,1 Yunjian Pan,1 Chao Cheng,1,3 Yihua Sun,1,3 Haiquan Chen1–4 1Department of Thoracic Surgery, Fudan University Shanghai Cancer Center, Shanghai, China; 2Cancer Research Center, Fudan University Shanghai Cancer Center, Shanghai, China; 3Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China; 4Institutes of Biomedical Sciences, Fudan University, Shanghai, China Purpose: To identify novel oncogenic mutations in non-small cell lung cancer patient specimens that lack mutations in known targetable genes (“pan-negative” patients.Methods: Comprehensive mutational analyses were performed on 1,356 lung adenocarcinoma specimens. In this cohort of patients, common lung cancer oncogenic driver mutations were detected in the epidermal growth factor receptor (EGFR kinase domain, the human epidermal growth factor receptor 2 kinase domain, as well as the KRAS, BRAF, ALK, ROS1 and RET genes. A sub-cohort of pan-negative patient specimens was assayed for mutations in the EGFR extracellular domain (ECD. Additionally, EGFR mutant NIH-3T3 stable cell lines were constructed and assessed for protein content, anchorage-independent growth, and tumor formation in xenograft models to identify oncogenic mutations. BaF3 lymphocytes were also used to test sensitivities of the mutations to tyrosine kinase inhibitors.Results: In pan-negative lung adenocarcinoma cases, a novel oncogenic EGFR ECD mutation was identified (M277E. EGFR M277E mutations encoded oncoproteins that transformed NIH-3T3 cells to grow in the absence of exogenous epidermal growth factor. Transformation was further evidenced by anchorage-independent growth and tumor formation in immunocompromised xenograft mouse models. Finally, as seen in the canonical EGFR L858R mutation, the M277E mutation conferred sensitivity to both erlotinib and cetuximab in BaF3 cell lines and to erlotinib in xenograft models.Conclusion: Here, a new EGFR driver mutation, M277E

  1. Ubiquitin Ligase Cbl-b Is Involved in Icotinib (BPI-2009H)-Induced Apoptosis and G1 Phase Arrest of EGFR Mutation-Positive Non-Small-Cell Lung Cancer

    OpenAIRE

    Mu, Xiaodong; Zhang, Ye; Qu, Xiujuan; Hou, Kezuo; Kang, Jian; Hu, Xuejun; Liu, Yunpeng

    2013-01-01

    Epidermal growth factor receptor (EGFR) is one of the most promising targets for non-small-cell lung cancer (NSCLC). Icotinib, a highly selective EGFR tyrosine kinase inhibitor (EGFR-TKI), has shown promising clinical efficacy and safety in patients with NSCLC. The exact molecular mechanism of icotinib remains unclear. In this study, we first investigated the antiproliferative effect of icotinib on NSCLC cells. Icotinib significantly inhibited proliferation of the EGFR-mutated lung cancer HCC...

  2. Fully automated synthesis of (phosphopeptide arrays in microtiter plate wells provides efficient access to protein tyrosine kinase characterization

    Directory of Open Access Journals (Sweden)

    Goldstein David J

    2005-01-01

    Full Text Available Abstract Background Synthetic peptides have played a useful role in studies of protein kinase substrates and interaction domains. Synthetic peptide arrays and libraries, in particular, have accelerated the process. Several factors have hindered or limited the applicability of various techniques, such as the need for deconvolution of combinatorial libraries, the inability or impracticality of achieving full automation using two-dimensional or pin solid phases, the lack of convenient interfacing with standard analytical platforms, or the difficulty of compartmentalization of a planar surface when contact between assay components needs to be avoided. This paper describes a process for synthesis of peptides and phosphopeptides on microtiter plate wells that overcomes previous limitations and demonstrates utility in determination of the epitope of an autophosphorylation site phospho-motif antibody and utility in substrate utilization assays of the protein tyrosine kinase, p60c-src. Results The overall reproducibility of phospho-peptide synthesis and multiplexed EGF receptor (EGFR autophosphorylation site (pY1173 antibody ELISA (9H2 was within 5.5 to 8.0%. Mass spectrometric analyses of the released (phosphopeptides showed homogeneous peaks of the expected molecular weights. An overlapping peptide array of the complete EGFR cytoplasmic sequence revealed a high redundancy of 9H2 reactive sites. The eight reactive phospopeptides were structurally related and interestingly, the most conserved antibody reactive peptide motif coincided with a subset of other known EGFR autophosphorylation and SH2 binding motifs and an EGFR optimal substrate motif. Finally, peptides based on known substrate specificities of c-src and related enzymes were synthesized in microtiter plate array format and were phosphorylated by c-Src with the predicted specificities. The level of phosphorylation was proportional to c-Src concentration with sensitivities below 0.1 Units of

  3. Association between BIM deletion polymorphism and clinical outcome of EGFR-mutated NSCLC patient with EGFR-TKI therapy: A meta-analysis.

    Science.gov (United States)

    Ma, Ji-Yong; Yan, Hai-Jun; Gu, Wei

    2015-01-01

    BIM deletion polymorphism was deemed to be associated with downregulation of BIM, resulting in a decreased apoptosis induced by epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) in EGFR mutation-positive non-small cell lung cancer (NSCLC). However, accumulating evidences concerning the association between BIM deletion polymorphism and efficacy of EGFR-TKI and survival in EGFR-mutation-driven NSCLC patient reported contradictory results. A meta-analysis was conducted by combing six original eligible studies including 871 NSCLC patients. Our study showed that BIM deletion polymorphism was significantly associated with poor response to EGFR-TKI therapy in mutant EGFRNSCLC patients (P(h) = 0.309, P(z) = 0.001, OR = 0.39, 95% confidence interval (CI) = 0.23-0.67). Disease control rate (DCR) in mutant EGFRNSCLC patient with treatment of EGFR-TKI was significantly decreased in patients with BIM deletion polymorphism comparing to patients harbored BIM wild variant (P(h) = 0.583, P(Z) = 0.007, OR = 0.46, 95%CI = 0.25-0.85). EGFR mutation-derived NSCLC patient carrying BIM deletion polymorphism had a shorter progression-free survival (PFS; P(h) deletion polymorphism might be a cause that contributes to primary EGFR-TKI resistance, and it could be used as a genetic predictor for EGFR-TKI outcome and an independent prognostic factor of EGFR mutation-driven NSCLC patient.

  4. Tyrosine-phosphorylation of AAV2 vectors and its consequences on viral intracellular trafficking and transgene expression

    OpenAIRE

    Zhong, Li; Li, Baozheng; Jayandharan, Giridhararao; Mah, Cathryn S.; Govindasamy, Lakshmanan; Agbandje-McKenna, Mavis; Herzog, Roland W.; Weigel-Van Aken, Kirsten A.; Hobbs, Jacqueline A.; Zolotukhin, Sergei; Muzyczka, Nicholas; Srivastava, Arun

    2008-01-01

    We have documented that epidermal growth factor receptor protein tyrosine kinase (EGFR-PTK) signaling negatively affects intracellular trafficking and transduction efficiency of recombinant adeno-associated virus 2 (AAV2) vectors. Specifically, inhibition of EGFR-PTK signaling leads to decreased ubiquitination of AAV2 capsid proteins, which in turn, facilitates viral nuclear transport by limiting proteasome-mediated degradation of AAV2 vectors. In the present studies, we observed that AAV cap...

  5. δ-Tocopherol inhibits receptor tyrosine kinase-induced AKT activation in prostate cancer cells.

    Science.gov (United States)

    Wang, Hong; Hong, Jungil; Yang, Chung S

    2016-11-01

    The cancer preventive activity of vitamin E is suggested by epidemiological studies and supported by animal studies with vitamin E forms, γ-tocopherol and δ-tocopherol (δ-T). Several recent large-scale cancer prevention trials with high dose of α-tocopherol, however, yielded disappointing results. Whether vitamin E prevents or promotes cancer is a serious concern. A better understanding of the molecular mechanisms of action of the different forms of tocopherols would enhance our understanding of this topic. In this study, we demonstrated that δ-T was the most effective tocopherol form in inhibiting prostate cancer cell growth, by inducing cell cycle arrest and apoptosis. By profiling the effects of δ-T on the cell signaling using the phospho-kinase array, we found that the most inhibited target was the phosphorylation of AKT on T308. Further study on the activation of AKT by EGFR and IGFR revealed that δ-T attenuated the EGF/IGF-induced activation of AKT (via the phosphorylation of AKT on T308 induced by the activation of PIK3). Expression of dominant active PIK3 and AKT in prostate cancer cell line DU145 in which PIK3, AKT, and PTEN are wild type caused the cells to be reflectory to the inhibition of δ-T, supporting that δ-T inhibits the PIK3-mediated activation of AKT. Our data also suggest that δ-T interferes with the EGF-induced EGFR internalization, which leads to the inhibition of the receptor tyrosine kinase-dependent activation of AKT. In summary, our results revealed a novel mechanism of δ-T in inhibiting prostate cancer cell growth, supporting the cancer preventive activity δ-T. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

  6. Role of Bruton's tyrosine kinase in B cells and malignancies

    NARCIS (Netherlands)

    Pal Singh, S. (Simar); F. Dammeijer (Floris); R.W. Hendriks (Rudi)

    2018-01-01

    textabstractBruton's tyrosine kinase (BTK) is a non-receptor kinase that plays a crucial role in oncogenic signaling that is critical for proliferation and survival of leukemic cells in many B cell malignancies. BTK was initially shown to be defective in the primary immunodeficiency X-linked

  7. Characterization and response of newly developed high-grade glioma cultures to the tyrosine kinase inhibitors, erlotinib, gefitinib and imatinib

    International Nuclear Information System (INIS)

    Kinsella, Paula; Howley, Rachel; Doolan, Padraig; Clarke, Colin; Madden, Stephen F.; Clynes, Martin; Farrell, Michael; Amberger-Murphy, Verena

    2012-01-01

    High-grade gliomas (HGG), are the most common aggressive brain tumours in adults. Inhibitors targeting growth factor signalling pathways in glioma have shown a low clinical response rate. To accurately evaluate response to targeted therapies further in vitro studies are necessary. Growth factor pathway expression using epidermal growth factor receptor (EGFR), mutant EGFR (EGFRvIII), platelet derived growth factor receptor (PDGFR), C-Kit and C-Abl together with phosphatase and tensin homolog (PTEN) expression and downstream activation of AKT and phosphorylated ribosomal protein S6 (P70S6K) was analysed in 26 primary glioma cultures treated with the tyrosine kinase inhibitors (TKIs) erlotinib, gefitinib and imatinib. Response to TKIs was assessed using 50% inhibitory concentrations (IC 50 ). Response for each culture was compared with the EGFR/PDGFR immunocytochemical pathway profile using hierarchical cluster analysis (HCA) and principal component analysis (PCA). Erlotinib response was not strongly associated with high expression of the growth factor pathway components. PTEN expression did not correlate with response to any of the three TKIs. Increased EGFR expression was associated with gefitinib response; increased PDGFR-α expression was associated with imatinib response. The results of this in vitro study suggest gefitinib and imatinib may have therapeutic potential in HGG tumours with a corresponding growth factor receptor expression profile. -- Highlights: ► Non-responders had low EGFR expression, high PDGFR-β, and a low proliferation rate. ► PTEN is not indicative of response to a TKI. ► Erlotinib response was not associated with expression of the proteins examined. ► Imatinib-response correlated with expression of PDGFR-α. ► Gefitinib response correlated with increased expression of EGFR.

  8. Collagen type I induces EGFR-TKI resistance in EGFR-mutated cancer cells by mTOR activation through Akt-independent pathway.

    Science.gov (United States)

    Yamazaki, Shota; Higuchi, Youichi; Ishibashi, Masayuki; Hashimoto, Hiroko; Yasunaga, Masahiro; Matsumura, Yasuhiro; Tsuchihara, Katsuya; Tsuboi, Masahiro; Goto, Koichi; Ochiai, Atsushi; Ishii, Genichiro

    2018-06-01

    Primary resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) is a serious problem in lung adenocarcinoma patients harboring EGFR mutations. The aim of this study was to examine whether and how collagen type I (Col I), the most abundantly deposited matrix in tumor stroma, affects EGFR-TKI sensitivity in EGFR-mutant cells. We evaluated the EGFR-TKI sensitivity of EGFR-mutated cancer cells cultured with Col I. Changes in the activation of downstream signaling molecules of EGFR were analyzed. We also examined the association between the Col I expression in tumor stroma in surgical specimens and EGFR-TKI response of postoperative recurrence patients with EGFR mutations. Compared to cancer cells without Col I, the survival rate of cancer cells cultured with Col I was significantly higher after EGFR-TKI treatment. In cancer cells cultured with and without Col I, EGFR-TKI suppressed the levels of phosphorylated (p-)EGFR, p-ERK1/2, and p-Akt. When compared to cancer cells without Col I, expression of p-P70S6K, a hallmark of mTOR activation, was dramatically upregulated in cancer cells with Col I. This activation was maintained even after EGFR-TKI treatment. Simultaneous treatment with EGFR-TKI and mTOR inhibitor abrogated Col I-induced resistance to EGFR-TKI. Patients with Col I-rich stroma had a significantly shorter progression-free survival time after EGFR-TKI therapy (238 days vs 404 days; P Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  9. Genetic Association of Curative and Adverse Reactions to Tyrosine Kinase Inhibitors in Chinese advanced Non-Small Cell Lung Cancer patients

    Science.gov (United States)

    Ruan, Yunfeng; Jiang, Jie; Guo, Liang; Li, Yan; Huang, Hailiang; Shen, Lu; Luan, Mengqi; Li, Mo; Du, Huihui; Ma, Cheng; He, Lin; Zhang, Xiaoqing; Qin, Shengying

    2016-01-01

    Epidermal growth factor receptor (EGFR) Tyrosine kinase inhibitor (TKI) is an effective targeted therapy for advanced non-small cell lung cancer (NSCLC) but also causes adverse drug reactions (ADRs) e.g., skin rash and diarrhea. SNPs in the EGFR signal pathway, drug metabolism/ transport pathways and miRNA might contribute to the interpersonal difference in ADRs but biomarkers for therapeutic responses and ADRs to TKIs in Chinese population are yet to be fully investigated. We recruited 226 Chinese advanced NSCLC patients who received TKIs erlotinib, gefitinib and icotinib hydrochloride and systematically studied the genetic factors associated with therapeutic responses and ADRs. Rs884225 (T > C) in EGFR 3′ UTR was significantly associated with lower risk of ADRs to erlotinib (p value = 0.0010, adjusted p value = 0.042). A multivariant interaction four-SNP model (rs884225 in EGFR 3′UTR, rs7787082 in ABCB1 intron, rs38845 in MET intron and rs3803300 in AKT1 5′UTR) was associated with ADRs in general and the more specific drug induced skin injury. The SNPs associated with both therapeutic responses and ADRs indicates they might share a common genetic basis. Our study provided potential biomarkers and clues for further research of biomarkers for therapeutic responses and ADRs in Chinese NSCLC patients. PMID:26988277

  10. Interplay between TGF-β signaling and receptor tyrosine kinases in tumor development.

    Science.gov (United States)

    Shi, Qiaoni; Chen, Ye-Guang

    2017-10-01

    Transforming growth factor-β (TGF-β) signaling regulates cell proliferation, differentiation, migration and death, and plays a critical role in embryogenesis and tissue homeostasis. Its deregulation results in various diseases including tumor formation. Receptor tyrosine kinases (RTKs), such as epidermal growth factor receptor (EGFR), fibroblast growth factor receptor (FGFR), vascular endothelial growth factor receptor (VEGFR) and platelet-derived growth factor receptor (PDGFR), also play key roles in the development and progression of many types of tumors. It has been realized that TGF-β signaling and RTK pathways interact with each other and their interplay is important for cancer development. They are mutually regulated and cooperatively modulate cell survival and migration, epithelial-mesenchymal transition, and tumor microenvironment to accelerate tumorigenesis and tumor metastasis. RTKs can modulate Smad-dependent transcription or cooperate with TGF-β to potentiate its oncogenic activity, while TGF-β signaling can in turn control RTK signaling by regulating their activities or expression. This review summarizes current understandings of the interplay between TGF-β signaling and RTKs and its influence on tumor development.

  11. Predictive efficacy of low burden EGFR mutation detected by next-generation sequencing on response to EGFR tyrosine kinase inhibitors in non-small-cell lung carcinoma.

    Directory of Open Access Journals (Sweden)

    Hye Sook Kim

    Full Text Available Direct sequencing remains the most widely used method for the detection of epidermal growth factor receptor (EGFR mutations in lung cancer; however, its relatively low sensitivity limits its clinical use. The objective of this study was to investigate the sensitivity of detecting an epidermal growth factor receptor (EGFR mutation from peptide nucleic acid-locked nucleic acid polymerase chain reaction (PNA-LNA PCR clamp and Ion Torrent Personal Genome Machine (PGM techniques compared to that by direct sequencing. Furthermore, the predictive efficacy of EGFR mutations detected by PNA-LNA PCR clamp was evaluated. EGFR mutational status was assessed by direct sequencing, PNA-LNA PCR clamp, and Ion Torrent PGM in 57 patients with non-small cell lung cancer (NSCLC. We evaluated the predictive efficacy of PNA-LNA PCR clamp on the EGFR-TKI treatment in 36 patients with advanced NSCLC retrospectively. Compared to direct sequencing (16/57, 28.1%, PNA-LNA PCR clamp (27/57, 47.4% and Ion Torrent PGM (26/57, 45.6% detected more EGFR mutations. EGFR mutant patients had significantly longer progressive free survival (14.31 vs. 21.61 months, P = 0.003 than that of EGFR wild patients when tested with PNA-LNA PCR clamp. However, no difference in response rate to EGFR TKIs (75.0% vs. 82.4%, P = 0.195 or overall survival (34.39 vs. 44.10 months, P = 0.422 was observed between the EGFR mutations by direct sequencing or PNA-LNA PCR clamp. Our results demonstrate firstly that patients with EGFR mutations were detected more frequently by PNA-LNA PCR clamp and Ion Torrent PGM than those by direct sequencing. EGFR mutations detected by PNA-LNA PCR clamp may be as a predicative factor for EGFR TKI response in patients with NSCLC.

  12. DMPD: Macrophage-stimulating protein and RON receptor tyrosine kinase: potentialregulators of macrophage inflammatory activities. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 12472665 Macrophage-stimulating protein and RON receptor tyrosine kinase: potential...:545-53. (.png) (.svg) (.html) (.csml) Show Macrophage-stimulating protein and RON receptor tyrosine kinase:...le Macrophage-stimulating protein and RON receptor tyrosine kinase: potentialregulators of macrophage inflam

  13. Exploring the Hypersensitivity of PTEN Deleted Prostate Cancer Stem Cells to WEE1 Tyrosine Kinase Inhibitors

    Science.gov (United States)

    2015-12-01

    AWARD NUMBER: W81XWH-14-1-0251 TITLE: Exploring the Hypersensitivity of PTEN Deleted Prostate Cancer Stem Cells to WEE1 Tyrosine Kinase... Tyrosine Kinase Inhibitors 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-14-1-0251 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) Kiran Mahajan 5d...ABSTRACT Central to all cycling cells-including prostate cancer stem cells- is the expression of WEE1 tyrosine kinase. WEE1 monitors duplication of

  14. Genomic activation of the EGFR and HER2-neu genes in a significant proportion of invasive epithelial ovarian cancers

    Directory of Open Access Journals (Sweden)

    Ghislain Vanessa

    2008-01-01

    Full Text Available Abstract Background The status of the EGFR and HER2-neu genes has not been fully defined in ovarian cancer. An integrated analysis of both genes could help define the proportion of patients that would potentially benefit from targeted therapies. Methods We determined the tumour mutation status of the entire tyrosine kinase (TK domain of the EGFR and HER2-neu genes in a cohort of 52 patients with invasive epithelial ovarian cancer as well as the gene copy number and protein expression of both genes in 31 of these patients by DGGE and direct sequecing, immunohistochemistry and Fluorescent in Situ Hybridisation (FISH. Results The EGFR was expressed in 59% of the cases, with a 2+/3+ staining intensity in 38%. HER2-neu expression was found in 35%, with a 2/3+ staining in 18%. No mutations were found in exons 18–24 of the TK domains of EGFR and HER2-neu. High polysomy of the EGFR gene was observed in 13% of the invasive epthelial cancers and amplification of the HER2-neu gene was found in 10% and correlated with a high expression level by immunohistochemistry. Mutations within the tyrosine kinase domain were not found in the entire TK domain of both genes, but have been found in very rare cases by others. Conclusion Genomic alteration of the HER2-neu and EGFR genes is frequent (25% in ovarian cancer. EGFR/HER2-neu targeted therapies should be investigated prospectively and specifically in that subset of patients.

  15. Oncogenic Receptor Tyrosine Kinases Directly Phosphorylate Focal Adhesion Kinase (FAK) as a Resistance Mechanism to FAK-Kinase Inhibitors.

    Science.gov (United States)

    Marlowe, Timothy A; Lenzo, Felicia L; Figel, Sheila A; Grapes, Abigail T; Cance, William G

    2016-12-01

    Focal adhesion kinase (FAK) is a major drug target in cancer and current inhibitors targeted to the ATP-binding pocket of the kinase domain have entered clinical trials. However, preliminary results have shown limited single-agent efficacy in patients. Despite these unfavorable data, the molecular mechanisms that drive intrinsic and acquired resistance to FAK-kinase inhibitors are largely unknown. We have demonstrated that receptor tyrosine kinases (RTK) can directly bypass FAK-kinase inhibition in cancer cells through phosphorylation of FAK's critical tyrosine 397 (Y397). We also showed that HER2 forms a direct protein-protein interaction with the FAK-FERM-F1 lobe, promoting direct phosphorylation of Y397. In addition, FAK-kinase inhibition induced two forms of compensatory RTK reprogramming: (i) the rapid phosphorylation and activation of RTK signaling pathways in RTK High cells and (ii) the long-term acquisition of RTKs novel to the parental cell line in RTK Low cells. Finally, HER2 +: cancer cells displayed resistance to FAK-kinase inhibition in 3D growth assays using a HER2 isogenic system and HER2 + cancer cell lines. Our data indicate a novel drug resistance mechanism to FAK-kinase inhibitors whereby HER2 and other RTKs can rescue and maintain FAK activation (pY397) even in the presence of FAK-kinase inhibition. These data may have important ramifications for existing clinical trials of FAK inhibitors and suggest that individual tumor stratification by RTK expression would be important to predict patient response to FAK-kinase inhibitors. Mol Cancer Ther; 15(12); 3028-39. ©2016 AACR. ©2016 American Association for Cancer Research.

  16. BCR-ABL1 tyrosine kinase inhibitors for the treatment of chronic myeloid leukemia.

    Science.gov (United States)

    Cuellar, Sandra; Vozniak, Michael; Rhodes, Jill; Forcello, Nicholas; Olszta, Daniel

    2017-01-01

    The management of chronic myeloid leukemia with BCR-ABL1 tyrosine kinase inhibitors has evolved chronic myeloid leukemia into a chronic, manageable disease. A patient-centered approach is important for the appropriate management of chronic myeloid leukemia and optimization of long-term treatment outcomes. The pharmacist plays a key role in treatment selection, monitoring drug-drug interactions, identification and management of adverse events, and educating patients on adherence. The combination of tyrosine kinase inhibitors with unique safety profiles and individual patients with unique medical histories can make managing treatment difficult. This review will provide up-to-date information regarding tyrosine kinase inhibitor-based treatment of patients with chronic myeloid leukemia. Management strategies for adverse events and considerations for drug-drug interactions will not only vary among patients but also across tyrosine kinase inhibitors. Drug-drug interactions can be mild to severe. In instances where co-administration of concomitant medications cannot be avoided, it is critical to understand how drug levels are impacted and how subsequent dose modifications ensure therapeutic drug levels are maintained. An important component of patient-centered management of chronic myeloid leukemia also includes educating patients on the significance of early and regular monitoring of therapeutic milestones, emphasizing the importance of adhering to treatment in achieving these targets, and appropriately modifying treatment if these clinical goals are not being met. Overall, staying apprised of current research, utilizing the close pharmacist-patient relationship, and having regular interactions with patients, will help achieve successful long-term treatment of chronic myeloid leukemia in the age of BCR-ABL1 tyrosine kinase inhibitors.

  17. Chemotherapeutics-resistance "arms" race: An update on mechanisms involved in resistance limiting EGFR inhibitors in lung cancer.

    Science.gov (United States)

    Singh, Pankaj Kumar; Silakari, Om

    2017-10-01

    Clinical reports suggest that EGFR-mutated lung cancer usually respond significantly towards small molecule tyrosine kinase inhibitors. Same studies also report the eventual development of acquired resistance within a median time interval of 9 to 14months. One of the major mechanisms involved in this acquired resistance was found to be a secondary point mutation at gate-keeper residue, EGFR T790M. However, there are other recent studies which disclose the role of few other novel key players such as, ZEB1, TOPK etc., in the development of tolerance towards the EGFR TKI's, along with other commonly known mechanisms, such as amplification of signalling pathways such as, c-MET, Erbb2, AXL, additional acquired secondary mutations (PIK3CA, BRAF), or phenotypic transformation (small cell or epithelial to mesenchymal transitions). Interestingly, a recent study showed development of resistance via another point mutation, C797S, in case of tumors which were previously resistant and were administered agents capable of overcoming T790M gatekeeper mutation based resistance. Thus, raising serious concern over the direction of drug development involving tyrosine kinases such as EGFR. Current approaches focussing on development of third generation inhibitors, dual inhibitors or inhibitors of HSP90 have shown significant activity but do not answer the long term question of resistance. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Structural basis for the regulation mechanism of the tyrosine kinase CapB from Staphylococcus aureus

    DEFF Research Database (Denmark)

    Olivares-Illana, Vanesa; Meyer, Philippe; Bechet, Emmanuelle

    2008-01-01

    Bacteria were thought to be devoid of tyrosine-phosphorylating enzymes. However, several tyrosine kinases without similarity to their eukaryotic counterparts have recently been identified in bacteria. They are involved in many physiological processes, but their accurate functions remain poorly...... understood due to slow progress in their structural characterization. They have been best characterized as copolymerases involved in the synthesis and export of extracellular polysaccharides. These compounds play critical roles in the virulence of pathogenic bacteria, and bacterial tyrosine kinases can thus...... be considered as potential therapeutic targets. Here, we present the crystal structures of the phosphorylated and unphosphorylated states of the tyrosine kinase CapB from the human pathogen Staphylococcus aureus together with the activator domain of its cognate transmembrane modulator CapA. This first high...

  19. Receptor protein tyrosine phosphatase alpha activates Src-family kinases and controls integrin-mediated responses in fibroblasts

    DEFF Research Database (Denmark)

    Su, J; Muranjan, M; Sap, J

    1999-01-01

    of tyrosine kinases, the activity of which is tightly controlled by inhibitory phosphorylation of a carboxyterminal tyrosine residue (Tyr527 in chicken c-Src); this phosphorylation induces the kinases to form an inactive conformation. Whereas the identity of such inhibitory Tyr527 kinases has been well...... established, no corresponding phosphatases have been identified that, under physiological conditions, function as positive regulators of c-Src and Fyn in fibroblasts. RESULTS: Receptor protein tyrosine phosphatase alpha (RPTPalpha) was inactivated by homologous recombination. Fibroblasts derived from...... these RPTPalpha-/- mice had impaired tyrosine kinase activity of both c-Src and Fyn, and this was accompanied by a concomitant increase in c-Src Tyr527 phosphorylation. RPTPalpha-/- fibroblasts also showed a reduction in the rate of spreading on fibronectin substrates, a trait that is a phenocopy of the effect...

  20. Navigating into the binding pockets of the HER family protein kinases: discovery of novel EGFR inhibitor as antitumor agent.

    Science.gov (United States)

    Liu, Wei; Ning, Jin-Feng; Meng, Qing-Wei; Hu, Jing; Zhao, Yan-Bin; Liu, Chao; Cai, Li

    2015-01-01

    The epidermal growth factor receptor (EGFR) family has been validated as a successful antitumor drug target for decades. Known EGFR inhibitors were exposed to distinct drug resistance against the various EGFR mutants within non-small-cell lung cancer (NSCLC), particularly the T790M mutation. Although so far a number of studies have been reported on the development of third-generation EGFR inhibitors for overcoming the resistance issue, the design procedure largely depends on the intuition of medicinal chemists. Here we retrospectively make a detailed analysis of the 42 EGFR family protein crystal complexes deposited in the Protein Data Bank (PDB). Based on the analysis of inhibitor binding modes in the kinase catalytic cleft, we identified a potent EGFR inhibitor (compound A-10) against drug-resistant EGFR through fragment-based drug design. This compound showed at least 30-fold more potency against EGFR T790M than the two control molecules erlotinib and gefitinib in vitro. Moreover, it could exhibit potent HER2 inhibitory activities as well as tumor growth inhibitory activity. Molecular docking studies revealed a structural basis for the increased potency and mutant selectivity of this compound. Compound A-10 may be selected as a promising candidate in further preclinical studies. In addition, our findings could provide a powerful strategy to identify novel selective kinase inhibitors on the basis of detailed kinase-ligand interaction space in the PDB.

  1. The mir-279/996 cluster represses receptor tyrosine kinase signaling to determine cell fates in the Drosophila eye.

    Science.gov (United States)

    Duan, Hong; de Navas, Luis F; Hu, Fuqu; Sun, Kailiang; Mavromatakis, Yannis E; Viets, Kayla; Zhou, Cyrus; Kavaler, Joshua; Johnston, Robert J; Tomlinson, Andrew; Lai, Eric C

    2018-04-09

    Photoreceptors in the crystalline Drosophila eye are recruited by receptor tyrosine kinase (RTK)/Ras signaling mediated by Epidermal growth factor receptor (EGFR) and the Sevenless (Sev) receptor. Analyses of an allelic deletion series of the mir-279/996 locus, along with a panel of modified genomic rescue transgenes, show that Drosophila eye patterning depends on both miRNAs. Transcriptional reporter and activity sensor transgenes reveal expression and function of miR-279/996 in non-neural cells of the developing eye. Moreover, mir-279/996 mutants exhibit substantial numbers of ectopic photoreceptors, particularly of R7, and cone cell loss. These miRNAs restrict RTK signaling in the eye, since mir-279/996 nulls are dominantly suppressed by positive components of the EGFR pathway and enhanced by heterozygosity for an EGFR repressor. miR-279/996 limit photoreceptor recruitment by targeting multiple positive RTK/Ras signaling components that promote photoreceptor/R7 specification. Strikingly, deletion of mir-279/996 sufficiently derepresses RTK/Ras signaling so as to rescue a population of R7 cells in R7-specific RTK null mutants boss and sev , which otherwise completely lack this cell fate. Altogether, we reveal a rare setting of developmental cell specification that involves substantial miRNA control. © 2018. Published by The Company of Biologists Ltd.

  2. Cross-phosphorylation of bacterial serine/threonine and tyrosine protein kinases on key regulatory residues

    Directory of Open Access Journals (Sweden)

    Lei eShi

    2014-09-01

    Full Text Available Bacteria possess protein serine/threonine and tyrosine kinases which resemble eukaryal kinases in their capacity to phosphorylate multiple substrates. We hypothesized that the analogy might extend further, and bacterial kinases may also undergo mutual phosphorylation and activation, which is currently considered as a hallmark of eukaryal kinase networks. In order to test this hypothesis, we explored the capacity of all members of four different classes of serine/threonine and tyrosine kinases present in the firmicute model organism Bacillus subtilis to phosphorylate each other in vitro and interact with each other in vivo. The interactomics data suggested a high degree of connectivity among all types of kinases, while phosphorylation assays revealed equally wide-spread cross-phosphorylation events. Our findings suggest that the Hanks-type kinases PrkC, PrkD and YabT exhibit the highest capacity to phosphorylate other B. subtilis kinases, while the BY-kinase PtkA and the two-component-like kinases RsbW and SpoIIAB show the highest propensity to be phosphorylated by other kinases. Analysis of phosphorylated residues on several selected recipient kinases suggests that most cross-phosphorylation events concern key regulatory residues. Therefore, cross-phosphorylation events are very likely to influence the capacity of recipient kinases to phosphorylate substrates downstream in the signal transduction cascade. We therefore conclude that bacterial serine/threonine and tyrosine kinases probably engage in a network-type behavior previously described only in eukaryal cells.

  3. Sapanisertib and Osimertinib in Treating Patients With Stage IV EGFR Mutation Positive Non-small Cell Lung Cancer After Progression on a Previous EGFR Tyrosine Kinase Inhibitor

    Science.gov (United States)

    2018-04-25

    EGFR Activating Mutation; EGFR Exon 19 Deletion Mutation; EGFR NP_005219.2:p.G719X; EGFR NP_005219.2:p.L858R; EGFR NP_005219.2:p.L861Q; EGFR T790M Mutation Negative; Recurrent Non-Small Cell Lung Carcinoma; Stage III Non-Small Cell Lung Cancer AJCC v7; Stage IIIA Non-Small Cell Lung Cancer AJCC v7; Stage IIIB Non-Small Cell Lung Cancer AJCC v7; Stage IV Non-Small Cell Lung Cancer AJCC v7

  4. On the nanotoxicity of PAMAM dendrimers: Superfect® stimulates the EGFR-ERK1/2 signal transduction pathway via an oxidative stress-dependent mechanism in HEK 293 cells.

    Science.gov (United States)

    Akhtar, Saghir; Chandrasekhar, Bindu; Attur, Sreeja; Yousif, Mariam H M; Benter, Ibrahim F

    2013-05-01

    Polyamidoamine (PAMAM) dendrimers are cationic branch-like macromolecules that may serve as drug delivery systems for gene-based therapies such as RNA interference. For their safe use in the clinic, they should ideally only enhance drug delivery to target tissues and exhibit no adverse effects. However, little is known about their toxicological profiles in terms of their interactions with cellular signal transduction pathways such as the epidermal growth factor receptor (EGFR). The EGFR is an important signaling cascade that regulates cell growth, differentiation, migration, survival and apoptosis. Here, we investigated the impact of naked, unmodified Superfect (SF), a commercially available generation 6 PAMAM dendrimer, on the epidermal growth factor receptor (EGFR) tyrosine kinase-extracellular-regulated kinase 1/2 (ERK1/2) signaling pathway in human embryonic kidney (HEK 293) cells. At concentrations routinely used for transfection, SF exhibited time and dose-dependent stimulation of EGFR and ERK1/2 phosphorylation whereas AG1478, a selective EGFR tyrosine kinase antagonist, inhibited EGFR-ERK1/2 signaling. SF-induced phosphorylation of EGFR for 1h was partly reversible upon removal of the dendrimer and examination of cells 24 later. Co-treatment of SF with epidermal growth factor (EGF) ligand resulted in greater EGFR stimulation than either agent alone implying that the stimulatory effects of SF and the ligand are synergistic. Dendrimer-induced stimulation of EGFR-ERK1/2 signaling could be attenuated by the antioxidants apocynin, catalase and tempol implying that an oxidative stress dependent mechanism was involved. These results show for the first time that PAMAM dendrimers, aside from their ability to improve drug delivery, can modulate the important EGFR-ERK1/2 cellular signal transduction pathway - a novel finding that may have a bearing on their safe application as drug delivery systems. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Mutational status of EGFR and KIT in thymoma and thymic carcinoma.

    Science.gov (United States)

    Yoh, Kiyotaka; Nishiwaki, Yutaka; Ishii, Genichiro; Goto, Koichi; Kubota, Kaoru; Ohmatsu, Hironobu; Niho, Seiji; Nagai, Kanji; Saijo, Nagahiro

    2008-12-01

    This study was conducted to evaluate the prevalence of EGFR and KIT mutations in thymomas and thymic carcinomas as a means of exploring the potential for molecularly targeted therapy with tyrosine kinase inhibitors. Genomic DNA was isolated from 41 paraffin-embedded tumor samples obtained from 24 thymomas and 17 thymic carcinomas. EGFR exons 18, 19, and 21, and KIT exons 9, 11, 13, and 17, were analyzed for mutations by PCR and direct sequencing. Protein expression of EGFR and KIT was evaluated immunohistochemically. EGFR mutations were detected in 2 of 20 thymomas, but not in any of the thymic carcinomas. All of the EGFR mutations detected were missense mutations (L858R and G863D) in exon 21. EGFR protein was expressed in 71% of the thymomas and 53% of the thymic carcinomas. The mutational analysis of KIT revealed only a missense mutation (L576P) in exon 11 of one thymic carcinoma. KIT protein was expressed in 88% of the thymic carcinomas and 0% of the thymomas. The results of this study indicate that EGFR and KIT mutations in thymomas and thymic carcinomas are rare, but that many of the tumors express EGFR or KIT protein.

  6. EGFR and KRAS mutation coexistence in lung adenocarcinomas

    Directory of Open Access Journals (Sweden)

    Vitor Manuel Leitão de Sousa

    2015-04-01

    Full Text Available Lung cancer is one of the most common causes of cancer deaths. The development of EGFR targeted therapies, including monoclonal antibodies and tyrosine kinase inhibitors have generated an interest in the molecular characterization of these tumours. KRAS mutations are associated with resistance to EGFR TKIs. EGFR and KRAS mutations have been considered as mutually exclusive. This paper presents three bronchial-pulmonary carcinomas, two adenocarcinomas and one pleomorphic sarcomatoid carcinoma, harboring EGFR and KRAS mutations. Case 1 corresponded to an adenocarcinoma with EGFR exon 21 mutation (L858R and KRAS codon 12 point mutation (G12V; case 2, a  mucinous adenocarcinoma expressed coexistence of EGFR exon 21 mutation (L858R and KRAS codon 12 point mutation (G12V; and case 3 a sarcomatoid carcinoma with EGFR exon 19 deletion – del 9bp and KRAS codon 12 point mutation (G12C - cysteine. Based on our experience and on the literature, we conclude that EGFR and KRAS mutations can indeed coexist in the same bronchial-pulmonary carcinoma, either in the same histological type or in different patterns. The biological implications of this coexistence are still poorly understood mainly because these cases are not frequent or currently searched. It is therefore necessary to study larger series of cases with the two mutations to better understand the biological, clinical and therapeutic implications.

  7. MHC-I-induced apoptosis in human B-lymphoma cells is dependent on protein tyrosine and serine/threonine kinases

    DEFF Research Database (Denmark)

    Pedersen, Anders Elm; Bregenholt, S; Johansen, B

    1999-01-01

    B lymphoma cells, is dependent on protein tyrosine kinases and the phosphatidylinositol 3 (PI-3) kinase. Functional studies showed that MHC-I crosslinking induced almost complete inhibition of the spontaneous proliferation of the B lymphoma cells as early as 6 h post-crosslinking and apoptosis 24 h...... post-crosslinking. Preincubation with either protein tyrosine kinase or protein serine/threonine kinase inhibitors reduced the MHC-I-induced apoptosis to background levels, whereas inhibition of PI-3 kinase had no effect. These data demonstrate a pivotal role for protein tyrosine and serine...

  8. Protein tyrosine kinase and mitogen-activated protein kinase signalling pathways contribute to differences in heterophil-mediated innate immune responsiveness between two lines of broilers

    Science.gov (United States)

    Protein tyrosine phosphorylation mediates signal transduction of cellular processes, with protein tyrosine kinases (PTKs) regulating virtually all signaling events. The mitogen-activated protein kinase (MAPK) super-family consists of three conserved pathways that convert receptor activation into ce...

  9. A new receptor tyrosine kinase inhibitor, icotinib, for patients with lung adenocarcinoma cancer without indication for chemotherapy.

    Science.gov (United States)

    Zheng, Xiao; Liu, Guan; Wang, Shengye; Zhang, Yunli; Bao, Wenlong; Deng, Dehou; Mao, Weiming; Fang, Meiyu

    2014-10-01

    Epidermal growth factor receptor (EGFR) is an important therapeutic target in lung cancer. Gefitinib and erlotinib, two reversible EGFR receptor tyrosine kinases inhibitors (TKIs), have been approved for the treatment of patients with metastatic non small-cell lung cancer. Icotinib, which is a selective EGFR-TKI, provides a similar efficacy to gefitinib. The present study aimed to investigate the survival and safety of icotinib in patients with lung adenocarcinoma with a poor performance status (PS). A total of 42 cases of lung adenocarcinoma, including 35 females and 7 males, were enrolled. Icotinib was used as the first-line of treatment due to poor PS of the patient or a more advanced age. Icotinib (125 mg) was orally administered three times per day. The overall response rate and disease control rates were 33.3 and 85.7%, respectively. The median survival time was 13.0 months (95% CI, 5.6-20.4), The median progression-free survival time was 7.0 months, and the 1-year survival rate was 71.4%. A total of 79% of patients had an improved PS following icotinib treatment. Grade 1 to 2 rashes and diarrhea were the most frequent side effects. One patient succumbed during the study due to interstitial pneumonia. In conclusion, this is the first study indicating that patients with lung adenocarcinoma and poor PS may benefit from first-line icotinib therapy, but should be cautious of the occurrence of interstitial lung disease.

  10. Acquired resistance L747S mutation in an epidermal growth factor receptor-tyrosine kinase inhibitor-naïve patient: A report of three cases.

    Science.gov (United States)

    Yamaguchi, Fumihiro; Fukuchi, Kunihiko; Yamazaki, Yohei; Takayasu, Hiromi; Tazawa, Sakiko; Tateno, Hidetsugu; Kato, Eisuke; Wakabayashi, Aya; Fujimori, Mami; Iwasaki, Takuya; Hayashi, Makoto; Tsuchiya, Yutaka; Yamashita, Jun; Takeda, Norikazu; Kokubu, Fumio

    2014-02-01

    The purpose of the present study was to report cases of epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI)-naïve patients carrying a mutation associated with acquired resistance to the drug. Gene alterations in 77 lung carcinoma patients were analyzed by collecting and studying curette lavage fluid at the time of diagnosis. PCRs were performed to amplify mutation hotspot regions in EGFR genes. The PCR products were direct-sequenced and the mutations confirmed by resequencing using different primers. Case 1 was a 78-year-old Japanese male diagnosed with stage IB lung adenocarcinoma who was found to have two EGFR mutations, G719S and L747S. Case 2 was a 73-year-old Japanese male diagnosed with stage IV squamous cell lung carcinoma and bone metastasis who had the EGFR mutation, L747S. Case 3 was an 82-year-old Japanese male diagnosed with hyponatremia due to inappropriate secretion of antidiuretic hormone and stage IIIB small cell lung carcinoma (SCLC) who had the EGFR mutation, L747S. Thus, the EGFR mutation L747S associated with acquired EGFR-TKI resistance was detected in two non-small cell lung carcinoma (NSCLC) patients and one SCLC patient, none of whom had ever received EGFR-TKI. The patients were current smokers with stages at diagnosis ranging from IB to IV, and their initial tumors contained resistant clones carrying L747S. L747S may be associated with primary resistance. To the best of our knowledge, this study is the first report of an EGFR mutation associated with resistance to EGFR-TKI in SCLC patients. The early detection of EGFR-TKI resistance mutations may be beneficial in making treatment decisions for lung carcinoma patients, including those with SCLC.

  11. Resistance to EGFR inhibitors in non-small cell lung cancer: Clinical management and future perspectives.

    Science.gov (United States)

    Tomasello, Chiara; Baldessari, Cinzia; Napolitano, Martina; Orsi, Giulia; Grizzi, Giulia; Bertolini, Federica; Barbieri, Fausto; Cascinu, Stefano

    2018-03-01

    In the last few years, the development of targeted therapies for non-small cell lung cancer (NSCLC) expressing oncogenic driver mutations (e.g. EGFR) has changed the clinical management and the survival outcomes of this specific minority of patients. Several phase III trials demonstrated the superiority of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs) over chemotherapy in EGFR-mutant NSCLC patients. However, in the vast majority of cases EGFR TKIs lose their clinical activity within 8-12 months. Many genetic aberrations have been described as possible mechanisms of EGFR TKIs acquired resistance and can be clustered in four main sub-groups: 1. Development of secondary EGFR mutations; 2. Activation of parallel signaling pathways; 3. Histological transformation; 4. Activation of downstream signaling pathways. In this review we will describe the molecular alterations underlying each of these EGFR TKIs resistance mechanisms, focusing on the currently available and future therapeutic strategies to overcome these phenomena. Copyright © 2018 Elsevier B.V. All rights reserved.

  12. MITF Modulates Therapeutic Resistance through EGFR Signaling.

    Science.gov (United States)

    Ji, Zhenyu; Erin Chen, Yiyin; Kumar, Raj; Taylor, Michael; Jenny Njauw, Ching-Ni; Miao, Benchun; Frederick, Dennie T; Wargo, Jennifer A; Flaherty, Keith T; Jönsson, Göran; Tsao, Hensin

    2015-07-01

    Response to targeted therapies varies significantly despite shared oncogenic mutations. Nowhere is this more apparent than in BRAF (V600E)-mutated melanomas where initial drug response can be striking and yet relapse is commonplace. Resistance to BRAF inhibitors have been attributed to the activation of various receptor tyrosine kinases (RTKs), although the underlying mechanisms have been largely uncharacterized. Here, we found that EGFR-induced vemurafenib resistance is ligand dependent. We employed whole-genome expression analysis and discovered that vemurafenib resistance correlated with the loss of microphthalmia-associated transcription factor (MITF), along with its melanocyte lineage program, and with the activation of EGFR signaling. An inverse relationship between MITF, vemurafenib resistance, and EGFR was then observed in patient samples of recurrent melanoma and was conserved across melanoma cell lines and patients' tumor specimens. Functional studies revealed that MITF depletion activated EGFR signaling and consequently recapitulated the resistance phenotype. In contrast, forced expression of MITF in melanoma and colon cancer cells inhibited EGFR and conferred sensitivity to BRAF/MEK inhibitors. These findings indicate that an "autocrine drug resistance loop" is suppressed by melanocyte lineage signal(s), such as MITF. This resistance loop modulates drug response and could explain the unique sensitivity of melanomas to BRAF inhibition.

  13. Autoinhibition of Bruton's tyrosine kinase (Btk) and activation by soluble inositol hexakisphosphate

    Science.gov (United States)

    Wang, Qi; Vogan, Erik M; Nocka, Laura M; Rosen, Connor E; Zorn, Julie A; Harrison, Stephen C; Kuriyan, John

    2015-01-01

    Bruton's tyrosine kinase (Btk), a Tec-family tyrosine kinase, is essential for B-cell function. We present crystallographic and biochemical analyses of Btk, which together reveal molecular details of its autoinhibition and activation. Autoinhibited Btk adopts a compact conformation like that of inactive c-Src and c-Abl. A lipid-binding PH-TH module, unique to Tec kinases, acts in conjunction with the SH2 and SH3 domains to stabilize the inactive conformation. In addition to the expected activation of Btk by membranes containing phosphatidylinositol triphosphate (PIP3), we found that inositol hexakisphosphate (IP6), a soluble signaling molecule found in both animal and plant cells, also activates Btk. This activation is a consequence of a transient PH-TH dimerization induced by IP6, which promotes transphosphorylation of the kinase domains. Sequence comparisons with other Tec-family kinases suggest that activation by IP6 is unique to Btk. DOI: http://dx.doi.org/10.7554/eLife.06074.001 PMID:25699547

  14. Tumor-targeted Nanobullets: Anti-EGFR nanobody-liposomes loaded with anti-IGF-1R kinase inhibitor for cancer treatment.

    Science.gov (United States)

    van der Meel, Roy; Oliveira, Sabrina; Altintas, Isil; Haselberg, Rob; van der Veeken, Joris; Roovers, Rob C; van Bergen en Henegouwen, Paul M P; Storm, Gert; Hennink, Wim E; Schiffelers, Raymond M; Kok, Robbert J

    2012-04-30

    The epidermal growth factor receptor (EGFR) is a validated target for anti-cancer therapy and several EGFR inhibitors are used in the clinic. Over the years, an increasing number of studies have reported on the crosstalk between EGFR and other receptors that can contribute to accelerated cancer development or even acquisition of resistance to anti-EGFR therapies. Combined targeting of EGFR and insulin-like growth factor 1 receptor (IGF-1R) is a rational strategy to potentiate anti-cancer treatment and possibly retard resistance development. In the present study, we have pursued this by encapsulating the kinase inhibitor AG538 in anti-EGFR nanobody-liposomes. The thus developed dual-active nanobody-liposomes associated with EGFR-(over)expressing cells in an EGFR-specific manner and blocked both EGFR and IGF-1R activation, due to the presence of the EGFR-blocking nanobody EGa1 and the anti-IGF-1R kinase inhibitor AG538 respectively. AG538-loaded nanobody-liposomes induced a strong inhibition of tumor cell proliferation even upon short-term exposure followed by a drug-free wash-out period. Therefore, AG538-loaded nanobody-liposomes are a promising anti-cancer formulation due to efficient intracellular delivery of AG538 in combination with antagonistic and downregulating properties of the EGa1 nanobody-liposomes. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. Bosutinib induced pleural effusions: Case report and review of tyrosine kinase inhibitors induced pulmonary toxicity

    Directory of Open Access Journals (Sweden)

    Natalia I. Moguillansky, MD

    2017-01-01

    Full Text Available Tyrosine kinase inhibitors are known to cause pulmonary complications. We report a case of bosutinib related bilateral pleural effusions in a patient with chronic myeloid leukemia. Characteristics of the pleural fluid are presented. We also discuss other tyrosine kinase inhibitors induced pulmonary toxicities, including pulmonary hypertension and interstitial lung disease.

  16. Regulation of hematopoietic cell function by protein tyrosine kinase-encoding oncogenes, a review

    NARCIS (Netherlands)

    Punt, C. J.

    1992-01-01

    Tyrosine phosphorylation of proteins by protein tyrosine kinases (PTKs) is an important mechanism in the regulation of various cellular processes such as proliferation, differentiation, and transformation. Accumulating data implicate PTKs as essential intermediates in the transduction of

  17. Function of Bruton's tyrosine kinase during B cell development is partially independent of its catalytic activity

    NARCIS (Netherlands)

    S. Middendorp; G.M. Dingjan (Gemma); A. Maas (Alex); K. Dahlenborg; R.W. Hendriks (Rudi)

    2003-01-01

    textabstractThe Tec family member Bruton's tyrosine kinase (Btk) is a cytoplasmic protein tyrosine kinase that transduces signals from the pre-B and B cell receptor (BCR). Btk is involved in pre-B cell maturation by regulating IL-7 responsiveness, cell surface phenotype changes,

  18. DMPD: Receptor tyrosine kinases and the regulation of macrophage activation. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 14726496 Receptor tyrosine kinases and the regulation of macrophage activation. Cor...osine kinases and the regulation of macrophage activation. PubmedID 14726496 Title Receptor tyrosine...rell PH, Morrison AC, Lutz MA. J Leukoc Biol. 2004 May;75(5):731-7. Epub 2004 Jan 14. (.png) (.svg) (.html) (.csml) Show Receptor tyr

  19. Osimertinib and Necitumumab in Treating Patients With EGFR-Mutant Stage IV or Recurrent Non-small Cell Lung Cancer Who Have Progressed on a Previous EGFR Tyrosine Kinase Inhibitor

    Science.gov (United States)

    2018-03-07

    EGFR Exon 19 Deletion Mutation; EGFR Exon 20 Insertion Mutation; EGFR NP_005219.2:p.G719X; EGFR NP_005219.2:p.L858R; EGFR NP_005219.2:p.L861Q; EGFR NP_005219.2:p.T790M; EGFR T790M Mutation Negative; Recurrent Non-Small Cell Lung Carcinoma; Stage IV Non-Small Cell Lung Cancer AJCC v7

  20. CCR 20th anniversary commentary: a chimeric antibody, C225, inhibits EGFR activation and tumor growth.

    Science.gov (United States)

    Mendelsohn, John; Prewett, Marie; Rockwell, Patricia; Goldstein, Neil I

    2015-01-15

    Murine mAb 225 was effective against the EGFR tyrosine kinase and inhibited tumor growth in preclinical studies. A phase I trial showed safety, tumor localization, and satisfactory pharmacokinetics. Human:murine chimeric C225 retained biologic activity, which was essential for the conduct of subsequent combination therapy trials and eventual regulatory approval. ©2015 American Association for Cancer Research.

  1. Band 3 tyrosine kinase in avian erythrocyte plasma membrane is immunologically related to pp60c-src

    International Nuclear Information System (INIS)

    Hillsgrove, D.; Shores, C.G.; Parker, J.C.; Maness, P.F.

    1987-01-01

    The authors have identified in the plasma membrane of the chicken erythrocyte a 60-kDa tyrosine-specific protein kinase immunologically related to the transforming protein pp60 v-src of Rous sarcoma virus. The erythrocyte protein kinase phosphorylated heavy chains of tumor-bearing rabbit (TBR) antibodies reactive with pp60 c-src at tyrosine in immune complex protein kinase assays. The kinase was identified as a 60-kDa protein by [ 35 S]methionine labeling of erythrocytes and by autophosphorylation in immune complexes. The kinase migrated on two-dimensional gel electrophoresis with an apparent pI and molecular mass similar to pp60 c-src . A plasma membrane-enriched fraction isolated from chicken red cells contained the majority of the kinase activity. Incubation of the plasma membrane fraction with [ 32 P]ATP resulted in tyrosine phosphorylation of the anion transport protein band 3. Band 3 phosphorylation was blocked by TBR antibodies, indicting that the kinase recognized by pp60 c-src antibodies was responsible for band 3 phosphorylation. These results demonstrate that the avian erythrocyte plasma membrane contains a tightly bound tyrosine-specific protein kinase identical or closely related to pp60 c-src and that this kinase is responsible for band 3 phosphorylation in vitro

  2. Free radical-mediated stimulation of tyrosine-specific protein kinase in rat liver plasma membrane

    International Nuclear Information System (INIS)

    Chan, T.M.; Tatoyan, A.; Cheng, E.; Shargill, N.S.; Pleta, M.

    1986-01-01

    Incorporation of 32 P from (γ- 32 P)-ATP into endogenous proteins of plasma membranes isolated from rat liver was significantly increased by several naphthoquinones including menadione. This apparent stimulation of membrane-associated protein kinase activity by these compounds was most striking (up to 6-7 fold) when the synthetic copolymers containing glutamate and tyrosine residues (4:1) was used as substrate. Since tyrosine residues are the only possible phosphate acceptor in the copolymers, the quinone-stimulated liver membrane protein kinase is most likely tyrosine specific. Although not required for protein kinase activity, dithiothreitol (DTT) was necessary for its stimulation by these quinonoid compounds. Hydrolysis of ATP was not significantly affected by quinones under the experimental conditions. Both menadione and vitamin k 5 increased phosphorylation of plasma membrane proteins of molecular weight 45 and 60 kd. The stimulatory effect of menadione on protein phosphorylation was prevented by the addition of superoxide dismutase. Dihydroxyfumerate, which spontaneously produces various radical species, and H 2 O 2 , also stimulated tyrosine-specific protein phosphorylation. DTT was also required for their full effect. It, therefore, appears that quinonone stimulation of tyrosine-specific protein phosphorylation is mediated by oxygen radicals

  3. Pharmacogenetics of telatinib, a VEGFR-2 and VEGFR-3 tyrosine kinase inhibitor, used in patients with solid tumors

    NARCIS (Netherlands)

    N. Steeghs (Neeltje); A.J. Gelderblom (Hans); J.A.M. Wessels (Judith); F.A.L.M. Eskens (Ferry); N. de Bont (Natasja); J.W. Nortier (Johan); H.J. Guchelaar (Henk Jan)

    2011-01-01

    textabstractSummary: Purpose Telatinib is an orally active small-molecule tyrosine kinase inhibitor of kinase insert domain receptor (KDR; VEGFR-2) and fms-related tyrosine kinase 4 (FLT4; VEGFR-3). This study aims at the identification of relationships between single nucleotide polymorphisms (SNPs)

  4. Elucidation of the critical epitope of an anti-EGFR monoclonal antibody EMab-134.

    Science.gov (United States)

    Kaneko, Mika K; Yamada, Shinji; Itai, Shunsuke; Chang, Yao-Wen; Nakamura, Takuro; Yanaka, Miyuki; Kato, Yukinari

    2018-07-01

    The epidermal growth factor receptor (EGFR) is a type-1 transmembrane receptor tyrosine kinase, which activates the downstream signaling cascades in many tumors, such as oral and lung cancers. We previously developed EMab-134, a novel anti-EGFR monoclonal antibody (mAb), which reacts with endogenous EGFR-expressing cancer cell lines and normal cells independent of glycosylation in Western blotting, flow cytometry, and immunohistochemical analysis. EMab-134 showed very high sensitivity (94.7%) to oral squamous cell carcinomas in immunohistochemical analysis. In this study, we performed enzyme-linked immunosorbent assay (ELISA), flow cytometry, and immunohistochemical analysis to determine the epitope of EMab-134. A blocking peptide (375-394 amino acids of EGFR) neutralized the EMab-134 reaction against oral cancer cells in flow cytometry and immunohistochemistry. The minimum epitope of EMab-134 was found to be the 377- RGDSFTHTPP -386 sequence. Our findings can be applied for the production of more functional anti-EGFR mAbs that in turn can be used for antitumor treatments.

  5. Tyrosine phosphorylation of AAV2 vectors and its consequences on viral intracellular trafficking and transgene expression

    Science.gov (United States)

    Zhong, Li; Li, Baozheng; Jayandharan, Giridhararao; Mah, Cathryn S.; Govindasamy, Lakshmanan; Agbandje-McKenna, Mavis; Herzog, Roland W.; Weigel-Van Aken, Kirsten A.; Hobbs, Jacqueline A.; Zolotukhin, Sergei; Muzyczka, Nicholas; Srivastava, Arun

    2008-01-01

    We have documented that epidermal growth factor receptor protein tyrosine kinase (EGFR-PTK) signaling negatively affects intracellular trafficking and transduction efficiency of recombinant adeno-associated virus 2 (AAV2) vectors. Specifically, inhibition of EGFR-PTK signaling leads to decreased ubiquitination of AAV2 capsid proteins, which in turn, facilitates viral nuclear transport by limiting proteasome-mediated degradation of AAV2 vectors. In the present studies, we observed that AAV capsids can indeed be phosphorylated at tyrosine residues by EGFR-PTK in in vitro phosphorylation assays and that phosphorylated AAV capsids retain their structural integrity. However, although phosphorylated AAV vectors enter cells as efficiently as their unphosphorylated counterparts, their transduction efficiency is significantly reduced. This reduction is not due to impaired viral second-strand DNA synthesis since transduction efficiency of both single-stranded AAV (ssAAV) and self-complementary AAV (scAAV) vectors is decreased by ~68% and ~74%, respectively. We also observed that intracellular trafficking of tyrosine-phosphorylated AAV vectors from cytoplasm to nucleus is significantly decreased, which leads to ubiquitination of AAV capsids followed by proteasome-mediated degradation, although downstream consequences of capsid ubiquitination may also be affected by tyrosine-phosphorylation. These studies provide new insights into the role of tyrosine-phosphorylation of AAV capsids in various steps in the virus life cycle, which has implications in the optimal use of recombinant AAV vectors in human gene therapy. PMID:18834608

  6. Tyrosine-phosphorylation of AAV2 vectors and its consequences on viral intracellular trafficking and transgene expression

    International Nuclear Information System (INIS)

    Zhong Li; Li Baozheng; Jayandharan, Giridhararao; Mah, Cathryn S.; Govindasamy, Lakshmanan; Agbandje-McKenna, Mavis; Herzog, Roland W.

    2008-01-01

    We have documented that epidermal growth factor receptor protein tyrosine kinase (EGFR-PTK) signaling negatively affects intracellular trafficking and transduction efficiency of recombinant adeno-associated virus 2 (AAV2) vectors. Specifically, inhibition of EGFR-PTK signaling leads to decreased ubiquitination of AAV2 capsid proteins, which in turn, facilitates viral nuclear transport by limiting proteasome-mediated degradation of AAV2 vectors. In the present studies, we observed that AAV capsids can indeed be phosphorylated at tyrosine residues by EGFR-PTK in in vitro phosphorylation assays and that phosphorylated AAV capsids retain their structural integrity. However, although phosphorylated AAV vectors enter cells as efficiently as their unphosphorylated counterparts, their transduction efficiency is significantly reduced. This reduction is not due to impaired viral second-strand DNA synthesis since transduction efficiency of both single-stranded AAV (ssAAV) and self-complementary AAV (scAAV) vectors is decreased by ∼ 68% and ∼ 74%, respectively. We also observed that intracellular trafficking of tyrosine-phosphorylated AAV vectors from cytoplasm to nucleus is significantly decreased, which results from ubiquitination of AAV capsids followed by proteasome-mediated degradation, although downstream consequences of capsid ubiquitination may also be affected by tyrosine-phosphorylation. These studies provide new insights into the role of tyrosine-phosphorylation of AAV capsids in various steps in the virus life cycle, which has implications in the optimal use of recombinant AAV vectors in human gene therapy

  7. Efficacy and mechanism of action of the tyrosine kinase inhibitors gefitinib, lapatinib and neratinib in the treatment of HER2-positive breast cancer: preclinical and clinical evidence.

    Science.gov (United States)

    Segovia-Mendoza, Mariana; González-González, María E; Barrera, David; Díaz, Lorenza; García-Becerra, Rocío

    2015-01-01

    An increasing number of tumors, including breast cancer, overexpress proteins of the epidermal growth factor receptor (EGFR) family. The interaction between family members activates signaling pathways that promote tumor progression and resistance to treatment. Human epidermal growth factor receptor type II (HER2) positive breast cancer represents a clinical challenge for current therapy. It has motivated the development of novel and more effective therapeutic EGFR family target drugs, such as tyrosine kinase inhibitors (TKIs). This review focuses on the effects of three TKIs mostly studied in HER2- positive breast cancer, lapatinib, gefitinib and neratinib. Herein, we discuss the mechanism of action, therapeutic advantages and clinical applications of these TKIs. To date, TKIs seem to be promising therapeutic agents for the treatment of HER2-overexpressing breast tumors, either as monotherapy or combined with other pharmacological agents.

  8. The biological activity of the human epidermal growth factor receptor is positively regulated by its C-terminal tyrosines

    DEFF Research Database (Denmark)

    Helin, K; Velu, T; Martin, P

    1991-01-01

    mutants in the full length receptor. EGF-dependent transforming ability of the single point mutants is similar to that of the wild type, while that of double mutants is decreased and an even lower activity is present in the triple mutant. In each bioassay, including EGF-dependent focal transformation...... biologically. The EGF-R kinase activity is affected by tyrosine substitution since in vitro phosphorylation of exogenous substrates is reduced in the double and triple mutants. Autophosphorylation, in vivo and in vitro, is also reduced, but not totally abolished in the triple point mutant and Dc123 indicating......The epidermal growth factor receptor (EGF-R) C-terminus contains three conserved tyrosines (Y-1068, Y-1148, Y-1173) which are phosphorylated upon EGF activation. To clarify the functional role of these tyrosines, each has been mutated to phenylalanine and studied as single, double and triple...

  9. 2-Triazenoazaindoles: α novel class of triazenes inducing transcriptional down-regulation of EGFR and HER-2 in human pancreatic cancer cells.

    Science.gov (United States)

    Kreutzer, Jan N; Salvador, Alessia; Diana, Patrizia; Cirrincione, Girolamo; Vedaldi, Daniela; Litchfield, David W; Issinger, Olaf-Georg; Guerra, Barbara

    2012-04-01

    Pancreatic cancer is a complex malignancy arising from the accumulation of genetic and epigenetic defects in the affected cells. Standard chemotherapy for patients with advanced disease shows only modest effects and is associated with considerable toxicity. Overexpression or aberrant activation of members of the epidermal growth factor receptor tyrosine kinase family, which includes EGFR and HER-2, occurs frequently and is associated with multiple drug resistance and decreased patient survival. In this study, we have investigated the therapeutic potential of AS104, a novel compound of the triazene class, with potential inhibitory effects on EGFR. We found that treatment of cells with AS104 causes significant reduction of cell growth and metabolic activity in four human pancreatic cancer cell lines. Furthermore, we show that the AS104-mediated induction of apoptotic cell death is associated with stimulation of autophagy in a dose-dependent manner. Treatment of cells with AS104 results in significant down-regulation of EGFR and HER-2 expression and activity and subsequent inhibition of downstream signaling proteins. Quantitative RT-PCR analysis and assays with proteasome inhibitors revealed that AS104 regulates the expression of EGFR and HER-2 at the transcriptional level. These findings provide for the first time experimental evidence for efficacy of AS104 in the simultaneous transcriptional repression of EGFR and HER-2 genes and suggest that AS104 may have therapeutic potential in the treatment of pancreatic cancers that express high levels of the aforementioned receptor tyrosine kinases.

  10. 2-Triazenoazaindoles: A novel class of triazenes inducing transcriptional down-regulation of EGFR and HER-2 in human pancreatic cancer cells

    Science.gov (United States)

    KREUTZER, JAN N.; SALVADOR, ALESSIA; DIANA, PATRIZIA; CIRRINCIONE, GIROLAMO; VEDALDI, DANIELA; LITCHFIELD, DAVID W.; ISSINGER, OLAF-GEORG; GUERRA, BARBARA

    2012-01-01

    Pancreatic cancer is a complex malignancy arising from the accumulation of genetic and epigenetic defects in the affected cells. Standard chemotherapy for patients with advanced disease shows only modest effects and is associated with considerable toxicity. Overexpression or aberrant activation of members of the epidermal growth factor receptor tyrosine kinase family, which includes EGFR and HER-2, occurs frequently and is associated with multiple drug resistance and decreased patient survival. In this study, we have investigated the therapeutic potential of AS104, a novel compound of the triazene class, with potential inhibitory effects on EGFR. We found that treatment of cells with AS104 causes significant reduction of cell growth and metabolic activity in four human pancreatic cancer cell lines. Furthermore, we show that the AS104-mediated induction of apoptotic cell death is associated with stimulation of autophagy in a dose-dependent manner. Treatment of cells with AS104 results in significant down-regulation of EGFR and HER-2 expression and activity and subsequent inhibition of downstream signaling proteins. Quantitative RT-PCR analysis and assays with proteasome inhibitors revealed that AS104 regulates the expression of EGFR and HER-2 at the transcriptional level. These findings provide for the first time experimental evidence for efficacy of AS104 in the simultaneous transcriptional repression of EGFR and HER-2 genes and suggest that AS104 may have therapeutic potential in the treatment of pancreatic cancers that express high levels of the aforementioned receptor tyrosine kinases. PMID:22134789

  11. Bacillus subtilis strain deficient for the protein-tyrosine kinase PtkA exhibits impaired DNA replication

    DEFF Research Database (Denmark)

    Petranovic, Dina; Michelsen, Ole; Zahradka, K

    2007-01-01

    Bacillus subtilis has recently come into the focus of research on bacterial protein-tyrosine phosphorylation, with several proteins kinases, phosphatases and their substrates identified in this Gram-positive model organism. B. subtilis protein-tyrosine phosphorylation system Ptk...... microscopy. B. subtilis cells lacking the kinase PtkA accumulated extra chromosome equivalents, exhibited aberrant initiation mass for DNA replication and an unusually long D period....

  12. Src-family-tyrosine kinase Lyn is critical for TLR2-mediated NF-κB activation through the PI 3-kinase signaling pathway.

    Science.gov (United States)

    Toubiana, Julie; Rossi, Anne-Lise; Belaidouni, Nadia; Grimaldi, David; Pene, Frederic; Chafey, Philippe; Comba, Béatrice; Camoin, Luc; Bismuth, Georges; Claessens, Yann-Erick; Mira, Jean-Paul; Chiche, Jean-Daniel

    2015-10-01

    TLR2 has a prominent role in host defense against a wide variety of pathogens. Stimulation of TLR2 triggers MyD88-dependent signaling to induce NF-κB translocation, and activates a Rac1-PI 3-kinase dependent pathway that leads to transactivation of NF-κB through phosphorylation of the P65 NF-κB subunit. This transactivation pathway involves tyrosine phosphorylations. The role of the tyrosine kinases in TLR signaling is controversial, with discrepancies between studies using only chemical inhibitors and knockout mice. Here, we show the involvement of the tyrosine-kinase Lyn in TLR2-dependent activation of NF-κB in human cellular models, by using complementary inhibition strategies. Stimulation of TLR2 induces the formation of an activation cluster involving TLR2, CD14, PI 3-kinase and Lyn, and leads to the activation of AKT. Lyn-dependent phosphorylation of the p110 catalytic subunit of PI 3-kinase is essential to the control of PI 3-kinase biological activity upstream of AKT and thereby to the transactivation of NF-κB. Thus, Lyn kinase activity is crucial in TLR2-mediated activation of the innate immune response in human mononuclear cells. © The Author(s) 2015.

  13. Variability in bioavailability of small molecular tyrosine kinase inhibitors

    NARCIS (Netherlands)

    Herbrink, Maikel; Nuijen, Bastiaan; Schellens, Jan H M; Beijnen, Jos H.

    2015-01-01

    Small molecular tyrosine kinase inhibitors (smTKIs) are in the centre of the very quickly expanding area of personalized chemotherapy and oral applicability thereof. The number of drugs in this class is rapidly growing, with twenty current approvals by both the European Medicines Agency (EMA) and

  14. Role of Receptor Tyrosine Kinase Signaling in Renal Fibrosis

    Directory of Open Access Journals (Sweden)

    Feng Liu

    2016-06-01

    Full Text Available Renal fibrosis can be induced in different renal diseases, but ultimately progresses to end stage renal disease. Although the pathophysiologic process of renal fibrosis have not been fully elucidated, it is characterized by glomerulosclerosis and/or tubular interstitial fibrosis, and is believed to be caused by the proliferation of renal inherent cells, including glomerular epithelial cells, mesangial cells, and endothelial cells, along with defective kidney repair, renal interstitial fibroblasts activation, and extracellular matrix deposition. Receptor tyrosine kinases (RTKs regulate a variety of cell physiological processes, including metabolism, growth, differentiation, and survival. Many studies from in vitro and animal models have provided evidence that RTKs play important roles in the pathogenic process of renal fibrosis. It is also showed that tyrosine kinases inhibitors (TKIs have anti-fibrotic effects in basic research and clinical trials. In this review, we summarize the evidence for involvement of specific RTKs in renal fibrosis process and the employment of TKIs as a therapeutic approach for renal fibrosis.

  15. Lipopolysaccharide induces VCAM-1 expression and neutrophil adhesion to human tracheal smooth muscle cells: Involvement of Src/EGFR/PI3-K/Akt pathway

    International Nuclear Information System (INIS)

    Lin, W.-N.; Luo, S.-F.; Wu, C.-B.; Lin, C.-C.; Yang, C.-M.

    2008-01-01

    In our previous study, LPS has been shown to induce vascular cell adhesion molecule-1(VCAM-1) expression through MAPKs and NF-κB in human tracheal smooth muscle cells (HTSMCs). In addition to these pathways, the non-receptor tyrosine kinases (Src), EGF receptor (EGFR), and phosphatidylinositol 3-kinase (PI3K) have been shown to be implicated in the expression of several inflammatory target proteins. Here, we reported that LPS-induced up-regulation of VCAM-1 enhanced the adhesion of neutrophils onto HTSMC monolayer, which was inhibited by LY294002 and wortmannin. LPS stimulated phosphorylation of protein tyrosine kinases including Src, PYK2, and EGFR, which were further confirmed using specific anti-phospho-Src, PYK2, or EGFR Ab, respectively, revealed by Western blotting. LPS-stimulated Src, PYK2, EGFR, and Akt phosphorylation and VCAM-1 expression were attenuated by the inhibitors of Src (PP1), EGFR (AG1478), PI3-K (LY294002 and wortmannin), and Akt (SH-5), respectively, or transfection with siRNAs of Src or Akt and shRNA of p110. LPS-induced VCAM-1 expression was also blocked by pretreatment with curcumin (a p300 inhibitor) or transfection with p300 siRNA. LPS-stimulated Akt activation translocated into nucleus and associated with p300 and VCAM-1 promoter region was further confirmed by immunofluorescence, immunoprecipitation, and chromatin immunoprecipitation assays. This association of Akt and p300 to VCAM-1 promoter was inhibited by pretreatment with PP1, AG1478, wortmannin, and SH-5. LPS-induced p300 activation enhanced VCAM-1 promoter activity and VCAM-1 mRNA expression. These results suggested that in HTSMCs, Akt phosphorylation mediated through transactivation of Src/PYK2/EGFR promoted the transcriptional p300 activity and eventually led to VCAM-1 expression induced by LPS

  16. Complexes of γ-tubulin with nonreceptor protein tyrosine kinases Src and Fyn in differentiating P19 embryonal carcinoma cells

    International Nuclear Information System (INIS)

    Kukharskyy, Vitaliy; Sulimenko, Vadym; Macurek, Libor; Sulimenko, Tetyana; Draberova, Eduarda; Draber, Pavel

    2004-01-01

    Nonreceptor protein tyrosine kinases of the Src family have been shown to play an important role in signal transduction as well as in regulation of microtubule protein interactions. Here we show that γ-tubulin (γ-Tb) in P19 embryonal carcinoma cells undergoing neuronal differentiation is phosphorylated and forms complexes with protein tyrosine kinases of the Src family, Src and Fyn. Elevated expression of both kinases during differentiation corresponded with increased level of proteins phosphorylated on tyrosine. Immunoprecipitation experiments with antibodies against Src, Fyn, γ-tubulin, and with anti-phosphotyrosine antibody revealed that γ-tubulin appeared in complexes with these kinases. In vitro kinase assays showed tyrosine phosphorylation of proteins in γ-tubulin complexes isolated from differentiated cells. Pretreatment of cells with Src family selective tyrosine kinase inhibitor PP2 reduced the amount of phosphorylated γ-tubulin in the complexes. Binding experiments with recombinant SH2 and SH3 domains of Src and Fyn kinases revealed that protein complexes containing γ-tubulin bound to SH2 domains and that these interactions were of SH2-phosphotyrosine type. The combined data suggest that Src family kinases might have an important role in the regulation of γ-tubulin interaction with tubulin dimers or other proteins during neurogenesis

  17. Dual targeting of EGFR and focal adhesion kinase in 3D grown HNSCC cell cultures

    International Nuclear Information System (INIS)

    Eke, Iris; Cordes, Nils

    2011-01-01

    Purpose: Epidermal growth factor receptor (EGFR) and focal adhesion kinase (FAK) show frequent overexpression and hyperactivity in various human malignancies including head and neck squamous cell carcinomas (HNSCC). To examine effects of dual EGFR/FAK inhibition on cellular radiosensitivity of HNSCC cells in a more physiological environment, we employed a previously established laminin-rich extracellular matrix (lrECM) based three-dimensional (3D) cell culture model. Materials and methods: UTSCC15 and SAS HNSCC cell lines stably transfected with EGFR-CFP or CFP were used. Single or combined EGFR (Cetuximab, siRNA) and FAK (TAE226, siRNA) inhibition were accomplished prior to measuring clonogenic survival and protein expression and phosphorylation. Immunofluorescence enabled visualization of EGFR-CFP and FAK. Results: Cetuximab resulted in higher radiosensitization in EGFR-CFP overexpressing cell lines than CFP controls. Single EGFR or FAK inhibition mediated radiosensitization, while dual EGFR/FAK targeting further augmented this effect. Despite signaling alterations upon Cetuximab and siRNA knockdown, analysis of protein expression and phosphorylation indicates EGFR and FAK signaling coexistence without obvious overlap. Conclusions: Combined EGFR/FAK targeting yielded stronger radiosensitization than either approach alone, which might be based on non-overlapping downstream signaling. Whether dual targeting of EGFR and FAK can reasonably be combined with radiotherapy and chemotherapy needs clarification.

  18. Ubiquitin ligase Cbl-b is involved in icotinib (BPI-2009H)-induced apoptosis and G1 phase arrest of EGFR mutation-positive non-small-cell lung cancer.

    Science.gov (United States)

    Mu, Xiaodong; Zhang, Ye; Qu, Xiujuan; Hou, Kezuo; Kang, Jian; Hu, Xuejun; Liu, Yunpeng

    2013-01-01

    Epidermal growth factor receptor (EGFR) is one of the most promising targets for non-small-cell lung cancer (NSCLC). Icotinib, a highly selective EGFR tyrosine kinase inhibitor (EGFR-TKI), has shown promising clinical efficacy and safety in patients with NSCLC. The exact molecular mechanism of icotinib remains unclear. In this study, we first investigated the antiproliferative effect of icotinib on NSCLC cells. Icotinib significantly inhibited proliferation of the EGFR-mutated lung cancer HCC827 cells. The IC50 values at 48 and 72 h were 0.67 and 0.07 μ M, respectively. Flow cytometric analysis showed that icotinib caused the G1 phase arrest and increased the rate of apoptosis in HCC827 cells. The levels of cyclin D1 and cyclin A2 were decreased. The apoptotic process was associated with activation of caspase-3, -8, and poly(ADP-ribose) polymerase (PARP). Further study revealed that icotinib inhibited phosphorylation of EGFR, Akt, and extracellular signal-regulated kinase. In addition, icotinib upregulated ubiquitin ligase Cbl-b expression. These observations suggest that icotinib-induced upregulation of Cbl-b is responsible, at least in part, for the antitumor effect of icotinib via the inhibition of phosphoinositide 3-kinase (PI3K)/Akt and mitogen-activated protein kinase pathways in EGFR-mutated NSCLC cells.

  19. A willingness-to-pay study of oral epidermal growth factor tyrosine kinase inhibitors in advanced non-small cell lung cancer.

    Science.gov (United States)

    Leighl, Natasha B; Tsao, W Stephen; Zawisza, Dianne L; Nematollahi, Mahsan; Shepherd, Frances A

    2006-01-01

    Oral epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs) are new agents in the treatment of advanced non-small cell lung cancer (NSCLC). Phase II studies demonstrate objective tumor responses and symptom improvement, combined with minimal toxicity and the convenience of an oral agent. We evaluated patient utility through willingness-to-pay (WTP) for these agents in the treatment of advanced NSCLC in Canada. Advanced NSCLC patients and healthy subjects participated in a structured interview and bidding exercise, reviewing current evidence supporting EGFR TKI therapy in advanced NSCLC and patient willingness-to-pay for treatment. Fifty-seven patients and 54 healthy subjects participated. The median amount both groups were willing to pay for a month of oral EGFR TKI therapy was $100 CAD (range $0-5000 per month). A minority of NSCLC patients received employment income, the majority relying on disability, pension income, and social assistance for financial support. Affordability of these agents was a key concern for both advanced NSCLC patients and healthy subjects. Univariate predictors of WTP included marital status, prior chemotherapy treatment, receiving pension income or financial social assistance. In multivariate analysis, only prior chemotherapy remained a significant predictor of WTP (p=0.049). Both advanced NSCLC patients and healthy subjects feel oral EGFR TKIs are worth paying for in the treatment of advanced NSCLC, but are willing to pay only a fraction of the market price. As many advanced NSCLC patients are financially disadvantaged, the potential for restricted access to newer therapies is of concern.

  20. HER2-targeted therapy in breast cancer. Monoclonal antibodies and tyrosine kinase inhibitors

    DEFF Research Database (Denmark)

    Nielsen, Dorte Lisbet; Andersson, Michael; Kamby, Claus

    2008-01-01

    There is strong clinical evidence that trastuzumab, a monoclonal antibody targeting the human epidermal growth factor receptor (HER) two tyrosine kinase receptor, is an important component of first-line treatment of patients with HER2-positive metastatic breast cancer. In particular the combination...... of trastuzumab to chemotherapy improves disease-free and overall survival. The use of lapatinib, a dual tyrosine kinase inhibitor of both HER1 and HER2, in combination with capecitabine in the second-line treatment of HER2-positive patients with metastatic breast cancer previously treated with trastuzumab has...

  1. Impact of the putative cancer stem cell markers and growth factor receptor expression on the sensitivity of ovarian cancer cells to treatment with various forms of small molecule tyrosine kinase inhibitors and cytotoxic drugs

    OpenAIRE

    Puvanenthiran, Soozana; Essapen, Sharadah; Seddon, Alan M.; Modjtahedi, Helmout

    2016-01-01

    Increased expression and activation of human epidermal growth factor receptor (EGFR) and HER-2 have been reported in numerous cancers. The aim of this study was to determine the sensitivity of a large panel of human ovarian cancer cell lines (OCCLs) to treatment with various forms of small molecule tyrosine kinase inhibitors (TKIs) and cytotoxic drugs. The aim was to see if there was any association between the protein expression of various biomarkers including three putative ovarian cancer s...

  2. Ibrutinib: a first in class covalent inhibitor of Bruton's tyrosine kinase.

    Science.gov (United States)

    Davids, Matthew S; Brown, Jennifer R

    2014-05-01

    Ibrutinib (formerly PCI-32765) is a potent, covalent inhibitor of Bruton's tyrosine kinase, a kinase downstream of the B-cell receptor that is critical for B-cell survival and proliferation. In preclinical studies, ibrutinib bound to Bruton's tyrosine kinase with high affinity, leading to inhibition of B-cell receptor signaling, decreased B-cell activation and induction of apoptosis. In clinical studies, ibrutinib has been well-tolerated and has demonstrated profound anti-tumor activity in a variety of hematologic malignancies, most notably chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL), leading to US FDA approval for relapsed CLL and MCL. Ongoing studies are evaluating ibrutinib in other types of non-Hodgkin's lymphoma, such as diffuse large B-cell lymphoma and Waldenström's macrogobulinemia, in larger Phase III studies in CLL and MCL, and in combination studies with monoclonal antibodies and chemotherapy. Future studies will combine ibrutinib with other promising novel agents currently in development in hematologic malignancies.

  3. Oligonucleotide aptamers against tyrosine kinase receptors: Prospect for anticancer applications.

    Science.gov (United States)

    Camorani, Simona; Crescenzi, Elvira; Fedele, Monica; Cerchia, Laura

    2018-04-01

    Transmembrane receptor tyrosine kinases (RTKs) play crucial roles in cancer cell proliferation, survival, migration and differentiation. Area of intense research is searching for effective anticancer therapies targeting these receptors and, to date, several monoclonal antibodies and small-molecule tyrosine kinase inhibitors have entered the clinic. However, some of these drugs show limited efficacy and give rise to acquired resistance. Emerging highly selective compounds for anticancer therapy are oligonucleotide aptamers that interact with their targets by recognizing a specific three-dimensional structure. Because of their nucleic acid nature, the rational design of advanced strategies to manipulate aptamers for both diagnostic and therapeutic applications is greatly simplified over antibodies. In this manuscript, we will provide a comprehensive overview of oligonucleotide aptamers as next generation strategies to efficiently target RTKs in human cancers. Copyright © 2018 Elsevier B.V. All rights reserved.

  4. Expression, purification and preliminary crystallographic studies on the catalytic region of the nonreceptor tyrosine kinase Fes

    International Nuclear Information System (INIS)

    Gnemmi, Ilaria; Scotti, Claudia; Cappelletti, Donata; Canonico, Pier Luigi; Condorelli, Fabrizio; Rosano, Camillo

    2006-01-01

    The catalytic domain of human Fes tyrosine kinase has been cloned, expressed, purified and crystallized. The proto-oncogene tyrosine protein kinase c-fps/fes encodes a structurally unique protein (Fes) of the nonreceptor protein-tyrosine kinase (PTK) family. Its expression has been demonstrated in myeloid haematopoietic cells, vascular endothelial cells and in neurons. In human-derived and murine-derived cell lines, the activated form of this kinase can induce cellular transformation; moreover, it has been shown that Fes is involved in the regulation of cell–cell and cell–matrix interactions mediated by adherens junctions and focal adhesions. The N-terminus of Fes contains the FCH (Fps/Fes/Fer/CIP4 homology) domain, which is unique to the Fes/Fer kinase family. It is followed by three coiled-coil domains and an SH2 (Src-homology 2) domain. The catalytic region (Fes-CR) is located at the C-terminus of the protein. The successful expression, purification and crystallization of the catalytic part of Fes (Fes-CR) are described

  5. Uncommon EGFR mutations in cytological specimens of 1,874 newly diagnosed Indonesian lung cancer patients

    Science.gov (United States)

    Syahruddin, Elisna; Wulandari, Laksmi; Sri Muktiati, Nunuk; Rima, Ana; Soeroso, Noni; Ermayanti, Sabrina; Levi, Michael; Hidajat, Heriawaty; Widjajahakim, Grace; Utomo, Ahmad Rusdan Handoyo

    2018-01-01

    Purpose We aimed to evaluate the distribution of individual epidermal growth factor receptor (EGFR) mutation subtypes found in routine cytological specimens. Patients and methods A retrospective audit was performed on EGFR testing results of 1,874 consecutive cytological samples of newly diagnosed or treatment-naïve Indonesian lung cancer patients (years 2015–2016). Testing was performed by ISO15189 accredited central laboratory. Results Overall test failure rate was 5.1%, with the highest failure (7.1%) observed in pleural effusion and lowest (1.6%) in needle aspiration samples. EGFR mutation frequency was 44.4%. Tyrosine kinase inhibitor (TKI)-sensitive common EGFR mutations (ins/dels exon 19, L858R) and uncommon mutations (G719X, T790M, L861Q) contributed 57.1% and 29%, respectively. Approximately 13.9% of mutation-positive patients carried a mixture of common and uncommon mutations. Women had higher EGFR mutation rate (52.9%) vs men (39.1%; pcytological techniques yielded similar success rate to detect EGFR mutations. Uncommon EGFR mutations were frequent events in Indonesian lung cancer patients. PMID:29615847

  6. Expression of tyrosine kinase gene in mouse thymic stromal cells

    NARCIS (Netherlands)

    Rinke de Wit, T. F.; Izon, D. J.; Revilla, C.; Oosterwegel, M.; Bakker, A. Q.; van Ewijk, W.; Kruisbeek, A. M.

    1996-01-01

    Amongst the most important signal transduction molecules involved in regulating growth and differentiation are the protein tyrosine kinases (PTK). Since T cell development is a consequence of interactions between thymic stromal cells (TSC) and thymocytes, identification of the PTK in both

  7. Signaling by Kit protein-tyrosine kinase--the stem cell factor receptor.

    Science.gov (United States)

    Roskoski, Robert

    2005-11-11

    Signaling by stem cell factor and Kit, its receptor, plays important roles in gametogenesis, hematopoiesis, mast cell development and function, and melanogenesis. Moreover, human and mouse embryonic stem cells express Kit transcripts. Stem cell factor exists as both a soluble and a membrane-bound glycoprotein while Kit is a receptor protein-tyrosine kinase. The complete absence of stem cell factor or Kit is lethal. Deficiencies of either produce defects in red and white blood cell production, hypopigmentation, and sterility. Gain-of-function mutations of Kit are associated with several human neoplasms including acute myelogenous leukemia, gastrointestinal stromal tumors, and mastocytomas. Kit consists of an extracellular domain, a transmembrane segment, a juxtamembrane segment, and a protein kinase domain that contains an insert of about 80 amino acid residues. Binding of stem cell factor to Kit results in receptor dimerization and activation of protein kinase activity. The activated receptor becomes autophosphorylated at tyrosine residues that serve as docking sites for signal transduction molecules containing SH2 domains. The adaptor protein APS, Src family kinases, and Shp2 tyrosyl phosphatase bind to phosphotyrosine 568. Shp1 tyrosyl phosphatase and the adaptor protein Shc bind to phosphotyrosine 570. C-terminal Src kinase homologous kinase and the adaptor Shc bind to both phosphotyrosines 568 and 570. These residues occur in the juxtamembrane segment of Kit. Three residues in the kinase insert domain are phosphorylated and attract the adaptor protein Grb2 (Tyr703), phosphatidylinositol 3-kinase (Tyr721), and phospholipase Cgamma (Tyr730). Phosphotyrosine 900 in the distal kinase domain binds phosphatidylinositol 3-kinase which in turn binds the adaptor protein Crk. Phosphotyrosine 936, also in the distal kinase domain, binds the adaptor proteins APS, Grb2, and Grb7. Kit has the potential to participate in multiple signal transduction pathways as a result of

  8. 4-Hydroxynonenal activates Src through a non-canonical pathway that involves EGFR/PTP1B

    Science.gov (United States)

    Zhang, Hongqiao; Forman, Henry Jay

    2015-01-01

    Src, a non-receptor protein tyrosine kinase involved in many biological processes, can be activated through both redox-dependent and independent mechanisms. 4-Hydroxy-2-nonenal (HNE) is a lipid peroxidation product that is increased in pathophysiological conditions associated with Src activation. This study examined how HNE activates human c-Src. In the canonical pathway Src activation is initiated by dephosphorylation of pTyr530 followed by conformational change that causes Src auto-phosphorylation at Tyr419 and its activation. HNE increased Src activation in both dose- and time-dependent manner, while it also increased Src phosphorylation at Tyr530 (pTyr530 Src), suggesting that HNE activated Src via a non-canonical mechanism. Protein tyrosine phosphatase 1B inhibitor (539741), at concentrations that increased basal pTyr530 Src, also increased basal Src activity and significantly reduced HNE-mediated Src activation. The EGFR inhibitor, AG1478, and EGFR silencing, abrogated HNE-mediated EGFR activation and inhibited basal and HNE-induced Src activity. In addition, AG1478 also eliminated the increase of basal Src activation by a PTP1B inhibitor. Taken together these data suggest that HNE can activate Src partly through a non-canonical pathway involving activation of EGFR and inhibition of PTP1B. PMID:26453921

  9. Engineering of kinase-based protein interacting devices: active expression of tyrosine kinase domains

    KAUST Repository

    Diaz Galicia, Miriam Escarlet

    2018-05-01

    Protein-protein interactions modulate cellular processes in health and disease. However, tracing weak or rare associations or dissociations of proteins is not a trivial task. Kinases are often regulated through interaction partners and, at the same time, themselves regulate cellular interaction networks. The use of kinase domains for creating a synthetic sensor device that reads low concentration protein-protein interactions and amplifies them to a higher concentration interaction which is then translated into a FRET (Fluorescence Resonance Energy Transfer) signal is here proposed. To this end, DNA constructs for interaction amplification (split kinases), positive controls (intact kinase domains), scaffolding proteins and phosphopeptide - SH2-domain modules for the reading of kinase activity were assembled and expression protocols for fusion proteins containing Lyn, Src, and Fak kinase domains in bacterial and in cell-free systems were optimized. Also, two non-overlapping methods for measuring the kinase activity of these proteins were stablished and, finally, a protein-fragment complementation assay with the split-kinase constructs was tested. In conclusion, it has been demonstrated that features such as codon optimization, vector design and expression conditions have an impact on the expression yield and activity of kinase-based proteins. Furthermore, it has been found that the defined PURE cell-free system is insufficient for the active expression of catalytic kinase domains. In contrast, the bacterial co-expression with phosphatases produced active kinase fusion proteins for two out of the three tested Tyrosine kinase domains.

  10. Phosphopeptide occupancy and photoaffinity cross-linking of the v-Src SH2 domain attenuates tyrosine kinase activity.

    Science.gov (United States)

    Garcia, P; Shoelson, S E; Drew, J S; Miller, W T

    1994-12-02

    Phosphorylation of c-Src at carboxyl-terminal Tyr-527 suppresses tyrosine kinase activity and transforming potential, presumably by facilitating the intramolecular interaction of the C terminus of Src with its SH2 domain. In addition, it has been shown previously that occupancy of the c-Src SH2 domain with a phosphopeptide stimulates c-Src kinase catalytic activity. We have performed analogous studies with v-Src, the transforming protein from Rous sarcoma virus, which has extensive homology with c-Src. v-Src lacks an autoregulatory phosphorylation site, and its kinase domain is constitutively active. Phosphopeptides corresponding to the sequences surrounding c-Src Tyr-527 and a Tyr-Glu-Glu-Ile motif from the hamster polyoma virus middle T antigen inhibit tyrosine kinase activity of baculovirus-expressed v-Src 2- and 4-fold, respectively. To determine the mechanism of this regulation, the Tyr-527 phosphopeptide was substituted with the photoactive amino acid p-benzoylphenylalanine at the adjacent positions (N- and C-terminal) to phosphotyrosine. These peptides photoinactivate the v-Src tyrosine kinase 5-fold in a time- and concentration-dependent manner. Furthermore, the peptides cross-link an isolated Src SH2 domain with similar rates and specificity. These data indicate that occupancy of the v-Src SH2 domain induces a conformational change that is transmitted to the kinase domain and attenuates tyrosine kinase activity.

  11. Clinical efficacy of icotinib in lung cancer patients with different EGFR mutation status: a meta-analysis.

    Science.gov (United States)

    Qu, Jian; Wang, Ya-Nan; Xu, Ping; Xiang, Da-Xiong; Yang, Rui; Wei, Wei; Qu, Qiang

    2017-05-16

    Icotinib is a novel and the third listed epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs), which exerts a good anti-tumor efficacy on non-small cell lung cancer (NSCLC). The efficacy of EGFR-TKIs has been shown to be associated with the EGFR mutation status, especially exon 19 deletion (19Del) and exon 21 L858R mutation. Therefore, a meta-analysis was performed to assess the efficacy of icotinib in NSCLC patients harboring EGFR mutations (19Del or L858R) and wild type (19Del and L858R loci wild type). A total of 24 studies were included for comparing the objective response rate (ORR) in the EGFR wild type and mutant patients treated with icotinib. The ORRs of EGFR mutant patients (19Del or L858R) are better than those of EGFR wild type patients (OR = 7.03(5.09-9.71), P icotinib treatment; EGFR 19Del patients treated with icotinib have better ORRs than EGFR L858R patients. EGFR mutation status is a useful biomarker for the evaluation of icotinib efficacy in NSCLC patients.

  12. Elucidation of the critical epitope of an anti-EGFR monoclonal antibody EMab-134

    Directory of Open Access Journals (Sweden)

    Mika K. Kaneko

    2018-07-01

    Full Text Available The epidermal growth factor receptor (EGFR is a type-1 transmembrane receptor tyrosine kinase, which activates the downstream signaling cascades in many tumors, such as oral and lung cancers. We previously developed EMab-134, a novel anti-EGFR monoclonal antibody (mAb, which reacts with endogenous EGFR-expressing cancer cell lines and normal cells independent of glycosylation in Western blotting, flow cytometry, and immunohistochemical analysis. EMab-134 showed very high sensitivity (94.7% to oral squamous cell carcinomas in immunohistochemical analysis. In this study, we performed enzyme-linked immunosorbent assay (ELISA, flow cytometry, and immunohistochemical analysis to determine the epitope of EMab-134. A blocking peptide (375–394 amino acids of EGFR neutralized the EMab-134 reaction against oral cancer cells in flow cytometry and immunohistochemistry. The minimum epitope of EMab-134 was found to be the 377-RGDSFTHTPP−386 sequence. Our findings can be applied for the production of more functional anti-EGFR mAbs that in turn can be used for antitumor treatments.

  13. Navigating into the binding pockets of the HER family protein kinases: discovery of novel EGFR inhibitor as antitumor agent

    Directory of Open Access Journals (Sweden)

    Liu W

    2015-07-01

    Full Text Available Wei Liu,1,* Jin-Feng Ning,2,* Qing-Wei Meng,1 Jing Hu,1 Yan-Bin Zhao,1 Chao Liu,3 Li Cai11The Fourth Department of Medical Oncology, Harbin Medical University Cancer Hospital, 2The Thoracic Surgery Department, Harbin Medical University Cancer Hospital, Harbin, People’s Republic of China; 3General Surgery Department, Mudanjiang Guanliju Central Hospital, Mishan, Heilongjiang Province, People’s Republic of China*These authors contributed equally to this workAbstract: The epidermal growth factor receptor (EGFR family has been validated as a successful antitumor drug target for decades. Known EGFR inhibitors were exposed to distinct drug resistance against the various EGFR mutants within non-small-cell lung cancer (NSCLC, particularly the T790M mutation. Although so far a number of studies have been reported on the development of third-generation EGFR inhibitors for overcoming the resistance issue, the design procedure largely depends on the intuition of medicinal chemists. Here we retrospectively make a detailed analysis of the 42 EGFR family protein crystal complexes deposited in the Protein Data Bank (PDB. Based on the analysis of inhibitor binding modes in the kinase catalytic cleft, we identified a potent EGFR inhibitor (compound A-10 against drug-resistant EGFR through fragment-based drug design. This compound showed at least 30-fold more potency against EGFR T790M than the two control molecules erlotinib and gefitinib in vitro. Moreover, it could exhibit potent HER2 inhibitory activities as well as tumor growth inhibitory activity. Molecular docking studies revealed a structural basis for the increased potency and mutant selectivity of this compound. Compound A-10 may be selected as a promising candidate in further preclinical studies. In addition, our findings could provide a powerful strategy to identify novel selective kinase inhibitors on the basis of detailed kinase–ligand interaction space in the PDB.Keywords: EGFR, kinase

  14. Efficacy of ponatinib against ABL tyrosine kinase inhibitor-resistant leukemia cells

    International Nuclear Information System (INIS)

    Okabe, Seiichi; Tauchi, Tetsuzo; Tanaka, Yuko; Ohyashiki, Kazuma

    2013-01-01

    Highlights: •Efficacy of ponatinib against ABL tyrosine kinase inhibitor-resistant leukemia cells okabe et al. •Imatinib or nilotinib resistance was involved Src family kinase. •The BCR-ABL point mutation (E334V) was highly resistant to imatinib or nilotinib. •Ponatinib was a powerful strategy against imatinib or nilotinib resistant Ph-positive cells. -- Abstract: Because a substantial number of patients with chronic myeloid leukemia acquire resistance to ABL tyrosine kinase inhibitors (TKIs), their management remains a challenge. Ponatinib, also known as AP24534, is an oral multi-targeted TKI. Ponatinib is currently being investigated in a pivotal phase 2 clinical trial. In the present study, we analyzed the molecular and functional consequences of ponatinib against imatinib- or nilotinib-resistant (R) K562 and Ba/F3 cells. The proliferation of imatinib- or nilotinib-resistant K562 cells did not decrease after treatment with imatinib or nilotinib. Src family kinase Lyn was activated. Point mutation Ba/F3 cells (E334 V) were also highly resistant to imatinib and nilotinib. Treatment with ponatinib for 72 h inhibited the growth of imatinib- and nilotinib-resistant cells. The phosphorylation of BCR-ABL, Lyn, and Crk-L was reduced. This study demonstrates that ponatinib has an anti-leukemia effect by reducing ABL and Lyn kinase activity and this information may be of therapeutic relevance

  15. Efficacy of ponatinib against ABL tyrosine kinase inhibitor-resistant leukemia cells

    Energy Technology Data Exchange (ETDEWEB)

    Okabe, Seiichi, E-mail: okabe@tokyo-med.ac.jp; Tauchi, Tetsuzo; Tanaka, Yuko; Ohyashiki, Kazuma

    2013-06-07

    Highlights: •Efficacy of ponatinib against ABL tyrosine kinase inhibitor-resistant leukemia cells okabe et al. •Imatinib or nilotinib resistance was involved Src family kinase. •The BCR-ABL point mutation (E334V) was highly resistant to imatinib or nilotinib. •Ponatinib was a powerful strategy against imatinib or nilotinib resistant Ph-positive cells. -- Abstract: Because a substantial number of patients with chronic myeloid leukemia acquire resistance to ABL tyrosine kinase inhibitors (TKIs), their management remains a challenge. Ponatinib, also known as AP24534, is an oral multi-targeted TKI. Ponatinib is currently being investigated in a pivotal phase 2 clinical trial. In the present study, we analyzed the molecular and functional consequences of ponatinib against imatinib- or nilotinib-resistant (R) K562 and Ba/F3 cells. The proliferation of imatinib- or nilotinib-resistant K562 cells did not decrease after treatment with imatinib or nilotinib. Src family kinase Lyn was activated. Point mutation Ba/F3 cells (E334 V) were also highly resistant to imatinib and nilotinib. Treatment with ponatinib for 72 h inhibited the growth of imatinib- and nilotinib-resistant cells. The phosphorylation of BCR-ABL, Lyn, and Crk-L was reduced. This study demonstrates that ponatinib has an anti-leukemia effect by reducing ABL and Lyn kinase activity and this information may be of therapeutic relevance.

  16. Interaction between focal adhesion kinase and Crk-associated tyrosine kinase substrate p130Cas.

    Science.gov (United States)

    Polte, T R; Hanks, S K

    1995-11-07

    The focal adhesion kinase (FAK) has been implicated in integrin-mediated signaling events and in the mechanism of cell transformation by the v-Src and v-Crk oncoproteins. To gain further insight into FAK signaling pathways, we used a two-hybrid screen to identify proteins that interact with mouse FAK. The screen identified two proteins that interact with FAK via their Src homology 3 (SH3) domains: a v-Crk-associated tyrosine kinase substrate (Cas), p130Cas, and a still uncharacterized protein, FIPSH3-2, which contains an SH3 domain closely related to that of p130Cas. These SH3 domains bind to the same proline-rich region of FAK (APPKPSR) encompassing residues 711-717. The mouse p130Cas amino acid sequence was deduced from cDNA clones, revealing an overall high degree of similarity to the recently reported rat sequence. Coimmunoprecipitation experiments confirmed that p130Cas and FAK are associated in mouse fibroblasts. The stable interaction between p130Cas and FAK emerges as a likely key element in integrin-mediated signal transduction and further represents a direct molecular link between the v-Src and v-Crk oncoproteins. The Src family kinase Fyn, whose Src homology 2 (SH2) domain binds to the major FAK autophosphorylation site (tyrosine 397), was also identified in the two-hybrid screen.

  17. Staurosporine scaffold-based rational discovery of the wild-type sparing reversible inhibitors of EGFR T790M gatekeeper mutant in lung cancer with analog-sensitive kinase technology.

    Science.gov (United States)

    Song, Xiaoyun; Liu, Xingcai; Ding, Xi

    2017-04-01

    The human epidermal growth factor receptor (EGFR) has been established as an attractive target for lung cancer therapy. However, an acquired EGFR T790M gatekeeper mutation is frequently observed in patients treated with first-line anticancer agents such as gefitinib and erlotinib to cause drug resistance, largely limiting the application of small-molecule kinase inhibitors in EGFR-targeted chemotherapy. Previously, the reversible pan-kinase inhibitor staurosporine and its several analogs such as Gö6976 and K252a have been reported to selectively inhibit the EGFR T790M mutant (EGFR T790M ) over wild-type kinase (EGFR WT ), suggesting that the staurosporine scaffold is potentially to develop the wild-type sparing reversible inhibitors of EGFR T790M . Here, we systematically evaluated the inhibitor response of 28 staurosporine scaffold-based compounds to EGFR T790M mutation at structural, energetic, and molecular levels by using an integrated in silico-in vitro analog-sensitive (AS) kinase technology. With the strategy, we were able to identify 4 novel wild-type sparing inhibitors UCN-01, UCN-02, AFN941, and SB-218078 with high or moderate selectivity of 30-, 45-, 5-, and 8-fold for EGFR T790M over EGFR WT , respectively, which are comparable with or even better than that of the parent compound staurosporine (24-fold). Molecular modeling and structural analysis revealed that van der Waals contacts and hydrophobic forces can form between the side chain of mutated residue Met790 and the pyrrolidinone moiety of inhibitor ligand UCN-02, which may simultaneously improve the favorable interaction energy between the kinase and inhibitor, and reduce the unfavorable desolvation penalty upon the kinase-inhibitor binding. A hydroxyl group of UCN-02 additional to staurosporine locates at the pyrrolidinone moiety, which can largely alter the electronic distribution of pyrrolidinone moiety and thus promote the intermolecular interaction with Met790 residue. This can well explain

  18. Ubiquitin Ligase Cbl-b Is Involved in Icotinib (BPI-2009H-Induced Apoptosis and G1 Phase Arrest of EGFR Mutation-Positive Non-Small-Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Xiaodong Mu

    2013-01-01

    Full Text Available Epidermal growth factor receptor (EGFR is one of the most promising targets for non-small-cell lung cancer (NSCLC. Icotinib, a highly selective EGFR tyrosine kinase inhibitor (EGFR-TKI, has shown promising clinical efficacy and safety in patients with NSCLC. The exact molecular mechanism of icotinib remains unclear. In this study, we first investigated the antiproliferative effect of icotinib on NSCLC cells. Icotinib significantly inhibited proliferation of the EGFR-mutated lung cancer HCC827 cells. The IC50 values at 48 and 72 h were 0.67 and 0.07 μM, respectively. Flow cytometric analysis showed that icotinib caused the G1 phase arrest and increased the rate of apoptosis in HCC827 cells. The levels of cyclin D1 and cyclin A2 were decreased. The apoptotic process was associated with activation of caspase-3, -8, and poly(ADP-ribose polymerase (PARP. Further study revealed that icotinib inhibited phosphorylation of EGFR, Akt, and extracellular signal-regulated kinase. In addition, icotinib upregulated ubiquitin ligase Cbl-b expression. These observations suggest that icotinib-induced upregulation of Cbl-b is responsible, at least in part, for the antitumor effect of icotinib via the inhibition of phosphoinositide 3-kinase (PI3K/Akt and mitogen-activated protein kinase pathways in EGFR-mutated NSCLC cells.

  19. Tyrosine kinase inhibition: A therapeutic target for the management of chronic-phase chronic myeloid leukemia

    Science.gov (United States)

    Jabbour, Elias J; Cortes, Jorge E; Kantarjian, Hagop M

    2014-01-01

    Chronic myeloid leukemia (CML) is a hematologic neoplasm with a progressive, ultimately terminal, disease course. In most cases, CML arises owing to the aberrant formation of a chimeric gene for a constitutively active tyrosine kinase. Inhibition of the signaling activity of this kinase has proved to be a highly successful treatment target transforming the prognosis of patients with CML. New tyrosine kinase inhibitors (TKIs) continue to improve the management of CML, offering alternative options for those resistant to or intolerant of standard TKIs. Here we review the pathobiology of CML and explore emerging strategies to optimize the management of chronic-phase CML, particularly first-line treatment. PMID:24236822

  20. Ibrutinib: a first in class covalent inhibitor of Bruton’s tyrosine kinase

    Science.gov (United States)

    Davids, Matthew S; Brown, Jennifer R

    2015-01-01

    Ibrutinib (formerly PCI-32765) is a potent, covalent inhibitor of Bruton’s tyrosine kinase, a kinase downstream of the B-cell receptor that is critical for B-cell survival and proliferation. In preclinical studies, ibrutinib bound to Bruton’s tyrosine kinase with high affinity, leading to inhibition of B-cell receptor signaling, decreased B-cell activation and induction of apoptosis. In clinical studies, ibrutinib has been well-tolerated and has demonstrated profound anti-tumor activity in a variety of hematologic malignancies, most notably chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL), leading to US FDA approval for relapsed CLL and MCL. Ongoing studies are evaluating ibrutinib in other types of non-Hodgkin’s lymphoma, such as diffuse large B-cell lymphoma and Waldenström’s macrogobulinemia, in larger Phase III studies in CLL and MCL, and in combination studies with monoclonal antibodies and chemotherapy. Future studies will combine ibrutinib with other promising novel agents currently in development in hematologic malignancies. PMID:24941982

  1. Effects of tyrosine kinase and phosphatase inhibitors on mitosis progression in synchronized tobacco BY-2 cells.

    Science.gov (United States)

    Sheremet, Ya A; Yemets, A I; Azmi, A; Vissenberg, K; Verbelen, J P; Blume, Ya B

    2012-01-01

    To test whether reversible tubulin phosphorylation plays any role in the process of plant mitosis the effects of inhibitors of tyrosine kinases, herbimycin A, genistein and tyrphostin AG 18, and of an inhibitor of tyrosine phosphatases, sodium orthovanadate, on microtubule organization and mitosis progression in a synchronized BY-2 culture has been investigated. It was found that treatment with inhibitors of tyrosine kinases of BY-2 cells at the G2/M transition did not lead to visible disturbances of mitotic microtubule structures, while it did reduce the frequency of their appearance. We assume that a decreased tyrosine phosphorylation level could alter the microtubule dynamic instability parameters during interphase/prophase transition. All types of tyrosine kinase inhibitors used caused a prophase delay: herbimycin A and genistein for 2 h, and tyrphostin AG18 for 1 h. Thereafter the peak of mitosis was displaced for 1 h by herbimycin A or genistein exposure, but after tyrphostin AG18 treatment the timing of the mitosis-peak was comparable to that in control cells. Enhancement of tyrosine phosphorylation induced by the tyrosine phosphatase inhibitor resulted in the opposite effect on BY-2 mitosis transition. Culture treatment with sodium orthovanadate during 1 h resulted in an accelerated start of the prophase and did not lead to the alteration in time of the mitotic index peak formation, as compared to control cells. We suppose that the reversible tyrosine phosphorylation can be involved in the regulation of interphase to M phase transition possibly through regulation of microtubule dynamics in plant cells.

  2. Suppressor of cytokine signaling 1 interacts with oncogenic lymphocyte-specific protein tyrosine kinase.

    Science.gov (United States)

    Venkitachalam, Srividya; Chueh, Fu-Yu; Leong, King-Fu; Pabich, Samantha; Yu, Chao-Lan

    2011-03-01

    Lymphocyte-specific protein tyrosine kinase (Lck) plays a key role in T cell signal transduction and is tightly regulated by phosphorylation and dephosphorylation. Lck can function as an oncoprotein when overexpressed or constantly activated by mutations. Our previous studies showed that Lck-induced cellular transformation could be suppressed by enforced expression of suppressor of cytokine signaling 1 (SOCS1), a SOCS family member involved in the negative feedback control of cytokine signaling. We observed attenuated Lck kinase activity in SOCS1-expressing cells, suggesting an important role of SOCS in regulating Lck functions. It remains largely unknown whether and how SOCS proteins interact with the oncogenic Lck kinase. Here, we report that among four SOCS family proteins, SOCS1, SOCS2, SOCS3 and CIS (cytokine-inducible SH2 domain containing protein), SOCS1 has the highest affinity in binding to the oncogenic Lck kinase. We identified the positive regulatory phosphotyrosine 394 residue in the kinase domain as the key interacting determinant in Lck. Additionally, the Lck kinase domain alone is sufficient to bind SOCS1. While the SH2 domain in SOCS1 is important in its association with the oncogenic Lck kinase, other functional domains may also contribute to overall binding affinity. These findings provide important mechanistic insights into the role of SOCS proteins as tumor suppressors in cells transformed by oncogenic protein tyrosine kinases.

  3. Intramolecular regulatory switch in ZAP-70: analogy with receptor tyrosine kinases

    Czech Academy of Sciences Publication Activity Database

    Brdička, Tomáš; Kadlecek, T.A.; Roose, J.P.; Pastuszak, A.W.; Weiss, A.

    2005-01-01

    Roč. 25, č. 12 (2005), s. 4924-4933 ISSN 0270-7306 Institutional research plan: CEZ:AV0Z50520514 Keywords : protein tyrosin- kinase * phosphorylation * ZAP-70 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.093, year: 2005

  4. Small cell lung cancer transformation from EGFR-mutated lung adenocarcinoma: A case report and literatures review.

    Science.gov (United States)

    Liu, Yangyang

    2018-06-03

    Epithelial growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) have markedly improved the response of non-small cell lung cancer (NSCLC) with EGFR-mutant patients. However, these patients inevitably come cross acquired resistance to EGFR-TKIs. The transformation of lung adenocarcinoma to small cell lung cancer (SCLC) following treatment with EGFR-TKIs is rare, which leads to resistance to EGFR-TKIs. The present case concerns a case of a 38-year-old man presenting with cough and dyspnea. Radical resection was performed and confirmed an EGFR exon 21 L858R lung adenocarcinoma. However, the patient suffered pleural metastasis after successful treatment with surgery and adjuvant treatment. So, erlotinib was administered with 18 months. Because of enlarged pleural nodule, repeat biopsy identified an SCLC and chemotherapy was started. However, despite the brief success of chemotherapy, our patient suffered brain metastasis. Our case emaphsizes both the profile of transformation from NSCLC to SCLC and the importance of repeat biopsy dealing with drug resistance. We also summarize the clinical characteristics, mechanisms, predictors of SCLC transformation, treatment after transformation and other types of transformation to SCLC.

  5. Frequent activation of EGFR in advanced chordomas

    Directory of Open Access Journals (Sweden)

    Dewaele Barbara

    2011-07-01

    Full Text Available Abstract Background Chordomas are rare neoplasms, arising from notochordal remnants in the midline skeletal axis, for which the current treatment is limited to surgery and radiotherapy. Recent reports suggest that receptor tyrosine kinases (RTK might be essential for the survival or proliferation of chordoma cells, providing a rationale for RTK targeted therapy. Nevertheless, the reported data are conflicting, most likely due to the assorted tumor specimens used for the studies and the heterogeneous methodological approaches. In the present study, we performed a comprehensive characterization of this rare entity using a wide range of assays in search for relevant therapeutic targets. Methods Histopathological features of 42 chordoma specimens, 21 primary and 21 advanced, were assessed by immunohistochemistry and fluorescent in situ hybridization (FISH using PDGFRB, CSF1R, and EGFR probes. Twenty-two of these cases, for which frozen material was available (nine primary and 13 advanced tumors, were selectively analyzed using the whole-genome 4.3 K TK-CGH-array, phospho-kinase antibody array or Western immunoblotting. The study was supplemented by direct sequencing of KIT, PDGFRB, CSF1R and EGFR. Results We demonstrated that EGFR is frequently and the most significantly activated RTK in chordomas. Furthermore, concurrent to EGFR activation, the tumors commonly reveal co-activation of alternative RTK. The consistent activation of AKT, the frequent loss of the tumor suppressor PTEN allele, the recurrent activation of upstream RTK and of downstream effectors like p70S6K and mTOR, all indicate the PI3K/AKT pathway as an important mediator of transformation in chordomas. Conclusions Given the complexity of the signaling in chordomas, combined treatment regimens targeting multiple RTK and downstream effectors are likely to be the most effective in these tumors. Personalized therapy with careful selection of the patients, based on the molecular profile of

  6. Non-small-cell lung cancer cells combat epidermal growth factor receptor tyrosine kinase inhibition through immediate adhesion-related responses

    Directory of Open Access Journals (Sweden)

    Wang HY

    2016-05-01

    Full Text Available Hsian-Yu Wang,1,2 Min-Kung Hsu,3,4 Kai-Hsuan Wang,1 Ching-Ping Tseng,2,4 Feng-Chi Chen,3,4 John T-A Hsu1,4 1Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes (NHRI, Zhunan, Miaoli County, 2Institute of Molecular Medicine and Bioengineering, National Chiao Tung University (NCTU, Hsinchu, 3Division of Biostatistics and Bioinformatics, Institute of Population Health Sciences, National Health Research Institutes (NHRI, Zhunan, Miaoli County, 4Department of Biological Science and Technology, National Chiao Tung University (NCTU, Hsinchu, Taiwan, Republic of China Background: Epidermal growth factor receptor (EGFR tyrosine kinase inhibitors (TKIs, such as gefitinib, erlotinib, and afatinib, have greatly improved treatment efficacy in non-small cell lung cancer (NSCLC patients with drug-sensitive EGFR mutations. However, in some TKI responders, the benefits of such targeted therapies are limited by the rapid development of resistance, and strategies to overcome this resistance are urgently needed. Studies of drug resistance in cancer cells typically involve long term in vitro induction to obtain stably acquired drug-resistant cells followed by elucidation of resistance mechanisms, but the immediate responses of cancer cells upon drug treatment have been ignored. The aim of this study was to investigate the immediate responses of NSCLC cells upon treatment with EGFR TKIs.Results: Both NSCLC cells, ie, PC9 and H1975, showed immediate enhanced adhesion-related responses as an apoptosis-countering mechanism upon first-time TKI treatment. By gene expression and pathway analysis, adhesion-related pathways were enriched in gefitinib-treated PC9 cells. Pathway inhibition by small-hairpin RNAs or small-molecule drugs revealed that within hours of EGFR TKI treatment, NSCLC cells used adhesion-related responses to combat the drugs. Importantly, we show here that the Src family inhibitor, dasatinib, dramatically inhibits

  7. Phorbol ester-induced serine phosphorylation of the insulin receptor decreases its tyrosine kinase activity.

    Science.gov (United States)

    Takayama, S; White, M F; Kahn, C R

    1988-03-05

    The effect of 12-O-tetradecanoylphorbol-13-acetate (TPA) on the function of the insulin receptor was examined in intact hepatoma cells (Fao) and in solubilized extracts purified by wheat germ agglutinin chromatography. Incubation of ortho[32P]phosphate-labeled Fao cells with TPA increased the phosphorylation of the insulin receptor 2-fold after 30 min. Analysis of tryptic phosphopeptides from the beta-subunit of the receptor by reverse-phase high performance liquid chromatography and determination of their phosphoamino acid composition suggested that TPA predominantly stimulated phosphorylation of serine residues in a single tryptic peptide. Incubation of the Fao cells with insulin (100 nM) for 1 min stimulated 4-fold the phosphorylation of the beta-subunit of the insulin receptor. Prior treatment of the cells with TPA inhibited the insulin-stimulated tyrosine phosphorylation by 50%. The receptors extracted with Triton X-100 from TPA-treated Fao cells and purified on immobilized wheat germ agglutinin retained the alteration in kinase activity and exhibited a 50% decrease in insulin-stimulated tyrosine autophosphorylation and phosphotransferase activity toward exogenous substrates. This was due primarily to a decrease in the Vmax for these reactions. TPA treatment also decreased the Km of the insulin receptor for ATP. Incubation of the insulin receptor purified from TPA-treated cells with alkaline phosphatase decreased the phosphate content of the beta-subunit to the control level and reversed the inhibition, suggesting that the serine phosphorylation of the beta-subunit was responsible for the decreased tyrosine kinase activity. Our results support the notion that the insulin receptor is a substrate for protein kinase C in the Fao cell and that the increase in serine phosphorylation of the beta-subunit of the receptor produced by TPA treatment inhibited tyrosine kinase activity in vivo and in vitro. These data suggest that protein kinase C may regulate the function

  8. [Recent Advances and Prospect of Advanced Non-small Cell Lung Cancer Targeted 
Therapy: Focus on Small Molecular Tyrosine Kinase Inhibitors].

    Science.gov (United States)

    Zhang, Guowei; Wang, Huijuan; Ma, Zhiyong

    2017-04-20

    At present the treatment of advanced non-small cell lung cancer enters a targeted era and develops rapidly. New drugs appear constantly. Small molecular tyrosine kinase inhibitors have occupied the biggest piece of the territory, which commonly have a clear biomarker as predictor, and show remarkable effect in specific molecular classification of patients. The epidermal growth factor tyrosine kinase inhibitors such as gefitinib, erlotinib, icotinib and anaplastic lymphoma kinase tyrosine kinase inhibitors crizotinib have brought a milestone advance. In recent years new generations of tyrosine kinase inhibitors have achieved a great success in patients with acquired resistance to the above two kinds of drugs. At the same time new therapeutic targets are constantly emerging. So in this paper, we reviewed and summarized the important drugs and clinical trails on this topic, and made a prospect of the future development.

  9. Recent Advances and Prospect of Advanced Non-small Cell Lung Cancer Targeted 
Therapy: Focus on Small Molecular Tyrosine Kinase Inhibitors

    Directory of Open Access Journals (Sweden)

    Guowei ZHANG

    2017-04-01

    Full Text Available At present the treatment of advanced non-small cell lung cancer enters a targeted era and develops rapidly. New drugs appear constantly. Small molecular tyrosine kinase inhibitors have occupied the biggest piece of the territory, which commonly have a clear biomarker as predictor, and show remarkable effect in specific molecular classification of patients. The epidermal growth factor tyrosine kinase inhibitors such as gefitinib, erlotinib, icotinib and anaplastic lymphoma kinase tyrosine kinase inhibitors crizotinib have brought a milestone advance. In recent years new generations of tyrosine kinase inhibitors have achieved a great success in patients with acquired resistance to the above two kinds of drugs. At the same time new therapeutic targets are constantly emerging. So in this paper, we reviewed and summarized the important drugs and clinical trails on this topic, and made a prospect of the future development.

  10. LC-MS/MS assay for the quantitation of the tyrosine kinase inhibitor neratinib in human plasma.

    Science.gov (United States)

    Kiesel, Brian F; Parise, Robert A; Wong, Alvin; Keyvanjah, Kiana; Jacobs, Samuel; Beumer, Jan H

    2017-02-05

    Neratinib is an orally available tyrosine kinase inhibitor targeting HER2 (ERBB2) and EGFR (ERBB). It is being clinically evaluated for the treatment of breast and other solid tumors types as a single agent or in combination with other chemotherapies. In support of several phase I/II clinical trials investigating neratinib combinations, we developed and validated a novel LC-MS/MS assay for the quantification of neratinib in 100μL of human plasma with a stable isotopic internal standard. Analytes were extracted from plasma using protein precipitation and evaporation of the resulting supernatant followed by resuspension. Chromatographic separation was achieved using an Acquity UPLC BEH Shield RP18 column and a gradient methanol-water mobile phase containing 10% ammonium acetate. An ABI 4000 mass spectrometer and electrospray positive mode ionization were used for detection. The assay was linear from 2 to 1,000ng/mL and proved to be accurate (98.9-106.5%) and precise (neratinib. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Targeting the epidermal growth factor receptor in non-small cell lung cancer cells: the effect of combining RNA interference with tyrosine kinase inhibitors or cetuximab

    Directory of Open Access Journals (Sweden)

    Chen Gang

    2012-03-01

    Full Text Available Abstract Background The epidermal growth factor receptor (EGFR is a validated therapeutic target in non-small cell lung cancer (NSCLC. However, current single agent receptor targeting does not achieve a maximal therapeutic effect, and some mutations confer resistance to current available agents. In the current study we have examined, in different NSCLC cell lines, the combined effect of RNA interference targeting the EGFR mRNA, and inactivation of EGFR signaling using different receptor tyrosine kinase inhibitors (TKIs or a monoclonal antibody cetuximab. Methods NSCLC cells (cell lines HCC827, H292, H358, H1650, and H1975 were transfected with EGFR siRNA and/or treated with the TKIs gefitinib, erlotinib, and afatinib, and/or with the monoclonal antibody cetuximab. The reduction of EGFR mRNA expression was measured by real-time quantitative RT-PCR. The down-regulation of EGFR protein expression was measured by western blot, and the proliferation, viability, caspase3/7 activity, and apoptotic morphology were monitored by spectrophotometry, fluorimetry, and fluorescence microscopy. The combined effect of EGFR siRNA and different drugs was evaluated using a combination index. Results EGFR-specific siRNA strongly inhibited EGFR protein expression almost equally in all cell lines and inhibited cell growth and induced cell apoptosis in all NSCLC cell lines studied, albeit with a different magnitude. The effects on growth obtained with siRNA was strikingly different from the effects obtained with TKIs. The effects of siRNA probably correlate with the overall oncogenic significance of the receptor, which is only partly inhibited by the TKIs. The cells which showed weak response to TKIs, such as the H1975 cell line containing the T790M resistance mutation, were found to be responsive to siRNA knockdown of EGFR, as were cell lines with downstream TKI resistance mutations. The cell line HCC827, harboring an exon 19 deletion mutation, was more than 10-fold

  12. Urokinase receptor expression involves tyrosine phosphorylation of phosphoglycerate kinase.

    Science.gov (United States)

    Shetty, Praveenkumar; Velusamy, Thirunavukkarasu; Bhandary, Yashodhar P; Liu, Ming C; Shetty, Sreerama

    2010-02-01

    The interaction of urokinase-type plasminogen activator (uPA) with its receptor, uPAR, plays a central role in several pathophysiological processes, including cancer. uPA induces its own cell surface receptor expression through stabilization of uPAR mRNA. The mechanism involves binding of a 51 nt uPAR mRNA coding sequence with phosphoglycerate kinase (PGK) to down regulate cell surface uPAR expression. Tyrosine phosphorylation of PGK mediated by uPA treatment enhances uPAR mRNA stabilization. In contrast, inhibition of tyrosine phosphorylation augments PGK binding to uPAR mRNA and attenuates uPA-induced uPAR expression. Mapping the specific peptide region of PGK indicated that its first quarter (amino acids 1-100) interacts with uPAR mRNA. To determine if uPAR expression by uPA is regulated through activation of tyrosine residues of PGK, we mutated the specific tyrosine residue and tested mutant PGK for its ability to interfere with uPAR expression. Inhibition of tyrosine phosphorylation by mutating Y76 residue abolished uPAR expression induced by uPA treatment. These findings collectively demonstrate that Y76 residue present in the first quarter of the PGK molecule is involved in lung epithelial cell surface uPAR expression. This region can effectively mimic the function of a whole PGK molecule in inhibiting tumor cell growth.

  13. Bruton tyrosine kinase inhibition in chronic lymphocytic leukemia.

    Science.gov (United States)

    Maddocks, Kami; Jones, Jeffrey A

    2016-04-01

    Chronic lymphocytic leukemia (CLL) is the most common adult leukemia and remains incurable outside of the setting of allogeneic stem cell transplant. While the standard therapy for both initial and relapsed CLL has traditionally included monoclonal antibody therapy in combination with chemotherapy, there are patients with high-risk disease features including unmutated IgVH, del(11q22) and del(17p13) that are associated with poor overall responses to these therapies with short time to relapse and shortened overall survival. Additionally, many of these therapies have a high rate of infectious toxicity in a population already at increased risk. Targeting the B-cell receptor (BCR) signaling pathway has emerged as a promising therapeutic advance in a variety of B-cell malignancies, including CLL. Bruton agammaglobulinemia tyrosine kinase (Btk) is a tyrosine kinase in the BCR pathway critical to the survival of both normal and malignant B cells and inhibition of this kinase has shown to block the progression of CLL. Ibrutinib, a first in class oral inhibitor of Btk, has shown promise as a very effective agent in the treatment of CLL-in both relapsed and upfront therapy, alone and in combination with other therapies, and in patients of all-risk disease-which has led to its approval in relapsed CLL and as frontline therapy in patients with the high-risk del(17p13) disease. Several studies are ongoing to evaluate the efficacy and safety of ibrutinib in combination with chemotherapy as frontline treatment for CLL and investigation into newer-generation Btk inhibitors is also underway. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. The Efficacy of Single-Agent Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor Therapy in Biologically Selected Patients with Non-Small-Cell Lung Cancer: A Meta-Analysis of 19 Randomized Controlled Trials.

    Science.gov (United States)

    Li, Guifang; Gao, Shunji; Sheng, Zhixin; Li, Bin

    2016-01-01

    To determine the efficacy of first-generation single-agent epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) therapy in advanced non-small-cell lung cancer patients with known EGFR mutation status, we undertook this pooled analysis. We searched for randomized controlled trials (RCTs) in Medline, Embase, the Cochrane Controlled Trials Register, the Science Citation Index, and the American Society of Clinical Oncology annual meetings. Out of 2,129 retrieved articles, 19 RCTs enrolling 2,016 patients with wild-type EGFR tumors and 1,034 patients with mutant EGFR tumors were identified. For these EGFR mutant patients, single-agent EGFR-TKI therapy improved progression-free survival (PFS) over chemotherapy: the summary hazard ratios (HRs) were 0.41 (p well as chemotherapy in the first-line setting (HR = 1.65, p = 0.03) and in the second-/third-line setting (HR = 1.27, p = 0.006). No statistically significant difference was observed in terms of overall survival (OS). Using platinum-based doublet chemotherapy as a common comparator, indirect comparison showed the superior efficacy of single-agent EGFR-TKI therapy over EGFR-TKIs added to chemotherapy in PFS [HR = 1.35 (1.03, 1.77), p = 0.03]. Additionally, a marginal trend towards the same direction was found in the OS analysis [HR = 1.16 (0.99, 1.35), p = 0.06]. Interestingly, for those EGFR wild-type tumors, single-agent EGFR-TKI therapy was inferior to EGFR-TKIs added to chemotherapy in PFS [HR = 0.38 (0.33, 0.44), p chemotherapy. However, single-agent EGFR-TKI therapy was inferior to chemotherapy in PFS for those EGFR wild-type patients. Single-agent EGFR-TKI therapy could improve PFS over the combination of EGFR-TKIs and chemotherapy in these EGFR mutant patients. However, EGFR-TKIs combined with chemotherapy could provide additive PFS and OS benefit over single-agent EGFR-TKI therapy in those EGFR wild-type patients. © 2016 S. Karger AG, Basel.

  15. Icotinib, a potent and specific EGFR tyrosine kinase inhibitor, inhibits growth of squamous cell carcinoma cell line A431 through negatively regulating AKT signaling.

    Science.gov (United States)

    Gao, Zhenzhen; Chen, Wei; Zhang, Xiaohua; Cai, Peifen; Fang, Xianying; Xu, Qiang; Sun, Yang; Gu, Yanhong

    2013-06-01

    Icotinib is a potent and specific epidermal growth factor receptor tyrosine kinase inhibitor. In this study, we reported that icotinib had the antitumor activity on human squamous cell carcinoma cell line A431 in vitro. Meanwhile, adhesion to fibronectin and expression of integrin α3 and β1 were significantly reduced in a dose-dependent manner after the treatment of icotinib. Moreover, icotinib induced cell cycle arrested and affected expression of various cell cycle related proteins in squamous cancer cell line A431, whereas it did not cause apoptosis. Furthermore, icotinib remarkably down-regulated phosphorylation of protein kinase B (AKT) though blocking the interaction between 3-phosphoinositide-dependent protein kinase-1 (PDK1) and AKT in A431 cells. Taken together, it is shown that the small molecular compound, icotinib, has an anti-squamous cell carcinoma activity in vitro and its antitumor mechanism is associated with the blockage of the interaction between PDK1 and AKT. These results provide a novel strategy for anti-squamous cell carcinoma therapy. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  16. Should EGFR mutations be tested in advanced lung squamous cell carcinomas to guide frontline treatment?

    Science.gov (United States)

    Chiu, Chao-Hua; Chou, Teh-Ying; Chiang, Chi-Lu; Tsai, Chun-Ming

    2014-10-01

    There is no argument over using epidermal growth factor receptor (EGFR) mutation status to guide the frontline treatment for advanced lung adenocarcinoma (LADC); however, the role of the testing in lung squamous cell carcinoma (LSQC) remains controversial. Currently, the guidelines/consensus statements regarding EGFR mutation testing in LSQC are not consistent among different oncology societies. American Society of Clinical Oncology recommends performing EGFR mutation testing in all patients; European Society for Medical Oncology, College of American Pathologists/International Association for the Study of Lung Cancer/Association for Molecular Pathology, and National Comprehensive Cancer Network suggest for some selected group. EGFR mutation is rarely found in LSQC; however, more importantly, it is not a valid predictive biomarker for EGFR tyrosine kinase inhibitors (EGFR-TKI) in LSQC as it has been shown in LADC. Available data showed that the response rate and progression-free survival in EGFR mutant LSQC patients treated with EGFR-TKI are not better than that observed in patients treated with platinum-doublet chemotherapy in the first-line setting. Therefore, in contrast to advanced LADC, EGFR mutation testing may not be necessarily performed upfront in advanced LSQC because not only the mutation rate is low, but also the predictive value is insufficient. For LSQC patients with known sensitizing-EGFR mutations, both conventional chemotherapy and EGFR-TKI are acceptable frontline treatment options.

  17. Feasibility Study of Sequentially Alternating EGFR-TKIs and Chemotherapy for Patients with Non-small Cell Lung Cancer.

    Science.gov (United States)

    Takemura, Yoshizumi; Chihara, Yusuke; Morimoto, Yoshie; Tanimura, Keiko; Imabayashi, Tatsuya; Seko, Yurie; Kaneko, Yoshiko; Date, Koji; Ueda, Mikio; Arimoto, Taichiro; Iwasaki, Yoshinobu; Takayama, Koichi

    2018-04-01

    The purpose of this trial was to evaluate the feasibility and efficacy of alternating platinum-based doublet chemotherapy with epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) in patients with EGFR-mutant non-small cell lung cancer (NSCLC). Chemotherapy-naive patients with advanced NSCLC harboring an EGFR mutation were enrolled. All patients underwent induction chemotherapy by sequentially alternating pemetrexed/cisplatin/bevacizumab and EGFR-TKIs followed by maintenance therapy with pemetrexed/bevacizumab and EGFR-TKIs. The primary outcome was the completion rate of the induction therapy. Eighteen eligible patients were enrolled between May 2011 and March 2016. The completion rate of induction therapy was 72.2% (13/18). Unfortunately, one patient developed grade 4 acute renal injury, but no other serious complications concerning this protocol were observed. Furthermore, diarrhea, rashes, and hematological adverse effects were mild. The completion rate of induction therapy was promising. Alternating chemotherapy and EGFR-TKIs should be further investigated regarding feasibility and efficacy. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  18. Survival significance of epidermal growth factor receptor tyrosine kinase inhibitors and current staging system for survival after recurrence in patients with completely resected lung adenocarcinoma

    Science.gov (United States)

    Saji, Hisashi; Sakai, Hiroki; Kimura, Hiroyuki; Miyazawa, Tomoyuki; Marushima, Hideki; Nakamura, Haruhiko

    2017-01-01

    Objective We previously reported that the staging system and epidermal growth factor receptor (EGFR) mutation status are key factors for treatment strategy and predicting survival. However, the significance of these factors as predictors of overall survival (OS) and postoperative recurrence survival (PRS) has not been sufficiently elucidated. The objective here was to investigate EGFR mutation status and p-stage, which affect PRS and OS in patients with completely resected lung adenocarcinoma, using a different database. Patients and methods We retrospectively reviewed 56 consecutive lung adenocarcinoma patients with disease recurrence in St. Marianna University Hospital between January 2010 and December 2014. Results EGFR mutants (M) were detected in 16/56 patients (29%). The patients with EGFR M had a better OS than those with EGFR wild-type (WT) status (5-year survival: 50.3% vs 43.1, P=0.133). There was no significant difference in the 3-year recurrence-free survival rate between patients with M and WT (6.3% vs 7.7%, P=0.656), and the patients with EGFR M had a significantly better 3-year PRS than those with WT (77.4% vs 51.7%, P=0.033). The 3-year PRS rate for patients with M/pathologic stage (p-stage) I–II (87.5%) was better than that for patients with M/p-stage III (60.0%), WT/p-stage I–II (52.7%), and WT/p-stage III (43.8%). There was a significant difference between patients with M/p-stage I and WT/p-stage I–II or WT/p-stage III (P=0.021 and 0.030, respectively). During the study period, of the 16 patients with mutants, 12 patients (75%) received EGFR-tyrosine kinase inhibitor (TKI) therapy and among the 40 patients with WT, no patient received EGFR-TKI therapy. Multivariate survival analysis showed that patients with EGFR-TKI therapy had a statistically significant association with favorable PRS (hazard ratio 0.271; 95% confidence interval 0.074–1.000; P=0.050). Conclusion EGFR status and p-stage were found to be essential prognostic factors for

  19. Interdisciplinary management of EGFR-inhibitor-induced skin reactions: a German expert opinion.

    Science.gov (United States)

    Potthoff, K; Hofheinz, R; Hassel, J C; Volkenandt, M; Lordick, F; Hartmann, J T; Karthaus, M; Riess, H; Lipp, H P; Hauschild, A; Trarbach, T; Wollenberg, A

    2011-03-01

    Anti-epidermal growth factor receptor treatment strategies, i.e. monoclonal antibodies such as cetuximab and panitumumab, or epidermal growth factor receptor (EGFR) small molecule tyrosine kinase inhibitors, such as erlotinib and gefitinib, have expanded the treatment options for different tumor types. Dermatologic toxic effects are the most common side-effects of EGFR inhibitor therapy. They can profoundly affect the patient's quality of life. The aim of this study was to provide interdisciplinary expert recommendations on how to treat patients with skin reactions undergoing anti-EGFR treatment. An expert panel from Germany with expertise in medical oncology, dermatology or clinical pharmacology was convened to develop expert recommendations based on published peer-reviewed literature. The expert recommendations for the state-of-the-art treatment of skin reactions induced by EGFR inhibitor therapy include recommendations for diagnostics and grading as well as grade-specific and stage-adapted treatment approaches and preventive measures. It was concluded that EGFR-inhibitor-related dermatologic reactions should always be treated combining basic care of the skin and a specific therapy adapted to stage and grade of skin reaction. For grade 2 and above, specific treatment recommendations for early- and later-stage skin reactions induced by EGFR-inhibitor therapy were proposed. This paper presents a German national expert opinion for the treatment of skin reactions in patients receiving EGFR inhibitor therapy.

  20. Adenocarcinoma of the lung with EGFR gene mutation and subsequent resistance mechanisms exploration: case report

    Directory of Open Access Journals (Sweden)

    Xu L

    2017-09-01

    Full Text Available Li Xu,1,2 Qian Z Wang,1,2 Lin Wu1,2 1Department of the Second Chest Medicine, Hunan Cancer Hospital, Changsha, Hunan, People’s Republic of China; 2Department of the Second Chest Medicine, The Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, People’s Republic of China Abstract: The treatment of lung cancer has made paradigm-shift advancements in the past decade with the development of therapies directed at specific genetic alterations, such as epidermal growth factor receptor (EGFR. Here, we present a rare case of lung adenocarcinoma harboring EGFR activating mutation and ALK overexpression. During the EGFR-tyrosine kinase inhibitors treatment, next-generation sequencing revealed phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin pathway amplifications in tumor specimen and subsequent T790M mutation via plasma circulating tumor DNA. In conclusion, this case illustrates the existence of concomitant resistance mechanisms and demonstrates that circulating tumor DNA can reflect tumor heterogeneity. Keywords: epidermal growth factor receptor, PI3K/Akt/mTOR pathway, T790M, next-generation sequencing, circulating tumor DNA

  1. Brief report: Afatinib and cetuximab in four patients with EGFR exon 20 insertion positive advanced non-small-cell lung cancer.

    Science.gov (United States)

    van Veggel, Bianca; de Langen, Adrianus J; Hashemi, Sayed; Monkhorst, Kim; Heideman, Daniëlle A M; Thunnissen, Erik; Smit, Egbert F

    2018-04-24

    Epidermal growth factor receptor (EGFR) exon 20 insertions comprise 4-9% of EGFR mutated non-small-cell lung cancer (NSCLC). Despite being an oncogenic driver, they are associated with primary resistance to EGFR tyrosine kinase inhibitors (TKIs). We hypothesized that dual EGFR blockade with afatinib, an irreversible EGFR TKI, and cetuximab, a monoclonal antibody against EGFR, could induce tumor responses. Four patients with EGFR exon 20 insertion positive NSCLC were treated with afatinib 40 mg once daily and cetuximab 250-500 mg/m 2 every two weeks. All patients had stage IV adenocarcinoma of the lung harboring an EGFR exon 20 insertion mutation. Previous lines of treatment consisted of platinum doublet chemotherapy (n=4) and EGFR TKI (n=2). Three out of four patients showed a partial response according to RECIST 1.1. Median progression-free survival was 5.4 months (95% confidence interval 0.0 - 14.2 months; range 2.7 - 17.6 months). Toxicity was manageable with appropriate skin management and dose reduction being required in two patients. Dual EGFR blockade with afatinib and cetuximab may induce tumor responses in patients with EGFR exon 20 insertion positive NSCLC. Copyright © 2018. Published by Elsevier Inc.

  2. Nuclear translocation of IGF1R by intracellular amphiregulin contributes to the resistance of lung tumour cells to EGFR-TKI.

    Science.gov (United States)

    Guerard, Marie; Robin, Thomas; Perron, Pascal; Hatat, Anne-Sophie; David-Boudet, Laurence; Vanwonterghem, Laetitia; Busser, Benoit; Coll, Jean-Luc; Lantuejoul, Sylvie; Eymin, Beatrice; Hurbin, Amandine; Gazzeri, Sylvie

    2018-04-28

    Many Receptor Tyrosine Kinases translocate from the cell surface to the nucleus in normal and pathological conditions, including cancer. Here we report the nuclear expression of insulin-like growth factor-1 receptor (IGF1R) in primary human lung tumours. Using lung cancer cell lines and lung tumour xenografts, we demonstrate that the epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) gefitinib induces the nuclear accumulation of IGF1R in mucinous lung adenocarcinoma by a mechanism involving the intracellular re-localization of the growth factor amphiregulin. Amphiregulin allows the binding of IGF1R to importin-β1 and promotes its nuclear transport. The nuclear accumulation of IGF1R by amphiregulin induces cell cycle arrest through p21 WAF1/CIP1 upregulation, and prevents the induction of apoptosis in response to gefitinib. These results identify amphiregulin as the first nuclear localization signal-containing protein that interacts with IGF1R and allows its nuclear translocation. Furthermore they indicate that nuclear expression of IGF1R contributes to EGFR-TKI resistance in lung cancer. Copyright © 2018 Elsevier B.V. All rights reserved.

  3. Sch proteins are localized on endoplasmic reticulum membranes and are redistributed after tyrosine kinase receptor activation

    DEFF Research Database (Denmark)

    Lotti, L V; Lanfrancone, L; Migliaccio, E

    1996-01-01

    area of the cell and mostly associated with the cytosolic side of rough endoplasmic reticulum membranes. Upon epidermal growth factor treatment and receptor tyrosine kinase activation, the immunolabeling became peripheral and was found to be associated with the cytosolic surface of the plasma membrane....... The rough endoplasmic reticulum localization of Shc proteins in unstimulated cells and their massive recruitment to the plasma membrane, endocytic structures, and peripheral cytosol following receptor tyrosine kinase activation could account for multiple putative functions of the adaptor protein....

  4. Therapeutic Efficacy Comparison of 5 Major EGFR-TKIs in Advanced EGFR-positive Non-Small-cell Lung Cancer: A Network Meta-analysis Based on Head-to-Head Trials.

    Science.gov (United States)

    Zhang, Yaxiong; Zhang, Zhonghan; Huang, Xiaodan; Kang, Shiyang; Chen, Gang; Wu, Manli; Miao, Siyu; Huang, Yan; Zhao, Hongyun; Zhang, Li

    2017-09-01

    Five major first- and second-generation epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs), including erlotinib, gefitinib, icotinib, afatinib, and dacomitinib, are currently optional for patients with advanced non-small-cell lung cancer (NSCLC) who harbor EGFR mutations. However, there was no head-to-head-based network meta-analysis among all the TKIs in EGFR-mutated populations. Eligible literature was searched from an electronic database. Data of objective response rate, disease control rate, progression-free survival, and overall survival were extracted from enrolled studies. Multiple treatment comparisons based on Bayesian network integrated the efficacy of all included treatments. Six phase III randomized trials involving 1055 EGFR-mutated patients with advanced NSCLC were enrolled. Multiple treatment comparisons showed that 5 different EGFR-TKIs shared equivalent therapeutic efficacy in terms of all outcome measures. Rank probabilities indicated that dacomitinib and afatinib had potentially better efficacy compared with erlotinib, gefitinib, and icotinib in the EGFR-mutated patients. When compared with other agents, potential survival benefits (progression-free and overall survival) were observed in dacomitinib, whereas afatinib showed a better rank probability in overall response rate and disease control rate. Our study indicated a preferable therapeutic efficacy in the second-generation TKIs (dacomitinib and afatinib) when compared with the first-generation TKIs (erlotinib, gefitinib, and icotinib). Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Tyrosine Kinase Inhibition in HPV-related Squamous Cell Carcinoma Reveals Beneficial Expression of cKIT and Src.

    Science.gov (United States)

    Kramer, Benedikt; Kneissle, Marcel; Birk, Richard; Rotter, Nicole; Aderhold, Christoph

    2018-05-01

    Therapeutic options of locally advanced or metastatic head and neck squamous cell carcinoma (HNSCC) are limited. Src and cKIT are key protein regulators for local tumor progression. The aim of the study was to investigate the therapeutic potential of targeted therapies in human squamous cell carcinoma (HNSCC) in vitro. Therefore, the influence of the selective tyrosine kinase inhibitors niotinib, dasatinib, erlotinib, gefitinib and afatinib on Src and cKIT expression in Human papilloma virus (HPV)-positive and HPV-negative squamous cancer cells (SCC) was analyzed in vitro. ELISA was performed to evaluate the expression of Src and cKIT under the influence of nilotinib, dasatinib, erlotinib, gefitinib and afatinib (10 μmol/l) in HPV-negative and HPV-positive SCC (24-96 h of incubation). Gefitinib significantly increased cKIT expression in HPV-positive and HPV-negative cells whereas nilotinib and afatinib decreased cKIT expression in HPV-positive SCC. The influence of tyrosine kinase inhibitors in HPV-negative SCC was marginal. Surprisingly, Src expression was significantly increased by all tested tyrosine kinase inhibitors in HPV-positive SCC. The results revealed beneficial and unexpected information concerning the interaction of selective tyrosine kinase inhibitors and the tumor biology of HNSCC. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  6. Prevention of Bronchial Hyperplasia by EGFR Pathway Inhibitors in an Organotypic Culture Model

    Science.gov (United States)

    Lee, Jangsoon; Ryu, Seung-Hee; Kang, Shin Myung; Chung, Wen-Cheng; Gold, Kathryn Ann; Kim, Edward S.; Hittelman, Walter N.; Hong, Waun Ki; Koo, Ja Seok

    2011-01-01

    Lung cancer is the leading cause of cancer-related mortality worldwide. Early detection or prevention strategies are urgently needed to increase survival. Hyperplasia is the first morphologic change that occurs in the bronchial epithelium during lung cancer development, followed by squamous metaplasia, dysplasia, carcinoma in situ, and invasive tumor. The current study was designed to determine the molecular mechanisms that control bronchial epithelium hyperplasia. Using primary normal human tracheobronchial epithelial (NHTBE) cells cultured using the 3-dimensional organotypic method, we found that the epidermal growth factor receptor (EGFR) ligands EGF, transforming growth factor-alpha, and amphiregulin induced hyperplasia, as determined by cell proliferation and multilayered epithelium formation. We also found that EGF induced increased cyclin D1 expression, which plays a critical role in bronchial hyperplasia; this overexpression was mediated by activating the mitogen-activated protein kinase pathway but not the phosphoinositide 3-kinase/Akt signaling pathway. Erlotinib, an EGFR tyrosine kinase inhibitor, and U0126, a MEK inhibitor, completely inhibited EGF-induced hyperplasia. Furthermore, a promoter analysis revealed that the activator protein-1 transcription factor regulates EGF-induced cyclin D1 overexpression. Activator protein-1 depletion using siRNA targeting its c-Jun component completely abrogated EGF-induced cyclin D1 expression. In conclusion, we demonstrated that bronchial hyperplasia can be modeled in vitro using primary NHTBE cells maintained in a 3-dimensional (3-D) organotypic culture. EGFR and MEK inhibitors completely blocked EGF-induced bronchial hyperplasia, suggesting that they have a chemopreventive role. PMID:21505178

  7. A role for the tyrosine kinase ACK1 in neurotrophin signaling and neuronal extension and branching

    Science.gov (United States)

    La Torre, A; del Mar Masdeu, M; Cotrufo, T; Moubarak, R S; del Río, J A; Comella, J X; Soriano, E; Ureña, J M

    2013-01-01

    Neurotrophins are involved in many crucial cellular functions, including neurite outgrowth, synapse formation, and plasticity. Although these events have long been known, the molecular determinants underlying neuritogenesis have not been fully characterized. Ack1 (activated Cdc42-associated tyrosine kinase) is a non-receptor tyrosine kinase that is highly expressed in the brain. Here, we demonstrate that Ack1 is a molecular constituent of neurotrophin signaling cascades in neurons and PC12 cells. We report that Ack1 interacts with Trk receptors and becomes tyrosine phosphorylated and its kinase activity is increased in response to neurotrophins. Moreover, our data indicate that Ack1 acts upstream of the Akt and MAPK pathways. We show that Ack1 overexpression induces neuritic outgrowth and promotes branching in neurotrophin-treated neuronal cells, whereas the expression of Ack1 dominant negatives or short-hairpin RNAs counteract neurotrophin-stimulated differentiation. Our results identify Ack1 as a novel regulator of neurotrophin-mediated events in primary neurons and in PC12 cells. PMID:23598414

  8. Ret function in muscle stem cells points to tyrosine kinase inhibitor therapy for facioscapulohumeral muscular dystrophy.

    Science.gov (United States)

    Moyle, Louise A; Blanc, Eric; Jaka, Oihane; Prueller, Johanna; Banerji, Christopher Rs; Tedesco, Francesco Saverio; Harridge, Stephen Dr; Knight, Robert D; Zammit, Peter S

    2016-11-14

    Facioscapulohumeral muscular dystrophy (FSHD) involves sporadic expression of DUX4, which inhibits myogenesis and is pro-apoptotic. To identify target genes, we over-expressed DUX4 in myoblasts and found that the receptor tyrosine kinase Ret was significantly up-regulated, suggesting a role in FSHD. RET is dynamically expressed during myogenic progression in mouse and human myoblasts. Constitutive expression of either RET9 or RET51 increased myoblast proliferation, whereas siRNA-mediated knockdown of Ret induced myogenic differentiation. Suppressing RET activity using Sunitinib, a clinically-approved tyrosine kinase inhibitor, rescued differentiation in both DUX4-expressing murine myoblasts and in FSHD patient-derived myoblasts. Importantly, Sunitinib also increased engraftment and differentiation of FSHD myoblasts in regenerating mouse muscle. Thus, DUX4-mediated activation of Ret prevents myogenic differentiation and could contribute to FSHD pathology by preventing satellite cell-mediated repair. Rescue of DUX4-induced pathology by Sunitinib highlights the therapeutic potential of tyrosine kinase inhibitors for treatment of FSHD.

  9. Tyrosine kinase inhibitors and mesenchymal stromal cells: effects on self-renewal, commitment and functions

    Science.gov (United States)

    Borriello, Adriana; Caldarelli, Ilaria; Bencivenga, Debora; Stampone, Emanuela; Perrotta, Silverio; Oliva, Adriana; Ragione, Fulvio Della

    2017-01-01

    The hope of selectively targeting cancer cells by therapy and eradicating definitively malignancies is based on the identification of pathways or metabolisms that clearly distinguish “normal” from “transformed” phenotypes. Some tyrosine kinase activities, specifically unregulated and potently activated in malignant cells, might represent important targets of therapy. Consequently, tyrosine kinase inhibitors (TKIs) might be thought as the “vanguard” of molecularly targeted therapy for human neoplasias. Imatinib and the successive generations of inhibitors of Bcr-Abl1 kinase, represent the major successful examples of TKI use in cancer treatment. Other tyrosine kinases have been selected as targets of therapy, but the efficacy of their inhibition, although evident, is less definite. Two major negative effects exist in this therapeutic strategy and are linked to the specificity of the drugs and to the role of the targeted kinase in non-malignant cells. In this review, we will discuss the data available on the TKIs effects on the metabolism and functions of mesenchymal stromal cells (MSCs). MSCs are widely distributed in human tissues and play key physiological roles; nevertheless, they might be responsible for important pathologies. At present, bone marrow (BM) MSCs have been studied in greater detail, for both embryological origins and functions. The available data are evocative of an unexpected degree of complexity and heterogeneity of BM-MSCs. It is conceivable that this grade of intricacy occurs also in MSCs of other organs. Therefore, in perspective, the negative effects of TKIs on MSCs might represent a critical problem in long-term cancer therapies based on such inhibitors. PMID:27750212

  10. Neratinib, an irreversible pan-ErbB receptor tyrosine kinase inhibitor: results of a phase II trial in patients with advanced non-small-cell lung cancer.

    Science.gov (United States)

    Sequist, Lecia V; Besse, Benjamin; Lynch, Thomas J; Miller, Vincent A; Wong, Kwok K; Gitlitz, Barbara; Eaton, Keith; Zacharchuk, Charles; Freyman, Amy; Powell, Christine; Ananthakrishnan, Revathi; Quinn, Susan; Soria, Jean-Charles

    2010-06-20

    Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have had a significant impact on non-small-cell lung cancer (NSCLC) outcomes, particularly for patients with EGFR mutations. Resistance emerges after 9 to 12 months, primarily mediated by the T790M resistance mutation. We studied neratinib, an irreversible pan-ErbB TKI that may overcome T790M. Patients with advanced NSCLC underwent EGFR sequencing of available tumor tissue at enrollment. Those with > or = 12 weeks of prior TKI therapy were placed in arm A if they were EGFR mutation positive or arm B if they were wild-type. Arm C included TKI-naïve patients with adenocarcinoma and light smoking histories (neratinib, initially at 320 mg but subsequently reduced to 240 mg because of excessive diarrhea. The primary end point was objective response rate (RR). One-hundred sixty-seven patients were treated: 91 in arm A, 48 in arm B, and 28 in arm C. Diarrhea was the most common toxicity; grade 3 incidence was 50% at 320 mg but improved to 25% after dose reduction. The RR was 3% in arm A and zero in arms B and C. No patients with known T790M responded. Notably, three of four patients with an exon 18 G719X EGFR mutation had a partial response and the fourth had stable disease lasting 40 weeks. Neratinib had low activity in patients with prior benefit from TKIs and in TKI-naïve patients, potentially because of insufficient bioavailability from diarrhea-imposed dose limitation. Responses were seen in patients with the rare G719X EGFR mutation, highlighting the importance of obtaining comprehensive genetic information on trials of targeted agents.

  11. The Effect of Gene Alterations and Tyrosine Kinase Inhibition on Survival and Cause of Death in Patients With Adenocarcinoma of the Lung and Brain Metastases

    Energy Technology Data Exchange (ETDEWEB)

    Sperduto, Paul W., E-mail: psperduto@mropa.com [Minneapolis Radiation Oncology and University of Minnesota Gamma Knife Center, Minneapolis, Minnesota (United States); Yang, T. Jonathan; Beal, Kathryn [Sloan Kettering Cancer Center, New York, New York (United States); Pan, Hubert; Brown, Paul D. [MD Anderson Cancer Center, Houston, Texas (United States); Bangdiwala, Ananta; Shanley, Ryan [University of Minnesota, Masonic Cancer Center, Biostatistics, Minneapolis, Minnesota (United States); Yeh, Norman; Gaspar, Laurie E. [University of Colorado–Denver, Denver, Colorado (United States); Braunstein, Steve; Sneed, Penny [University of California–San Francisco, San Francisco, California (United States); Boyle, John; Kirkpatrick, John P. [Duke University, Durham, North Carolina (United States); Mak, Kimberley S.; Shih, Helen A. [Massachusetts General Hospital, Boston, Massachusetts (United States); Engelman, Alex [University of Maryland, Baltimore, Maryland (United States); Roberge, David [CHUM, University of Montreal, Montreal, Quebec (Canada); Arvold, Nils D.; Alexander, Brian; Awad, Mark M. [Dana Farber/Brigham and Women' s Cancer Center, Boston, Massachusetts (United States); and others

    2016-10-01

    Purpose: Lung cancer remains the most common cause of both cancer mortality and brain metastases (BM). The purpose of this study was to assess the effect of gene alterations and tyrosine kinase inhibition (TKI) on median survival (MS) and cause of death (CoD) in patients with BM from lung adenocarcinoma (L-adeno). Methods: A multi-institutional retrospective database of patients with L-adeno and newly diagnosed BM between 2006 and 2014 was created. Demographics, gene alterations, treatment, MS, and CoD were analyzed. The treatment patterns and outcomes were compared with those in prior trials. Results: Of 1521 L-adeno patients, 816 (54%) had known alteration status. The gene alteration rates were 29%, 10%, and 26% for EGFR, ALK, and KRAS, respectively. The time from primary diagnosis to BM for EGFR−/+ was 10/15 months (P=.02) and for ALK−/+ was 10/20 months (P<.01), respectively. The MS for the group overall (n=1521) was 15 months. The MS from first treatment for BM for EGFR and ALK−, EGFR+, ALK+ were 14, 23 (P<.01), and 45 (P<.0001) months, respectively. The MS after BM for EGFR+ patients who did/did not receive TKI before BM was 17/30 months (P<.01), respectively, but the risk of death was not statistically different between TKI-naïve patients who did/did not receive TKI after the diagnosis of BM (EGFR/ALK hazard ratios: 1.06 [P=.84]/1.60 [P=.45], respectively). The CoD was nonneurologic in 82% of patients with known CoD. Conclusion: EGFR and ALK gene alterations are associated with delayed onset of BM and longer MS relative to patients without these alterations. The CoD was overwhelmingly nonneurologic in patients with known CoD.

  12. A novel spleen tyrosine kinase inhibitor blocks c-Jun N-terminal kinase-mediated gene expression in synoviocytes

    NARCIS (Netherlands)

    Cha, Hoon-Suk; Boyle, David L.; Inoue, Tomoyuki; Schoot, Reineke; Tak, Paul P.; Pine, Polly; Firestein, Gary S.

    2006-01-01

    Spleen tyrosine kinase (Syk) is a key regulator of cell signaling induced by cytokines or Fc receptor engagement. However, the role of Syk in rheumatoid arthritis (RA) is not known yet. We investigated the pathways activated by Syk in tumor necrosis factor-alpha (TNFalpha)-stimulated fibroblast-like

  13. Heterogeneous resistance mechanisms in an EGFR exon 19-mutated non-small cell lung cancer patient treated with erlotinib

    DEFF Research Database (Denmark)

    Santoni-Rugiu, Eric; Grauslund, Morten; Melchior, Linea C.

    2017-01-01

    Patients with epidermal growth factor receptor (EGFR) gene-mutated non-small cell lung cancer (NSCLC) obtain substantial clinical benefit from EGFR tyrosine-kinase inhibitors (TKIs), but will ultimately develop TKI-resistance resulting in median progression-free survival of 9–15 months during first......-line TKI-therapy. However, type and timing of TKI-resistance cannot be predicted and several mechanisms may simultaneously/subsequently occur during TKI-treatment. In this respect, we present a 49 year-old Caucasian male ex-smoker with metastatic pulmonary adenocarcinoma (ADC) that concomitantly harbored...... for SCLC combined with erlotinib continuation was implemented obtaining significant objective response. However, after completing 6 cycles of this combination, new pulmonary and hepatic metastases appeared and showed persistence of the original EGFR- and FGFR3-mutated ADC phenotype together...

  14. Wild-type EGFR Is Stabilized by Direct Interaction with HSP90 in Cancer Cells and Tumors

    Directory of Open Access Journals (Sweden)

    Aarif Ahsan

    2012-08-01

    Full Text Available The epidermal growth factor receptor (EGFR has been targeted for inhibition using tyrosine kinase inhibitors and monoclonal antibodies, with improvement in outcome in subsets of patients with head and neck, lung, and colorectal carcinomas. We have previously found that EGFR stability plays a key role in cell survival after chemotherapy and radiotherapy. Heat shock protein 90 (HSP90 is known to stabilize mutant EGFR and ErbB2, but its role in cancers with wild-type (WT WT-EGFR is unclear. In this report, we demonstrate that fully mature, membrane-bound WT-EGFR interacts with HSP90 independent of ErbB2. Further, the HSP90 inhibitors geldanamycin (GA and AT13387 cause a decrease in WT-EGFR in cultured head and neck cancer cells. This decrease results from a significantly reduced half-life of WT-EGFR. WT-EGFR was also lost in head and neck xenograft specimens after treatment with AT13387 under conditions that inhibited tumor growth and prolonged survival of the mice. Our findings demonstrate that WT-EGFR is a client protein of HSP90 and that their interaction is critical for maintaining both the stability of the receptor as well as the growth of EGFR-dependent cancers. Furthermore, these findings support the search for specific agents that disrupt HSP90's ability to act as an EGFR chaperone.

  15. Phosphotyrosine phosphatase and tyrosine kinase inhibition modulate airway pressure-induced lung injury.

    Science.gov (United States)

    Parker, J C; Ivey, C L; Tucker, A

    1998-11-01

    We determined whether drugs which modulate the state of protein tyrosine phosphorylation could alter the threshold for high airway pressure-induced microvascular injury in isolated perfused rat lungs. Lungs were ventilated for successive 30-min periods with peak inflation pressures (PIP) of 7, 20, 30, and 35 cmH2O followed by measurement of the capillary filtration coefficient (Kfc), a sensitive index of hydraulic conductance. In untreated control lungs, Kfc increased by 1.3- and 3.3-fold relative to baseline (7 cmH2O PIP) after ventilation with 30 and 35 cmH2O PIP. However, in lungs treated with 100 microM phenylarsine oxide (a phosphotyrosine phosphatase inhibitor), Kfc increased by 4.7- and 16.4-fold relative to baseline at these PIP values. In lungs treated with 50 microM genistein (a tyrosine kinase inhibitor), Kfc increased significantly only at 35 cmH2O PIP, and the three groups were significantly different from each other. Thus phosphotyrosine phosphatase inhibition increased the susceptibility of rat lungs to high-PIP injury, and tyrosine kinase inhibition attenuated the injury relative to the high-PIP control lungs.

  16. Combination of EGFR/HER2 tyrosine kinase inhibition by BIBW 2992 and BIBW 2669 with irradiation in FaDu human squamous cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Schuetze, C.; Doerfler, A.; Zips, D.; Krause, M. [Univ. of Technology, Dresden (Germany). Dept. of Radiation Oncology; Eicheler, W.; Baumann, M. [Univ. of Technology, Dresden (Germany). Dept. of Radiation Oncology; Univ. of Technology, Dresden (Germany). Experimental Center; Hering, S. [Univ. of Technology, Dresden (Germany). Inst. of Legal Medicine; Solca, F. [Boehringer Ingelheim Austria, Vienna (Austria)

    2007-05-15

    Purpose: To investigate the effect of the dual EGFR/HER2 (ErbB2) tyrosine kinase inhibitors BIBW 2992 and BIBW 2669 on proliferation and clonogenic cell survival of FaDu human squamous cell carcinoma in vitro, and on tumor growth after single-dose irradiation in nude mice. Material and Methods: Cell proliferation, cell-cycle distribution and clonogenic cell survival after irradiation were assayed with and without BIBW 2992 or BIBW 2669 (3, 30, and 300 nM) in vitro. Tumor volume and tumor growth delay (GD{sub V2}) were determined in tumors growing in NMRI (nu/nu) nude mice, treated with (a) BIBW 2992 (20 mg kg{sup -1} body weight orally), BIBW 2669 (3-4 mg kg{sup -1} body weight orally) or carrier until a final tumor diameter of 15 mm, or, (b) 3 days before a 20-Gy single-dose irradiation or, (c) after a 20-Gy single-dose irradiation until reaching the final tumor diameter. Results: BIBW 2992 and BIBW 2669 significantly increased the doubling time of FaDu cells in vitro. A marked dose-dependent antiproliferative effect with blockade of the cells in G0/G1-phase of the cell cycle was found. Incubation with BIBW 2669 or BIBW 2992 for 3 days marginally increased radiosensitivity of FaDu cells in vitro. For BIBW 2992, this effect was statistically significant (p = 0.006). Daily oral application of BIBW 2669 or BIBW 2992 in mice bearing unirradiated FaDu tumors showed a marked antiproliferative effect with a significant prolongation of tumor growth delay (p < 0.0001). After drug application for 3 days, followed by 20-Gy single-dose irradiation, a slight effect of both drugs on tumor growth delay was seen. For BIBW 2669, this effect was statistically significant (p = 0.007). However, this effect disappeared when tumor volumes were normalized to the time point of irradiation suggesting that both drugs showed no or only a slight radiosensitizing effect in vivo. Daily application of BIBW 2669 or BIBW 2992 after a single-dose irradiation showed a clear inhibition of tumor

  17. Design, Synthesis and Biological Evaluation of Benzohydrazide Derivatives Containing Dihydropyrazoles as Potential EGFR Kinase Inhibitors

    Directory of Open Access Journals (Sweden)

    Hai-Chao Wang

    2016-08-01

    Full Text Available A series of novel benzohydrazide derivatives containing dihydropyrazoles have been synthesized as potential epidermal growth factor receptor (EGFR kinase inhibitors and their biological activities as potential antiproliferative agents have been evaluated. Among these compounds, compound H20 exhibited the most potent antiproliferative activity against four cancer cell line variants (A549, MCF-7, HeLa, HepG2 with IC50 values of 0.46, 0.29, 0.15 and 0.21 μM respectively, which showed the most potent EGFR inhibition activities (IC50 = 0.08 μM for EGFR. Molecular modeling simulation studies were performed in order to predict the biological activity and activity relationship (SAR of these benzohydrazide derivatives. These results suggested that compound H20 may be a promising anticancer agent.

  18. Radiotherapy for asymptomatic brain metastasis in epidermal growth factor receptor mutant non-small cell lung cancer without prior tyrosine kinase inhibitors treatment: a retrospective clinical study

    International Nuclear Information System (INIS)

    Liu, SongRan; Qiu, Bo; Chen, LiKun; Wang, Fang; Liang, Ying; Cai, PeiQiang; Zhang, Li; Chen, ZhaoLin; Liu, ShiLiang; Liu, MengZhong; Liu, Hui

    2015-01-01

    Non-small cell lung cancer (NSCLC) with brain metastasis (BM) harboring an epidermal growth factor receptor (EGFR) mutation shows good response to tyrosine kinase inhibitors (TKIs). This study is to assess the appropriate timing of brain radiotherapy (RT) for asymptomatic BM in EGFR mutant NSCLC patients. There were 628 patients diagnosed with EGFR mutant NSCLC between October 2005 and December 2011. Treatment outcomes had been retrospectively evaluated in 96 patients with asymptomatic BM without prior TKI treatment. 39 patients received first-line brain RT, 23 patients received delayed brain RT, and 34 patients did not receive brain RT. With a median follow-up of 26 months, the 2-year OS was 40.6 %. Univariate analyses revealed that ECOG performance status (p = 0.006), other distant metastases (p = 0.002) and first line systemic treatment (p = 0.032) were significantly associated with overall survival (OS). Multivariate analyses revealed that other sites of distant metastases (p = 0.030) were prognostic factor. The timing of brain RT was not significantly related to OS (p = 0.246). The 2-year BM progression-free survival (PFS) was 26.9 %. Brain RT as first-line therapy failed to demonstrate a significant association with BM PFS (p = 0.643). First-line brain RT failed to improve long-term survival in TKI-naïve EGFR mutant NSCLC patients with asymptomatic BM. Prospective studies are needed to validate these clinical findings

  19. Review of current classification, molecular alterations, and tyrosine kinase inhibitor therapies in myeloproliferative disorders with hypereosinophilia

    Directory of Open Access Journals (Sweden)

    Havelange V

    2013-08-01

    Full Text Available Violaine Havelange,1,2 Jean-Baptiste Demoulin1 1de Duve Institute, Université catholique de Louvain, Brussels, Belgium; 2Department of Hematology, Cliniques universitaires Saint-Luc, Université catholique de Louvain, Brussels, Belgium Abstract: Recent advances in our understanding of the molecular mechanisms underlying hypereosinophilia have led to the development of a 'molecular' classification of myeloproliferative disorders with eosinophilia. The revised 2008 World Health Organization classification of myeloid neoplasms included a new category called “myeloid and lymphoid neoplasms with eosinophilia and abnormalities of PDGFRA, PDGFRB or FGFR1.” Despite the molecular heterogeneity of PDGFR (platelet-derived growth factor receptor rearrangements, tyrosine kinase inhibitors at low dose induce rapid and complete hematological remission in the majority of these patients. Other kinase inhibitors are promising. Further discoveries of new molecular alterations will direct the development of new specific inhibitors. In this review, an update of the classifications of myeloproliferative disorders associated with hypereosinophilia is discussed together with open and controversial questions. Molecular mechanisms and promising results of tyrosine kinase inhibitor treatments are reviewed. Keywords: hypereosinophilia, classification, myeloproliferative disorders, molecular alterations, tyrosine kinase inhibitor

  20. Tyrosine kinase chromosomal translocations mediate distinct and overlapping gene regulation events

    International Nuclear Information System (INIS)

    Kim, Hani; Gillis, Lisa C; Jarvis, Jordan D; Yang, Stuart; Huang, Kai; Der, Sandy; Barber, Dwayne L

    2011-01-01

    Leukemia is a heterogeneous disease commonly associated with recurrent chromosomal translocations that involve tyrosine kinases including BCR-ABL, TEL-PDGFRB and TEL-JAK2. Most studies on the activated tyrosine kinases have focused on proximal signaling events, but little is known about gene transcription regulated by these fusions. Oligonucleotide microarray was performed to compare mRNA changes attributable to BCR-ABL, TEL-PDGFRB and TEL-JAK2 after 1 week of activation of each fusion in Ba/F3 cell lines. Imatinib was used to control the activation of BCR-ABL and TEL-PDGFRB, and TEL-JAK2-mediated gene expression was examined 1 week after Ba/F3-TEL-JAK2 cells were switched to factor-independent conditions. Microarray analysis revealed between 800 to 2000 genes induced or suppressed by two-fold or greater by each tyrosine kinase, with a subset of these genes commonly induced or suppressed among the three fusions. Validation by Quantitative PCR confirmed that eight genes (Dok2, Mrvi1, Isg20, Id1, gp49b, Cxcl10, Scinderin, and collagen Vα1(Col5a1)) displayed an overlapping regulation among the three tested fusion proteins. Stat1 and Gbp1 were induced uniquely by TEL-PDGFRB. Our results suggest that BCR-ABL, TEL-PDGFRB and TEL-JAK2 regulate distinct and overlapping gene transcription profiles. Many of the genes identified are known to be involved in processes associated with leukemogenesis, including cell migration, proliferation and differentiation. This study offers the basis for further work that could lead to an understanding of the specificity of diseases caused by these three chromosomal translocations

  1. An SH2 domain-based tyrosine kinase assay using biotin ligase modified with a terbium(III) complex.

    Science.gov (United States)

    Sueda, Shinji; Shinboku, Yuki; Kusaba, Takeshi

    2013-01-01

    Src homology 2 (SH2) domains are modules of approximately 100 amino acids and are known to bind phosphotyrosine-containing sequences with high affinity and specificity. In the present work, we developed an SH2 domain-based assay for Src tyrosine kinase using a unique biotinylation reaction from archaeon Sulfolobus tokodaii. S. tokodaii biotinylation has a unique property that biotin protein ligase (BPL) forms a stable complex with its biotinylated substrate protein (BCCP). Here, an SH2 domain from lymphocyte-specific tyrosine kinase was genetically fused to a truncated BCCP, and the resulting fusion protein was labeled through biotinylation with BPL carrying multiple copies of a luminescent Tb(3+) complex. The labeled SH2 fusion proteins were employed to detect a phosphorylated peptide immobilized on the surface of the microtiter plate, where the phosphorylated peptide was produced by phosphorylation to the substrate peptide by Src tyrosine kinase. Our assay allows for a reliable determination of the activity of Src kinase lower than 10 pg/μL by a simple procedure.

  2. Long-term treatment with EGFR inhibitor erlotinib attenuates renal inflammatory cytokines but not nephropathy in Alport syndrome mouse model.

    Science.gov (United States)

    Omachi, Kohei; Miyakita, Rui; Fukuda, Ryosuke; Kai, Yukari; Suico, Mary Ann; Yokota, Tsubasa; Kamura, Misato; Shuto, Tsuyoshi; Kai, Hirofumi

    2017-12-01

    Alport syndrome (AS) is a hereditary kidney disease caused by mutation of type IV collagen. Loss of collagen network induces collapse of glomerular basement membrane (GBM) structure. The previous studies showed that upregulation of some tyrosine kinase receptors signaling accompanied GBM disorder in AS mouse model. EGFR signaling is one of the well-known receptor kinase signaling that is involved in glomerular diseases. However, whether EGFR signaling is relevant to AS progression is still uninvestigated. Here, we determined the involvement of EGFR in AS and the effect of suppressing EGFR signaling by erlotinib treatment on AS progression. Phosphorylated EGFR expression was investigated by Western blotting analysis and immunostaining of kidney tissues of Col4a5 mutant mice (a mouse model of X-linked AS). To check the effect of blocking EGFR signaling in AS, we administered erlotinib to AS mice once a day (10 mg/kg/day) orally for 18 weeks. Renal function parameters (proteinuria, serum creatinine, and BUN) and renal histology were assessed, and the gene expressions of inflammatory cytokines were analyzed in renal tissues. Phosphorylated EGFR expression was upregulated in AS mice kidney tissues. Erlotinib slightly reduced the urinary protein and suppressed the expression of renal injury markers (Lcn2, Lysozyme) and inflammatory cytokines (Il-6, Il-1β and KC). Erlotinib did not improve renal pathology, such as glomerular sclerosis and fibrosis. These findings suggest that EGFR signaling is upregulated in kidney, but although inhibiting this signaling pathway suppressed renal inflammatory cytokines, it did not ameliorate renal dysfunction in AS mouse model.

  3. Development of freeze-dried kit for direct 99mTc -labeling of nimotuzumab to diagnose human EGFR positive tumors

    International Nuclear Information System (INIS)

    Toledo, Darien; Figueiras, Jenneby; Rojas, Gertrudis; León, Kalet; Gongora Bravo, Magdiel; Miguel Martínez, Antonio; Michel Alonso, Luis; Hernández, Ignacio; León, Mariela; Leyva, René; Hernández, Gerardo Ramses

    2016-01-01

    Epidermal growth factor receptor (EGFR), a 170 kDa transmembrane tyrosine kinase receptor which specifically binds epidermal growth factor (EGF) and transforming growth factor-a (TGFa) that are crucial in signaling cell proliferation, differentiation, and survival1. Overexpression of EGFR has been observed in breast cancer2, colorectal cancer, ovarian cancer, squamous cell lung carcinoma3 head and neck cancer4 and bladder cancer. Molecular imaging using radiopharmaceuticals directed towards EGFR could characterize the receptor status of tumors and thereby predict response to anti-EGFR agents for the treatment of cancer. Nimotuzumab is a humanized anti-EGFR monoclonal antibody5 designed to reduce immunogenicity and rate of clearance from the body6. Radiolabelled formulations of nimotuzumab would have applications for non-invasive imaging in order to characterize EGFR-positive tumors and thus to select patient populations that could benefit from therapy7, 8. The studies described in this work were designed to develop and evaluate the in vitro and in vivo the properties of a radiolabeled freeze dried kit of nimotuzumab and to determine its potential for radio immunodiagnostic applications. (author)

  4. TYROSINE KINASE INHIBITORS AND PREGNANCY

    Directory of Open Access Journals (Sweden)

    Elisabetta Abruzzese

    2014-04-01

    Full Text Available The management of patients with chronic myeloid leukemia (CML during pregnancy has became recently a matter of continuous debate.  The introduction of the Tyrosine Kinase Inhibitors (TKIs in clinical practice has dramatically changed the prognosis of CML patients.  Patients diagnosed in chronic phase can reasonably expect many years of excellent disease control and good quality of life, as well as a normal life expectancy.  This fact has come the necessity to address issues relating to fertility and pregnancy. Physicians are not infrequently being asked for advice regarding the need for, and or the appropriateness of, stopping treatment in order to conceive. In this report we will review the data published in terms of fertility, conception, pregnancy, pregnancy outcome and illness control for all the approved TKIs, as well as suggest how to manage a planned and/or unplanned pregnancy.

  5. Phosphorylated EGFR expression may predict outcome of EGFR-TKIs therapy for the advanced NSCLC patients with wild-type EGFR

    Directory of Open Access Journals (Sweden)

    Wang Fen

    2012-08-01

    Full Text Available Abstract Background EGFR mutation is a strong predictive factor of EGFR-TKIs therapy. However, at least 10% of patients with EGFR wild-type are responsive to TKIs, suggesting that other determinants of outcome besides EGFR mutation might exist. We hypothesized that activation of phosphorylated EGFR could be a potential predictive biomarker to EGFR-TKIs treatment among patients in wild-type EGFR. Method Total of 205 stage IIIb and IV NSCLC patients, tissue samples of whom were available for molecular analysis, were enrolled in this study. The phosphorylation of EGFR at tyrosine 1068 (pTyr1068 and 1173 (pTyr1173 were assessed by immunohistochemistry, and EGFR mutations were detected by denaturing high performance liquid chromatograph (DHPLC. Results Among 205 patients assessable for EGFR mutation and phosphorylation analysis, 92 (44.9% were EGFR mutant and 165 patients (57.6% had pTyr1173 expression. Superior progression-free survival (PFS was seen after EGFR-TKIs therapy in patients with pTyr1068 expression compared to pTyr1068 negative ones (median PFS 7.0 months vs. 1.2 months, P P = 0.016. In subgroup of patients with wild-type EGFR, pTyr1068 expression positive ones had a significantly prolonged PFS (4.2 months vs.1.2 months P  Conclusion pTyr1068 may be a predictive biomarker for screening the population for clinical response to EGFR-TKIs treatment; especially for patients with wild-type EGFR.

  6. Functional and structural stability of the epidermal growth factor receptor in detergent micelles and phospholipid nanodiscs

    DEFF Research Database (Denmark)

    Mi, Li-Zhi; Grey, Michael J; Nishida, Noritaka

    2008-01-01

    Cellular signaling mediated by the epidermal growth factor receptor (EGFR or ErbB) family of receptor tyrosine kinases plays an important role in regulating normal and oncogenic cellular physiology. While structures of isolated EGFR extracellular domains and intracellular protein tyrosine kinase...... differential functional stability in Triton X-100 versus dodecyl maltoside. Furthermore, the kinase activity can be significantly stabilized by reconstituting purified EGF-bound EGFR dimers in phospholipid nanodiscs or vesicles, suggesting that the environment around the hydrophobic transmembrane...

  7. Identifying activating mutations in the EGFR gene: prognostic and therapeutic implications in non-small cell lung cancer.

    Science.gov (United States)

    Lopes, Gabriel Lima; Vattimo, Edoardo Filippo de Queiroz; Castro Junior, Gilberto de

    2015-01-01

    Lung cancer is the leading cause of cancer-related deaths worldwide. Promising new therapies have recently emerged from the development of molecular targeted drugs; particularly promising are those blocking the signal transduction machinery of cancer cells. One of the most widely studied cell signaling pathways is that of EGFR, which leads to uncontrolled cell proliferation, increased cell angiogenesis, and greater cell invasiveness. Activating mutations in the EGFR gene (deletions in exon 19 and mutation L858R in exon 21), first described in 2004, have been detected in approximately 10% of all non-squamous non-small cell lung cancer (NSCLC) patients in Western countries and are the most important predictors of a response to EGFR tyrosine-kinase inhibitors (EGFR-TKIs). Studies of the EGFR-TKIs gefitinib, erlotinib, and afatinib, in comparison with platinum-based regimens, as first-line treatments in chemotherapy-naïve patients have shown that the EGFR-TKIs produce gains in progression-free survival and overall response rates, although only in patients whose tumors harbor activating mutations in the EGFR gene. Clinical trials have also shown EGFR-TKIs to be effective as second- and third-line therapies in advanced NSCLC. Here, we review the main aspects of EGFR pathway activation in NSCLC, underscore the importance of correctly identifying activating mutations in the EGFR gene, and discuss the main outcomes of EGFR-TKI treatment in NSCLC.

  8. Identifying activating mutations in the EGFR gene: prognostic and therapeutic implications in non-small cell lung cancer *

    Science.gov (United States)

    Lopes, Gabriel Lima; Vattimo, Edoardo Filippo de Queiroz; de Castro, Gilberto

    2015-01-01

    Abstract Lung cancer is the leading cause of cancer-related deaths worldwide. Promising new therapies have recently emerged from the development of molecular targeted drugs; particularly promising are those blocking the signal transduction machinery of cancer cells. One of the most widely studied cell signaling pathways is that of EGFR, which leads to uncontrolled cell proliferation, increased cell angiogenesis, and greater cell invasiveness. Activating mutations in the EGFR gene (deletions in exon 19 and mutation L858R in exon 21), first described in 2004, have been detected in approximately 10% of all non-squamous non-small cell lung cancer (NSCLC) patients in Western countries and are the most important predictors of a response to EGFR tyrosine-kinase inhibitors (EGFR-TKIs). Studies of the EGFR-TKIs gefitinib, erlotinib, and afatinib, in comparison with platinum-based regimens, as first-line treatments in chemotherapy-naïve patients have shown that the EGFR-TKIs produce gains in progression-free survival and overall response rates, although only in patients whose tumors harbor activating mutations in the EGFR gene. Clinical trials have also shown EGFR-TKIs to be effective as second- and third-line therapies in advanced NSCLC. Here, we review the main aspects of EGFR pathway activation in NSCLC, underscore the importance of correctly identifying activating mutations in the EGFR gene, and discuss the main outcomes of EGFR-TKI treatment in NSCLC. PMID:26398757

  9. A protein-tyrosine phosphatase with sequence similarity to the SH2 domain of the protein-tyrosine kinases.

    Science.gov (United States)

    Shen, S H; Bastien, L; Posner, B I; Chrétien, P

    1991-08-22

    The phosphorylation of proteins at tyrosine residues is critical in cellular signal transduction, neoplastic transformation and control of the mitotic cycle. These mechanisms are regulated by the activities of both protein-tyrosine kinases (PTKs) and protein-tyrosine phosphatases (PTPases). As in the PTKs, there are two classes of PTPases: membrane associated, receptor-like enzymes and soluble proteins. Here we report the isolation of a complementary DNA clone encoding a new form of soluble PTPase, PTP1C. The enzyme possesses a large noncatalytic region at the N terminus which unexpectedly contains two adjacent copies of the Src homology region 2 (the SH2 domain) found in various nonreceptor PTKs and other cytoplasmic signalling proteins. As with other SH2 sequences, the SH2 domains of PTP1C formed high-affinity complexes with the activated epidermal growth factor receptor and other phosphotyrosine-containing proteins. These results suggest that the SH2 regions in PTP1C may interact with other cellular components to modulate its own phosphatase activity against interacting substrates. PTPase activity may thus directly link growth factor receptors and other signalling proteins through protein-tyrosine phosphorylation.

  10. Sequential treatment of icotinib after first-line pemetrexed in advanced lung adenocarcinoma with unknown EGFR gene status.

    Science.gov (United States)

    Zheng, Yulong; Fang, Weijia; Deng, Jing; Zhao, Peng; Xu, Nong; Zhou, Jianying

    2014-07-01

    In non-small cell lung cancer (NSCLC), the well-developed epidermal growth factor receptor (EGFR) is an important therapeutic target. EGFR activating gene mutations have been proved strongly predictive of response to EGFR-tyrosine kinase inhibitors (TKI) in NSCLC. However, both in daily clinical practice and clinical trials, patients with unknown EGFR gene status (UN-EGFR-GS) are very common. In this study, we assessed efficacy and tolerability of sequential treatment of first-line pemetrexed followed by icotinib in Chinese advanced lung adenocarcinoma with UN-EGFR-GS. We analyzed 38 patients with advanced lung adenocarcinoma with UN-EGFR-GS treated with first-line pemetrexed-based chemotherapy followed by icotinib as maintenance or second-line therapy. The response rates to pemetrexed and icotinib were 21.1% and 42.1%, respectively. The median overall survival was 27.0 months (95% CI, 19.7-34.2 months). The 12-month overall survival probability was 68.4%. The most common toxicities observed in icotinib phase were rashes, diarrheas, and elevated aminotransferase. Subgroup analysis indicated that the overall survival is correlated with response to icotinib. The sequence of first-line pemetrexed-based chemotherapy followed by icotinib treatment is a promising option for advanced lung adenocarcinoma with UN-EGFR-GS in China.

  11. Ret receptor tyrosine kinase sustains proliferation and tissue maturation in intestinal epithelia

    DEFF Research Database (Denmark)

    Perea, Daniel; Guiu, Jordi; Hudry, Bruno

    2017-01-01

    Expression of the Ret receptor tyrosine kinase is a defining feature of enteric neurons. Its importance is underscored by the effects of its mutation in Hirschsprung disease, leading to absence of gut innervation and severe gastrointestinal symptoms. We report a new and physiologically significant...

  12. Metformin as a prevention and treatment for preeclampsia: effects on soluble fms-like tyrosine kinase 1 and soluble endoglin secretion and endothelial dysfunction.

    Science.gov (United States)

    Brownfoot, Fiona C; Hastie, Roxanne; Hannan, Natalie J; Cannon, Ping; Tuohey, Laura; Parry, Laura J; Senadheera, Sevvandi; Illanes, Sebastian E; Kaitu'u-Lino, Tu'uhevaha J; Tong, Stephen

    2016-03-01

    Preeclampsia is associated with placental ischemia/hypoxia and secretion of soluble fms-like tyrosine kinase 1 and soluble endoglin into the maternal circulation. This causes widespread endothelial dysfunction that manifests clinically as hypertension and multisystem organ injury. Recently, small molecule inhibitors of hypoxic inducible factor 1α have been found to reduce soluble fms-like tyrosine kinase 1 and soluble endoglin secretion. However, their safety profile in pregnancy is unknown. Metformin is safe in pregnancy and is also reported to inhibit hypoxic inducible factor 1α by reducing mitochondrial electron transport chain activity. The purposes of this study were to determine (1) the effects of metformin on placental soluble fms-like tyrosine kinase 1 and soluble endoglin secretion, (2) to investigate whether the effects of metformin on soluble fms-like tyrosine kinase 1 and soluble endoglin secretion are regulated through the mitochondrial electron transport chain, and (3) to examine its effects on endothelial dysfunction, maternal blood vessel vasodilation, and angiogenesis. We performed functional (in vitro and ex vivo) experiments using primary human tissues to examine the effects of metformin on soluble fms-like tyrosine kinase 1 and soluble endoglin secretion from placenta, endothelial cells, and placental villous explants. We used succinate, mitochondrial complex II substrate, to examine whether the effects of metformin on soluble fms-like tyrosine kinase 1 and soluble endoglin secretion were mediated through the mitochondria. We also isolated mitochondria from preterm preeclamptic placentas and gestationally matched control subjects and measured mitochondrial electron transport chain activity using kinetic spectrophotometric assays. Endothelial cells or whole maternal vessels were incubated with metformin to determine whether it rescued endothelial dysfunction induced by either tumor necrosis factor-α (to endothelial cells) or placenta villous

  13. Physical and functional interactions between SH2 and SH3 domains of the Src family protein tyrosine kinase p59fyn

    NARCIS (Netherlands)

    Panchamoorthy, G.; Fukazawa, T.; Stolz, L.; Payne, G.; Reedquist, K.; Shoelson, S.; Songyang, Z.; Cantley, L.; Walsh, C.; Band, H.

    1994-01-01

    The Src family protein tyrosine kinases participate in signalling through cell surface receptors that lack intrinsic tyrosine kinase domains. All nine members of this family possess adjacent Src homology (SH2 and SH3) domains, both of which are essential for repression of the enzymatic activity. The

  14. Tyrosine Kinase Inhibitors Induced Thyroid Dysfunction: A Review of Its Incidence, Pathophysiology, Clinical Relevance, and Treatment

    Directory of Open Access Journals (Sweden)

    Hala Ahmadieh

    2013-01-01

    Full Text Available Tyrosine kinase inhibitors (TKI belong to a new class of molecular multitargeted anticancer therapy which targets different growth factor receptors and hence attenuates cancer cell survival and growth. Since their introduction as adjunct treatment for renal cell carcinoma and gastrointestinal stromal tumors (GIST, a number of reports have demonstrated that TKI can induce thyroid dysfunction which was especially more common with sunitinib maleate. Many mechanisms with respect to this adverse effect of tyrosine kinase inhibitors have been proposed including their induction of thyroiditis, capillary regression in the thyroid gland, antithyroid peroxidase antibody production, and their ability to decrease iodine uptake by the thyroid gland. Of interest is the observation that TKI-induced thyroid dysfunction may actually be protective as it was shown to improve overall survival, and it was suggested that it may have a prognostic value. Followup on thyroid function tests while patients are maintained on tyrosine kinase inhibitor is strongly recommended. When thyroid dysfunction occurs, appropriate treatment should be individualized depending on patients symptoms and thyroid stimulating hormone level.

  15. Is There a Survival Benefit of First-Line Epidermal Growth Factor Receptor Tyrosine-Kinase Inhibitor Monotherapy Versus Chemotherapy in Patients with Advanced Non-Small-Cell Lung Cancer?: A Meta-Analysis.

    Science.gov (United States)

    Guetz, Gaetan Des; Landre, Thierry; Uzzan, Bernard; Chouahnia, Kader; Nicolas, Patrick; Morere, Jean-François

    2016-02-01

    Tyrosine-kinase inhibitors (TKIs) markedly improve progression-free survival (PFS) of patients with advanced non-small-cell lung cancer (NSCLC) mutated for epidermal growth factor receptor (EGFR). Results on overall survival (OS) are less clear-cut. We performed a publication-based meta-analysis to address further this issue. We did a PubMed query using keywords simultaneously (lung neoplasm, tyrosine kinase inhibitors, epidermal growth factor receptor mutation, survival, and randomized controlled trials). We also searched for relevant abstracts in annual proceedings of ASCO, ESMO, and WCLC meetings. We cross-checked all references from all eligible articles. Only phase III randomized controlled trials comparing TKI monotherapy and platinum-based doublet chemotherapy in first-line treatment of metastatic or advanced NSCLC were included. We used EasyMA software to perform statistical analyses. A random effect model was used in case of heterogeneity between studies (and a fixed effect model in absence of heterogeneity). The eight eligible studies included 2962 patients (780 males, 2182 females, mostly Asian, median age 60 years), 2909 adenocarcinomas (98 %), 1739 mutated tumors (897 exon 19 deletion, 699 L858 mutation), 448 stage IIIB, and 2222 stage IV (75 %) tumours and 2453 never smokers (83 %). Four studies assessed gefitinib, two studies assessed erlotinib, and two studies assessed afatinib. Chemotherapies were doublets including a platinum salt. All studies included patients with EGFR mutations, but six studies included only EGFR mutated patients. OS was similar among patients who first received TKI or chemotherapy (HR 0.98, 95 % CI 0.87-1.10, fixed effect model). Conversely, compared with chemotherapy, EGFR TKIs significantly improved PFS in patients with EGFR-mutated tumours (HR 0.37, 95 % CI 0.29-0.49, random effect model). Concerning side effects, rash (RR 6.29, 95 % CI 4.05-9.77), diarrhoea (RR 3.51, 95 % CI 2.15-5.75), stomatitis (RR 3.57, 95 % CI 1

  16. Membrane depolarization-induced RhoA/Rho-associated kinase activation and sustained contraction of rat caudal arterial smooth muscle involves genistein-sensitive tyrosine phosphorylation

    Science.gov (United States)

    Mita, Mitsuo; Tanaka, Hitoshi; Yanagihara, Hayato; Nakagawa, Jun-ichi; Hishinuma, Shigeru; Sutherland, Cindy; Walsh, Michael P.; Shoji, Masaru

    2013-01-01

    Rho-associated kinase (ROK) activation plays an important role in K+-induced contraction of rat caudal arterial smooth muscle (Mita et al., Biochem J. 2002; 364: 431–40). The present study investigated a potential role for tyrosine kinase activity in K+-induced RhoA activation and contraction. The non-selective tyrosine kinase inhibitor genistein, but not the src family tyrosine kinase inhibitor PP2, inhibited K+-induced sustained contraction (IC50 = 11.3 ± 2.4 µM). Genistein (10 µM) inhibited the K+-induced increase in myosin light chain (LC20) phosphorylation without affecting the Ca2+ transient. The tyrosine phosphatase inhibitor vanadate induced contraction that was reversed by genistein (IC50 = 6.5 ± 2.3 µM) and the ROK inhibitor Y-27632 (IC50 = 0.27 ± 0.04 µM). Vanadate also increased LC20 phosphorylation in a genistein- and Y-27632-dependent manner. K+ stimulation induced translocation of RhoA to the membrane, which was inhibited by genistein. Phosphorylation of MYPT1 (myosin-targeting subunit of myosin light chain phosphatase) was significantly increased at Thr855 and Thr697 by K+ stimulation in a genistein- and Y-27632-sensitive manner. Finally, K+ stimulation induced genistein-sensitive tyrosine phosphorylation of proteins of ∼55, 70 and 113 kDa. We conclude that a genistein-sensitive tyrosine kinase, activated by the membrane depolarization-induced increase in [Ca2+]i, is involved in the RhoA/ROK activation and sustained contraction induced by K+. Ca2+ sensitization, myosin light chain phosphatase, RhoA, Rho-associated kinase, tyrosine kinase PMID:24133693

  17. High Level of Soluble FMS-Like Tyrosine Kinase-1 (sFlt-1 Serum in Pregnancy as a Risk Factor of Preeclampsia

    Directory of Open Access Journals (Sweden)

    I Gede Mega Putra

    2016-07-01

    Full Text Available Background: Recently, etiology and pathogenesis of preeclampsia remain unknown. One of the theory indicating that hypoxia and ischemic placenta caused by abnormal cytotrophoblast invasion in preeclampsia. Soluble Fms-like tyrosine kinase-1 (sFlt-1 serum as a laboratory marker of hypoxia condition that contributes to the occurrence of endothelial damage and clinical manifestations in preeclampsia. Objective: This study was aimed at proving that high level of soluble Fms-like tyrosine kinase-1 (sFlt-1 serum in pregnancy as a risk factor for preeclampsia. Methods: This study was a case control. Among 58 pregnant women studied, 29 women with preeclampsia as a case group and 29 women with normal pregnancy as a control group. Soluble Fms-like tyrosine kinase-1 (sFlt-1 serum was analyzed in the Prodia Laboratory. Collected data were tested for normality using Kolmogorov-Smirnov, then analyzed with independent sample test. Chi-Square test used to determine soluble Fms-like tyrosine kinase-1 (sFlt-1 serum level in preeclampsia. Results: This research concluded that the average level of soluble Fms-like tyrosine kinase-1 (sFlt-1 serum in preeclampsia were 11231.00 ± 8390.3 pg/mL and 3981.62 ± 4921.5 pg/mL in normal pregnancy. Analysis of significance with independent t-test concluded that the value of t = 4.01 and p = 0.001. This mean the average levels of soluble Fms-like tyrosine kinase-1 (sFlt-1serum levels in both groups were difference significantly (p <0.05. Based on the cut-off point of sFlt-1 serum levels was 4505.50 pg/mL with 79.3% sensitivity and 82.8% specificity, the relative risk of preeclampsia was 18 times (OR = 18.40, IK 95% = 4.93 to 68.70, p = 0.001. Conclusion:  Based on this research, high levels of soluble Fms-like tyrosine kinase-1 (sFlt-1 in pregnancy was proved as a risk factor for preeclampsia.

  18. Effect of tyrosine kinase blockade on norepinephrine-induced cytosolic calcium response in rat afferent arterioles

    DEFF Research Database (Denmark)

    Salomonsson, Max; Arendshorst, William J

    2004-01-01

    We used genistein (Gen) and tyrphostin 23 (Tyr-23) to evaluate the importance of tyrosine phosphorylation in norepinephrine (NE)-induced changes in intracellular free calcium concentration ([Ca(2+)](i)) in rat afferent arterioles. [Ca(2+)](i) was measured in microdissected arterioles using...... ratiometric photometry of fura 2 fluorescence. The control [Ca(2+)](i) response to NE (1 microM) consisted of a rapid initial peak followed by a plateau phase sustained above baseline. Pretreatment with the tyrosine kinase inhibitor Tyr-23 (50 microM, 10 min) caused a slow 40% increase in baseline [Ca(2+)](i...... of nifedipine and Tyr-23 were not additive. Nifedipine had no inhibitory effect after Tyr-23 pretreatment, indicating Tyr-23 inhibition of Ca(2+) entry. Another tyrosine kinase inhibitor, Gen (5 and 50 microM), did not affect baseline [Ca(2+)](i). High-dose Gen inhibited the peak and plateau response to NE...

  19. The protist, Monosiga brevicollis, has a tyrosine kinase signaling network more elaborate and diverse than found in any known metazoan.

    Science.gov (United States)

    Manning, Gerard; Young, Susan L; Miller, W Todd; Zhai, Yufeng

    2008-07-15

    Tyrosine kinase signaling has long been considered a hallmark of intercellular communication, unique to multicellular animals. Our genomic analysis of the unicellular choanoflagellate Monosiga brevicollis discovers a remarkable count of 128 tyrosine kinases, 38 tyrosine phosphatases, and 123 phosphotyrosine (pTyr)-binding SH2 proteins, all higher counts than seen in any metazoan. This elaborate signaling network shows little orthology to metazoan counterparts yet displays many innovations reminiscent of metazoans. These include extracellular domains structurally related to those of metazoan receptor kinases, alternative methods for membrane anchoring and phosphotyrosine interaction in cytoplasmic kinases, and domain combinations that link kinases to small GTPase signaling and transcription. These proteins also display a wealth of combinations of known signaling domains. This uniquely divergent and elaborate signaling network illuminates the early evolution of pTyr signaling, explores innovative ways to traverse the cellular signaling circuitry, and shows extensive convergent evolution, highlighting pervasive constraints on pTyr signaling.

  20. Protein-tyrosine phosphatase SHP2 contributes to GDNF neurotrophic activity through direct binding to phospho-Tyr687 in the RET receptor tyrosine kinase.

    Science.gov (United States)

    Perrinjaquet, Maurice; Vilar, Marçal; Ibáñez, Carlos F

    2010-10-08

    The signaling mechanisms by which neurotrophic receptors regulate neuronal survival and axonal growth are still incompletely understood. In the receptor tyrosine kinase RET, a receptor for GDNF (glial cell line-derived neurotrophic factor), the functions of the majority of tyrosine residues that become phosphorylated are still unknown. Here we have identified the protein-tyrosine phosphatase SHP2 as a novel direct interactor of RET and the first effector known to bind to phosphorylated Tyr(687) in the juxtamembrane region of the receptor. We show that SHP2 is recruited to RET upon ligand binding in a cooperative fashion, such that both interaction with Tyr(687) and association with components of the Tyr(1062) signaling complex are required for stable recruitment of SHP2 to the receptor. SHP2 recruitment contributes to the ability of RET to activate the PI3K/AKT pathway and promote survival and neurite outgrowth in primary neurons. Furthermore, we find that activation of protein kinase A (PKA) by forskolin reduces the recruitment of SHP2 to RET and negatively affects ligand-mediated neurite outgrowth. In agreement with this, mutation of Ser(696), a known PKA phosphorylation site in RET, enhances SHP2 binding to the receptor and eliminates the effect of forskolin on ligand-induced outgrowth. Together, these findings establish SHP2 as a novel positive regulator of the neurotrophic activities of RET and reveal Tyr(687) as a critical platform for integration of RET and PKA signals. We anticipate that several other phosphotyrosines of unknown function in neuronal receptor tyrosine kinases will also support similar regulatory functions.

  1. How tyrosine kinase inhibitors impair metabolism and endocrine system function: a systematic updated review.

    Science.gov (United States)

    Breccia, Massimo; Molica, Matteo; Alimena, Giuliana

    2014-12-01

    Tyrosine kinase inhibitors (TKIs) advent has deeply changed the outcome of chronic myeloid leukemia (CML) patients, with improved rates of response and overall survival. However, for this success some patients paid the price of a number of peculiar side effects, the so-called off-target side effects, specific for each one TKI. These effects are due to non-selective inhibition of other tyrosine kinase receptors, such as PDGFR, c-KIT, Src, VEGF. Consequences of this inhibition, some metabolic changes during the treatment with TKIs are reported. Aim of present review is to report metabolic changes and potential mechanisms involved in the pathogenesis related to imatinib, second (nilotinib and dasatinib) and third generation (bosutinib and ponatinib) TKIs. Copyright © 2014 Elsevier Ltd. All rights reserved.

  2. Phospho-kinase profile of triple negative breast cancer and androgen receptor signaling

    International Nuclear Information System (INIS)

    Cuenca-López, María D; Montero, Juan C; Morales, Jorge C; Prat, Aleix; Pandiella, Atanasio; Ocana, Alberto

    2014-01-01

    The androgen receptor (AR) plays a central role in the oncogenesis of different tumors, as is the case in prostate cancer. In triple negative breast cancer (TNBC) a gene expression classification has described different subgroups including a luminal androgen subtype. The AR can be controlled by several mechanisms like the activation of membrane tyrosine kinases and downstream signaling pathways. However little is known in TNBC about how the AR is modulated by these mechanisms and the potential therapeutic strategists to inhibit its expression. We used human samples to evaluate the expression of AR by western-blot and phospho-proteomic kinase arrays that recognize membrane tyrosine kinase receptors and downstream mediators. Western-blots in human cell lines were carried out to analyze the expression and activation of individual proteins. Drugs against these kinases in different conditions were used to measure the expression of the androgen receptor. PCR experiments were performed to assess changes in the AR gene after therapeutic modulation of these pathways. AR is present in a subset of TNBC and its expression correlates with activated membrane receptor kinases-EGFR and PDGFRβ in human samples and cell lines. Inhibition of the PI3K/mTOR pathway in TNBC cell lines decreased notably the expression of the AR. Concomitant administration of the anti-androgen bicalutamide with the EGFR, PDGFRβ and Erk1/2 inhibitors, decreased the amount of AR compared to each agent given alone, and had an additive anti-proliferative effect. Administration of dihydrotestosterone augmented the expression of AR that was not modified by the inhibition of the PI3K/mTOR or Erk1/2 pathways. AR expression was posttranscriptionally regulated by PI3K or Erk1/2 inhibition. Our results describe the expression of the AR in TNBC as a druggable target and further suggest the combination of bicalutamide with inhibitors of EGFR, PDGFRβ or Erk1/2 for future development

  3. Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase

    International Nuclear Information System (INIS)

    Mattila, Elina; Marttila, Heidi; Sahlberg, Niko; Kohonen, Pekka; Tähtinen, Siri; Halonen, Pasi; Perälä, Merja; Ivaska, Johanna

    2010-01-01

    T-cell protein tyrosine phosphatase (TCPTP/TC45) is a ubiquitously expressed intra-cellular non-receptor protein tyrosine phosphatase involved in the negative regulation of several cancer relevant cellular signalling pathways. We have previously shown that interaction between the α-cytoplasmic tail of α1β1 integrin and TCPTP activates TCPTP by disrupting an inhibitory intra-molecular bond in TCPTP. Thus, inhibition of the regulatory interaction in TCPTP is a desirable strategy for TCPTP activation and attenuation of oncogenic RTK signalling. However, this is challenging with low molecular weight compounds. We developed a high-throughput compatible assay to analyse activity of recombinant TCPTP in vitro. Using this assay we have screened 64280 small molecules to identify novel agonists for TCPTP. Dose-dependent response to TCPTP agonist was performed using the in vitro assay. Inhibition effects and specificity of TCPTP agonists were evaluated using TCPTP expressing and null mouse embryonic fibroblasts. Western blot analysis was used to evaluate attenuation of PDGFRβ and EGFR phosphorylation. Inhibition of VEGF signalling was analysed with VEGF-induced endothelial cell sprouting assays. From the screen we identified six TCPTP agonists. Two compounds competed with α1-cytoplasmic domain for binding to TCPTP, suggesting that they activate TCPTP similar to α1-cyt by disrupting the intra-molecular bond in TCPTP. Importantly, one of the compounds (spermidine) displayed specificity towards TCPTP in cells, since TCPTP -/- cells were 43-fold more resistant to the compound than TCPTP expressing cells. This compound attenuates PDGFRβ and VEGFR2 signalling in cells in a TCPTP-dependent manner and functions as a negative regulator of EGFR phosphorylation in cancer cells. In this study we showed that small molecules mimicking TCPTP-α1 interaction can be used as TCPTP agonists. These data provide the first proof-of-concept description of the use of high-throughput screening

  4. FDG-PET/CT response evaluation during EGFR-TKI treatment in patients with NSCLC

    Institute of Scientific and Technical Information of China (English)

    Matthijs; H; van; Gool; Tjeerd; S; Aukema; Koen; J; Hartemink; Renato; A; Valdés; Olmos; Houke; M; Klomp; Harm; van; Tinteren

    2014-01-01

    Over recent years,[18F]-fluorodeoxyglucose positron emission tomography acquired together with low dose computed tomography(FDG-PET/CT)has proven its role as a staging modality in patients with non-small cell lung cancer(NSCLC).The purpose of this review was to present the evidence to use FDG-PET/CT for response evaluation in patients with NSCLC,treated with epidermal growth factor receptor(EGFR)-tyrosine kinase inhibitors(TKI).All published articles from 1November 2003 to 1 November 2013 reporting on 18FFDG-PET response evaluation during EGFR-TKI treatment in patients with NSCLC were collected.In total 7studies,including data of 210 patients were eligible for analyses.Our report shows that FDG-PET/CT responseduring EGFR-TKI therapy has potential in targeted treatment for NSCLC.FDG-PET/CT response is associated with clinical and radiologic response and with survival.Furthermore FDG-PET/CT response monitoring can be performed as early as 1-2 wk after initiation of EGFR-TKI treatment.Patients with substantial decrease of metabolic activity during EGFR-TKI treatment will probably benefit from continued treatment.If metabolic response does not occur within the first weeks of EGFR-TKI treatment,patients may be spared(further)unnecessary toxicity of ineffective treatment.Refining FDG-PET response criteria may help the clinician to decide on continuation or discontinuation of targeted treatment.

  5. Tyrosine phosphorylation of Eps15 is required for ligand-regulated, but not constitutive, endocytosis

    DEFF Research Database (Denmark)

    Confalonieri, S; Salcini, A E; Puri, C

    2000-01-01

    for endocytosis of the epidermal growth factor receptor (EGFR), the prototypical ligand-inducible receptor, but not of the transferrin receptor (TfR), the prototypical constitutively internalized receptor. Eps15, an endocytic protein that is tyrosine phosphorylated by EGFR, is a candidate for such a function....... Here, we show that tyrosine phosphorylation of Eps15 is necessary for internalization of the EGFR, but not of the TfR. We mapped Tyr 850 as the major in vivo tyrosine phosphorylation site of Eps15. A phosphorylation-negative mutant of Eps15 acted as a dominant negative on the internalization...... of the EGFR, but not of the TfR. A phosphopeptide, corresponding to the phosphorylated sequence of Eps15, inhibited EGFR endocytosis, suggesting that phosphotyrosine in Eps15 serves as a docking site for a phosphotyrosine binding protein. Thus, tyrosine phosphorylation of Eps15 represents the first molecular...

  6. Downstream of tyrosine kinase/docking protein 6, as a novel substrate of tropomyosin-related kinase C receptor, is involved in neurotrophin 3-mediated neurite outgrowth in mouse cortex neurons

    Directory of Open Access Journals (Sweden)

    Yuan Jian

    2010-06-01

    Full Text Available Abstract Background The downstream of tyrosine kinase/docking protein (Dok adaptor protein family has seven members, Dok1 to Dok7, that act as substrates of multiple receptor tyrosine kinase and non-receptor tyrosine kinase. The tropomyosin-related kinase (Trk receptor family, which has three members (TrkA, TrkB and TrkC, are receptor tyrosine kinases that play pivotal roles in many stages of nervous system development, such as differentiation, migration, axon and dendrite projection and neuron patterning. Upon related neurotrophin growth factor stimulation, dimerisation and autophosphorylation of Trk receptors can occur, recruiting adaptor proteins to mediate signal transduction. Results In this report, by using yeast two-hybrid assays, glutathione S-transferase (GST precipitation assays and coimmunoprecipitation (Co-IP experiments, we demonstrate that Dok6 selectively binds to the NPQY motif of TrkC through its phosphotyrosine-binding (PTB domain in a kinase activity-dependent manner. We further confirmed their interaction by coimmunoprecipitation and colocalisation in E18.5 mouse cortex neurons, which provided more in vivo evidence. Next, we demonstrated that Dok6 is involved in neurite outgrowth in mouse cortex neurons via the RNAi method. Knockdown of Dok6 decreased neurite outgrowth in cortical neurons upon neurotrophin 3 (NT-3 stimulation. Conclusions We conclude that Dok6 interacts with the NPQY motif of the TrkC receptor through its PTB domain in a kinase activity-dependent manner, and works as a novel substrate of the TrkC receptor involved in NT-3-mediated neurite outgrowth in mouse cortex neurons.

  7. Multi-lobulation of the nucleus in prolonged S phase by nuclear expression of Chk tyrosine kinase.

    Science.gov (United States)

    Nakayama, Yuji; Yamaguchi, Naoto

    2005-04-01

    Chk tyrosine kinase phosphorylates Src-family tyrosine kinases and suppresses their kinase activity. We recently showed that Chk localizes to the nucleus as well as the cytoplasm and inhibits cell proliferation. To investigate the role of nuclear Chk in proliferation, various Chk mutants were constructed and expressed. Nuclear localization of Chk-induced dynamic multi-lobulation of the nucleus and prolonged S phase of the cell cycle. The N-terminal domain of Chk and a portion of its kinase domain but not the kinase activity were responsible for induction of the multi-lobulation. Cell sorting analysis revealed that nuclear multi-lobulated cells were enriched in late S phase. Multi-lobulated nuclei were surrounded with lamin B1 that was particularly concentrated in concave regions of the nuclei. Furthermore, treatment with nocodazole or taxol disrupted multi-lobulation of the nucleus. These results suggest that nuclear multi-lobulation in late S phase, which is dependent on polymerization and depolymerization of microtubules, may be involved in nuclear Chk-induced inhibition of proliferation.

  8. Multi-lobulation of the nucleus in prolonged S phase by nuclear expression of Chk tyrosine kinase

    International Nuclear Information System (INIS)

    Nakayama, Yuji; Yamaguchi, Naoto

    2005-01-01

    Chk tyrosine kinase phosphorylates Src-family tyrosine kinases and suppresses their kinase activity. We recently showed that Chk localizes to the nucleus as well as the cytoplasm and inhibits cell proliferation. To investigate the role of nuclear Chk in proliferation, various Chk mutants were constructed and expressed. Nuclear localization of Chk-induced dynamic multi-lobulation of the nucleus and prolonged S phase of the cell cycle. The N-terminal domain of Chk and a portion of its kinase domain but not the kinase activity were responsible for induction of the multi-lobulation. Cell sorting analysis revealed that nuclear multi-lobulated cells were enriched in late S phase. Multi-lobulated nuclei were surrounded with lamin B1 that was particularly concentrated in concave regions of the nuclei. Furthermore, treatment with nocodazole or taxol disrupted multi-lobulation of the nucleus. These results suggest that nuclear multi-lobulation in late S phase, which is dependent on polymerization and depolymerization of microtubules, may be involved in nuclear Chk-induced inhibition of proliferation

  9. Tyrosine kinases, drugs, and Shigella flexneri dissemination.

    Science.gov (United States)

    Dragoi, Ana-Maria; Agaisse, Hervé

    2014-01-01

    Shigella flexneri is an enteropathogenic bacterium responsible for approximately 100 million cases of severe dysentery each year. S. flexneri colonization of the human colonic epithelium is supported by direct spread from cell to cell, which relies on actin-based motility. We have recently uncovered that, in intestinal epithelial cells, S. flexneri actin-based motility is regulated by the Bruton's tyrosine kinase (Btk). Consequently, treatment with Ibrutinib, a specific Btk inhibitor currently used in the treatment of B-cell malignancies, effectively impaired S. flexneri spread from cell to cell. Thus, therapeutic intervention capitalizing on drugs interfering with host factors supporting the infection process may represent an effective alternative to treatments with antimicrobial compounds.

  10. Baseline and Trend of Lymphocyte-to-Monocyte Ratio as Prognostic Factors in Epidermal Growth Factor Receptor Mutant Non-Small Cell Lung Cancer Patients Treated with First-Line Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitors.

    Directory of Open Access Journals (Sweden)

    Yu-Mu Chen

    Full Text Available Patients with early-stage lung cancer who have a high baseline lymphocyte-to-monocyte ratio (LMR have a favorable prognosis. However, the prognostic significance of LMR in patients with advanced-stage EGFR-mutant non-small cell lung cancer (NSCLC receiving first-line epidermal growth factor receptor (EGFR-tyrosine kinase inhibitors (TKIs has not been established. We conducted a retrospective analysis to investigate the influence of LMR on clinical outcomes including progression-free survival (PFS and overall survival (OS in EGFR-mutant patients with NSCLC.Of 1310 lung cancer patients diagnosed between January 2011 and October 2013, 253 patients receiving first-line EGFR-TKIs for EGFR-mutant NSCLC were included. The cut-off values for baseline and the 1-month-to-baseline ratio of LMR (MBR, determined by using receiver operating characteristic curves, were 3.29 and 0.63, respectively. Patients were divided into 3 prognostic groups: high LMR and MBR, high LMR or MBR, and low LMR and MBR.The mean patient age was 65.2 years, and 41% were men. The median PFS and OS were 10.3 and 22.0 months, respectively. The PFS in patients with high LMR and MBR, high LMR or MBR, and low LMR and MBR were 15.4, 7.1, and 2.0 months, respectively (p < 0.001, whereas the OS were 32.6, 13.7, and 5.1 months, respectively (p < 0.001.A combination of baseline and trend of LMR can be used to identify patients with a high mortality risk in EGFR-mutant NSCLC patients receiving first-line EGFR-TKIs.

  11. The radioprotector O-phospho-tyrosine stimulates DNA-repair via epidermal growth factor receptor- and DNA-dependent kinase phosphorylation

    International Nuclear Information System (INIS)

    Dittmann, Klaus; Mayer, Claus; Wanner, Gabriele; Kehlbach, Rainer; Rodemann, H. Peter

    2007-01-01

    Background and purpose: Purpose of the study was to elucidate the underlying molecular mechanism of the radioprotector O-phospho-tyrosine (P-Tyr). Methods: Molecular effects of P-Tyr at the level of EGFR responses were investigated in vitro with bronchial carcinoma cell line A549. Nuclear EGFR transport and DNA-PK activation were quantified after Western blotting. Residual DNA-damages were quantified by help of γH 2 AX focus assay. Results: As determined by dose-response curves, treatment of cells with P-Tyr for 16 h before irradiation results in radioprotection. Simultaneous treatment with EGFR blocking antibody Cetuximab abolished P-Tyr associated radioprotection. At the molecular level P-Tyr mediated a general phosphorylation of EGFR and a pronounced phosphorylation of nuclear EGFR at residue Thr No. 654, also observed after treatment with ionizing radiation. This phosphorylation was associated with nuclear EGFR accumulation. Moreover, P-Tyr-triggered EGFR nuclear accumulation was associated with phosphorylation of DNA-PK at Thr 2609. This activated form of DNA-PK was not DNA associated, but after radiation, DNA binding increased, particularly after P-Tyr pre-treatment. These molecular effects of P-Tyr resulted in a reduction of residual DNA-damage after irradiation. Conclusions: Radioprotection by P-Tyr is mediated through its stimulation of nuclear EGFR transport and concurrent, but DNA-damage independent, activation of DNA-PK. Thus, subsequent irradiation results in increased binding of DNA-PK to DNA, improved DNA-repair and increased cell survival

  12. Protein-tyrosine Phosphatase SHP2 Contributes to GDNF Neurotrophic Activity through Direct Binding to Phospho-Tyr687 in the RET Receptor Tyrosine Kinase*

    Science.gov (United States)

    Perrinjaquet, Maurice; Vilar, Marçal; Ibáñez, Carlos F.

    2010-01-01

    The signaling mechanisms by which neurotrophic receptors regulate neuronal survival and axonal growth are still incompletely understood. In the receptor tyrosine kinase RET, a receptor for GDNF (glial cell line-derived neurotrophic factor), the functions of the majority of tyrosine residues that become phosphorylated are still unknown. Here we have identified the protein-tyrosine phosphatase SHP2 as a novel direct interactor of RET and the first effector known to bind to phosphorylated Tyr687 in the juxtamembrane region of the receptor. We show that SHP2 is recruited to RET upon ligand binding in a cooperative fashion, such that both interaction with Tyr687 and association with components of the Tyr1062 signaling complex are required for stable recruitment of SHP2 to the receptor. SHP2 recruitment contributes to the ability of RET to activate the PI3K/AKT pathway and promote survival and neurite outgrowth in primary neurons. Furthermore, we find that activation of protein kinase A (PKA) by forskolin reduces the recruitment of SHP2 to RET and negatively affects ligand-mediated neurite outgrowth. In agreement with this, mutation of Ser696, a known PKA phosphorylation site in RET, enhances SHP2 binding to the receptor and eliminates the effect of forskolin on ligand-induced outgrowth. Together, these findings establish SHP2 as a novel positive regulator of the neurotrophic activities of RET and reveal Tyr687 as a critical platform for integration of RET and PKA signals. We anticipate that several other phosphotyrosines of unknown function in neuronal receptor tyrosine kinases will also support similar regulatory functions. PMID:20682772

  13. EGFR-TKI therapy for patients with brain metastases from non-small-cell lung cancer: a pooled analysis of published data

    Directory of Open Access Journals (Sweden)

    Fan Y

    2014-11-01

    Full Text Available Yun Fan,1,2 Xiaoling Xu,3 Conghua Xie4 1Zhongnan Hospital of Wuhan University, Department of Radiation Oncology, Wuhan, People's Republic of China; 2Department of Chemotherapy, Zhejiang Cancer Hospital, Hangzhou, People’s Republic of China; 3Zhejiang Cancer Hospital, Hangzhou, People’s Republic of China; 4Zhongnan Hospital of Wuhan University, Department of Radiation Oncology, Wuhan, People’s Republic of China Introduction: Brain metastases are one of the leading causes of death from non-small-cell lung cancer (NSCLC. The use of epidermal growth factor receptor (EGFR tyrosine kinase inhibitors (TKIs to treat brain metastases remains controversial. Thus, we performed a pooled analysis of published data to evaluate the efficacy of EGFR-TKIs in NSCLC patients with brain metastases, particularly for tumors with activating EGFR mutations. Methods: Several data sources were searched, including PubMed, Web of Science, and ASCO Annual Meetings databases. The end points were intracranial overall response rate (ORR, disease control rate (DCR, progression-free survival (PFS, overall survival (OS, and adverse events. The pooled ORR, DCR, PFS, and OS with 95% confidence intervals (CIs were calculated employing fixed- or random-effect models, depending on the heterogeneity of the included studies. Results: Sixteen published studies were included in this analysis, with a total of 464 enrolled patients. The EGFR mutational status was unknown for 362 (unselected group, and 102 had activating EGFR mutations. The pooled intracranial ORR and DCR were 51.8% (95% CI: 45.8%–57.8% and 75.7% (95% CI: 70.3%–80.5%, respectively. A higher ORR was observed in the EGFR mutation group than in the unselected group (85.0% vs 45.1%; a similar trend was observed for the DCR (94.6% vs 71.3%. The pooled median PFS and OS were 7.4 months (95% CI, 4.9–9.9 and 11.9 months (95% CI, 7.7–16.2, respectively, with longer PFS (12.3 months vs 5.9 months and OS (16.2 months vs

  14. The role of Ryk and Ror receptor tyrosine kinases in Wnt signal transduction

    NARCIS (Netherlands)

    Green, J.; Nusse, R.; van Amerongen, R.

    2014-01-01

    Receptor tyrosine kinases of the Ryk and Ror families were initially classified as orphan receptors because their ligands were unknown. They are now known to contain functional extracellular Wnt-binding domains and are implicated in Wnt-signal transduction in multiple species. Although their

  15. Identifying activating mutations in the EGFR gene: prognostic and therapeutic implications in non-small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Gabriel Lima Lopes

    2015-08-01

    Full Text Available AbstractLung cancer is the leading cause of cancer-related deaths worldwide. Promising new therapies have recently emerged from the development of molecular targeted drugs; particularly promising are those blocking the signal transduction machinery of cancer cells. One of the most widely studied cell signaling pathways is that of EGFR, which leads to uncontrolled cell proliferation, increased cell angiogenesis, and greater cell invasiveness. Activating mutations in the EGFR gene (deletions in exon 19 and mutation L858R in exon 21, first described in 2004, have been detected in approximately 10% of all non-squamous non-small cell lung cancer (NSCLC patients in Western countries and are the most important predictors of a response to EGFR tyrosine-kinase inhibitors (EGFR-TKIs. Studies of the EGFR-TKIs gefitinib, erlotinib, and afatinib, in comparison with platinum-based regimens, as first-line treatments in chemotherapy-naïve patients have shown that the EGFR-TKIs produce gains in progression-free survival and overall response rates, although only in patients whose tumors harbor activating mutations in the EGFR gene. Clinical trials have also shown EGFR-TKIs to be effective as second- and third-line therapies in advanced NSCLC. Here, we review the main aspects of EGFR pathway activation in NSCLC, underscore the importance of correctly identifying activating mutations in the EGFR gene, and discuss the main outcomes of EGFR-TKI treatment in NSCLC.

  16. All-trans-retinoic acid enhances apoptosis induction by tyrosine kinase inhibitors in the eosinophilic leukemia-derived EoL-1 cell line.

    Science.gov (United States)

    Robert, Carine; Apàti, Agota; Chomienne, Christine; Papp, Béla

    2008-02-01

    Imatinib and retinoids induce apoptosis in FIP1L1/PDGFRalpha-positive EoL-1 leukemia cells. Although imatinib induces complete remission in most FIP1L1/PDGFRalpha-positive patients, response to imatinib is sometimes suboptimal. In order to enhance the potency of the molecularly targeted therapy of eosinophilic leukemia, we investigated the effect of retinoids combined with tyrosine kinase inhibitors on EoL-1 cells. We demonstrate that retinoids combined with tyrosine kinase inhibitors lead to enhanced apoptosis induction in EoL-1 cells. Our results suggest that tyrosine kinase inhibitors combined with retinoids may constitute a valuable therapeutic approach for sensitive neoplasias that may display enhanced anti-leukemic potency when compared to single drug treatments.

  17. Anti-EGFR-Targeted Therapy for Esophageal and Gastric Cancers: An Evolving Concept

    Directory of Open Access Journals (Sweden)

    Tomislav Dragovich

    2009-01-01

    Full Text Available Cancers of the esophagus and stomach present a major health burden worldwide. In the past 30 years we have witnessed some interesting shifts in terms of epidemiology of esophago gastric cancers. Regardless of a world region, the majority of patients diagnosed with esophageal or gastric cancers die from progression or recurrence of their disease. While there are many active cytotoxic agents for esophageal and stomach cancers, their impact on the disease course has been modest at best. Median survival for patients with advanced gastroesophageal cancer is still less than a year. Therefore, novel strategies, based on our understanding of biology and genetics, are desperately needed. Epidermal growth factor receptor (EGFR pathway has been implicated in pathophysiology of many epithelial malignancies, including esophageal and stomach cancers. EGFR inhibitors, small molecule tyrosine kinase inhibitors and monoclonal antibodies, have been explored in patients with esophageal and gastric cancers. It appears that tumors of the distal esophagus and gastroesophageal junction (GEJ may be more sensitive to EGFR blockade than distal gastric adenocarcinomas. Investigations looking into potential molecular predictors of sensitivity to EGFR inhibitors for patients with esophageal and GEJ cancers are ongoing. While we are still searching for those predictors, it is clear that they will be different from ones identified in lung and colorectal cancers. Further development of EGFR inhibitors for esophageal and GEJ cancers should be driven by better understanding of EGFR pathway disregulation that drives cancer progression in a sensitive patient population.

  18. Predictive value of K-ras and PIK3CA in non-small cell lung cancer patients treated with EGFR-TKIs: a systemic review and meta-analysis

    International Nuclear Information System (INIS)

    Chen, Jie-Ying; Cheng, Ya-Nan; Han, Lei; Wei, Feng; Yu, Wen-Wen; Zhang, Xin-Wei; Cao, Shui; Yu, Jin-Pu

    2015-01-01

    A meta-analysis was performed to augment the insufficient data on the impact of mutative EGFR downstream phosphatidylinositol-3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) pathways on the clinical efficiency of epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) treatment of non-small cell lung cancer (NSCLC) patients. Network databases were explored in April, 2015. Papers that investigated the clinical outcomes of NSCLC patients treated with EGFR-TKIs according to the status of K-ras and/or PIK3CA gene mutation were included. A quantitative meta-analysis was conducted using standard statistical methods. Odds ratios (ORs) for objective response rate (ORR) and hazard ratios (HRs) for progression-free survival (PFS) and overall survival (OS) were calculated. Mutation in K-ras significantly predicted poor ORR [OR =0.22; 95% confidence interval (CI), 0.13-0.35], shorter PFS (HR =1.56; 95% CI, 1.27-1.92), and shorter OS (HR =1.59; 95% CI, 1.33-1.91) in NSCLC patients treated with EGFR-TKIs. Mutant PIK3CA significantly predicted shorter OS (HR =1.83; 95% CI, 1.05-3.20), showed poor ORR (OR =0.70; 95% CI, 0.22-2.18), and shorter PFS (HR =1.79; 95% CI, 0.91-3.53) in NSCLC patients treated with EGFR-TKIs. K-ras mutation adversely affected the clinical response and survival of NSCLC patients treated with EGFR-TKIs. PIK3CA mutation showed similar trends. In addition to EGFR, adding K-ras and PIK3CA as routine gene biomarkers in clinical genetic analysis is valuable to optimize the effectiveness of EGFR-TKI regimens and identify optimal patients who will benefit from EGFR-TKI treatment

  19. Identification of a functional interaction between Kv4.3 channels and c-Src tyrosine kinase.

    Science.gov (United States)

    Gomes, Pedro; Saito, Tomoaki; Del Corsso, Cris; Alioua, Abderrahmane; Eghbali, Mansoureh; Toro, Ligia; Stefani, Enrico

    2008-10-01

    Voltage-gated K(+) (Kv) channels are key determinants of cardiac and neuronal excitability. A substantial body of evidence has accumulated in support of a role for Src family tyrosine kinases in the regulation of Kv channels. In this study, we examined the possibility that c-Src tyrosine kinase participates in the modulation of the transient voltage-dependent K(+) channel Kv4.3. Supporting a mechanistic link between Kv4.3 and c-Src, confocal microscopy analysis of HEK293 cells stably transfected with Kv4.3 showed high degree of co-localization of the two proteins at the plasma membrane. Our results further demonstrate an association between Kv4.3 and c-Src by co-immunoprecipitation and GST pull-down assays, this interaction being mediated by the SH2 and SH3 domains of c-Src. Furthermore, we show that Kv4.3 is tyrosine phosphorylated under basal conditions. The functional relevance of the observed interaction between Kv4.3 and c-Src was established in patch-clamp experiments, where application of the Src inhibitor PP2 caused a decrease in Kv4.3 peak current amplitude, but not the inactive structural analogue PP3. Conversely, intracellular application of recombinant c-Src kinase or the protein tyrosine phosphatase inhibitor bpV(phen) increased Kv4.3 peak current amplitude. In conclusion, our findings provide evidence that c-Src-induced Kv4.3 channel activation involves their association in a macromolecular complex and suggest a role for c-Src-Kv4.3 pathway in regulating cardiac and neuronal excitability.

  20. Survival significance of epidermal growth factor receptor tyrosine kinase inhibitors and current staging system for survival after recurrence in patients with completely resected lung adenocarcinoma

    Directory of Open Access Journals (Sweden)

    Saji H

    2017-08-01

    Full Text Available Hisashi Saji,1,2 Hiroki Sakai,1 Hiroyuki Kimura,1 Tomoyuki Miyazawa,1 Hideki Marushima,1 Haruhiko Nakamura1 1Department of Chest Surgery, St Marianna University School of Medicine, Miyamae-ku, Kawasaki, Kanagawa, Japan; 2Department of Thoracic Surgery, Tokyo Medical University, Shinjuku-ku, Tokyo, Japan Objective: We previously reported that the staging system and epidermal growth factor receptor (EGFR mutation status are key factors for treatment strategy and predicting survival. However, the significance of these factors as predictors of overall survival (OS and postoperative recurrence survival (PRS has not been sufficiently elucidated. The objective here was to investigate EGFR mutation status and p-stage, which affect PRS and OS in patients with completely resected lung adenocarcinoma, using a different database.Patients and methods: We retrospectively reviewed 56 consecutive lung adenocarcinoma patients with disease recurrence in St. Marianna University Hospital between January 2010 and December 2014.Results: EGFR mutants (M were detected in 16/56 patients (29%. The patients with EGFR M had a better OS than those with EGFR wild-type (WT status (5-year survival: 50.3% vs 43.1, P=0.133. There was no significant difference in the 3-year recurrence-free survival rate between patients with M and WT (6.3% vs 7.7%, P=0.656, and the patients with EGFR M had a significantly better 3-year PRS than those with WT (77.4% vs 51.7%, P=0.033. The 3-year PRS rate for patients with M/pathologic stage (p-stage I–II (87.5% was better than that for patients with M/p-stage III (60.0%, WT/p-stage I–II (52.7%, and WT/p-stage III (43.8%. There was a significant difference between patients with M/p-stage I and WT/p-stage I–II or WT/p-stage III (P=0.021 and 0.030, respectively. During the study period, of the 16 patients with mutants, 12 patients (75% received EGFR-tyrosine kinase inhibitor (TKI therapy and among the 40 patients with WT, no patient received

  1. Vorinostat and metformin sensitize EGFR-TKI resistant NSCLC cells via BIM-dependent apoptosis induction.

    Science.gov (United States)

    Chen, Hengyi; Wang, Yubo; Lin, Caiyu; Lu, Conghua; Han, Rui; Jiao, Lin; Li, Li; He, Yong

    2017-11-07

    There is a close relationship between low expression of BIM and resistance to epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI). Vorinostat is a pan-histone deacetylase inhibitor (HDACi) that augments BIM expression in various types of tumor cells, however, this effect is attenuated by the high expression of anti-apoptotic proteins in EGFR-TKI resistant non-small cell lung cancer (NSCLC) cells. Vorinostat in combination with metformin - a compound that can inhibit anti-apoptotic proteins expression, might cooperate to activate apoptotic signaling and overcome EGFR-TKI resistance. This study aimed to investigate the cooperative effect and evaluate possible molecular mechanisms. The results showed that vorinostat combined with gefitinib augmented BIM expression and increased the sensitivity of EGFR-TKI resistant NSCLC cells to gefitinib, adding metformin simultaneously could obviously inhibit the expression of anti-apoptotic proteins, and further increased expression levels of BIM and BAX, and as a result, further improved the sensitivity of gefitinib both on the NSCLC cells with intrinsic and acquired resistance to EGFR-TKI. In addition, autophagy induced by gefitinib and vorinostat could be significantly suppressed by metformin, which might also contribute to enhance apoptosis and improve sensitivity of gefitinib. These results suggested that the combination of vorinostat and metformin might represent a novel strategy to overcome EGFR-TKI resistance associated with BIM-dependent apoptosis in larger heterogeneous populations.

  2. Partial response to carboplatin in an RRx-001 pretreated patient with EGFR-inhibitor-resistance and T790M-negative NSCLC.

    Science.gov (United States)

    Carter, Corey A; Oronsky, Bryan; Caroen, Scott; Scicinski, Jan; Cabrales, Pedro; Degesys, Aiste; Brzezniak, Christina

    2016-01-01

    Few therapeutic options are available for T790M-negative non-small cell lung cancer (NSCLC) after failure of primary epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) and chemotherapy. This report presents the case of a 71-year-old Asian female never smoker with EGFR mutated T790M negative non squamous cell lung cancer (NSCLC) pre-treated with the experimental epi-immunotherapeutic agent, RRx-001, that re-responded to single agent carboplatin after failure of platinum doublets, TKIs, pemetrexed and nivolumab. The management of advanced EGFR mutation-positive NSCLC is briefly reviewed herein and the emerging paradigm of episensitization, which contradicts the long-standing and widely accepted tenet about the immutability of resistance and the futility of therapeutic rechallenge, is introduced as a strategy to avert treatment failure and thereby stave off deterioration and death.

  3. KEAP1 loss modulates sensitivity to kinase targeted therapy in lung cancer. | Office of Cancer Genomics

    Science.gov (United States)

    Inhibitors that target the receptor tyrosine kinase (RTK)/Ras/mitogen-activated protein kinase (MAPK) pathway have led to clinical responses in lung and other cancers, but some patients fail to respond and in those that do resistance inevitably occurs (Balak et al., 2006; Kosaka et al., 2006; Rudin et al., 2013; Wagle et al., 2011). To understand intrinsic and acquired resistance to inhibition of MAPK signaling, we performed CRISPR-Cas9 gene deletion screens in the setting of BRAF, MEK, EGFR, and ALK inhibition.

  4. Structure-based network analysis of activation mechanisms in the ErbB family of receptor tyrosine kinases: the regulatory spine residues are global mediators of structural stability and allosteric interactions.

    Directory of Open Access Journals (Sweden)

    Kevin A James

    Full Text Available The ErbB protein tyrosine kinases are among the most important cell signaling families and mutation-induced modulation of their activity is associated with diverse functions in biological networks and human disease. We have combined molecular dynamics simulations of the ErbB kinases with the protein structure network modeling to characterize the reorganization of the residue interaction networks during conformational equilibrium changes in the normal and oncogenic forms. Structural stability and network analyses have identified local communities integrated around high centrality sites that correspond to the regulatory spine residues. This analysis has provided a quantitative insight to the mechanism of mutation-induced "superacceptor" activity in oncogenic EGFR dimers. We have found that kinase activation may be determined by allosteric interactions between modules of structurally stable residues that synchronize the dynamics in the nucleotide binding site and the αC-helix with the collective motions of the integrating αF-helix and the substrate binding site. The results of this study have pointed to a central role of the conserved His-Arg-Asp (HRD motif in the catalytic loop and the Asp-Phe-Gly (DFG motif as key mediators of structural stability and allosteric communications in the ErbB kinases. We have determined that residues that are indispensable for kinase regulation and catalysis often corresponded to the high centrality nodes within the protein structure network and could be distinguished by their unique network signatures. The optimal communication pathways are also controlled by these nodes and may ensure efficient allosteric signaling in the functional kinase state. Structure-based network analysis has quantified subtle effects of ATP binding on conformational dynamics and stability of the EGFR structures. Consistent with the NMR studies, we have found that nucleotide-induced modulation of the residue interaction networks is not

  5. MHC class I signaling in T cells leads to tyrosine kinase activity and PLC-gamma 1 phosphorylation

    DEFF Research Database (Denmark)

    Skov, S; Odum, Niels; Claesson, M H

    1995-01-01

    phosphorylation and the subsequent calcium response. The early tyrosine kinase activity was found to be dependent on expression of the TCR/CD3 complex and the CD45 molecule on the surface of the T cells. Furthermore, MHC-I cross-linking was shown to tyrosine phosphorylate PLC-gamma 1 (phospholipase C-gamma 1...

  6. Functional interaction between nonreceptor tyrosine kinase c-Abl and SR-Rich protein RBM39

    International Nuclear Information System (INIS)

    Mai, Sanyue; Qu, Xiuhua; Li, Ping; Ma, Qingjun; Liu, Xuan; Cao, Cheng

    2016-01-01

    RBM39, also known as splicing factor HCC1.4, acts as a transcriptional coactivator for the steroid nuclear receptors JUN/AP-1, ESR1/ER-α and ESR2/ER-β. RBM39 is involved in the regulation of the transcriptional responses of these steroid nuclear receptors and promotes transcriptional initiation. In this paper, we report that RBM39 interacts with the nonreceptor tyrosine kinase c-Abl. Both the Src homology (SH) 2 and SH3 domains of c-Abl interact with RBM39. The major tyrosine phosphorylation sites on RBM39 that are phosphorylated by c-Abl are Y95 and Y99, as demonstrated by liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS) and mutational analysis. c-Abl was shown boost the transcriptional coactivation activity of RBM39 for ERα and PRβ in a tyrosine kinase-dependent manner. The results suggest that mammalian c-Abl plays an important role in steroid hormone receptor-mediated transcription by regulating RBM39. - Highlights: • c-Abl interacts with RBM39. • RBM39 is phosphorylated by c-Abl. • c-Abl regulates transcriptional coactivation activity of RBM39 on the ERα and PRβ.

  7. Functional interaction between nonreceptor tyrosine kinase c-Abl and SR-Rich protein RBM39

    Energy Technology Data Exchange (ETDEWEB)

    Mai, Sanyue [Beijing Institute of Biotechnology, 27 Taiping Rd, Haidian District, Beijing 100850 (China); Qu, Xiuhua [General Navy Hospital of PLA, 6 Fucheng Rd, Haidian District, Beijing 100037 (China); Li, Ping; Ma, Qingjun [Beijing Institute of Biotechnology, 27 Taiping Rd, Haidian District, Beijing 100850 (China); Liu, Xuan, E-mail: liux931932@163.com [Beijing Institute of Biotechnology, 27 Taiping Rd, Haidian District, Beijing 100850 (China); Cao, Cheng, E-mail: cao_c@sohu.com [Beijing Institute of Biotechnology, 27 Taiping Rd, Haidian District, Beijing 100850 (China)

    2016-04-22

    RBM39, also known as splicing factor HCC1.4, acts as a transcriptional coactivator for the steroid nuclear receptors JUN/AP-1, ESR1/ER-α and ESR2/ER-β. RBM39 is involved in the regulation of the transcriptional responses of these steroid nuclear receptors and promotes transcriptional initiation. In this paper, we report that RBM39 interacts with the nonreceptor tyrosine kinase c-Abl. Both the Src homology (SH) 2 and SH3 domains of c-Abl interact with RBM39. The major tyrosine phosphorylation sites on RBM39 that are phosphorylated by c-Abl are Y95 and Y99, as demonstrated by liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS) and mutational analysis. c-Abl was shown boost the transcriptional coactivation activity of RBM39 for ERα and PRβ in a tyrosine kinase-dependent manner. The results suggest that mammalian c-Abl plays an important role in steroid hormone receptor-mediated transcription by regulating RBM39. - Highlights: • c-Abl interacts with RBM39. • RBM39 is phosphorylated by c-Abl. • c-Abl regulates transcriptional coactivation activity of RBM39 on the ERα and PRβ.

  8. Effects of EGFR Inhibitor on Helicobacter pylori Induced Gastric Epithelial Pathology in Vivo

    Directory of Open Access Journals (Sweden)

    Philip A. Robinson

    2013-10-01

    Full Text Available Helicobacter pylori transactivates the Epidermal Growth Factor Receptor (EGFR and predisposes to gastric cancer development in humans and animal models. To examine the importance of EGFR signalling to gastric pathology, this study investigated whether treatment of Mongolian gerbils with a selective EGFR tyrosine kinase inhibitor, EKB-569, altered gastric pathology in chronic H. pylori infection. Gerbils were infected with H. pylori and six weeks later received either EKB-569-supplemented, or control diet, for 32 weeks prior to sacrifice. EKB-569-treated H. pylori-infected gerbils had no difference in H. pylori colonisation or inflammation scores compared to infected animals on control diet, but showed significantly less corpus atrophy, mucous metaplasia and submucosal glandular herniations along with markedly reduced antral and corpus epithelial proliferation to apoptosis ratios. EKB-569-treated infected gerbils had significantly decreased abundance of Cox-2, Adam17 and Egfr gastric transcripts relative to infected animals on control diet. EGFR inhibition by EKB-569 therefore reduced the severity of pre-neoplastic gastric pathology in chronically H. pylori-infected gerbils. EKB-569 increased gastric epithelial apoptosis in H. pylori-infected gerbils which counteracted some of the consequences of increased gastric epithelial cell proliferation. Similar chemopreventative strategies may be useful in humans who are at high risk of developing H.pylori-induced gastric adenocarcinoma.

  9. rse, a novel receptor-type tyrosine kinase with homology to Axl/Ufo, is expressed at high levels in the brain.

    Science.gov (United States)

    Mark, M R; Scadden, D T; Wang, Z; Gu, Q; Goddard, A; Godowski, P J

    1994-04-08

    We have isolated cDNA clones that encode the human and murine forms of a novel receptor-type tyrosine kinase termed Rse. Sequence analysis indicates that human Rse contains 890 amino acids, with an extracellular region composed of two immunoglobulin-like domains followed by two fibronectin type III domains. Murine Rse contains 880 amino acids and shares 90% amino acid identity with its human counterpart. Rse is structurally similar to the receptor-type tyrosine kinase Axl/Ufo, and the two proteins have 35 and 63% sequence identity in their extracellular and intracellular domains, respectively. To study the synthesis and activation of this putative receptor-type tyrosine kinase, we constructed a version of Rse (termed gD-Rse, where gD represents glycoprotein D) that contains an NH2-terminal epitope tag. NIH3T3 cells were engineered to express gD-Rse, which could be detected at the cell surface by fluorescence-activated cell sorting. Moreover, gD-Rse was rapidly phosphorylated on tyrosine residues upon incubation of the cells with an antibody directed against the epitope tag, suggesting that rse encodes an active tyrosine kinase. In the human tissues we examined, the highest level of expression of rse mRNA was observed in the brain; rse mRNA was also detected in the premegakaryocytopoietic cell lines CMK11-5 and Dami. The gene for rse was localized to human chromosome 15.

  10. Domains of the growth hormone receptor required for association and activation of JAK2 tyrosine kinase

    DEFF Research Database (Denmark)

    VanderKuur, J A; Wang, X; Zhang, L

    1994-01-01

    Growth hormone (GH) has recently been shown to activate the GH receptor (GHR)-associated tyrosine kinase JAK2. In the present study, regions of the GHR required for JAK2 association with GHR were identified. GH-dependent JAK2 association with GHR was detected in Chinese hamster ovary (CHO) cells...... and RIN-5AH cells, the ability of JAK2 to associate with the mutated GHR was found to correlate with GH-dependent activation of JAK2, tyrosyl phosphorylation of GHR (in the case of GHR1-638 and GHR1-454), and the ability of the GHR to copurify with tyrosine kinase activity. In CHO cells expressing mutated......, and that tyrosines in the N-terminal half of the cytoplasmic domain of the GHR are phosphorylated by JAK2. The finding that a specific interaction with the C-terminal half of GHR appears to be necessary for p97 phosphorylation indicates that while JAK2 activation may be necessary for a full biological response to GH...

  11. Levels of active tyrosine kinase receptor determine the tumor response to Zalypsis

    International Nuclear Information System (INIS)

    Moneo, Victoria; Serelde, Beatriz G; Blanco-Aparicio, Carmen; Diaz-Uriarte, Ramon; Avilés, Pablo; Santamaría, Gemma; Tercero, Juan C; Cuevas, Carmen; Carnero, Amancio

    2014-01-01

    Zalypsis® is a marine compound in phase II clinical trials for multiple myeloma, cervical and endometrial cancer, and Ewing’s sarcoma. However, the determinants of the response to Zalypsis are not well known. The identification of biomarkers for Zalypsis activity would also contribute to broaden the spectrum of tumors by selecting those patients more likely to respond to this therapy. Using in vitro drug sensitivity data coupled with a set of molecular data from a panel of sarcoma cell lines, we developed molecular signatures that predict sensitivity to Zalypsis. We verified these results in culture and in vivo xenograft studies. Zalypsis resistance was dependent on the expression levels of PDGFRα or constitutive phosphorylation of c-Kit, indicating that the activation of tyrosine kinase receptors (TKRs) may determine resistance to Zalypsis. To validate our observation, we measured the levels of total and active (phosphorylated) forms of the RTKs PDGFRα/β, c-Kit, and EGFR in a new panel of diverse solid tumor cell lines and found that the IC50 to the drug correlated with RTK activation in this new panel. We further tested our predictions about Zalypsis determinants for response in vivo in xenograft models. All cells lines expressing low levels of RTK signaling were sensitive to Zalypsis in vivo, whereas all cell lines except two with high levels of RTK signaling were resistant to the drug. RTK activation might provide important signals to overcome the cytotoxicity of Zalypsis and should be taken into consideration in current and future clinical trials

  12. Tumor suppressor function of Bruton tyrosine kinase is independent of its catalytic activity

    NARCIS (Netherlands)

    S. Middendorp; A.J.E. Zijlstra (Esther); R. Kersseboom (Rogier); G.M. Dingjan (Gemma); H. Jumaa; R.W. Hendriks (Rudi)

    2005-01-01

    textabstractDuring B-cell development in the mouse, Bruton tyrosine kinase (Btk) and the adaptor protein SLP-65 (Src homology 2 [SH2] domain-containing leukocyte protein of 65 kDa) limit the expansion and promote the differentiation of pre-B cells. Btk is thought to mainly function

  13. Three Paths to Better Tyrosine Kinase Inhibition Behind the Blood-Brain Barrier in Treating Chronic Myelogenous Leukemia and Glioblastoma with Imatinib

    Science.gov (United States)

    Kast, Richard E; Focosi, Daniele

    2010-01-01

    Chronic myelogenous leukemia (CML) can be controlled for years with the tyrosine kinase inhibitor imatinib but because imatinib poorly penetrates the blood-brain barrier (BBB), on occasion, the CML clone will thrive and evolve to an accelerated phase in the resulting imatinib sanctuary within the central nervous system. In this, CML resembles glioblastoma in that imatinib, which otherwise may be effective, cannot get to the tumor. Although a common street drug of abuse, methamphetamine is Food and Drug Administration-approved and marketed as a pharmaceutical drug to treat attention-deficit disorders. It has shown the ability to open the BBB in rodents. We have some clinical hints that it may do so in humans as well. This short note presents three new points potentially leading to better tyrosine kinase inhibition behind the BBB: 1) Pharmaceutical methamphetamine may have a useful role in treating both CML and glioblastoma by allowing higher imatinib concentrations behind the BBB. 2) The old antidepressant and monoamine oxidase inhibitor selegiline, used to treat Parkinson disease, is catabolized to methamphetamine. Selegiline, as a nonscheduled drug,may therefore be an easier way to open the BBB, allowing more effective chemotherapy with tyrosine kinases. 3) Dasatinib is a tyrosine kinase inhibitor with a spectrum of inhibition only partially overlapping that of imatinib and a mechanism of tyrosine kinase inhibition that is different from that of imatinib. The two should be additive. In addition, dasatinib crosses the BBB poorly, and it can therefore be expected to benefit from methamphetamine-assisted entry. PMID:20165690

  14. Combined Targeting of BCL-2 and BCR-ABL Tyrosine Kinase Eradicates Chronic Myeloid Leukemia Stem Cells

    Science.gov (United States)

    Mak, Po Yee; Mu, Hong; Zhou, Hongsheng; Mak, Duncan H.; Schober, Wendy; Leverson, Joel D.; Zhang, Bin; Bhatia, Ravi; Huang, Xuelin; Cortes, Jorge; Kantarjian, Hagop; Konopleva, Marina

    2016-01-01

    BCR-ABL tyrosine kinase inhibitors (TKIs) are effective against chronic myeloid leukemia (CML), but they rarely eliminate CML stem cells. Disease relapse is common upon therapy cessation, even in patients with complete molecular responses. Furthermore, once CML progresses to blast crisis (BC), treatment outcomes are dismal. We hypothesized that concomitant targeting of BCL-2 and BCR-ABL tyrosine kinase could overcome these limitations. We demonstrate increased BCL-2 expression at the protein level in bone marrow cells, particularly in Lin−Sca-1+cKit+ cells of inducible CML in mice as determined by CyTOF mass cytometry. Further, selective inhibition of BCL-2, aided by TKI-mediated MCL-1 and BCL-XL inhibition, markedly decreased leukemic Lin−Sca-1+cKit+ cell numbers and long-term stem cell frequency, and prolonged survival in a murine CML model. Additionally, this combination effectively eradicated CD34+CD38−, CD34+CD38+, and quiescent stem/progenitor CD34+ cells from BC CML patient samples. Our results suggest that BCL-2 is a key survival factor for CML stem/progenitor cells and that combined inhibition of BCL-2 and BCR-ABL tyrosine kinase has the potential to significantly improve depth of response and cure rates of chronic phase and BC CML. PMID:27605552

  15. PET imaging-based phenotyping as a predictive biomarker of response to tyrosine kinase inhibitor therapy in non-small cell lung cancer: Are we there yet?

    Energy Technology Data Exchange (ETDEWEB)

    Gerbaudo, Victor H.; Kim, Chun K. [Div. of Nuclear Medicine and Molecular Imaging, Dept. of Radiology,Brigham and Women' s Hospital and Harvard Medical School, Boston (United States)

    2017-03-15

    The increased understanding of the molecular pathology of different malignancies, especially lung cancer, has directed investigational efforts to center on the identification of different molecular targets and on the development of targeted therapies against these targets. A good representative is the epidermal growth factor receptor (EGFR); a major driver of non-small cell lung cancer tumorigenesis. Today, tumor growth inhibition is possible after treating lung tumors expressing somatic mutations of the EGFR gene with tyrosine kinase inhibitors (TKI). This opened the doors to biomarker-directed precision or personalized treatments for lung cancer patients. The success of these targeted anticancer therapies depends in part on being able to identify biomarkers and their patho-molecular make-up in order to select patients that could respond to specific therapeutic agents. While the identification of reliable biomarkers is crucial to predict response to treatment before it begins, it is also essential to be able to monitor treatment early during therapy to avoid the toxicity and morbidity of futile treatment in non-responding patients. In this context, we share our perspective on the role of PET imaging-based phenotyping in the personalized care of lung cancer patients to non-invasively direct and monitor the treatment efficacy of TKIs in clinical practice.

  16. Activation of the protein tyrosine phosphatase SHP2 via the interleukin-6 signal transducing receptor protein gp130 requires tyrosine kinase Jak1 and limits acute-phase protein expression.

    Science.gov (United States)

    Schaper, F; Gendo, C; Eck, M; Schmitz, J; Grimm, C; Anhuf, D; Kerr, I M; Heinrich, P C

    1998-11-01

    Stimulation of the interleukin-6 (IL-6) signalling pathway occurs via the IL-6 receptor-glycoprotein 130 (IL-6R-gp130) receptor complex and results in the regulation of acute-phase protein genes in liver cells. Ligand binding to the receptor complex leads to tyrosine phosphorylation and activation of Janus kinases (Jak), phosphorylation of the signal transducing subunit gp130, followed by recruitment and phosphorylation of the signal transducer and activator of transcription factors STAT3 and STAT1 and the src homology domain (SH2)-containing protein tyrosine phosphatase (SHP2). The tyrosine phosphorylated STAT factors dissociate from the receptor, dimerize and translocate to the nucleus where they bind to enhancer sequences of IL-6 target genes. Phosphorylated SHP2 is able to bind growth factor receptor bound protein (grb2) and thus might link the Jak/STAT pathway to the ras/raf/mitogen-activated protein kinase pathway. Here we present data on the dose-dependence, kinetics and kinase requirements for SHP2 phosphorylation after the activation of the signal transducer, gp130, of the IL-6-type family receptor complex. When human fibrosarcoma cell lines deficient in Jak1, Jak2 or tyrosine kinase 2 (Tyk2) were stimulated with IL-6-soluble IL-6R complexes it was found that only in Jak1-, but not in Jak 2- or Tyk2-deficient cells, SHP2 activation was greatly impaired. It is concluded that Jak1 is required for the tyrosine phosphorylation of SHP2. This phosphorylation depends on Tyr-759 in the cytoplasmatic domain of gp130, since a Tyr-759-->Phe exchange abrogates SHP2 activation and in turn leads to elevated and prolonged STAT3 and STAT1 activation as well as enhanced acute-phase protein gene induction. Therefore, SHP2 plays an important role in acute-phase gene regulation.

  17. Bioinformatics in cancer therapy and drug design

    International Nuclear Information System (INIS)

    Horbach, D.Y.; Usanov, S.A.

    2005-01-01

    One of the mechanisms of external signal transduction (ionizing radiation, toxicants, stress) to the target cell is the existence of membrane and intracellular proteins with intrinsic tyrosine kinase activity. No wonder that etiology of malignant growth links to abnormalities in signal transduction through tyrosine kinases. The epidermal growth factor receptor (EGFR) tyrosine kinases play fundamental roles in development, proliferation and differentiation of tissues of epithelial, mesenchymal and neuronal origin. There are four types of EGFR: EGF receptor (ErbB1/HER1), ErbB2/Neu/HER2, ErbB3/HER3 and ErbB4/HER4. Abnormal expression of EGFR, appearance of receptor mutants with changed ability to protein-protein interactions or increased tyrosine kinase activity have been implicated in the malignancy of different types of human tumors. Bioinformatics is currently using in investigation on design and selection of drugs that can make alterations in structure or competitively bind with receptors and so display antagonistic characteristics. (authors)

  18. Bioinformatics in cancer therapy and drug design

    Energy Technology Data Exchange (ETDEWEB)

    Horbach, D Y [International A. Sakharov environmental univ., Minsk (Belarus); Usanov, S A [Inst. of bioorganic chemistry, National academy of sciences of Belarus, Minsk (Belarus)

    2005-05-15

    One of the mechanisms of external signal transduction (ionizing radiation, toxicants, stress) to the target cell is the existence of membrane and intracellular proteins with intrinsic tyrosine kinase activity. No wonder that etiology of malignant growth links to abnormalities in signal transduction through tyrosine kinases. The epidermal growth factor receptor (EGFR) tyrosine kinases play fundamental roles in development, proliferation and differentiation of tissues of epithelial, mesenchymal and neuronal origin. There are four types of EGFR: EGF receptor (ErbB1/HER1), ErbB2/Neu/HER2, ErbB3/HER3 and ErbB4/HER4. Abnormal expression of EGFR, appearance of receptor mutants with changed ability to protein-protein interactions or increased tyrosine kinase activity have been implicated in the malignancy of different types of human tumors. Bioinformatics is currently using in investigation on design and selection of drugs that can make alterations in structure or competitively bind with receptors and so display antagonistic characteristics. (authors)

  19. Tyrosine phosphorylation of the BRI1 receptor kinase occurs via a posttranslational modification and is activated by the juxtamembrane domain

    Science.gov (United States)

    In metazoans, receptor kinases control many essential processes related to growth and development and response to the environment. The receptor kinases in plants and animals are structurally similar but evolutionarily distinct from one another, and thus while most animal receptor kinases are tyrosin...

  20. Metazoan-like signaling in a unicellular receptor tyrosine kinase

    Directory of Open Access Journals (Sweden)

    Schultheiss Kira P

    2013-02-01

    Full Text Available Abstract Background Receptor tyrosine kinases (RTKs are crucial components of signal transduction systems in multicellular animals. Surprisingly, numerous RTKs have been identified in the genomes of unicellular choanoflagellates and other protists. Here, we report the first biochemical study of a unicellular RTK, namely RTKB2 from Monosiga brevicollis. Results We cloned, expressed, and purified the RTKB2 kinase, and showed that it is enzymatically active. The activity of RTKB2 is controlled by autophosphorylation, as in metazoan RTKs. RTKB2 possesses six copies of a unique domain (designated RM2 in its C-terminal tail. An isolated RM2 domain (or a synthetic peptide derived from the RM2 sequence served as a substrate for RTKB2 kinase. When phosphorylated, the RM2 domain bound to the Src homology 2 domain of MbSrc1 from M. brevicollis. NMR structural studies of the RM2 domain indicated that it is disordered in solution. Conclusions Our results are consistent with a model in which RTKB2 activation stimulates receptor autophosphorylation within the RM2 domains. This leads to recruitment of Src-like kinases (and potentially other M. brevicollis proteins and further phosphorylation, which may serve to increase or dampen downstream signals. Thus, crucial features of signal transduction circuitry were established prior to the evolution of metazoans from their unicellular ancestors.

  1. Association of BIM Deletion Polymorphism and BIM-γ RNA Expression in NSCLC with EGFR Mutation.

    Science.gov (United States)

    Isobe, Kazutoshi; Kakimoto, Atsushi; Mikami, Tetsuo; Kaburaki, Kyohei; Kobayashi, Hiroshi; Yoshizawa, Takahiro; Makino, Takashi; Otsuka, Hajime; Sano, G O; Sugino, Keishi; Sakamoto, Susumu; Takai, Yujiro; Tochigi, Naobumi; Iyoda, Akira; Homma, Sakae

    This pilot study assessed the association of BIM deletion polymorphism and BIM RNA isoform in patients with EGFR-positive non-small cell lung cancer (NSCLC). The study included 33 patients with EGFR-positive NSCLC treated with gefitinib. BIM deletion polymorphism and BIM RNA isoform (EL/L/S/γ) were determined by polymerase chain reaction (PCR). BIM-γ expression was significantly higher in patients with BIM deletion polymorphism than among those without BIM deletion polymorphism inside tumors (p=0.038) and around tumors (p=0.0024). Relative BIM-γ expression was significantly higher in patients with BIM deletion polymorphism than among those without BIM deletion polymorphism (p=0.0017). Patients with BIM-γ had significantly shorter progression-free survival than those without BIM-γ (median: 304 vs. 732 days; p=0.023). Expression of BIM-γ mRNA and BIM deletion polymorphism were strongly associated. BIM-γ overexpression may have a role in apoptosis related to EGFR-tyrosine kinase inhibitor. Copyright© 2016, International Institute of Anticancer Research (Dr. John G. Delinasios), All rights reserved.

  2. Progranulin and the receptor tyrosine kinase EphA2, partners in crime?

    Science.gov (United States)

    Chitramuthu, Babykumari; Bateman, Andrew

    2016-01-01

    Progranulin is a secreted protein with roles in tumorigenesis, inflammation, and neurobiology, but its signaling receptors have remained unclear. In this issue, Neill et al. (2016. J. Cell Biol. https://doi.org/10.1083/jcb.201603079) identify the tyrosine kinase EphA2 as a strong candidate for such a receptor, providing insight into progranulin and EphA2 signaling. PMID:27903608

  3. LONG-TERM RESULTS OF TARGET THERAPY WITH FIRST AND * SECOND-LINE TYROSINE KINASE INHIBITORS IN PATIENTS WITH CHRONIC MYELOID LEUKEMIA

    Directory of Open Access Journals (Sweden)

    L. L. Vysotskaya

    2015-01-01

    Full Text Available Aim: To assess long-term efficacy of firstand second-line tyrosine kinase inhibitors in non-selected patients with chronic myeloid leukemia in a real-life clinical setting.Materials and methods: The assessment is based on long-term results of a prospective single center comparative clinical trial that was based on non-selected groups of 116 patients with various stages of chronic myeloid leukemia being treated with a first generation tyrosine kinase inhibitor imatinib, and of 44 patients being treated with a second generation tyrosine kinase inhibitor nilotinib. We analyzed all-cause mortality, progression-free survival from April 2005 to April 2013, with a median of the follow-up of 128 months.Results: In 116 patients with chronic myeloid leukemia treated with imatinib, the Kaplan-Meier survival estimate was 120 months. In 44 patients at an early chronic phase, 5-year overall survival and progression-free survival was 93.2% and 8-year overall and progression-free survival was 79.5%. In 44 patients at a late chronic stage, 5-year overall and progression-free survival was 95.5%, 8-year overall and progression-free survival, 72.7%. In 28 patients at acceleration phase, 5-years overall survival was 78.6% and 8-year overall survival, 46%. Median of overall survival in patients treated with nilotinib was not reached. During 78.6 months of combination treatment with cytotoxic agents, tyrosine kinase inhibitors of the first (imatinib and second line (nilotinib, overall survival was 100%.Conclusion: In clinical practice, inclusion of patients with chronic myeloid leukemia and imatinib resistance (disease relapse or imatinib intolerance into the treatment program with frontline therapy with general cytotoxic agents and thereafter with firstand second-line tyrosine kinase inhibitors significantly improves overall survival.

  4. An in vivo C. elegans model system for screening EGFR-inhibiting anti-cancer drugs.

    Directory of Open Access Journals (Sweden)

    Young-Ki Bae

    Full Text Available The epidermal growth factor receptor (EGFR is a well-established target for cancer treatment. EGFR tyrosine kinase (TK inhibitors, such as gefinitib and erlotinib, have been developed as anti-cancer drugs. Although non-small cell lung carcinoma with an activating EGFR mutation, L858R, responds well to gefinitib and erlotinib, tumors with a doubly mutated EGFR, T790M-L858R, acquire resistance to these drugs. The C. elegans EGFR homolog LET-23 and its downstream signaling pathway have been studied extensively to provide insight into regulatory mechanisms conserved from C. elegans to humans. To develop an in vivo screening system for potential cancer drugs targeting specific EGFR mutants, we expressed three LET-23 chimeras in which the TK domain was replaced with either the human wild-type TK domain (LET-23::hEGFR-TK, a TK domain with the L858R mutation (LET-23::hEGFR-TK[L858R], or a TK domain with the T790M-L858R mutations (LET-23::hEGFR-TK[T790M-L858R] in C. elegans vulval cells using the let-23 promoter. The wild-type hEGFR-TK chimeric protein rescued the let-23 mutant phenotype, and the activating mutant hEGFR-TK chimeras induced a multivulva (Muv phenotype in a wild-type C. elegans background. The anti-cancer drugs gefitinib and erlotinib suppressed the Muv phenotype in LET-23::hEGFR-TK[L858R]-expressing transgenic animals, but not in LET-23::hEGFR-TK[T790M-L858R] transgenic animals. As a pilot screen, 8,960 small chemicals were tested for Muv suppression, and AG1478 (an EGFR-TK inhibitor and U0126 (a MEK inhibitor were identified as potential inhibitors of EGFR-mediated biological function. In conclusion, transgenic C. elegans expressing chimeric LET-23::hEGFR-TK proteins are a model system that can be used in mutation-specific screens for new anti-cancer drugs.

  5. Inhibiting Src family tyrosine kinase activity blocks glutamate signalling to ERK1/2 and Akt/PKB but not JNK in cultured striatal neurones.

    Science.gov (United States)

    Crossthwaite, Andrew J; Valli, Haseeb; Williams, Robert J

    2004-03-01

    Glutamate receptor activation of mitogen-activated protein (MAP) kinase signalling cascades has been implicated in diverse neuronal functions such as synaptic plasticity, development and excitotoxicity. We have previously shown that Ca2+-influx through NMDA receptors in cultured striatal neurones mediates the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt/protein kinase B (PKB) through a phosphatidylinositol 3-kinase (PI 3-kinase)-dependent pathway. Exposing neurones to the Src family tyrosine kinase inhibitor PP2, but not the inactive analogue PP3, inhibited NMDA receptor-induced phosphorylation of ERK1/2 and Akt/PKB in a concentration-dependent manner, and reduced cAMP response element-binding protein (CREB) phosphorylation. To establish a link between Src family tyrosine kinase-mediated phosphorylation and PI 3-kinase signalling, affinity precipitation experiments were performed with the SH2 domains of the PI 3-kinase regulatory subunit p85. This revealed a Src-dependent phosphorylation of a focal adhesion kinase (FAK)-p85 complex on glutamate stimulation. Demonstrating that PI3-kinase is not ubiquitously involved in NMDA receptor signal transduction, the PI 3-kinase inhibitors wortmannin and LY294002 did not prevent NMDA receptor Ca2+-dependent phosphorylation of c-Jun N-terminal kinase 1/2 (JNK1/2). Further, inhibiting Src family kinases increased NMDA receptor-dependent JNK1/2 phosphorylation, suggesting that Src family kinase-dependent cascades may physiologically limit signalling to JNK. These results demonstrate that Src family tyrosine kinases and PI3-kinase are pivotal regulators of NMDA receptor signalling to ERK/Akt and JNK in striatal neurones.

  6. Icotinib in Patients with Pretreated Advanced Esophageal Squamous Cell Carcinoma with EGFR Overexpression or EGFR Gene Amplification: A Single-Arm, Multicenter Phase 2 Study.

    Science.gov (United States)

    Huang, Jing; Fan, Qingxia; Lu, Ping; Ying, Jianming; Ma, Changwu; Liu, Wei; Liu, Ying; Tan, Fenlai; Sun, Yan

    2016-06-01

    Epidermal growth factor receptor (EGFR) has been reported to be overexpressed and amplified in a high percentage of patients with esophageal squamous cell carcinoma (ESCC). The activity of icotinib, an EGFR tyrosine kinase inhibitor, was assessed in previously treated ESCC with EGFR overexpression or amplification. For this phase 2, single-arm, multicenter trial undertaken at six hospitals in China, we included Chinese patients with previously treated, histologically confirmed advanced ESCC and EGFR overexpression (immunohistochemical staining sore of 3+) or amplification (positive fluorescence in situ hybridization result). These patients received oral icotinib (250 mg, three times daily).The primary end point was the proportion of patients with objective responses as assessed by an independent radiology review committee. Between December 5, 2013, and May 28, 2015, a total of 281 patients were screened. Fifty-four eligible patients were enrolled. Nine responses were observed, including one complete response and eight partial responses, and 16 patients had stable disease, resulting in a 16.7% objective response rate (95% confidence interval [CI]: 6.7-26.6) and 46.3% disease control rate (95% CI: 33.0-59.6). The median progression-free survival and overall survival times were 52 (95% CI: 40-95) days and 153 (95% CI: 139-218) days, respectively. A total of 43 patients experienced at least one adverse event, but most were only grade 1 to 2 in severity. The most frequent was rash (48.1%), followed by diarrhea (22.2%). Icotinib showed favorable activity in patients with advanced, previously treated ESCC with EGFR overexpression or amplification. These findings suggest further research into EGFR overexpression or amplification for selecting responsive patients. Copyright © 2016 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.

  7. PGE2 mediates EGFR internalization and nuclear translocation via caveolin endocytosis promoting its transcriptional activity and proliferation in human NSCLC cells.

    Science.gov (United States)

    Bazzani, Lorenzo; Donnini, Sandra; Giachetti, Antonio; Christofori, Gerhard; Ziche, Marina

    2018-03-13

    Prostaglandin E 2 (PGE 2 ) contributes to tumor progression by promoting cancer cell growth, invasion and by creating a favorable pro-tumor microenvironment. PGE 2 has been reported to transactivate and internalize into the nucleus receptor tyrosine kinases such as Epidermal growth factor receptor (EGFR), thereby supporting tumor progression. Here we demonstrate that in non-small cell lung carcinoma (NSCLC) cells, PGE 2 induces EGFR nuclear translocation via different dynamin-dependent endocytic pathways, promotes the formation of an EGFR-STAT3 complex, affects nuclear EGFR target gene expression and mediates tumor cell proliferation. Indeed, we find that PGE 2 induces EGFR internalization and consequent nuclear import through Clathrin- and Caveolin-mediated endocytosis and through the interaction of EGFR with Importin β1. Within the nucleus, EGFR forms a complex with STAT3, an event blocked by ablation of Clathrin Heavy Chain or Caveolin-1. The combination of EGF and PGE 2 prolongs nuclear EGFR transcriptional activity manifested by the upregulation of CCND1 , PTGS2 , MYC and NOS2 mRNA levels and potentiates nuclear EGFR-induced NSCLC cell proliferation. Additionally, NSCLC patients with high expression of a nuclear EGFR gene signature display shorter survival times than those with low expression, thus showing a putative correlation between nuclear EGFR and poor prognosis in NSCLC. Together, our findings indicate a complex mechanism underlying PGE 2 -induced EGF/EGFR signaling and transcriptional control, which plays a key role in cancer progression.

  8. EGFR targeted therapy in non-small cell lung cancer: potential role of cetuximab

    Directory of Open Access Journals (Sweden)

    Chad A Reade

    2009-05-01

    Full Text Available Chad A Reade1, Apar Kishor Ganti1,21Department of Internal Medicine, University of Nebraska Medical Center, Omaha, NE, USA; 2Section of Oncology-Hematology, Department of internal Medicine, VA Medical Center, Omaha, NE, USAAbstract: Chemotherapy alone has limited ability to significantly improve survival in non-small lung cancer (NSCLC beyond what has already been achieved. The epidermal growth factor (EGF pathway plays a vital role in the pathogenesis and progression of NSCLC. Two classes of drugs inhibit the EGF receptor (EGFR pathway: small molecules that inhibit the intracellular tyrosine kinase activity of the receptor, and monoclonal antibodies that target the extracellular domain in the ligand-binding region. Cetuximab is a human – mouse chimeric immunoglobulin G1 class monoclonal antibody directed against EGFR. Preclinical studies with cetuximab suggested that there was inhibition of growth of human NSCLC cell lines. Cetuximab is currently the focus of intense investigation in various patient populations with NSCLC. This review focuses on clinical trials of cetuximab in NSCLC and identifies future directions with this agent.Keywords: non-small cell lung cancer, EGFR, cetuximab, monoclonal antibodies

  9. Receptor tyrosine kinase structure and function in health and disease

    Directory of Open Access Journals (Sweden)

    Oleg A. Karpov

    2015-09-01

    Full Text Available Receptor tyrosine kinases (RTKs are membrane proteins that control the flow of information through signal transduction pathways, impacting on different aspects of cell function. RTKs are characterized by a ligand-binding ectodomain, a single transmembrane α-helix, a cytosolic region comprising juxtamembrane and kinase domains followed by a flexible C-terminal tail. Somatic and germline RTK mutations can induce aberrant signal transduction to give rise to cardiovascular, developmental and oncogenic abnormalities. RTK overexpression occurs in certain cancers, correlating signal strength and disease incidence. Diverse RTK activation and signal transduction mechanisms are employed by cells during commitment to health or disease. Small molecule inhibitors are one means to target RTK function in disease initiation and progression. This review considers RTK structure, activation, and signal transduction and evaluates biological relevance to therapeutics and clinical outcomes.

  10. Inhibition of ErbB2 by receptor tyrosine kinase inhibitors causes myofibrillar structural damage without cell death in adult rat cardiomyocytes

    International Nuclear Information System (INIS)

    Pentassuglia, Laura; Graf, Michael; Lane, Heidi; Kuramochi, Yukio; Cote, Gregory; Timolati, Francesco; Sawyer, Douglas B.; Zuppinger, Christian; Suter, Thomas M.

    2009-01-01

    Inhibition of ErbB2 (HER2) with monoclonal antibodies, an effective therapy in some forms of breast cancer, is associated with cardiotoxicity, the pathophysiology of which is poorly understood. Recent data suggest, that dual inhibition of ErbB1 (EGFR) and ErbB2 signaling is more efficient in cancer therapy, however, cardiac safety of this therapeutic approach is unknown. We therefore tested an ErbB1-(CGP059326) and an ErbB1/ErbB2-(PKI166) tyrosine kinase inhibitor in an in-vitro system of adult rat ventricular cardiomyocytes and assessed their effects on 1. cell viability, 2. myofibrillar structure, 3. contractile function, and 4. MAPK- and Akt-signaling alone or in combination with Doxorubicin. Neither CGP nor PKI induced cardiomyocyte necrosis or apoptosis. PKI but not CGP caused myofibrillar structural damage that was additive to that induced by Doxorubicin at clinically relevant doses. These changes were associated with an inhibition of excitation-contraction coupling. PKI but not CGP decreased p-Erk1/2, suggesting a role for this MAP-kinase signaling pathway in the maintenance of myofibrils. These data indicate that the ErbB2 signaling pathway is critical for the maintenance of myofibrillar structure and function. Clinical studies using ErbB2-targeted inhibitors for the treatment of cancer should be designed to include careful monitoring for cardiac dysfunction.

  11. Acquired Resistance of EGFR-Mutant Lung Adenocarcinomas to Afatinib plus Cetuximab Is Associated with Activation of mTORC1

    Directory of Open Access Journals (Sweden)

    Valentina Pirazzoli

    2014-05-01

    Full Text Available Patients with EGFR-mutant lung adenocarcinomas (LUADs who initially respond to first-generation tyrosine kinase inhibitors (TKIs develop resistance to these drugs. A combination of the irreversible TKI afatinib and the EGFR antibody cetuximab can be used to overcome resistance to first-generation TKIs; however, resistance to this drug combination eventually emerges. We identified activation of the mTORC1 signaling pathway as a mechanism of resistance to dual inhibition of EGFR in mouse models. The addition of rapamycin reversed resistance in vivo. Analysis of afatinib-plus-cetuximab-resistant biopsy specimens revealed the presence of genomic alterations in genes that modulate mTORC1 signaling, including NF2 and TSC1. These findings pinpoint enhanced mTORC1 activation as a mechanism of resistance to afatinib plus cetuximab and identify genomic mechanisms that lead to activation of this pathway, revealing a potential therapeutic strategy for treating patients with resistance to these drugs.

  12. Itk tyrosine kinase substrate docking is mediated by a nonclassical SH2 domain surface of PLCgamma1.

    Science.gov (United States)

    Min, Lie; Joseph, Raji E; Fulton, D Bruce; Andreotti, Amy H

    2009-12-15

    Interleukin-2 tyrosine kinase (Itk) is a Tec family tyrosine kinase that mediates signaling processes after T cell receptor engagement. Activation of Itk requires recruitment to the membrane via its pleckstrin homology domain, phosphorylation of Itk by the Src kinase, Lck, and binding of Itk to the SLP-76/LAT adapter complex. After activation, Itk phosphorylates and activates phospholipase C-gamma1 (PLC-gamma1), leading to production of two second messengers, DAG and IP(3). We have previously shown that phosphorylation of PLC-gamma1 by Itk requires a direct, phosphotyrosine-independent interaction between the Src homology 2 (SH2) domain of PLC-gamma1 and the kinase domain of Itk. We now define this docking interface using a combination of mutagenesis and NMR spectroscopy and show that disruption of the Itk/PLCgamma1 docking interaction attenuates T cell signaling. The binding surface on PLCgamma1 that mediates recognition by Itk highlights a nonclassical binding activity of the well-studied SH2 domain providing further evidence that SH2 domains participate in important signaling interactions beyond recognition of phosphotyrosine.

  13. Targeting the epidermal growth factor receptor in solid tumor malignancies

    DEFF Research Database (Denmark)

    Nedergaard, Mette K; Hedegaard, Chris J; Poulsen, Hans S

    2012-01-01

    been proposed as valid targets in many cancer therapy settings. Different strategies have been developed in order to either inhibit EGFR/EGFRvIII activity or to ablate EGFR/EGFRvIII-positive tumor cells. Drugs that inhibit these receptors include monoclonal antibodies (mAbs) that bind...... to the extracellular part of EGFR, blocking the binding sites for the EGFR ligands, and intracellular tyrosine kinase inhibitors (TKIs) that block the ATP binding site of the tyrosine kinase domain. Besides an EGFRvIII-targeted vaccine, conjugated anti-EGFR mAbs have been used in different settings to deliver lethal...... agents to the EGFR/EGFRvIII-positive cells; among these are radio-labelled mAbs and immunotoxins. This article reviews the current status and efficacy of EGFR/EGFRvIII-targeted therapies....

  14. Crystal Structure of Human Dual-Specificity Tyrosine-Regulated Kinase 3 Reveals New Structural Features and Insights into its Auto-phosphorylation.

    Science.gov (United States)

    Kim, Kuglae; Cha, Jeong Seok; Cho, Yong-Soon; Kim, Hoyoung; Chang, Nienping; Kim, Hye-Jung; Cho, Hyun-Soo

    2018-04-07

    Dual-specificity tyrosine-regulated kinases (DYRKs) auto-phosphorylate a critical tyrosine residue in their activation loop and phosphorylate their substrate on serine and threonine residues. The auto-phosphorylation occurs intramolecularly and is a one-off event. DYRK3 is selectively expressed at a high level in hematopoietic cells and attenuates erythroblast development, leading to anemia. In the present study, we determined the crystal structure of the mature form of human DYRK3 in complex with harmine, an ATP competitive inhibitor. The crystal structure revealed a phosphorylation site, residue S350, whose phosphorylation increases the stability of DYRK3 and enhances its kinase activity. In addition, our structural and biochemical assays suggest that the N-terminal auto-phosphorylation accessory domain stabilizes the DYRK3 protein, followed by auto-phosphorylation of the tyrosine of the activation loop, which is important for kinase activity. Finally, our docking analysis provides information for the design of novel and potent therapeutics to treat anemia. Copyright © 2018 Elsevier Ltd. All rights reserved.

  15. Molecular mechanisms of the synergy between cysteinyl-leukotrienes and receptor tyrosine kinase growth factors on human bronchial fibroblast proliferation

    Directory of Open Access Journals (Sweden)

    Hajime Yoshisue

    2006-12-01

    Full Text Available We have reported that cysteinyl-leukotrienes (cys-LTs synergise not only with epidermal growth factor (EGF but also with platelet-derived growth factor (PDGF and fibroblast growth factor (FGF to induce mitogenesis in human bronchial fibroblasts. We now describe the molecular mechanisms underlying this synergism. Mitogenesis was assessed by incorporation of [3H]thymidine into DNA and changes in protein phosphorylation by Western blotting. Surprisingly, no CysLT receptor antagonists (MK-571, montelukast, BAY u9773 prevented the synergistic mitogenesis. LTD4 did not cause phosphorylation of EGFR nor did it augment EGF-induced phosphorylation of EGFR, and the synergy between LTD4 and EGF was not blocked by the metalloproteinase inhibitor GM6001 or by an HB-EGF neutralising antibody. The EGFR-selective kinase inhibitor, AG1478, suppressed the synergy by LTD4 and EGF, but had no effect on the synergy with PDGF and FGF. While inhibitors of mitogen-activated protein kinase, phosphatidylinositol 3-kinase and protein kinase C (PKC prevented the synergy, these drugs also inhibited mitogenesis elicited by EGF alone. In contrast, pertussis toxin (PTX efficiently inhibited the potentiating effect of LTD4 on EGF-induced mitogenesis, as well as that provoked by PDGF or FGF, but had no effect on mitogenesis elicited by the growth factors alone. Whereas LTD4 alone did not augment phosphorylation of extracellular signal-regulated kinase (Erk-1/2 and Akt, it increased phosphorylation of PKC in a Gi-dependent manner. Addition of LTD4 prolonged the duration of EGF-induced phosphorylation of Erk-1/2 and Akt, both of which were sensitive to PTX. The effect of cys-LTs involves a PTX-sensitive and PKC-mediated intracellular pathway leading to sustained growth factor-dependent phosphorylation of Erk-1/2 and Akt.

  16. Treatment Choice for Advanced Non-small Cell Lung Cancer Patients Who Had Gradual Progression After EGFR-TKIs: 32 Cases Report

    Directory of Open Access Journals (Sweden)

    Lin LIN

    2013-10-01

    Full Text Available Background and objective The epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs have been widely used in the treatment of the advanced non-small cell lung cancer (NSCLC, especially in the adenocarcinoma patients with activating EGFR mutations. But there is no published overview of the following treatment. This report through observing the efficacy, toxicity and overall survival of different treatments to the advanced NSCLC patients who had gradual progression after EGFR-TKIs, evaluates the influence of the continued treatment and switching chemotherapy. Methods Retrospective review is conducted on 32 cases of advanced NSCLC patients who experienced treatment failure of EGFR-TKIs. One group accepted the continued treatment and the other group accepted the switching chemotherapy. Results The median overall survival of the continued treatment group is 36.0 months. The respose rate of the switching chemotherapy group is 43.75%, and clinical benefit rate (complete and partial response and stable disease is 87.5%. The median overall survival is 15.5 months. The main toxicities are nausea, vomiting and hematological toxicities. Conclusion For the advanced NSCLC patients who had gradual progression after EGFR-TKIs, the continued treatment is one of the acceptable choices.

  17. Induction of matrix metalloproteinase-2 by tenascin-X deficiency is mediated through the c-Jun N-terminal kinase and protein tyrosine kinase phosphorylation pathway

    International Nuclear Information System (INIS)

    Matsumoto, Ken-ichi; Minamitani, Takeharu; Orba, Yasuko; Sato, Mami; Sawa, Hirofumi; Ariga, Hiroyoshi

    2004-01-01

    The results of our previous study showed that tumor invasion and metastasis are promoted in extracellular matrix (ECM) tenascin-X-deficient (TNX-/-) mice via increased expression of matrix metalloproteinases (MMPs). However, little is known about the relationship between TNX deficiency and activation of MMP genes. In this study, we investigated the molecular mechanism by which TNX deficiency activates the MMP-2 gene. We examined the intracellular signaling pathways that regulate gene expression of the proteinase in isolated fibroblasts. Results of gelatin zymography showed that MMP-2 was induced to a greater extent in TNX-/- fibroblasts embedded in type I collagen than in wild-type fibroblasts. RT-PCR analysis revealed that the increased level of MMP-2 expression was caused at the transcription level. Conversely, stable overexpression of TNX in a fibroblast cell line reduced MMP-2 expression and suppressed MMP-2 promoter activity. In addition, treatment of TNX-/- fibroblasts with SP600125, a c-Jun N-terminal kinase (JNK) inhibitor, and genistein, a tyrosine kinase inhibitor, suppressed the increased level of proMMP-2 and increased MMP-2 promoter activity in TNX-/- fibroblasts. Furthermore, increased activation of JNK and tyrosine phosphorylation of certain proteins were observed in TNX-/- fibroblasts. These findings suggest that induction of MMP-2 by TNX deficiency is mediated, at least in part, through the JNK and protein tyrosine kinase phosphorylation pathway

  18. EGFR, HER-2 and KRAS in canine gastric epithelial tumors: a potential human model?

    Directory of Open Access Journals (Sweden)

    Rossella Terragni

    Full Text Available Epidermal growth factor receptor (EGFR or HER-1 and its analog c-erbB-2 (HER-2 are protein tyrosine kinases correlated with prognosis and response to therapy in a variety of human cancers. KRAS mediates the transduction of signals between EGFR and the nucleus, and its mutation has been identified as a predictor of resistance to anti-EGFR drugs. In human oncology, the importance of the EGFR/HER-2/KRAS signalling pathway in gastric cancer is well established, and HER-2 testing is required before initiating therapy. Conversely, this pathway has never been investigated in canine gastric tumours. A total of 19 canine gastric epithelial neoplasms (5 adenomas and 14 carcinomas were retrospectively evaluated for EGFR/HER-2 immunohistochemical expression and KRAS mutational status. Five (35.7% carcinomas were classified as intestinal-type and 9 (64.3% as diffuse-type. EGFR was overexpressed (≥ 1+ in 8 (42.1% cases and HER-2 (3+ in 11 (57.9% cases, regardless of tumour location or biological behaviour. The percentage of EGFR-positive tumours was significantly higher in the intestinal-type (80% than in the diffuse-type (11.1%, p = 0.023. KRAS gene was wild type in 18 cases, whereas one mucinous carcinoma harboured a point mutation at codon 12 (G12R. EGFR and HER-2 may be promising prognostic and therapeutic targets in canine gastric epithelial neoplasms. The potential presence of KRAS mutation should be taken into account as a possible mechanism of drug resistance. Further studies are necessary to evaluate the role of dog as a model for human gastric cancer.

  19. [Clinical Observation of Icotinib Hydrochloride for Advanced Non-small Cell Lung Cancer Patients with EGFR Status Identified].

    Science.gov (United States)

    Li, Xi; Qin, Na; Wang, Jinghui; Yang, Xinjie; Zhang, Xinyong; Lv, Jialin; Wu, Yuhua; Zhang, Hui; Nong, Jingying; Zhang, Quan; Zhang, Shucai

    2015-12-01

    Icotinib is the first self-developed small molecular drug in China for targeted therapy of lung cancer. Compared to the other two commercially available epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors, gefitinib and erlotinib, icotinib is similar to them in chemical structure, mechanism of activity and therapeutic effects. To explore the efficacy and side effects of icotinib hydrochloride in the treatment of the advanced non-small cell lung cancer (NSCLC) patients with EGFR mutation and wild-type. Patients with advanced NSCLC who were treated with icotinib hydrochloride in Beijing Chest Hospital were retrospective analyzed from March 2009 to December 2014. The clinical data of 124 patients (99 with EGFR mutation and 25 with wild type) with advanced NSCLC were enrolled in this study. The patients' overall objective response rate (ORR) was 51.6 % and the disease control rate (DCR) was 79.8%; The patients with EGFR mutation, ORR was 63.6%, DCR was 93.9%. The ORR was 4.0% and the DCR was 24.0% in the wild-type patients. Median progression-free survival (PFS) with icotinib treatment in EGFR mutation patients was 10.5 months and 1.0 month in wild-type patients. The major adverse events were mild skin rash (30.6%) and diarrhea (16.1%). Monotherapy with icotinib hydrochloride is effective and tolerable for the advanced NSCLC EGFR mutation patients.


  20. Validation of the digital PCR system in tyrosine kinase inhibitor-resistant EGFR mutant non-small-cell lung cancer.

    Science.gov (United States)

    Masago, Katsuhiro; Fujita, Shiro; Hata, Akito; Okuda, Chiyuki; Yoshizumi, Yuko; Kaji, Reiko; Katakami, Nobuyuki; Hirata, Yukio; Yatabe, Yasushi

    2018-03-01

    The aim of this study was to compare the accuracy of the QuantStudio 3D Digital polymerase chain reaction (dPCR) system and a PCR-based next generation sequencing (NGS) system for detecting a secondary mutation in the epidermal growth factor receptor (EGFR) gene T790M in non-small cell lung cancer (NSCLC) patients previously diagnosed with EGFR-activating mutations. Twenty-five patients with NSCLC previously treated with EGFR-TKIs were examined. The patients were treated daily with either erlotinib or gefitinib. New biopsies, followed by DNA sequencing on an Ion Torrent systems using the Ion Torrent AmpliSeq Cancer Hotspot Panel and dPCR were performed. A comparison of NGS, sensitive PCR, and dPCR revealed that the sensitivities of NGS and dPCR were similar in this study. As well, T790M was detected in as low as about 5% of mutant allelic frequencies, which represented 5% of the total reads on site mapped reads in NGS and greater than 5% of the dPCR reads, which represented mutant and wild type copies. The strategy in which NGS sequencing is followed by revealed acquired mutation with dPCR may be a reasonable one. We demonstrated the utility of combining NGS and dPCR as a tool for monitoring T790M. NGS and dPCR with formalin-fixed paraffin-embedded (FFPE) specimens might become a standard genomic test for exploring acquired resistance to targeted molecular medicines. © 2018 Japanese Society of Pathology and John Wiley & Sons Australia, Ltd.

  1. Next generation of adeno-associated virus 2 vectors: Point mutations in tyrosines lead to high-efficiency transduction at lower doses

    OpenAIRE

    Zhong, Li; Li, Baozheng; Mah, Cathryn S.; Govindasamy, Lakshmanan; Agbandje-McKenna, Mavis; Cooper, Mario; Herzog, Roland W.; Zolotukhin, Irene; Warrington, Kenneth H.; Weigel-Van Aken, Kirsten A.; Hobbs, Jacqueline A.; Zolotukhin, Sergei; Muzyczka, Nicholas; Srivastava, Arun

    2008-01-01

    Recombinant adeno-associated virus 2 (AAV2) vectors are in use in several Phase I/II clinical trials, but relatively large vector doses are needed to achieve therapeutic benefits. Large vector doses also trigger an immune response as a significant fraction of the vectors fails to traffic efficiently to the nucleus and is targeted for degradation by the host cell proteasome machinery. We have reported that epidermal growth factor receptor protein tyrosine kinase (EGFR-PTK) signaling negatively...

  2. Activation of Bacillus subtilis Ugd by the BY-Kinase PtkA Proceeds via Phosphorylation of Its Residue Tyrosine 70

    DEFF Research Database (Denmark)

    Petranovic, Dina; Grangeasse, C.; Macek, B.

    2009-01-01

    -specific phosphoproteomic study indicated that tyrosine 70 is phosphorylated in the Bacillus subtilis UDP-glucose dehydrogenase Ugd. In this study we confirm that this tyrosine 70 is indeed the main residue phosphorylated by the cognate BY-kinase PtkA. Homology-based modeling of the Ugd structure using structures from UDP...

  3. The Syk protein tyrosine kinase can function independently of CD45 or Lck in T cell antigen receptor signaling

    NARCIS (Netherlands)

    Chu, D. H.; Spits, H.; Peyron, J. F.; Rowley, R. B.; Bolen, J. B.; Weiss, A.

    1996-01-01

    The protein tyrosine phosphatase CD45 is a critical component of the T cell antigen receptor (TCR) signaling pathway, acting as a positive regulator of Src family protein tyrosine kinases (PTKs) such as Lck. Most CD45-deficient human and murine T cell lines are unable to signal through their TCRs.

  4. A threshold model for receptor tyrosine kinase signaling specificity and cell fate determination [version 1; referees: 4 approved

    Directory of Open Access Journals (Sweden)

    Allen Zinkle

    2018-06-01

    Full Text Available Upon ligand engagement, the single-pass transmembrane receptor tyrosine kinases (RTKs dimerize to transmit qualitatively and quantitatively different intracellular signals that alter the transcriptional landscape and thereby determine the cellular response. The molecular mechanisms underlying these fundamental events are not well understood. Considering recent insights into the structural biology of fibroblast growth factor signaling, we propose a threshold model for RTK signaling specificity in which quantitative differences in the strength/longevity of ligand-induced receptor dimers on the cell surface lead to quantitative differences in the phosphorylation of activation loop (A-loop tyrosines as well as qualitative differences in the phosphorylation of tyrosines mediating substrate recruitment. In this model, quantitative differences on A-loop tyrosine phosphorylation result in gradations in kinase activation, leading to the generation of intracellular signals of varying amplitude/duration. In contrast, qualitative differences in the pattern of tyrosine phosphorylation on the receptor result in the recruitment/activation of distinct substrates/intracellular pathways. Commensurate with both the dynamics of the intracellular signal and the types of intracellular pathways activated, unique transcriptional signatures are established. Our model provides a framework for engineering clinically useful ligands that can tune receptor dimerization stability so as to bias the cellular transcriptome to achieve a desired cellular output.

  5. EGFR mutations in patients with lung adenocarcinoma in southwest China: are G719S/A and L861Q more likely detected in tumors derived from smokers?

    Directory of Open Access Journals (Sweden)

    Wang QS

    2013-07-01

    Full Text Available Qiushi Wang,1 Jianghong Mou,1 Xin Yang,1 Yong He,2 Zengpeng Li,1 Qingya Luo,1 Yanqing Li,1 Li Lin,1 Yu Ma,1 Hualiang Xiao11Department of Pathology, 2Department of Respiration, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, People’s Republic of ChinaBackground: The clinical characteristics of epidermal growth factor receptor (EGFR hotspot mutations, such as deletions in exon 19, substitution of L858R in exon 21, and mutations in exon 20, have been widely reported in nonsmall cell lung cancer. However, the clinical features of other low frequency EGFR mutations in these four exons (especially the relationship with smoking history, eg, substitutions of G719S/A/C in exon 18 and L861Q in exon 21, remain unclear. This study investigated the relationship between G719S/A/C and L861Q mutations (in exon 18 and 21 and smoking history.Methods: Specimens from 194 patients with lung adenocarcinoma were analyzed for EGFR mutations in exons 18–21 by high-resolution melting curve analysis and amplification refractory mutation technology to establish the relationship between G719S/A/C and L861Q mutations and smoking history.Results: Ninety-six of 194 tumors (49.5% were confirmed to be EGFR mutation-positive. Among these mutations, 71 of 104 (68.3% were from never smokers, six of 17 (35.3% were from former smokers, and 19 of 73 (26.0% were from current smokers (P < 0.001. The mutation rate in heavy smokers (5/23, 21.7% was significantly lower than in light smokers (20/67, 29.9% and never smokers (71/104, 68.3%, P < 0.001. Seven low frequency EGFR mutations (four substitutions of G719S, and G719 A, respectively, and three of L861Q in exon 21 were identified. Five of these mutations were derived from smokers (one former light smoker, one current heavy smoker, and three current light smokers. Four of these patients had been treated with tyrosine kinase inhibitors and all had a partial response, with median overall

  6. The Receptor Tyrosine Kinase AXL in Cancer Progression

    Directory of Open Access Journals (Sweden)

    Erinn B. Rankin

    2016-11-01

    Full Text Available The AXL receptor tyrosine kinase (AXL has emerged as a promising therapeutic target for cancer therapy. Recent studies have revealed a central role of AXL signaling in tumor proliferation, survival, stem cell phenotype, metastasis, and resistance to cancer therapy. Moreover, AXL is expressed within cellular components of the tumor microenvironment where AXL signaling contributes to the immunosuppressive and protumorigenic phenotypes. A variety of AXL inhibitors have been developed and are efficacious in preclinical studies. These agents offer new opportunities for therapeutic intervention in the prevention and treatment of advanced disease. Here we review the literature that has illuminated the cellular and molecular mechanisms by which AXL signaling promotes tumor progression and we will discuss the therapeutic potential of AXL inhibition for cancer therapy.

  7. Neonatal lethality in knockout mice expressing the kinase-dead form of the gefitinib target GAK is caused by pulmonary dysfunction.

    Directory of Open Access Journals (Sweden)

    Hiroe Tabara

    Full Text Available Gefitinib (Iressa is an inhibitor of the epidermal growth factor receptor (EGFR that has shown promising activity in the treatment of patients with non-small cell lung cancer (NSCLC. However, adverse side effects of gefitinib treatment, such as respiratory dysfunction, have limited the therapeutic benefit of this targeting strategy. The present results show that this adverse effect can be attributed to the inhibition of the novel gefitinib target GAK (Cyclin G-associated kinase, which is as potently inhibited by the drug as the tyrosine kinase activity of EGFR. Knockout mice expressing the kinase-dead form of GAK (GAK-kd died within 30 min after birth primarily due to respiratory dysfunction. Immunohistochemical analysis revealed that surfactant protein A (SP-A was abundant within alveolar spaces in GAK-kd(+/+ mice but not in GAK-kd(-/- pups. E-cadherin and phosphorylated EGFR signals were also abnormal, suggesting the presence of flat alveolar cells with thin junctions. These results suggest that inhibition of GAK by gefitinib may cause pulmonary alveolar dysfunction, and the present study may help prevent side effects associated with gefitinib therapy in NSCLC patients.

  8. Combining RNA interference and kinase inhibitors against cell signalling components involved in cancer

    International Nuclear Information System (INIS)

    O'Grady, Michael; Raha, Debasish; Hanson, Bonnie J; Bunting, Michaeline; Hanson, George T

    2005-01-01

    The transcription factor activator protein-1 (AP-1) has been implicated in a large variety of biological processes including oncogenic transformation. The tyrosine kinases of the epidermal growth factor receptor (EGFR) constitute the beginning of one signal transduction cascade leading to AP-1 activation and are known to control cell proliferation and differentiation. Drug discovery efforts targeting this receptor and other pathway components have centred on monoclonal antibodies and small molecule inhibitors. Resistance to such inhibitors has already been observed, guiding the prediction of their use in combination therapies with other targeted agents such as RNA interference (RNAi). This study examines the use of RNAi and kinase inhibitors for qualification of components involved in the EGFR/AP-1 pathway of ME180 cells, and their inhibitory effects when evaluated individually or in tandem against multiple components of this important disease-related pathway. AP-1 activation was assessed using an ME180 cell line stably transfected with a beta-lactamase reporter gene under the control of AP-1 response element following epidermal growth factor (EGF) stimulation. Immunocytochemistry allowed for further quantification of small molecule inhibition on a cellular protein level. RNAi and RT-qPCR experiments were performed to assess the amount of knockdown on an mRNA level, and immunocytochemistry was used to reveal cellular protein levels for the targeted pathway components. Increased potency of kinase inhibitors was shown by combining RNAi directed towards EGFR and small molecule inhibitors acting at proximal or distal points in the pathway. After cellular stimulation with EGF and analysis at the level of AP-1 activation using a β-lactamase reporter gene, a 10–12 fold shift or 2.5–3 fold shift toward greater potency in the IC 50 was observed for EGFR and MEK-1 inhibitors, respectively, in the presence of RNAi targeting EGFR. EGFR pathway components were qualified as

  9. Nuclear localization of Src-family tyrosine kinases is required for growth factor-induced euchromatinization

    International Nuclear Information System (INIS)

    Takahashi, Akinori; Obata, Yuuki; Fukumoto, Yasunori; Nakayama, Yuji; Kasahara, Kousuke; Kuga, Takahisa; Higashiyama, Yukihiro; Saito, Takashi; Yokoyama, Kazunari K.; Yamaguchi, Naoto

    2009-01-01

    Src-family kinases (SFKs), which participate in various signaling events, are found at not only the plasma membrane but also several subcellular compartments, including the nucleus. Nuclear structural changes are frequently observed during transcription, cell differentiation, senescence, tumorigenesis, and cell cycle. However, little is known about signal transduction in the alteration of chromatin texture. Here, we develop a pixel imaging method for quantitatively evaluating chromatin structural changes. Growth factor stimulation increases euchromatic hypocondensation and concomitant heterochromatic hypercondensation in G 1 phase, and the levels reach a plateau by 30 min, sustain for at least 5 h and return to the basal levels after 24 h. Serum-activated SFKs in the nucleus were more frequently detected in the euchromatin areas than the heterochromatin areas. Nuclear expression of kinase-active SFKs, but not unrelated Syk kinase, drastically increases both euchromatinization and heterochromatinization in a manner dependent on the levels of nuclear tyrosine phosphorylation. However, growth factor stimulation does not induce chromatin structural changes in SYF cells lacking SFKs, and reintroduction of one SFK member into SYF cells can, albeit insufficiently, induce chromatin structural changes. These results suggest that nuclear tyrosine phosphorylation by SFKs plays an important role in chromatin structural changes upon growth factor stimulation.

  10. Pemetrexed-carboplatin with intercalated icotinib in the treatment of patient with advanced EGFR wild-type lung adenocarcinoma

    Science.gov (United States)

    Xu, Tongpeng; Wu, Hao; Jin, Shidai; Min, Huang; Zhang, Zhihong; Shu, Yongqian; Wen, Wei; Guo, Renhua

    2017-01-01

    Abstract Rationale: Tyrosine kinase inhibitors (TKIs) are known to have greater efficacy in epidermal growth factor receptor (EGFR) mutation nonsmall cell lung cancer (NSCLC). However, about 10% of EGFR wild-type (wt) patients respond to TKIs. Patient concerns: Several strategies to increase the efficacy of TKIs in wt NSCLC are the subjects of ongoing investigations. One of them is combining EGFR TKI with intercalated chemotherapy. Diagnoses: We describe a patient with EGFR wt NSCLC, who was found with ovarian and lung metastasis, was treated with pemetrexed and intercalated icotinib. Interventions: In this case, we reported the successful long-term maintenance treatment of a patient with EGFR wt NSCLC with pemetrexed and Icotinib. The patient (40-year-old female) was found with ovarian masses and lung masses. Pathological, immunohistochemical, and amplification refractory mutation system (ARMS) assay examinations of ovarian specimen suggested the expression of metastatic lung adenocarcinoma with wt EGFR. After failure treatment with paclitaxel-carboplatin, the patient received 4 cycles of pemetrexed plus platinum with intercalated icotinib and then remained on pemetrexed and icotinib. Outcomes: A partial response was achieved after the treatment. The patient's condition had remained stable on pemetrexed and icotinib for more than 20 months, with no evidence of progression. Lessons: To our knowledge, this is the first report using the long-term maintenance treatment with pemetrexed and intercalated icotinib in EGFR wt patient. The therapeutic strategies warrant further exploration in selected populations of NSCLC. PMID:28816950

  11. The selectivity of receptor tyrosine kinase signaling is controlled by a secondary SH2 domain binding site.

    Science.gov (United States)

    Bae, Jae Hyun; Lew, Erin Denise; Yuzawa, Satoru; Tomé, Francisco; Lax, Irit; Schlessinger, Joseph

    2009-08-07

    SH2 domain-mediated interactions represent a crucial step in transmembrane signaling by receptor tyrosine kinases. SH2 domains recognize phosphotyrosine (pY) in the context of particular sequence motifs in receptor phosphorylation sites. However, the modest binding affinity of SH2 domains to pY containing peptides may not account for and likely represents an oversimplified mechanism for regulation of selectivity of signaling pathways in living cells. Here we describe the crystal structure of the activated tyrosine kinase domain of FGFR1 in complex with a phospholipase Cgamma fragment. The structural and biochemical data and experiments with cultured cells show that the selectivity of phospholipase Cgamma binding and signaling via activated FGFR1 are determined by interactions between a secondary binding site on an SH2 domain and a region in FGFR1 kinase domain in a phosphorylation independent manner. These experiments reveal a mechanism for how SH2 domain selectivity is regulated in vivo to mediate a specific cellular process.

  12. Identification of the zinc finger 216 (ZNF216) in human carcinoma cells: a potential regulator of EGFR activity

    Science.gov (United States)

    Mincione, Gabriella; Di Marcantonio, Maria Carmela; Tarantelli, Chiara; Savino, Luca; Ponti, Donatella; Marchisio, Marco; Lanuti, Paola; Sancilio, Silvia; Calogero, Antonella; Di Pietro, Roberta; Muraro, Raffaella

    2016-01-01

    Epidermal Growth Factor Receptor (EGFR), a member of the ErbB family of receptor tyrosine kinase (RTK) proteins, is aberrantly expressed or deregulated in tumors and plays pivotal roles in cancer onset and metastatic progression. ZNF216 gene has been identified as one of Immediate Early Genes (IEGs) induced by RTKs. Overexpression of ZNF216 protein sensitizes 293 cell line to TNF-α induced apoptosis. However, ZNF216 overexpression has been reported in medulloblastomas and metastatic nasopharyngeal carcinomas. Thus, the role of this protein is still not clearly understood. In this study, the inverse correlation between EGFR and ZNF216 expression was confirmed in various human cancer cell lines differently expressing EGFR. EGF treatment of NIH3T3 cells overexpressing both EGFR and ZNF216 (NIH3T3-EGFR/ZNF216), induced a long lasting activation of EGFR in the cytosolic fraction and an accumulation of phosphorylated EGFR (pEGFR) more in the nuclear than in the cytosolic fraction compared to NIH3T3-EGFR cells. Moreover, EGF was able to stimulate an increased expression of ZNF216 in the cytosolic compartment and its nuclear translocation in a time-dependent manner in NIH3T3-EGFR/ZNF216. A similar trend was observed in A431 cells endogenously expressing the EGFR and transfected with Znf216. The increased levels of pEGFR and ZNF216 in the nuclear fraction of NIH3T3-EGFR/ZNF216 cells were paralleled by increased levels of phospho-MAPK and phospho-Akt. Surprisingly, EGF treatment of NIH3T3-EGFR/ZNF216 cells induced a significant increase of apoptosis thus indicating that ZNF216 could sensitize cells to EGF-induced apoptosis and suggesting that it may be involved in the regulation and effects of EGFR signaling. PMID:27732953

  13. An Immunohistochemical Study of Anaplastic Lymphoma Kinase and Epidermal Growth Factor Receptor Mutation in Non-Small Cell Lung Carcinoma.

    Science.gov (United States)

    Verma, Sonal; Kumar, Madhu; Kumari, Malti; Mehrotra, Raj; Kushwaha, R A S; Goel, Madhumati; Kumar, Ashutosh; Kant, Surya

    2017-07-01

    Lung cancer is one of the leading causes of cancer related death. Targeted treatment for specific markers may help in reducing the cancer related morbidity and mortality. To study expression of Anaplastic Lymphoma Kinase (ALK)and Epidermal Growth Factor Receptor (EGFR) mutations in patients of Non-Small Cell Lung Cancer NSCLC, that are the targets for specific ALK inhibitors and EGFR tyrosine kinase inhibitors. Total 69 cases of histologically diagnosed NSCLC were examined retrospectively for immunohistochemical expression of EGFR and ALK, along with positive control of normal placental tissue and anaplastic large cell lymphoma respectively. Of the NSCLC, Squamous Cell Carcinoma (SCC) accounted for 71.0% and adenocarcinoma was 26.1%. ALK expression was seen in single case of 60-year-old female, non-smoker with adenocarcinoma histology. EGFR expression was seen in both SCC (59.18%) and adenocarcinoma in (77.78%) accounting for 63.77% of all cases. Both ALK and EGFR mutation were mutually exclusive. EGFR expression was seen in 63.77% of cases, highlighting the importance of its use in routine analysis, for targeted therapy and better treatment results. Although, ALK expression was seen in 1.45% of all cases, it is an important biomarker in targeted cancer therapy. Also, the mutually exclusive expression of these two markers need further studies to develop a diagnostic algorithm for NSCLC patients.

  14. Research progress on criteria for discontinuation of EGFR inhibitor therapy

    Directory of Open Access Journals (Sweden)

    Zhuang HQ

    2012-10-01

    Full Text Available Hong-qing Zhuang, Zhi-yong Yuan, Jun Wang, Ping Wang, Lu-jun Zhao, Bai-lin ZhangDepartment of Radiotherapy, Tianjin Medical University Cancer Institute and Hospital, Tianjin Key Laboratory of Cancer Prevention and Therapy, Tianjin Lung Cancer Center, Tianjin, People's Republic of ChinaAbstract: The clinical success of the epidermal growth factor receptor (EGFR tyrosine kinase inhibitors (TKI as therapeutic agents has prompted great interest in their further development and clinical testing for a wide variety of malignancies. However, most studies have focused on the efficacy of TKI, and few studies have been done on the criteria for their discontinuation. The current standard for drug discontinuation is “until progression”, based on change in tumor size. However, tumor size is not related to the gene expression which determines the efficacy of TKI in the final analysis, and it is also difficult to make a thorough and correct prediction based on tumor size when the TKI is discontinued. Nevertheless, clinical evaluation of the criteria for TKI discontinuation is still in its early days. Some promising findings have started to emerge. With the improving knowledge of EGFR and its inhibitors, it is expected that the criteria for discontinuation of EGFR inhibitor therapy will become clearer.Keywords: epidermal growth factor receptor, drug discontinuation, acquired drug-resistance

  15. Receptor Tyrosine Kinases as Targets for Treatment of Peripheral Nerve Sheath Tumors in NF 1 Patients

    National Research Council Canada - National Science Library

    Mautner, Victor-Felix

    2007-01-01

    .... The only available but unsatisfying therapy is surgical tumor resection. The purpose of this study is the preclinical testing of multiple available tyrosine kinase inhibitors for NF1-associated MPNST using in vitro and in vivo systems...

  16. Pemetrexed-carboplatin with intercalated icotinib in the treatment of patient with advanced EGFR wild-type lung adenocarcinoma: A case report.

    Science.gov (United States)

    Xu, Tongpeng; Wu, Hao; Jin, Shidai; Min, Huang; Zhang, Zhihong; Shu, Yongqian; Wen, Wei; Guo, Renhua

    2017-08-01

    Tyrosine kinase inhibitors (TKIs) are known to have greater efficacy in epidermal growth factor receptor (EGFR) mutation nonsmall cell lung cancer (NSCLC). However, about 10% of EGFR wild-type (wt) patients respond to TKIs. Several strategies to increase the efficacy of TKIs in wt NSCLC are the subjects of ongoing investigations. One of them is combining EGFR TKI with intercalated chemotherapy. We describe a patient with EGFR wt NSCLC, who was found with ovarian and lung metastasis, was treated with pemetrexed and intercalated icotinib. In this case, we reported the successful long-term maintenance treatment of a patient with EGFR wt NSCLC with pemetrexed and Icotinib. The patient (40-year-old female) was found with ovarian masses and lung masses. Pathological, immunohistochemical, and amplification refractory mutation system (ARMS) assay examinations of ovarian specimen suggested the expression of metastatic lung adenocarcinoma with wt EGFR. After failure treatment with paclitaxel-carboplatin, the patient received 4 cycles of pemetrexed plus platinum with intercalated icotinib and then remained on pemetrexed and icotinib. A partial response was achieved after the treatment. The patient's condition had remained stable on pemetrexed and icotinib for more than 20 months, with no evidence of progression. To our knowledge, this is the first report using the long-term maintenance treatment with pemetrexed and intercalated icotinib in EGFR wt patient. The therapeutic strategies warrant further exploration in selected populations of NSCLC.

  17. Tyrosine kinase inhibitors for brain metastases in HER2-positive breast cancer.

    Science.gov (United States)

    Duchnowska, Renata; Loibl, Sibylle; Jassem, Jacek

    2018-06-01

    Approximately 30-50% of advanced HER2-positive breast cancer patients will develop central nervous system (CNS) metastases, with an annual risk of around 10%, and a half of them will die from brain progression. An increased risk of brain metastases is also seen in patients with early HER2-positive breast cancer administered curative therapy. Brain metastases in HER2-positive breast cancer patients usually constitute the first site of recurrence. The administration of anti-HER2 monoclonal antibodies, trastuzumab and pertuzumab, considerably delays the onset of symptomatic brain disease: however, the limited penetration of these compounds into the CNS hinders their efficacy. The small-molecule tyrosine kinase inhibitors of epidermal growth factor receptors family have established activity in HER2-positive breast cancer in both advanced disease and neoadjuvant setting. Favorable physico-chemical properties of these compounds allow them for a more efficient penetration through the blood-brain barrier, and hold the promise for more effective prevention and treatment of brain metastases. In this article we review the role of currently available or investigational HER2 tyrosine kinase inhibitors: lapatinib, neratinib, afatinib and tucatinib in the treatment of brain metastases in HER2-positive breast cancer patients. Copyright © 2018 Elsevier Ltd. All rights reserved.

  18. Internalization and down-regulation of the human epidermal growth factor receptor are regulated by the carboxyl-terminal tyrosines

    DEFF Research Database (Denmark)

    Helin, K; Beguinot, L

    1991-01-01

    with receptors in which 1, 2, or all 3 tyrosines were changed to phenylalanines. The triple point mutant EGF-R, expressed in NIH-3T3, exhibited low autophosphorylation in vivo, low biological and reduced kinase activities. Single and double point mutants were down-regulated, as well as wild type EGF......-R in response to EGF showing a half-life of about 1 h. Degradation of the triple point mutant, however, was impaired and resulted in a half-life of 4 h in the presence of EGF. EGF-dependent down-regulation of surface receptors was decreased in the triple point mutant EGF-R as was internalization and degradation...... of EGF. The specific rate of internalization of the triple point mutant was reduced. By contrast, intracellular processing of ligand previously internalized at 20 degrees C was similar between wild type and mutant receptors. Taken together the data indicate that the delay in degradation observed in cells...

  19. Contribution of EGFR and ErbB-3 Heterodimerization to the EGFR Mutation-Induced Gefitinib- and Erlotinib-Resistance in Non-Small-Cell Lung Carcinoma Treatments.

    Directory of Open Access Journals (Sweden)

    Debby D Wang

    Full Text Available EGFR mutation-induced drug resistance has become a major threat to the treatment of non-small-cell lung carcinoma. Essentially, the resistance mechanism involves modifications of the intracellular signaling pathways. In our work, we separately investigated the EGFR and ErbB-3 heterodimerization, regarded as the origin of intracellular signaling pathways. On one hand, we combined the molecular interaction in EGFR heterodimerization with that between the EGFR tyrosine kinase and its inhibitor. For 168 clinical subjects, we characterized their corresponding EGFR mutations using molecular interactions, with three potential dimerization partners (ErbB-2, IGF-1R and c-Met of EGFR and two of its small molecule inhibitors (gefitinib and erlotinib. Based on molecular dynamics simulations and structural analysis, we modeled these mutant-partner or mutant-inhibitor interactions using binding free energy and its components. As a consequence, the mutant-partner interactions are amplified for mutants L858R and L858R_T790M, compared to the wild type EGFR. Mutant delL747_P753insS represents the largest difference between the mutant-IGF-1R interaction and the mutant-inhibitor interaction, which explains the shorter progression-free survival of an inhibitor to this mutant type. Besides, feature sets including different energy components were constructed, and efficient regression trees were applied to map these features to the progression-free survival of an inhibitor. On the other hand, we comparably examined the interactions between ErbB-3 and its partners (EGFR mutants, IGF-1R, ErbB-2 and c-Met. Compared to others, c-Met shows a remarkably-strong binding with ErbB-3, implying its significant role in regulating ErbB-3 signaling. Moreover, EGFR mutants corresponding to poor clinical outcomes, such as L858R_T790M, possess lower binding affinities with ErbB-3 than c-Met does. This may promote the communication between ErbB-3 and c-Met in these cancer cells. The

  20. Monitoring of high-density lipoprotein cholesterol level is predictive of EGFR mutation and efficacy of EGFR-TKI in patients with advanced lung adenocarcinoma

    Directory of Open Access Journals (Sweden)

    Lv Y

    2016-01-01

    Full Text Available Yang Lv,1,2 Li-Yun Miao,2 Qiu-Fang Chen,1 Yan Li,2 Zhi-Xiang Shi,1 Xuan-Sheng Ding1 1Department of Clinical Pharmacy, China Pharmaceutical University, Nanjing, Jiangsu, People’s Republic of China; 2Division of Respiratory Medicine, Department of Respiration, The Affiliated Drum Tower Hospital of Nanjing University Medical College, Nanjing University Medical School, Nanjing, Jiangsu, People’s Republic of China Abstract: High-density lipoprotein cholesterol (HDL-C has an inverse association with the incidence of lung cancer. However, whether it can be used as a predictive factor in advanced lung adenocarcinoma patients treated with epidermal growth factor receptor (EGFR tyrosine kinase inhibitors (TKI still remains undefined. This research aimed at studying the relationship of serum HDL-C baseline level and HDL-C kinetics to EGFR mutation, the efficacy of EGFR-TKI, and the predictive value of PFS. The presence of mutation rate in the 192 patients with lung adenocarcinoma was compared within stratified groups. Levels of baseline HDL-C and kinetics of HDL-C were analyzed retrospectively in patients treated with EGFR-TKI harboring EGFR mutation. Univariate and multivariate analyses were performed to investigate the prognostic value of HDL-C. EGFR mutation rate of HDL-C high-level group was significantly higher than that of low-level group (59.0% vs 35.6%, P=0.001. Multivariate logistic analysis showed that high-level HDL-C was an independent predictive factor for EGFR gene mutation (P=0.005; odds ratio =0.417; 95% confidence interval [CI], 0.227–0.768. Patients with a low level of HDL-C before therapy showed a progression of disease in most cases (P<0.001. According to HDL-C kinetics, patients who received EGFR-TKI treatment harboring EGFR mutation were divided into four groups. Univariate analysis showed that patients in nondecreased group had longer progression-free survival (P<0.001; hazard ratio =0.003; 95% CI, 0.001–0.018. Multivariate

  1. The significance of major and stable molecular responses in chronic myeloid leukemia in the tyrosine kinase inhibitor era

    Directory of Open Access Journals (Sweden)

    Ilana Zalcberg Renault

    2011-12-01

    Full Text Available Tyrosine kinase inhibitors have changed the management and outcomes of chronic myeloid leukemia patients. Quantitative polymerase chain reaction is used to monitor molecular responses to tyrosine kinase inhibitors. Molecular monitoring represents the most sensitive tool to judge chronic myeloid leukemia disease course and allows early detection of relapse. Evidence of achieving molecular response is important for several reasons: 1. early molecular response is associated with major molecular response rates at 18-24 months; 2. patients achieving major molecular response are less likely to lose their complete cytogenetic response; 3. a durable, stable major molecular response is associated with increased progression-free survival. However, standardization of molecular techniques is still challenging.

  2. The significance of major and stable molecular responses in chronic myeloid leukemia in the tyrosine kinase inhibitor era

    Science.gov (United States)

    Renault, Ilana Zalcberg; Scholl, Vanesa; Hassan, Rocio; Capelleti, Paola; de Lima, Marcos; Cortes, Jorge

    2011-01-01

    Tyrosine kinase inhibitors have changed the management and outcomes of chronic myeloid leukemia patients. Quantitative polymerase chain reaction is used to monitor molecular responses to tyrosine kinase inhibitors. Molecular monitoring represents the most sensitive tool to judge chronic myeloid leukemia disease course and allows early detection of relapse. Evidence of achieving molecular response is important for several reasons: 1. early molecular response is associated with major molecular response rates at 18-24 months; 2. patients achieving major molecular response are less likely to lose their complete cytogenetic response; 3. a durable, stable major molecular response is associated with increased progression-free survival. However, standardization of molecular techniques is still challenging. PMID:23049363

  3. Spectrum of EGFR gene mutations in Vietnamese patients with non-small cell lung cancer.

    Science.gov (United States)

    Vu, Hoang Anh; Xinh, Phan Thi; Ha, Hua Thi Ngoc; Hanh, Ngo Thi Tuyet; Bach, Nguyen Duc; Thao, Doan Thi Phuong; Dat, Ngo Quoc; Trung, Nguyen Sao

    2016-03-01

    Epidermal growth factor receptor (EGFR) mutational status is a crucial biomarker for prediction of response to tyrosine kinase inhibitors in patients with non-small cell lung cancer (NSCLC). Although these mutations have been well characterized in other countries, little is known about the frequency or spectrum of EGFR mutations in Vietnamese NSCLC patients. Using Sanger DNA sequencing, we investigated mutations in EGFR exons 18-21 from 332 patients diagnosed with NSCLC at University of Medicine and Pharmacy, Ho Chi Minh City, Vietnam. DNA was extracted from formalin-fixed, paraffin-embedded tissues, followed by PCR amplification and sequencing. EGFR mutations were detected in 135 samples (40.7%), of which eight samples carried double mutations. In total, 46 different types of EGFR mutations were found, including six novel mutations (p.K713E, p.K714R, p.P794S, p.R803W, p.P848S, and p.K867E). Among the four exons investigated, exon 19 was most frequently mutated (63 out of 332 patients, 19%), with the p.E746_A750del appearing in 43 samples. Exon 21 was mutated in 56 samples (16.9%), of which 47 were p.L858R. Each of exons 18 and 20 was mutated in 12 samples (3.6%). The frequency of EGFR mutations was higher in females than in males (48.9% vs 35%, P = 0.012), but not statistically different between adenocarcinomas and other histological types of NSCLC (41.3% vs 34.5%, P = 0.478). DNA sequencing detected EGFR mutations with high frequency and revealed a broad spectrum of mutation type in Vietnamese patients with NSCLC. © 2015 Wiley Publishing Asia Pty Ltd.

  4. Multi-tyrosine kinase inhibitors in preclinical studies for pediatric CNS AT/RT: Evidence for synergy with Topoisomerase-I inhibition

    Directory of Open Access Journals (Sweden)

    Jayanthan Aarthi

    2011-12-01

    Full Text Available Abstract Background Currently, Atypical Teratoid Rhabdoid Tumor (AT/RT constitutes one of the most difficult to treat malignancies in pediatrics. Hence, new knowledge of potential targets for therapeutics and the development of novel treatment approaches are urgently needed. We have evaluated the presence of cytokine pathways and the effects of two clinically available multi-tyrosine kinase inhibitors for cytotoxicity, target modulation and drug combinability against AT/RT cell lines. Results AT/RT cell lines expressed measurable quantities of VEGF, FGF, PDGF and SDF-1, although the absolute amounts varied between the cell lines. The targeted receptor tyrosine kinase inhibitor sorafenib inhibited the key signaling molecule Erk, which was activated following the addition of own conditioned media, suggesting the existence of autocrine/paracrine growth stimulatory pathways. The multi-tyrosine kinase inhibitors sorafenib and sunitinib also showed significant growth inhibition of AT/RT cells and their activity was enhanced by combination with the topoisomerase inhibitor, irinotecan. The loss of cytoplasmic NF-kappa-B in response to irinotecan was diminished by sorafenib, providing evidence for a possible benefit for this drug combination. Conclusions In addition to previously described involvement of insulin like growth factor (IGF family of cytokines, a multitude of other growth factors may contribute to the growth and survival of AT/RT cells. However, consistent with the heterogeneous nature of this tumor, quantitative and qualitative differences may exist among different tumor samples. Multi-tyrosine kinase inhibitors appear to have effective antitumor activity against all cell lines studied. In addition, the target modulation studies and drug combinability data provide the groundwork for additional studies and support the evaluation of these agents in future treatment protocols.

  5. Greater Sensitivity of Blood Pressure Than Renal Toxicity to Tyrosine Kinase Receptor Inhibition With Sunitinib

    DEFF Research Database (Denmark)

    Lankhorst, Stephanie; Baelde, Hans J; Kappers, Mariëtte H W

    2015-01-01

    Hypertension and renal injury are off-target effects of sunitinib, a tyrosine kinase receptor inhibitor used for the treatment of various tumor types. Importantly, these untoward effects are accompanied by activation of the endothelin system. Here, we set up a study to explore the dose dependency...

  6. A cross-talk between TrkB and Ret tyrosine kinases receptors mediates neuroblastoma cells differentiation.

    Directory of Open Access Journals (Sweden)

    Carla Lucia Esposito

    Full Text Available Understanding the interplay between intracellular signals initiated by multiple receptor tyrosine kinases (RTKs to give the final cell phenotype is a major pharmacological challenge. Retinoic acid (RA-treatment of neuroblastoma (NB cells implicates activation of Ret and TrkB RTKs as critical step to induce cell differentiation. By studying the signaling interplay between TrkB and Ret as paradigmatic example, here we demonstrate the existence of a cross-talk mechanism between the two unrelated receptors that is needed to induce the cell differentiation. Indeed, we show that TrkB receptor promotes Ret phosphorylation by a mechanism that does not require GDNF. This reveals to be a key mechanism, since blocking either TrkB or Ret by small interfering RNA causes a failure in NB biochemical and morphological differentiation. Our results provide the first evidence that a functional transactivation between distinct tyrosine kinases receptors is required for an important physiological process.

  7. Targeting Bruton Tyrosine Kinase: A novel strategy in the treatment of B-cell lymphomas

    Directory of Open Access Journals (Sweden)

    Sklavenitis-Pistofidis R.

    2017-06-01

    Full Text Available In normal B-cells, Bruton tyrosine kinase (Btk, a non-receptor tyrosine kinase involved in B-cell receptor (BCR signalling, is essential for cell survival and maturation. Not surprisingly, Btk is also implicated in the pathogenesis of B-cell lymphomas, like Chronic Lymphocytic Leukaemia/Small Lymphocytic Lymphoma (CLL/SLL, Mantle Cell Lymphoma (MCL and Waldenström’s Macroglobulinemia (WM, which are driven by aberrant BCR signalling. Thus, targeting Btk represents a promising therapeutic strategy in the treatment of B-cell lymphoma patients. Ibrutinib, a selective Btk inhibitor, has already been approved as second-line treatment of CLL/SLL, MCL and WM patients, while more clinical studies of ibrutinib and novel Btk inhibitors are currently under way. In light of results of the RESONATE-2 trial, the approval of ibrutinib as a first-line treatment of CLL/SLL may well be approaching. Herein, we review Btk’s role in normal and malignant BCR signalling, as well as ibrutinib’s performance in B-cell lymphoma treatment and prognosis.

  8. mTORC2 promotes type I insulin-like growth factor receptor and insulin receptor activation through the tyrosine kinase activity of mTOR.

    Science.gov (United States)

    Yin, Yancun; Hua, Hui; Li, Minjing; Liu, Shu; Kong, Qingbin; Shao, Ting; Wang, Jiao; Luo, Yuanming; Wang, Qian; Luo, Ting; Jiang, Yangfu

    2016-01-01

    Mammalian target of rapamycin (mTOR) is a core component of raptor-mTOR (mTORC1) and rictor-mTOR (mTORC2) complexes that control diverse cellular processes. Both mTORC1 and mTORC2 regulate several elements downstream of type I insulin-like growth factor receptor (IGF-IR) and insulin receptor (InsR). However, it is unknown whether and how mTOR regulates IGF-IR and InsR themselves. Here we show that mTOR possesses unexpected tyrosine kinase activity and activates IGF-IR/InsR. Rapamycin induces the tyrosine phosphorylation and activation of IGF-IR/InsR, which is largely dependent on rictor and mTOR. Moreover, mTORC2 promotes ligand-induced activation of IGF-IR/InsR. IGF- and insulin-induced IGF-IR/InsR phosphorylation is significantly compromised in rictor-null cells. Insulin receptor substrate (IRS) directly interacts with SIN1 thereby recruiting mTORC2 to IGF-IR/InsR and promoting rapamycin- or ligand-induced phosphorylation of IGF-IR/InsR. mTOR exhibits tyrosine kinase activity towards the general tyrosine kinase substrate poly(Glu-Tyr) and IGF-IR/InsR. Both recombinant mTOR and immunoprecipitated mTORC2 phosphorylate IGF-IR and InsR on Tyr1131/1136 and Tyr1146/1151, respectively. These effects are independent of the intrinsic kinase activity of IGF-IR/InsR, as determined by assays on kinase-dead IGF-IR/InsR mutants. While both rictor and mTOR immunoprecitates from rictor(+/+) MCF-10A cells exhibit tyrosine kinase activity towards IGF-IR and InsR, mTOR immunoprecipitates from rictor(-/-) MCF-10A cells do not induce IGF-IR and InsR phosphorylation. Phosphorylation-deficient mutation of residue Tyr1131 in IGF-IR or Tyr1146 in InsR abrogates the activation of IGF-IR/InsR by mTOR. Finally, overexpression of rictor promotes IGF-induced cell proliferation. Our work identifies mTOR as a dual-specificity kinase and clarifies how mTORC2 promotes IGF-IR/InsR activation.

  9. Prevention of pulmonary vascular and myocardial remodeling by the combined tyrosine and serine-/threonine kinase inhibitor, sorafenib, in pulmonary hypertension and right heart failure

    Directory of Open Access Journals (Sweden)

    M. Klein

    2008-06-01

    Full Text Available Inhibition of tyrosine kinases can reverse pulmonary hypertension but little is known about the role of serine-/threonine kinases in vascular and myocardial remodeling. We investigated the effects of sorafenib, an inhibitor of the tyrosine kinases VEGFR, PDGFR and c-kit as well as the serine-/threonine kinase Raf-1, in pulmonary hypertension and right ventricular (RV pressure overload. In monocrotaline treated rats, sorafenib (10 mg·kg–1·d–1 p.o. reduced pulmonary arterial pressure, pulmonary artery muscularization and RV hypertrophy, and improved systemic hemodynamics (table 1. Sorafenib prevented phosphorylation of Raf-1 and suppressed activation of downstream signaling pathways (Erk 1/2. After pulmonary banding, sorafenib, but not the PDGFR/c-KIT/ABL-inhibitor imatinib reduced RV mass and RV filling pressure significantly. Congruent with these results, sorafenib only prevented ERK phosphorylation and vasopressin induced hypertrophy of the cardiomyocyte cell line H9c2 dose dependently (IC50 = 300 nM. Combined inhibition of tyrosine and serine-/threonine kinases by sorafenib prevents vascular and cardiac remodeling in pulmonary hypertension, which is partly mediated via inhibition of the Raf kinase pathway.

  10. Mutational analysis of EGFR and related signaling pathway genes in lung adenocarcinomas identifies a novel somatic kinase domain mutation in FGFR4.

    Directory of Open Access Journals (Sweden)

    Jenifer L Marks

    2007-05-01

    Full Text Available Fifty percent of lung adenocarcinomas harbor somatic mutations in six genes that encode proteins in the EGFR signaling pathway, i.e., EGFR, HER2/ERBB2, HER4/ERBB4, PIK3CA, BRAF, and KRAS. We performed mutational profiling of a large cohort of lung adenocarcinomas to uncover other potential somatic mutations in genes of this signaling pathway that could contribute to lung tumorigenesis.We analyzed genomic DNA from a total of 261 resected, clinically annotated non-small cell lung cancer (NSCLC specimens. The coding sequences of 39 genes were screened for somatic mutations via high-throughput dideoxynucleotide sequencing of PCR-amplified gene products. Mutations were considered to be somatic only if they were found in an independent tumor-derived PCR product but not in matched normal tissue. Sequencing of 9MB of tumor sequence identified 239 putative genetic variants. We further examined 22 variants found in RAS family genes and 135 variants localized to exons encoding the kinase domain of respective proteins. We identified a total of 37 non-synonymous somatic mutations; 36 were found collectively in EGFR, KRAS, BRAF, and PIK3CA. One somatic mutation was a previously unreported mutation in the kinase domain (exon 16 of FGFR4 (Glu681Lys, identified in 1 of 158 tumors. The FGFR4 mutation is analogous to a reported tumor-specific somatic mutation in ERBB2 and is located in the same exon as a previously reported kinase domain mutation in FGFR4 (Pro712Thr in a lung adenocarcinoma cell line.This study is one of the first comprehensive mutational analyses of major genes in a specific signaling pathway in a sizeable cohort of lung adenocarcinomas. Our results suggest the majority of gain-of-function mutations within kinase genes in the EGFR signaling pathway have already been identified. Our findings also implicate FGFR4 in the pathogenesis of a subset of lung adenocarcinomas.

  11. Serum concentrations of nitrite and malondialdehyde as markers of oxidative stress in chronic myeloid leukemia patients treated with tyrosine kinase inhibitors

    Directory of Open Access Journals (Sweden)

    Maria Juracy Petrola

    2012-01-01

    Full Text Available BACKGROUND: Chronic myeloid leukemia is a neoplasm characterized by clonal expansion of hematopoietic progenitor cells resulting from the (9:22(q34,11 translocation. The tyrosine kinase abl fusion protein,the initial leukemogenic event in chronic myeloid leukemia, is constitutively activated thus inducing the production of reactive oxygen species. Of particular relevance is the fact that an increase in reactive oxygen species can facilitate genomic instability and may contribute to disease progression. OBJETIVE: To evaluate oxidative stress by determining the levels of malondialdehyde and nitrite in chronic myeloid leukemia patients under treatment with 1st and 2nd generation tyrosine kinase inhibitors monitored at a referral hospital in Fortaleza, Ceará. METHODS: A cross-sectional study was performed of 64 male and female adults. Patients were stratified according to treatment. The levels of malondialdehyde and nitrite were determined by spectrophotometry. Statistical differences between groups were observed using the Student t-test and Fisher's exact test. The results are expressed as mean ± standard error of mean. The significance level was set for a p-value < 0.05 in all analyses. RESULTS: The results show significantly higher mean concentrations of nitrite and malondialdehyde in chronic myeloid leukemia patients using second-generation tyrosine kinase inhibitors compared to patients on imatinib. Conclusion: It follows that chronic myeloid leukemia patients present higher oxidative activity and that the increases in oxidative damage markers can indicate resistance to 1st generation tyrosine kinase inhibitors.

  12. Network meta-analysis of erlotinib, gefitinib, afatinib and icotinib in patients with advanced non-small-cell lung cancer harboring EGFR mutations.

    Science.gov (United States)

    Liang, Wenhua; Wu, Xuan; Fang, Wenfeng; Zhao, Yuanyuan; Yang, Yunpeng; Hu, Zhihuang; Xue, Cong; Zhang, Jing; Zhang, Jianwei; Ma, Yuxiang; Zhou, Ting; Yan, Yue; Hou, Xue; Qin, Tao; Dinglin, Xiaoxiao; Tian, Ying; Huang, Peiyu; Huang, Yan; Zhao, Hongyun; Zhang, Li

    2014-01-01

    Several EGFR-tyrosine kinase inhibitors (EGFR-TKIs) including erlotinib, gefitinib, afatinib and icotinib are currently available as treatment for patients with advanced non-small-cell lung cancer (NSCLC) who harbor EGFR mutations. However, no head to head trials between these TKIs in mutated populations have been reported, which provides room for indirect and integrated comparisons. We searched electronic databases for eligible literatures. Pooled data on objective response rate (ORR), progression free survival (PFS), overall survival (OS) were calculated. Appropriate networks for different outcomes were established to incorporate all evidences. Multiple-treatments comparisons (MTCs) based on Bayesian network integrated the efficacy and specific toxicities of all included treatments. Twelve phase III RCTs that investigated EGFR-TKIs involving 1821 participants with EGFR mutation were included. For mutant patients, the weighted pooled ORR and 1-year PFS of EGFR-TKIs were significant superior to that of standard chemotherapy (ORR: 66.6% vs. 30.9%, OR 5.46, 95%CI 3.59 to 8.30, Picotinib (19%, 29%, NA, NA), respectively. However, afatinib and erlotinib showed significant severer rash and diarrhea compared with gefitinib and icotinib. The current study indicated that erlotinib, gefitinib, afatinib and icotinib shared equivalent efficacy but presented different efficacy-toxicity pattern for EGFR-mutated patients. Erlotinib and afatinib revealed potentially better efficacy but significant higher toxicities compared with gefitinib and icotinib.

  13. Tyrosine kinase signalling in breast cancer: Modulation of tyrosine kinase signalling in human breast cancer through altered expression of signalling intermediates

    International Nuclear Information System (INIS)

    Kairouz, Rania; Daly, Roger J

    2000-01-01

    The past decade has seen the definition of key signalling pathways downstream of receptor tyrosine kinases (RTKs) in terms of their components and the protein-protein interactions that facilitate signal transduction. Given the strong evidence that links signalling by certain families of RTKs to the progression of breast cancer, it is not surprising that the expression profile of key downstream signalling intermediates in this disease has also come under scrutiny, particularly because some exhibit transforming potential or amplify mitogenic signalling pathways when they are overexpressed. Reflecting the diverse cellular processes regulated by RTKs, it is now clear that altered expression of such signalling proteins in breast cancer may influence not only cellular proliferation (eg Grb2) but also the invasive properties of the cancer cells (eg EMS1/cortactin)

  14. Fragment-based lead discovery of small molecule inhibitors for the EPHA4 receptor tyrosine kinase

    NARCIS (Netherlands)

    van Linden, O.P.J.; Farenc, C; Zoutman, W.H.; Hameetman, L; Wijtmans, M.; Leurs, R.; Tensen, C.P.; Siegal, G.; de Esch, I.J.P.

    2011-01-01

    The in silico identification, optimization and crystallographic characterization of a 6,7,8,9-tetrahydro-3H-pyrazolo[3,4-c]isoquinolin-1-amine scaffold as an inhibitor for the EPHA4 receptor tyrosine kinase is described. A database containing commercially available compounds was subjected to an in

  15. Alternative signaling pathways as potential therapeutic targets for overcoming EGFR and c-Met inhibitor resistance in non-small cell lung cancer.

    Directory of Open Access Journals (Sweden)

    Jason T Fong

    Full Text Available The use of tyrosine kinase inhibitors (TKIs against EGFR/c-Met in non-small cell lung cancer (NSCLC has been shown to be effective in increasing patient progression free survival (PFS, but their efficacy is limited due to the development of resistance and tumor recurrence. Therefore, understanding the molecular mechanisms underlying development of drug resistance in NSCLC is necessary for developing novel and effective therapeutic approaches to improve patient outcome. This study aims to understand the mechanism of EGFR/c-Met tyrosine kinase inhibitor (TKI resistance in NSCLC. H2170 and H358 cell lines were made resistant to SU11274, a c-Met inhibitor, and erlotinib, an EGFR inhibitor, through step-wise increases in TKI exposure. The IC50 concentrations of resistant lines exhibited a 4-5 and 11-22-fold increase for SU11274 and erlotinib, respectively, when compared to parental lines. Furthermore, mTOR and Wnt signaling was studied in both cell lines to determine their roles in mediating TKI resistance. We observed a 2-4-fold upregulation of mTOR signaling proteins and a 2- to 8-fold upregulation of Wnt signaling proteins in H2170 erlotinib and SU11274 resistant cells. H2170 and H358 cells were further treated with the mTOR inhibitor everolimus and the Wnt inhibitor XAV939. H358 resistant cells were inhibited by 95% by a triple combination of everolimus, erlotinib and SU11274 in comparison to 34% by a double combination of these drugs. Parental H2170 cells displayed no sensitivity to XAV939, while resistant cells were significantly inhibited (39% by XAV939 as a single agent, as well as in combination with SU11274 and erlotinib. Similar results were obtained with H358 resistant cells. This study suggests a novel molecular mechanism of drug resistance in lung cancer.

  16. Downsizing Treatment with Tyrosine Kinase Inhibitors in Patients with Advanced Gastrointestinal Stromal Tumors Improved Resectability

    Science.gov (United States)

    Sjölund, Katarina; Andersson, Anna; Nilsson, Erik; Nilsson, Ola; Ahlman, Håkan

    2010-01-01

    Background Gastrointestinal stromal tumors (GISTs) express the receptor tyrosine kinase KIT. Most GISTs have mutations in the KIT or PDGFRA gene, causing activation of tyrosine kinase. Imatinib, a tyrosine kinase inhibitor (TKI), is the first-line palliative treatment for advanced GISTs. Sunitinib was introduced for patients with mutations not responsive to imatinib. The aim was to compare the survival of patients with high-risk resected GISTs treated with TKI prior to surgery with historical controls and to determine if organ-preserving surgery was facilitated. Methods Ten high-risk GIST-patients had downsizing/adjuvant TKI treatment: nine with imatinib and one with sunitinib. The patients were matched with historical controls (n = 89) treated with surgery alone, from our population-based series (n = 259). Mutational analysis of KIT and PDGFRA was performed in all cases. The progression-free survival was calculated. Results The primary tumors decreased in mean diameter from 20.4 cm to 10.5 cm on downsizing imatinib. Four patients with R0 resection and a period of adjuvant imatinib had no recurrences versus 67% in the historical control group. Four patients with residual liver metastases have stable disease on continuous imatinib treatment after surgery. One patient has undergone reoperation with liver resection. The downsizing treatment led to organ-preserving surgery in nine patients and improved preoperative nutritional status in one patient. Conclusions Downsizing TKI is recommended for patients with bulky tumors with invasion of adjacent organs. Sunitinib can be used for patients in case of imatinib resistance (e.g., wild-type GISTs), underlining the importance of mutational analysis for optimal surgical planning. PMID:20512492

  17. Next generation of adeno-associated virus 2 vectors: Point mutations in tyrosines lead to high-efficiency transduction at lower doses

    Science.gov (United States)

    Zhong, Li; Li, Baozheng; Mah, Cathryn S.; Govindasamy, Lakshmanan; Agbandje-McKenna, Mavis; Cooper, Mario; Herzog, Roland W.; Zolotukhin, Irene; Warrington, Kenneth H.; Weigel-Van Aken, Kirsten A.; Hobbs, Jacqueline A.; Zolotukhin, Sergei; Muzyczka, Nicholas; Srivastava, Arun

    2008-01-01

    Recombinant adeno-associated virus 2 (AAV2) vectors are in use in several Phase I/II clinical trials, but relatively large vector doses are needed to achieve therapeutic benefits. Large vector doses also trigger an immune response as a significant fraction of the vectors fails to traffic efficiently to the nucleus and is targeted for degradation by the host cell proteasome machinery. We have reported that epidermal growth factor receptor protein tyrosine kinase (EGFR-PTK) signaling negatively affects transduction by AAV2 vectors by impairing nuclear transport of the vectors. We have also observed that EGFR-PTK can phosphorylate AAV2 capsids at tyrosine residues. Tyrosine-phosphorylated AAV2 vectors enter cells efficiently but fail to transduce effectively, in part because of ubiquitination of AAV capsids followed by proteasome-mediated degradation. We reasoned that mutations of the surface-exposed tyrosine residues might allow the vectors to evade phosphorylation and subsequent ubiquitination and, thus, prevent proteasome-mediated degradation. Here, we document that site-directed mutagenesis of surface-exposed tyrosine residues leads to production of vectors that transduce HeLa cells ≈10-fold more efficiently in vitro and murine hepatocytes nearly 30-fold more efficiently in vivo at a log lower vector dose. Therapeutic levels of human Factor IX (F.IX) are also produced at an ≈10-fold reduced vector dose. The increased transduction efficiency of tyrosine-mutant vectors is due to lack of capsid ubiquitination and improved intracellular trafficking to the nucleus. These studies have led to the development of AAV vectors that are capable of high-efficiency transduction at lower doses, which has important implications in their use in human gene therapy. PMID:18511559

  18. EGFR Activation and Ultraviolet Light‐Induced Skin Carcinogenesis

    Directory of Open Access Journals (Sweden)

    Taghrid B. El-Abaseri

    2007-01-01

    Full Text Available The epidermal growth factor receptor (EGFR regulates the proliferation of keratinocytes through multiple mechanisms that differ depending on the localization of the cell within the skin. Ultraviolet (UV irradiation, the main etiologic factor in the development of skin cancer, also activates the receptor. In this review, we discuss how the UV-induced activation of EGFR regulates the response of the skin to UV. UV-induced EGFR activation increases keratinocyte proliferation, suppresses apoptosis, and augments and accelerates epidermal hyperplasia in response to UV. Pharmacological inhibition of the UV-induced activation of EGFR in a genetically initiated mouse skin tumorigenesis model suppresses tumorigenesis and the activation of mitogen-activated protein (MAP kinases and phosphatidyl inositol-3-kinase (PI3K/AKT signaling pathways. EGFR has pleiotropic, complex, and cell-type-specific functions in cutaneous keratinocytes; suggesting that the receptor is an appropriate target for the development of molecularly targeted therapies for skin cancer and other pathologies.

  19. Clinical Observation of Icotinib Hydrochloride for Advanced Non-small Cell Lung Cancer Patients with EGFR Status Identified

    Directory of Open Access Journals (Sweden)

    Xi LI

    2015-12-01

    Full Text Available Background and objective Icotinib is the first self-developed small molecular drug in China for targeted therapy of lung cancer. Compared to the other two commercially available epidermal growth factor receptor (EGFR tyrosine kinase inhibitors, gefitinib and erlotinib, icotinib is similar to them in chemical structure, mechanism of activity and therapeutic effects. To explore the efficacy and side effects of icotinib hydrochloride in the treatment of the advanced non-small cell lung cancer (NSCLC patients with EGFR mutation and wild-type. Methods Patients with advanced NSCLC who were treated with icotinib hydrochloride in Beijing Chest Hospital were retrospective analyzed from March 2009 to December 2014. Results The clinical data of 124 patients (99 with EGFR mutation and 25 with wild type with advanced NSCLC were enrolled in this study. The patients’ overall objective response rate (ORR was 51.6 % and the disease control rate (DCR was 79.8%; The patients with EGFR mutation, ORR was 63.6%, DCR was 93.9%. The ORR was 4.0% and the DCR was 24.0% in the wild-type patients. Median progression-free survival (PFS with icotinib treatment in EGFR mutation patients was 10.5 months and 1.0 month in wild-type patients. The major adverse events were mild skin rash (30.6% and diarrhea (16.1%. Conclusion Monotherapy with icotinib hydrochloride is effective and tolerable for the advanced NSCLC EGFR mutation patients.

  20. NF-κB signaling is activated and confers resistance to apoptosis in three-dimensionally cultured EGFR-mutant lung adenocarcinoma cells

    International Nuclear Information System (INIS)

    Sakuma, Yuji; Yamazaki, Yukiko; Nakamura, Yoshiyasu; Yoshihara, Mitsuyo; Matsukuma, Shoichi; Koizume, Shiro; Miyagi, Yohei

    2012-01-01

    Highlights: ► EGFR-mutant cells in 3D culture resist EGFR inhibition compared with suspended cells. ► Degradation of IκB and activation of NF-κB are observed in 3D-cultured cells. ► Inhibiting NF-κB enhances the efficacy of the EGFR inhibitor in 3D-cultured cells. -- Abstract: Epidermal growth factor receptor (EGFR)-mutant lung adenocarcinoma cells in suspension undergo apoptosis to a greater extent than adherent cells in a monolayer when EGFR autophosphorylation is inhibited by EGFR tyrosine kinase inhibitors (TKIs). This suggests that cell adhesion to a culture dish may activate an anti-apoptotic signaling pathway other than the EGFR pathway. Since the microenvironment of cells cultured in a monolayer are substantially different to that of cells existing in three-dimension (3D) in vivo, we assessed whether two EGFR-mutant lung adenocarcinoma cell lines, HCC827 and H1975, were more resistant to EGFR TKI-induced apoptosis when cultured in a 3D extracellular matrix (ECM) as compared with in suspension. The ECM-adherent EGFR-mutant cells in 3D were significantly less sensitive to treatment with WZ4002, an EGFR TKI, than the suspended cells. Further, a marked degradation of IκBα, the inhibitor of nuclear factor (NF)-κB, was observed only in the 3D-cultured cells, leading to an increase in the activation of NF-κB. Moreover, the inhibition of NF-κB with pharmacological inhibitors enhanced EGFR TKI-induced apoptosis in 3D-cultured EGFR-mutant cells. These results suggest that inhibition of NF-κB signaling would render ECM-adherent EGFR-mutant lung adenocarcinoma cells in vivo more susceptible to EGFR TKI-induced cell death.

  1. Dual Inhibition of Topoisomerase II and Tyrosine Kinases by the Novel Bis-Fluoroquinolone Chalcone-Like Derivative HMNE3 in Human Pancreatic Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Yong-Chao Ma

    Full Text Available Both tyrosine kinase and topoisomerase II (TopII are important anticancer targets, and their respective inhibitors are widely used in cancer therapy. However, some combinations of anticancer drugs could exhibit mutually antagonistic actions and drug resistance, which further limit their therapeutic efficacy. Here, we report that HMNE3, a novel bis-fluoroquinolone chalcone-like derivative that targets both tyrosine kinase and TopII, induces tumor cell proliferation and growth inhibition. The viabilities of 6 different cancer cell lines treated with a range of HMNE3 doses were detected using the 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay. Cellular apoptosis was determined using Hoechst 33258 fluorescence staining and the terminal deoxynucleotidyl transferase (TdT dUTP nick-end labeling (TUNEL assay. The expression of activated Caspase-3 was examined by immunocytochemistry. The tyrosine kinase activity was measured with a human receptor tyrosine kinase (RTK detection kit using a horseradish peroxidase (HRP-conjugated phosphotyrosine (pY20 antibody as the substrate. The topoisomerase II activity was measured using agarose gel electrophoresis with the DNA plasmid pBR322 as the substrate. The expression levels of the P53, Bax, Bcl-2, Caspase-3, -8, -9, p-cSrc, c-Src and topoisomerase II proteins were detected by western blot analysis. The proliferation of five of the six cancer cell lines was significantly inhibited by HMNE3 at 0.312 to 10 μmol/L in a time- and dose-dependent manner. Treatment of the Capan-1 and Panc-1 cells with 1.6 to 3.2 μM HMNE3 for 48 h significantly increased the percentage of apoptotic cells (P<0.05, and this effect was accompanied by a decrease in tyrosine kinase activity. HMNE3 potentially inhibited tyrosine kinase activity in vitro with an IC50 value of 0.64±0.34 μmol/L in Capan-1 cells and 3.1±0.86 μmol/L in Panc-1 cells. The activity of c-Src was significantly inhibited by HMNE3 in a dose

  2. Binding of a diphosphorylated-ITAM peptide to spleen tyrosine kinase (Syk) induces distal conformational changes : a hydrogen exchange mass spectrometry study

    NARCIS (Netherlands)

    Catalina, M Isabel; Fischer, Marcel J E; Liskamp, Rob M J; Heck, Albert J R; Dekker, Frank

    Structural flexibility plays a crucial role in protein function. To assess whether specific structural changes are associated with the binding of an immunoreceptor tyrosine-based activation motif (ITAM) to the tandem Src homology-2 domains (tSH2) of the spleen tyrosine kinase [EC 2.7.7.112] (Syk),

  3. Overall survival after immunotherapy, tyrosine kinase inhibitors and surgery in treatment of metastatic renal cell cancer

    DEFF Research Database (Denmark)

    de Lichtenberg, Trine Honnens; Hermann, Gregers G.; Rorth, Mikael

    2014-01-01

    Abstract Objective. The aim of this study was to evaluate overall survival (OS) after treatment of metastatic renal cell carcinoma (mRCC) following the introduction of tyrosine kinase inhibitors (TKIs) and mammalian target of rapamycin (mTOR) inhibitors. Material and methods. One-hundred and forty...

  4. Mutation pattern in the Bruton's tyrosine kinase gene in 26 unrelated patients with X-linked agammaglobulinemia

    DEFF Research Database (Denmark)

    Vorechovský, I; Luo, L; Hertz, Jens Michael

    1997-01-01

    Mutation pattern was characterized in the Bruton's tyrosine kinase gene (BTK) in 26 patients with X-linked agammaglobulinemia, the first described immunoglobulin deficiency, and was related to BTK expression. A total of 24 different mutations were identified. Most BTK mutations were found to result...

  5. Histone Deacetylase 3 Inhibition Overcomes BIM Deletion Polymorphism-Mediated Osimertinib Resistance in EGFR-Mutant Lung Cancer.

    Science.gov (United States)

    Tanimoto, Azusa; Takeuchi, Shinji; Arai, Sachiko; Fukuda, Koji; Yamada, Tadaaki; Roca, Xavier; Ong, S Tiong; Yano, Seiji

    2017-06-15

    Purpose: The BIM deletion polymorphism is associated with apoptosis resistance to EGFR tyrosine kinase inhibitors (EGFR-TKI), such as gefitinib and erlotinib, in non-small cell lung cancer (NSCLC) harboring EGFR mutations. Here, we investigated whether the BIM deletion polymorphism contributes to resistance against osimertinib, a third-generation EGFR-TKI. In addition, we determined the efficacy of a histone deacetylase (HDAC) inhibitor, vorinostat, against this form of resistance and elucidated the underlying mechanism. Experimental Design: We used EGFR -mutated NSCLC cell lines, which were either heterozygous or homozygous for the BIM deletion polymorphism, to evaluate the effect of osimertinib in vitro and in vivo Protein expression was examined by Western blotting. Alternative splicing of BIM mRNA was analyzed by RT-PCR. Results: EGFR -mutated NSCLC cell lines with the BIM deletion polymorphism exhibited apoptosis resistance to osimertinib in a polymorphism dosage-dependent manner, and this resistance was overcome by combined use with vorinostat. Experiments with homozygous BIM deletion-positive cells revealed that vorinostat affected the alternative splicing of BIM mRNA in the deletion allele, increased the expression of active BIM protein, and thereby induced apoptosis in osimertinib-treated cells. These effects were mediated predominantly by HDAC3 inhibition. In xenograft models, combined use of vorinostat with osimertinib could regress tumors in EGFR -mutated NSCLC cells homozygous for the BIM deletion polymorphism. Moreover, this combination could induce apoptosis even when tumor cells acquired EGFR -T790M mutations. Conclusions: These findings indicate the importance of developing HDAC3-selective inhibitors, and their combined use with osimertinib, for treating EGFR -mutated lung cancers carrying the BIM deletion polymorphism. Clin Cancer Res; 23(12); 3139-49. ©2016 AACR . ©2016 American Association for Cancer Research.

  6. Functional characterization of autophosphorylation sites of the activated insulin receptor-tyrosine kinase

    International Nuclear Information System (INIS)

    Flores-Riveros, J.R.; Lane, M.D.

    1987-01-01

    Insulin receptor, solubilized from 3T3-L1 cellular membranes and then purified, was autophosphorylated with [γ- 32 P]ATP in the absence or presence of insulin. Specific phosphopeptides generated by trypsin digestion of the 32 P-labeled β-subunit were identified and separated by reverse phase HPLC. In the absence of insulin, radioactivity of the phosphopeptides is evenly distributed among four major peaks designated as sites I, II, III and IV, according to their order of elution. This pattern is maintained for at least the first 30 min of autophosphorylation. When the reaction is carried out in the presence of insulin, > 50% of the total 32 P radioactivity is found in site I and the rate of 32 P incorporation into this site is markedly higher than into sites II, III and IV. Maximal activation of tyrosine kinase activity, as estimated by substrate phosphorylation, is coincident with the nearly complete phosphorylation of site I. Delayed activation of previously autophosphorylated receptor by insulin, but not by EGF or IGF-I, produced a similar pattern where phosphorylated site I predominates. These observations indicate that one major insulin-regulated autophosphorylation site in the β-subunit is responsible for activation of the insulin receptor tyrosine kinase. The isolation of this phosphopeptide on a preparative scale and its characterization are now in progress

  7. Exogenous Restoration of TUSC2 Expression Induces Responsiveness to Erlotinib in Wildtype Epidermal Growth Factor Receptor (EGFR Lung Cancer Cells through Context Specific Pathways Resulting in Enhanced Therapeutic Efficacy.

    Directory of Open Access Journals (Sweden)

    Bingbing Dai

    Full Text Available Expression of the tumor suppressor gene TUSC2 is reduced or absent in most lung cancers and is associated with worse overall survival. In this study, we restored TUSC2 gene expression in several wild type EGFR non-small cell lung cancer (NSCLC cell lines resistant to the epidermal growth factor receptor (EGFR tyrosine kinase inhibitor erlotinib and analyzed their sensitivity to erlotinib in vitro and in vivo. A significant inhibition of cell growth and colony formation was observed with TUSC2 transient and stable expression. TUSC2-erlotinib cooperativity in vitro could be reproduced in vivo in subcutaneous tumor growth and lung metastasis formation lung cancer xenograft mouse models. Combination treatment with intravenous TUSC2 nanovesicles and erlotinib synergistically inhibited tumor growth and metastasis, and increased apoptotic activity. High-throughput qRT-PCR array analysis enabling multi-parallel expression profile analysis of eighty six receptor and non-receptor tyrosine kinase genes revealed a significant decrease of FGFR2 expression level, suggesting a potential role of FGFR2 in TUSC2-enhanced sensitivity to erlotinib. Western blots showed inhibition of FGFR2 by TUSC2 transient transfection, and marked increase of PARP, an apoptotic marker, cleavage level after TUSC2-erlotinb combined treatment. Suppression of FGFR2 by AZD4547 or gene knockdown enhanced sensitivity to erlotinib in some but not all tested cell lines. TUSC2 inhibits mTOR activation and the latter cell lines were responsive to the mTOR inhibitor rapamycin combined with erlotinib. These results suggest that TUSC2 restoration in wild type EGFR NSCLC may overcome erlotinib resistance, and identify FGFR2 and mTOR as critical regulators of this activity in varying cellular contexts. The therapeutic activity of TUSC2 could extend the use of erlotinib to lung cancer patients with wildtype EGFR.

  8. Efficacy of Icotinib for Advanced Non-small Cell Lung Cancer Patients with EGFR Status Identified

    Directory of Open Access Journals (Sweden)

    Yiping ZHANG

    2013-03-01

    Full Text Available Background and objective As the first epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI in China, icotinib shows promising anticancer activity in vitro and vivo. The phase III clinical study (ICOGEN showed that icotinib has a good efficacy and tolerability in Chinese patients with advanced non-small cell lung cancer (NSCLC compared with gefitinib. This retrospective study aims to evaluate the efficacy and tolerability of icotinib monotherapy for advanced NSCLC patients with EGFR mutation and wild-type patients in our hospital. Methods Patients with advanced NSCLC who were treated with icotinib in Zhejiang Cancer Hospital were retrospectively analyzed from August, 2011 to August, 2012. Survival was estimated using Kaplan-Meier analysis and Log-rank tests. Results The clinical data of 49 patients (13 with wild-type and 36 with EGFR mutation with NSCLC were enrolled in the current study. The patients’ overall objective response rate (ORR was 58.3% and the disease control rate (DCR in 36 EGFR mutation patients was 88.9%. The ORR was 7.7% and DCR was 53.8% in the wild-type patients. Median progression-free survival (PFS with icotinib treatment in EGFR mutation patients was 9.5 months and 2.2 months in wild-type patients (P<0.001. Nineteen patients with EGFR mutation received icotinib as first-line and 17 in further-line treatment. The PFS was 9.5 months in the first-line and 8.5 months for second-line or further-line patients (P=0.41. Median overall survival (OS in EGFR mutation patients was not reached, but was 12.6 months in wild-type patients. Most of the drug-related adverse events were mild (grade I or II and reversible with no grade IV toxicity. Conclusion Icotinib monotherapy showed significant antitumor activity in advanced NSCLC EGFR mutation patients. The toxicity was well tolerated and acceptable.

  9. Impact of active smoking on survival of patients with metastatic lung adenocarcinoma harboring an epidermal growth factor receptor (EGFR) mutation.

    Science.gov (United States)

    Erdogan, Bulent; Kodaz, Hilmi; Karabulut, Senem; Cinkaya, Ahmet; Tozkir, Hilmi; Tanriverdi, Ozgur; Cabuk, Devrim; Hacioglu, Muhammed Bekir; Turkmen, Esma; Hacibekiroglu, Ilhan; Uzunoglu, Sernaz; Cicin, Irfan

    2016-11-10

    Lung cancer in smokers and non-smokers demonstrates distinct genetic profiles, and cigarette smoking affects epidermal growth factor receptor (EGFR) function and causes secondary EGFR tyrosine kinase resistance. We evaluated the effect of active smoking in patients with metastatic lung adenocarcinoma. A total of 132 metastatic lung adenocarcinoma patients, diagnosed between 2008 and 2013, with known EGFR mutation status, were evaluated retrospectively. Among these patients, 40 had an activating EGFR mutation. Patients who continued smoking during the treatment were defined as active smokers. Former smokers and never smokers were together defined as non-smokers. The outcomes of the treatment in relation to the EGFR mutation and smoking status were evaluated. The median follow-up time was 10.5 months. The overall response rate for the first-line therapy was significantly higher among the EGFR-mutant patients (p = 0.01), however, smoking status had no impact on the response rate (p = 0.1). The EGFR-mutant active smokers progressed earlier than the non-smokers (p non-smokers and patients treated with erlotinib was significantly longer (p = 0.02 and p = 0.01, respectively). Smoking status did not affect the OS in EGFR wild type tumors (p = 0.49) but EGFR-mutant non-smokers had a longer OS than the active smokers (p = 0.01).The active smokers treated with erlotinib had poorer survival than the non-smokers (p = 0.03). Multivariate analysis of EGFR-mutant patients showed that erlotinib treatment at any line and non-smoking were independent prognostic factors for the OS (p = 0.04 and p = 0.01, respectively). Smoking during treatment is a negative prognostic factor in metastatic lung adenocarcinoma with an EGFR mutation.

  10. Tyrosine Kinase Inhibitor Treatment for Newly Diagnosed Chronic Myeloid Leukemia.

    Science.gov (United States)

    Radich, Jerald P; Mauro, Michael J

    2017-08-01

    Chronic myeloid leukemia (CML) is a myeloproliferative disorder that accounts for approximately 10% of new cases of leukemia. The introduction of tyrosine kinase inhibitors has led to a reduction in mortalities. Thus, the estimated prevalence of CML is increasing. The National Comprehensive Cancer Network and the European Leukemia Net guidelines incorporate frequent molecular monitoring of the fusion BCR-ABL transcript to ensure that patients reach and keep treatment milestones. Most patients with CML are diagnosed in the chronic phase, and approximately 10% to 30% of these patients will at some time in their course meet definition criteria of resistance to imatinib. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Evidence for in vivo phosphorylation of the Grb2 SH2-domain binding site on focal adhesion kinase by Src-family protein-tyrosine kinases.

    OpenAIRE

    Schlaepfer, D D; Hunter, T

    1996-01-01

    Focal adhesion kinase (FAK) is a nonreceptor protein-tyrosine kinase (PTK) that associates with integrin receptors and participates in extracellular matrix-mediated signal transduction events. We showed previously that the c-Src nonreceptor PTK and the Grb2 SH2/SH3 adaptor protein bound directly to FAK after fibronectin stimulation (D. D. Schlaepfer, S.K. Hanks, T. Hunter, and P. van der Geer, Nature [London] 372:786-791, 1994). Here, we present evidence that c-Src association with FAK is req...

  12. Hepatitis C Virus Particle Assembly Involves Phosphorylation of NS5A by the c-Abl Tyrosine Kinase.

    Science.gov (United States)

    Yamauchi, Shota; Takeuchi, Kenji; Chihara, Kazuyasu; Sun, Xuedong; Honjoh, Chisato; Yoshiki, Hatsumi; Hotta, Hak; Sada, Kiyonao

    2015-09-04

    Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) is thought to regulate the replication of viral RNA and the assembly of virus particles in a serine/threonine phosphorylation-dependent manner. However, the host kinases that phosphorylate NS5A have not been fully identified. Here, we show that HCV particle assembly involves the phosphorylation of NS5A by the c-Abl tyrosine kinase. Pharmacological inhibition or knockdown of c-Abl reduces the production of infectious HCV (J6/JFH1) particles in Huh-7.5 cells without markedly affecting viral RNA translation and replication. NS5A is tyrosine-phosphorylated in HCV-infected cells, and this phosphorylation is also reduced by the knockdown of c-Abl. Mutational analysis reveals that NS5A tyrosine phosphorylation is dependent, at least in part, on Tyr(330) (Tyr(2306) in polyprotein numbering). Mutation of this residue to phenylalanine reduces the production of infectious HCV particles but does not affect the replication of the JFH1 subgenomic replicon. These findings suggest that c-Abl promotes HCV particle assembly by phosphorylating NS5A at Tyr(330). © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. The combi-targeting concept: synthesis of stable nitrosoureas designed to inhibit the epidermal growth factor receptor (EGFR).

    Science.gov (United States)

    Domarkas, Juozas; Dudouit, Fabienne; Williams, Christopher; Qiyu, Qiu; Banerjee, Ranjita; Brahimi, Fouad; Jean-Claude, Bertrand Jacques

    2006-06-15

    According to the "combi-targeting" concept, the EGFR tyrosine kinase (TK) inhibitory potency of compounds termed "combi-molecules" is critical for selective growth inhibition of tumor cells with disordered expression of EGFR or its closest family member erbB2. Here we report on the optimization of the EGFR TK inhibitory potency of the combi-molecules of the nitrosourea class by comparison with their aminoquinazoline and ureidoquinazoline precursors. This led to the discovery of a new structural parameter that influences their EGFR TK inhibitory potency, i.e., the torsion angle between the plane of the quinazoline ring and the ureido or the nitrosoureido moiety of the synthesized drugs. Compounds (3'-Cl and Br series) with small angles (0.5-3 degrees ) were generally stronger EGFR TK inhibitors than those with large angles (18-21 degrees ). This was further corroborated by ligand-receptor van der Waals interaction calculations that showed significant binding hindrance imposed by large torsion angles in the narrow ATP cleft of EGFR. Selective antiproliferative studies in a pair of mouse fibroblast NIH3T3 cells, one of which NIH3T3/neu being transfected with the erbB2 oncogene, showed that IC(50) values for inhibition of EGFR TK could be good predictors of their selective potency against the serum-stimulated growth of the erbB2-tranfected cell line (Pearson r = 0.8). On the basis of stability (t(1/2)), EGFR TK inhibitory potency (IC(50)), and selective erbB2 targeting, compound 23, a stable nitrosourea, was considered to have the structural requirements for further development.

  14. Non-Targeted Metabolomics Analysis of the Effects of Tyrosine Kinase Inhibitors Sunitinib and Erlotinib on Heart, Muscle, Liver and Serum Metabolism In Vivo

    Directory of Open Access Journals (Sweden)

    Brian C. Jensen

    2017-06-01

    Full Text Available Background: More than 90 tyrosine kinases have been implicated in the pathogenesis of malignant transformation and tumor angiogenesis. Tyrosine kinase inhibitors (TKIs have emerged as effective therapies in treating cancer by exploiting this kinase dependency. The TKI erlotinib targets the epidermal growth factor receptor (EGFR, whereas sunitinib targets primarily vascular endothelial growth factor receptor (VEGFR and platelet-derived growth factor receptor (PDGFR.TKIs that impact the function of non-malignant cells and have on- and off-target toxicities, including cardiotoxicities. Cardiotoxicity is very rare in patients treated with erlotinib, but considerably more common after sunitinib treatment. We hypothesized that the deleterious effects of TKIs on the heart were related to their impact on cardiac metabolism. Methods: Female FVB/N mice (10/group were treated with therapeutic doses of sunitinib (40 mg/kg, erlotinib (50 mg/kg, or vehicle daily for two weeks. Echocardiographic assessment of the heart in vivo was performed at baseline and on Day 14. Heart, skeletal muscle, liver and serum were flash frozen and prepped for non-targeted GC-MS metabolomics analysis. Results: Compared to vehicle-treated controls, sunitinib-treated mice had significant decreases in systolic function, whereas erlotinib-treated mice did not. Non-targeted metabolomics analysis of heart identified significant decreases in docosahexaenoic acid (DHA, arachidonic acid (AA/ eicosapentaenoic acid (EPA, O-phosphocolamine, and 6-hydroxynicotinic acid after sunitinib treatment. DHA was significantly decreased in skeletal muscle (quadriceps femoris, while elevated cholesterol was identified in liver and elevated ethanolamine identified in serum. In contrast, erlotinib affected only one metabolite (spermidine significantly increased. Conclusions: Mice treated with sunitinib exhibited systolic dysfunction within two weeks, with significantly lower heart and skeletal muscle

  15. SAM domain-dependent activity of PfTKL3, an essential tyrosine kinase-like kinase of the human malaria parasite Plasmodium falciparum.

    Science.gov (United States)

    Abdi, Abdirahman; Eschenlauer, Sylvain; Reininger, Luc; Doerig, Christian

    2010-10-01

    Over the last decade, several protein kinases inhibitors have reached the market for cancer chemotherapy. The kinomes of pathogens represent potentially attractive targets in infectious diseases. The functions of the majority of protein kinases of Plasmodium falciparum, the parasitic protist responsible for the most virulent form of human malaria, remain unknown. Here we present a thorough characterisation of PfTKL3 (PF13_0258), an enzyme that belongs to the tyrosine kinase-like kinase (TKL) group. We demonstrate by reverse genetics that PfTKL3 is essential for asexual parasite proliferation in human erythrocytes. PfTKL3 is expressed in both asexual and gametocytes stages, and in the latter the protein co-localises with cytoskeleton microtubules. Recombinant PfTKL3 displays in vitro autophosphorylation activity and is able to phosphorylate exogenous substrates, and both activities are dramatically dependent on the presence of an N-terminal "sterile alpha-motif" domain. This study identifies PfTKL3 as a validated drug target amenable to high-throughput screening.

  16. Detection of EGFR somatic mutations in non-small cell lung cancer (NSCLC) using a novel mutant-enriched liquidchip (MEL) technology.

    Science.gov (United States)

    Zhang, Li; Yang, Huiyi; Zhao, Yanwei; Liu, Wenchao; Wu, Shiyang; He, Jiaying; Luo, Xiaodi; Zhu, Zeyao; Xu, Jiasen; Zhou, Qinghua; Ren-Heidenreich, Lifen

    2012-09-01

    We have developed and standardized a novel technology, mutant-enriched liquidchip (MEL), for clinical detection of EGFR mutations. The MEL integrates a mutant-enriched PCR procedure with liquidchip technology for detections of EGFR exon 19 deletions and L858R mutation on both formalin-fixed and paraffin-embedded (FFPE) slides and plasma samples from patients with non-small cell lung cancer (NSCLC). The detection sensitivity was 0.1% of mutant DNA in the presence of its wild-type DNA. The cross-reaction rate was lower than 5%. To evaluate the MEL platform, the EGFR mutation status of 59 patients with advanced NSCLC treated with EGFRTKIs (Tyrosine Kinase Inhibitors) were tested on their FFPE samples. EGFR exon 19 deletions and L858R were detected in 21 patients (21/59) and 76.2% (16/21) of them had partial response to the EGFR-TKIs, while by sequencing method, only 4 (4/59) mutations were detected. Plasma samples from 627 patients with various stages of NSCLC were examined with the MEL and 22% of EGFR exon 19 deletions and L858R were detected. Furthermore, in patients with advanced disease there are more mutations detected in plasma samples than in patients with less advanced disease. In conclusion, the MEL is a sensitive, stable, and robust technology for detecting EGFR DNA mutations from both FFPE and plasma samples from patients with NSCLC and is now routinely used for clinical diagnosis.

  17. Genome-Wide Identification and Characterization of Tyrosine Kinases in the Silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Songzhen He

    2018-03-01

    Full Text Available The tyrosine kinases (TKs are important parts of metazoan signaling pathways and play significant roles in cell growth, development, apoptosis and disease. Genome-wide characterization of TKs has been conducted in many metazoans, however, systematic information about this family in Lepidoptera is still lacking. We retrieved 33 TK-encoding genes in silkworm and classified them into 25 subfamilies by sequence analysis, without members in AXL, FRK, PDGFR, STYK1 and TIE subfamilies. Although domain sequences in each subfamily are conserved, TKs in vertebrates tend to be remarkably conserved and stable. Our results of phylogenetic analysis supported the previous conclusion for the second major expansion of TK family. Gene-Ontology (GO analysis revealed that a higher proportion of BmTKs played roles in binding, catalysis, signal transduction, metabolism, biological regulation and response to stimulus, compared to all silkworm genes annotated in GO. Moreover, the expression profile analysis of BmTKs among multiple tissues and developmental stages demonstrated that many genes exhibited stage-specific and/or sex-related expression during embryogenesis, molting and metamorphosis, and that 8 BmTKs presented tissue-specific high expression. Our study provides systematic description of silkworm tyrosine kinases, and may also provide further insights into metazoan TKs and assist future studies addressing their functions.

  18. Characterization of the epidermal growth factor receptor associated with cytoskeletons of A431 cells

    International Nuclear Information System (INIS)

    Roy, L.M.; Gittinger, C.K.; Landreth, G.E.

    1989-01-01

    Epidermal growth factor receptors (EGF-R) have been shown to be associated with the detergent-insoluble cytoskeleton of A431 cells, where they retained both a functional ligand-binding domain and tyrosine kinase activity. In the present study we have characterized the tyrosine kinase and ligand binding activities of this cytoskeletally associated EGF-R. The tyrosine kinase activity of the cytoskeletally associated EGF-R was stimulated by EGF treatment of intact cells as evidenced by increased autophosphorylation and phosphorylation of the exogenous substrate angiotensin II (AII). The kinetic behavior of the EGF-R associated with cytoskeletons of EGF-treated cells was similar to that of purified receptors. The stimulation of the receptor kinase activity required EGF treatment of intact cells prior to Triton extraction. If cytoskeletons were prepared from untreated cells and then incubated with EGF, there was no stimulation of the detergent-insoluble receptor kinase activity, indicating that the immobilized receptor was unable to undergo EGF-stimulated activation. Comparison of peptide maps from soluble and cytoskeletally associated EGF-R revealed qualitatively similar patterns; however, they are distinguished by a prominent 46 kD band in digests of the cytoskeletal EGF-R. Saturable binding of 125I-EGF to A431 cytoskeletons prepared from adherent and suspended cells demonstrated the presence of specific receptors on the cytoskeleton. High-affinity EGF-R were preferentially retained upon detergent extraction of adherent cells, whereas both low- and high-affinity receptors were solubilized from the cytoskeletons of suspended cells. Suspension of cells resulted in the solubilization of an additional 15% of the EGF-R to that solubilized in adherent cells, indicating that EGF-R can reversibly associate with the structural elements of the cell

  19. Soluble TAM receptor tyrosine kinases in rheumatoid arthritis: correlation with disease activity and bone destruction.

    Science.gov (United States)

    Xu, L; Hu, F; Zhu, H; Liu, X; Shi, L; Li, Y; Zhong, H; Su, Y

    2018-04-01

    The TAM receptor tyrosine kinases (TAM RTK) are a subfamily of receptor tyrosine kinases, the role of which in autoimmune diseases such as systemic lupus erythematosus has been well explored, while their functions in rheumatoid arthritis (RA) remain largely unknown. In this study, we investigated the role of soluble TAM receptor tyrosine kinases (sAxl/sMer/sTyro3) in patients with RA. A total of 306 RA patients, 100 osteoarthritis (OA) patients and 120 healthy controls (HCs) were enrolled into this study. The serum concentrations of sAxl/sMer/sTyro3 were measured by enzyme-linked immunosorbent assay (ELISA), then the associations between sAxl/sMer/sTyro3 levels and clinical features of RA patients were analysed. We also investigated whether sTyro3 could promote osteoclast differentiation in vitro in RA patients. The results showed that compared with healthy controls (HCs), sTyro3 levels in the serum of RA patients were elevated remarkably and sMer levels were decreased significantly, whereas there was no difference between HCs and RA patients on sAxl levels. The sTyro3 levels were correlated weakly but positively with white blood cells (WBC), immunoglobulin (Ig)M, rheumatoid factor (RF), swollen joint counts, tender joint counts, total sharp scores and joint erosion scores. Conversely, there were no significant correlations between sMer levels and the above indices. Moreover, RA patients with high disease activity also showed higher sTyro3 levels. In-vitro osteoclast differentiation assay showed further that tartrate-resistant acid phosphatase (TRAP) + osteoclasts were increased significantly in the presence of sTyro3. Collectively, our study indicated that serum sTyro3 levels were elevated in RA patients and correlated positively with disease activity and bone destruction, which may serve as an important participant in RA pathogenesis. © 2017 British Society for Immunology.

  20. PROLACTIN-INDUCED TYROSINE PHOSPHORYLATION, ACTIVATION AND RECEPTOR ASSOCIATION OF FOCAL ADHESION KINASE (FAK) IN MAMMARY EPITHELIAL CELLS

    Science.gov (United States)

    Prolactin-Induced Tyrosine Phosphorylation, Activation and ReceptorAssociation of Focal Adhesion Kinase (FAK) in Mammary Epithelial Cells. Suzanne E. Fenton1 and Lewis G. Sheffield2. 1U.S. Environmental ProtectionAgency, MD-72, Research Triangle Park, NC 27711, and

  1. Combination therapy of apatinib with icotinib for primary acquired icotinib resistance in patients with advanced pulmonary adenocarcinoma with EGFR mutation.

    Science.gov (United States)

    Xia, Pinghui; Cao, Jinlin; Lv, Xiayi; Wang, Luming; Lv, Wang; Hu, Jian

    2018-05-01

    Multi-targeted agents represent the next generation of targeted therapies for solid tumors, and patients with acquired resistance to EGFR-tyrosine kinase inhibitors (TKIs) may also benefit from their combination with TKI therapy. Third-generation targeted drugs, such as osimertinib, are very expensive, thus a more economical solution is required. The aim of this study was to explore the use of apatinib combined with icotinib therapy for primary acquired resistance to icotinib in three patients with advanced pulmonary adenocarcinoma with EGFR mutations. We achieved favorable oncologic outcomes in all three patients, with progression-free survival of four to six months. Unfortunately, the patients ultimately had to cease combination therapy because of intolerable adverse effects of hand and foot syndrome and oral ulcers. Combination therapy of apatinib with icotinib for primary acquired resistance to icotinib may be an option for patients with advanced pulmonary adenocarcinoma with EGFR mutations, but physicians must also be aware of the side effects caused by such therapy. © 2018 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.

  2. The Risk of Herpes Zoster in Patients with Non-small Cell Lung Cancer according to Chemotherapy Regimens: Tyrosine Kinase Inhibitors versus Cytotoxic Chemotherapy.

    Science.gov (United States)

    Choi, Ji Young; Kim, Miso; Keam, Bhumsuk; Kim, Tae Min; Kim, Dong-Wan; Heo, Dae Seog; Jo, Seong Jin

    2018-04-05

    Despite the successful use of tyrosine kinase inhibitors (TKIs) in cancer patients, their effect on herpes zoster development has not been studied. The aim of this study was to evaluate and compare the effects of epidermal growth factor receptor (EGFR) TKI and cytotoxic chemotherapy on the risk of herpes zoster development in non-small cell lung cancer (NSCLC) patients. We conducted a medical review of all eligible NSCLC patients in Seoul National University hospital between 2002 and 2015. We classified patients based on whether they previously underwent EGFR TKI therapy into either the TKI group or the cytotoxic group. We compared the incidence rates of herpes zoster during TKI therapy and cytotoxic chemotherapy. Additionally, the longitudinal risk of herpes zoster from TKIs was analyzed using the incidence rate ratio (IRR) of the TKI group to the cytotoxic group and the log-rank test of the Kaplan-Meier method. Of the 2,981 NSCLC patients, 54 patients (1.54%) developed herpes zoster. In the TKI group (2,002 patients), the IRR of herpes zoster during TKI therapy compared to that during cytotoxic chemotherapy was 1.05 (95% confidence interval [CI], 0.53 to 2.09). The IRR of the TKI group compared to the cytotoxic group was 1.33 (95% CI, 0.64 to 2.76). The Kaplan-Meier cumulative risk of both groups was not significantly different. Our results show that the incidence rate of herpes zoster in the TKI group was not statistically different from the incidence in the cytotoxic group during and after chemotherapy in NSCLC patients.

  3. Detection and Analysis of EGFR and KRAS Mutations 
in the Patients with Lung Squamous Cell Carcinomas

    Directory of Open Access Journals (Sweden)

    Hui ZHANG

    2015-10-01

    Full Text Available Background and objective Activating mutations in epidermal growth factor receptor (EGFR and KRAS are important markers in non-small cell lung cancer. However, EGFR and KRAS gene mutations in lung squamous cell carcinoma are rarely reported. The aim of this study was to analyze EGFR and KRAS gene mutation rate and their relationship with clinical features in patients with lung squamous cell carcinomas. Methods A total of 139 patients undergoing treatment for naïve lung squamous cell carcinomas with tumor tissue samples available for testing were recruited. EGFR and KRAS mutation statuses of the tumor samples were detected using a mutant enriched liquid chip. Results Of the 139 cases of lung squamous cell carcinoma, EGFR mutations were detected in 25 cases (18%, KRAS mutations were detected in 7 cases (5%, and the presence of both EGFR and KRAS mutations was detected in 1 case (0.7%. EGFR mutations occurred more often in females than in males (33.3% vs 16.5% and in patients that never smoked than in those who smoke (29.6% vs 16.1%. However, the difference did not reach statistical significance (P>0.05. No significant differences were observed in age, stage, and different biopsy type. KRAS mutations occurred more often in males than in females (5.5% vs 0%, but the difference did not reach statistical significance (P>0.05. No significant differences were observed in age, stage, different biopsy type, and smoking status (P>0.05. Conclusion EGFR and KRAS mutations were low in lung squamous cell carcinomas, and had no significant correlation with clinical features. Before using tyrosine kinase inhibitor targeted therapy, EGFR and KRAS mutations should be detected in patients with lung squamous cell carcinomas.

  4. Neratinib: an oral, irreversible dual EGFR/HER2 inhibitor for breast and non-small cell lung cancer.

    Science.gov (United States)

    Bose, Prithviraj; Ozer, Howard

    2009-11-01

    The revolutionary success of imatinib, a specific inhibitor of the BCR-ABL tyrosine kinase (TK) in the treatment of chronic myelogenous leukemia ushered in the era of targeted therapies in cancer. The erythroblastic leukemia viral oncogene homolog family of receptor TKs, to which EGFR (HER1) and human epidermal growth factor receptor 2 (HER2)/neu TKs belong, has been implicated in a variety of cancers, and several agents that inhibit these TKs are in clinical use, with many more in various stages of development. To summarize current knowledge about neratinib (HKI-272), an oral, irreversible dual inhibitor of EGFR and HER2 and to define its future clinical role, especially in the context of related agents that are either available or in the pipeline. A Medline search using Pubmed was conducted using the keywords neratinib, HKI-272, EGFR, HER2, lapatinib, trastuzumab, erlotinib, gefitinib, cetuximab and panitumumab. Relevant abstracts presented at the American Society of Clinical Oncology and San Antonio Breast Cancer Symposium meetings were also reviewed. Both preclinical and human studies have shown that neratinib has promising activity in both advanced breast cancer and NSCLC with an acceptable safety profile. The data support its continued clinical development.

  5. The tyrosine kinase inhibitor imatinib mesylate suppresses uric acid crystal-induced acute gouty arthritis in mice.

    Directory of Open Access Journals (Sweden)

    Laurent L Reber

    Full Text Available Gouty arthritis is caused by the deposition of monosodium urate (MSU crystals in joints. Despite many treatment options for gout, there is a substantial need for alternative treatments for patients unresponsive to current therapies. Tyrosine kinase inhibitors have demonstrated therapeutic benefit in experimental models of antibody-dependent arthritis and in rheumatoid arthritis in humans, but to date, the potential effects of such inhibitors on gouty arthritis has not been evaluated. Here we demonstrate that treatment with the tyrosine kinase inhibitor imatinib mesylate (imatinib can suppress inflammation induced by injection of MSU crystals into subcutaneous air pouches or into the ankle joint of wild type mice. Moreover, imatinib treatment also largely abolished the lower levels of inflammation which developed in IL-1R1-/- or KitW-sh/W-sh mice, indicating that this drug can inhibit IL-1-independent pathways, as well as mast cell-independent pathways, contributing to pathology in this model. Imatinib treatment not only prevented ankle swelling and synovial inflammation when administered before MSU crystals but also diminished these features when administrated after the injection of MSU crystals, a therapeutic protocol more closely mimicking the clinical situation in which treatment occurs after the development of an acute gout flare. Finally, we also assessed the efficiency of local intra-articular injections of imatinib-loaded poly(lactic-co-glycolic acid (PLGA nanoparticles in this model of acute gout. Treatment with low doses of this long-acting imatinib:PLGA formulation was able to reduce ankle swelling in a therapeutic protocol. Altogether, these results raise the possibility that tyrosine kinase inhibitors might have utility in the treatment of acute gout in humans.

  6. Single molecule force spectroscopy for in-situ probing oridonin inhibited ROS-mediated EGF-EGFR interactions in living KYSE-150 cells.

    Science.gov (United States)

    Pi, Jiang; Jin, Hua; Jiang, Jinhuan; Yang, Fen; Cai, Huaihong; Yang, Peihui; Cai, Jiye; Chen, Zheng W

    2017-05-01

    As the active anticancer component of Rabdosia Rubescens, oridonin has been proved to show strong anticancer activity in cancer cells, which is also found to be closely related to its specific inhibition effects on the EGFR tyrosine kinase activity. In this study, atomic force microscopy based single molecule force spectroscopy (AFM-SMFS) was used for real-time and in-situ detection of EGF-EGFR interactions in living esophageal cancer KYSE-150 cells to evaluate the anticancer activity of oridonin for the first time. Oridonin was found to induce apoptosis and also reduce EGFR expression in KYSE-150 cells. AFM-SMFS results demonstrated that oridonin could inhibit the binding between EGF and EGFR in KYSE-150 cells by decreasing the unbinding force and binding probability for EGF-EGFR complexes, which was further proved to be closely associated with the intracellular ROS level. More precise mechanism studies based on AFM-SMFS demonstrated that oridonin treatment could decrease the energy barrier width, increase the dissociation off rate constant and decrease the activation energy of EGF-EGFR complexes in ROS dependent way, suggesting oridonin as a strong anticancer agent targeting EGF-EGFR interactions in cancer cells through ROS dependent mechanism. Our results not only suggested oridonin as a strong anticancer agent targeting EGF-EGFR interactions in ROS dependent mechanism, but also highlighted AFM-SMFS as a powerful technique for pharmacodynamic studies by detecting ligand-receptor interactions, which was also expected to be developed into a promising tool for the screening and mechanism studies of drugs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. HPV infection and EGFR activation/alteration in HIV-infected East African patients with conjunctival carcinoma.

    Directory of Open Access Journals (Sweden)

    Jing Jie Yu

    2010-05-01

    Full Text Available There has been substantial growth in the numbers of patients with conjunctival squamous cell carcinoma infected with HIV in East Africa. The natural history of the conjunctival squamous cell carcinoma appears to be unique in this region of the world, but the etiologic mechanism unclear and therapeutic options limited. This research was carried out to determine if conjunctival squamous cell carcinoma harbors human papillomavirus DNA and is associated with activation of the EGFR signaling pathway. Positive findings would identify etiologic causes and provide clinical guidance to improve treatment.Expression of p-MAPK/MAPK, p-Akt/Akt and p-EGFR/EGFR in cell nuclei and cytoplasm of 38 FFPE specimens were assessed by immunohistochemistry; HPV genotype was detected by qPCR assay; EGFR mutation was assessed by DNA sequencing analysis; and EGFR mRNA expression was measured using relative qPCR. Statistical analyses included two-sided Fisher exact test or chi-square test, Spearman correlation coefficient and ANOVA. HPV 18 was found in 61% of samples, with HPV 16 double-genotype in 6 patients (16%. Immunohistochemistry and qPCR data suggest that activation and expression of the EGFR signaling pathway is related to disease progression of conjunctival cancer. The associations between cytoplasmic p-MAPK, cytoplasmic p-Akt and tumor invasiveness were significant (p = 0.05 or 0.028. Nuclear p-EGFR appeared only in invasive tumors. A significant positive association between EGFR expression and disease invasiveness was observed (p = 0.01. A SNP in 10 patients and one missense mutation were found within EGFR tyrosine kinase domain. Statistical analysis indicates that patients with measurable EGFR expression more likely harbor EGFR mutations, compared to those with negative EGFR expression (35.3% vs. 0%.We conclude that HPV types 16/18 infection is frequent in East African patients with AIDS-associated squamous cell carcinoma of the conjunctiva. EGFR activation

  8. Insulin receptor binding and tyrosine kinase activity in skeletal muscle from normal pregnant women and women with gestational diabetes

    DEFF Research Database (Denmark)

    Damm, P.; Handberg, A.; Kühl, C.

    1993-01-01

    OBJECTIVE: To ascertain whether the decreased glucose tolerance and insulin resistance found in normal and gestational diabetic pregnancy might be associated with changes in insulin receptor function. METHODS: Eight nonpregnant healthy women (nonpregnant controls), eight healthy pregnant women...... (pregnant controls), and eight women with gestational diabetes were investigated. All were non-obese. Muscle biopsies were obtained from the vastus lateralis muscle, and insulin binding and tyrosine kinase activities in partially purified skeletal muscle insulin receptors were studied. The pregnant controls...... with gestational diabetes compared to nonpregnant controls (P pregnant women did not differ from the other two groups. Postpartum, no differences in insulin binding were found between the groups. Basal and maximal tyrosine kinase activities toward the exogenous substrate poly(Glu4Tyr1) were...

  9. SKLB188 inhibits the growth of head and neck squamous cell carcinoma by suppressing EGFR signalling.

    Science.gov (United States)

    Barzegar, Mansoureh; Ma, Shuang; Zhang, Chao; Chen, Xin; Gu, Ying; Shang, Chaowei; Jiang, Xiaojuan; Yang, Jiao; Nathan, Cherie-Ann; Yang, Shengyong; Huang, Shile

    2017-10-10

    Overexpression of epidermal growth factor receptor (EGFR) occurs in approximately 90% of head and neck squamous cell carcinoma (HNSCC), and is correlated with poor prognosis. Thus, targeting EGFR is a promising strategy for treatment of HNSCC. Several small molecule EGFR inhibitors have been tested in clinical trials for treatment of HNSCC, but none of them are more effective than the current chemotherapeutic drugs. Thus, it is urgently needed to develop novel EGFR inhibitors for HNSCC treatment. By screening an in-house focused library containing approximately 650 000 known kinase inhibitors and kinase inhibitor-like compounds containing common kinase inhibitor core scaffolds, we identified SKLB188 as a lead compound for inhibition of EGFR. The anticancer effects of SKLB188 on HNSCC cells were investigated by in vitro cell growth, cell cycle and apoptosis assays, as well as in vivo FaDu xenograft mouse model. Molecular docking, in vitro kinase profiling and western blotting were performed to characterise EGFR as the molecular target. SKLB188 inhibited HNSCC cell proliferation by inducing G 1 cell cycle arrest, which was associated with downregulating the expression of Cdc25A, cyclins D1/A and cyclin-dependent kinases (CDK2/4), and upregulating the expression of cyclin-dependent kinase (CDK) inhibitors (p21 Cip1 and p27 Kip1 ), leading to decreased phosphorylation of Rb. SKLB188 also induced caspase-dependent apoptosis of HNSCC cells by downregulating the expression of Mcl-1 and survivin. Molecular docking revealed that SKLB188 could bind to the kinase domain of EGFR through hydrogen bonds and hydrophobic interactions. In vitro kinase assay showed that SKLB188 inhibited the activity of a recombinant human EGFR very potently (IC 50 =5 nM). Western blot analysis demonstrated that SKLB188 inhibited the phosphorylation of EGFR and its downstream targets, extracellular signal-regulated protein kinases 1 and 2 (Erk1/2) and Akt in the cells. In addition, SKLB188 dose

  10. The APPLE Trial: Feasibility and Activity of AZD9291 (Osimertinib) Treatment on Positive PLasma T790M in EGFR-mutant NSCLC Patients. EORTC 1613.

    Science.gov (United States)

    Remon, Jordi; Menis, Jessica; Hasan, Baktiar; Peric, Aleksandra; De Maio, Eleonora; Novello, Silvia; Reck, Martin; Berghmans, Thierry; Wasag, Bartosz; Besse, Benjamin; Dziadziuszko, Rafal

    2017-09-01

    The AZD9291 (Osimertinib) Treatment on Positive PLasma T790M in EGFR-mutant NSCLC Patients (APPLE) trial is a randomized, open-label, multicenter, 3-arm, phase II study in advanced, epidermal growth factor receptor (EGFR)-mutant and EGFR tyrosine kinase inhibitor (TKI)-naive non-small-cell lung cancer (NSCLC) patients, to evaluate the best strategy for sequencing gefitinib and osimertinib treatment. Advanced EGFR-mutant NSCLC patients, with World Health Organization performance status 0-2 who are EGFR TKI treatment-naive and eligible to receive first-line treatment with EGFR TKI will be randomized to: In all arms, a plasmatic ctDNA T790M test will be performed by a central laboratory at the Medical University of Gdansk (Poland) but will be applied as a predictive marker for making treatment decisions only in arm B. The primary objective is to evaluate the best strategy for sequencing of treatment with gefitinib and osimertinib in advanced NSCLC patients with common EGFR mutations, and to understand the value of liquid biopsy for the decision-making process. The progression-free survival rate at 18 months is the primary end point of the trial. The activity of osimertinib versus gefitinib to prevent brain metastases will be evaluated. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  11. Lemur Tyrosine Kinase-3 Suppresses Growth of Prostate Cancer Via the AKT and MAPK Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Pengcheng Sun

    2017-08-01

    Full Text Available Background/Aims: Lemur tyrosine kinase (LMTK-3 is a member of the receptor tyrosine kinase (RTK family. Abnormal expression of LMTK-3 exists in various types of cancers, especially in endocrine-resistant breast cancers; however, the precise level of expression and the biological function in prostate cancer are poorly understood. Methods: In the present study, we determined the expression of LMTK-3 in prostate cancer using immunohistochemistry and Western blotting. We infected PC3 and LNCaP cells with lentivirus-LMTK-3 and observed the biologic characteristics of the PC3 and LNCaP cells in vitro with TUNEL, and migration and invasion assays, respectively. We also established a transplant tumor model of human prostate cancer with infected cells in 15 BALB/c-nu/nu nude mice. Results: LMTK-3 was expressed in prostate epithelial cells. There was a significant decline in the level of LMTK-3 expression in prostate cancers compared to normal tissues. LMTK-3 inhibited PC3 and LNCaP cell growth, migration, and invasion, and induced cell apoptosis in vitro. We also observed that LMTK-3 induced PC3 cell apoptosis in vivo. Further study showed that LMTK-3 inhibited phosphorylation of AKT and ERK, and promoted phosphorylation and activation of p38 kinase and Jun kinase (JNK. Conclusion: Recombinant lentivirus with enhanced expression of LMTK-3 inhibited prostate cancer cell growth and induced apoptosis in vitro and in vivo. AKT and MAPK signaling pathways may contribute to the process.

  12. Alignment independent 3D-QSAR, quantum calculations and molecular docking of Mer specific tyrosine kinase inhibitors as anticancer drugs

    Directory of Open Access Journals (Sweden)

    Fereshteh Shiri

    2016-03-01

    Full Text Available Mer receptor tyrosine kinase is a promising novel cancer therapeutic target in many human cancers, because abnormal activation of Mer has been implicated in survival signaling and chemoresistance. 3D-QSAR analyses based on alignment independent descriptors were performed on a series of 81 Mer specific tyrosine kinase inhibitors. The fractional factorial design (FFD and the enhanced replacement method (ERM were applied and tested as variable selection algorithms for the selection of optimal subsets of molecular descriptors from a much greater pool of such regression variables. The data set was split into 65 molecules as the training set and 16 compounds as the test set. All descriptors were generated by using the GRid INdependent descriptors (GRIND approach. After variable selection, GRIND were correlated with activity values (pIC50 by PLS regression. Of the two applied variable selection methods, ERM had a noticeable improvement on the statistical parameters of PLS model, and yielded a q2 value of 0.77, an rpred2 of 0.94, and a low RMSEP value of 0.25. The GRIND information contents influencing the affinity on Mer specific tyrosine kinase were also confirmed by docking studies. In a quantum calculation study, the energy difference between HOMO and LUMO (gap implied the high interaction of the most active molecule in the active site of the protein. In addition, the molecular electrostatic potential energy at DFT level confirmed results obtained from the molecular docking. The identified key features obtained from the molecular modeling, enabled us to design novel kinase inhibitors.

  13. Alignment independent 3D-QSAR, quantum calculations and molecular docking of Mer specific tyrosine kinase inhibitors as anticancer drugs.

    Science.gov (United States)

    Shiri, Fereshteh; Pirhadi, Somayeh; Ghasemi, Jahan B

    2016-03-01

    Mer receptor tyrosine kinase is a promising novel cancer therapeutic target in many human cancers, because abnormal activation of Mer has been implicated in survival signaling and chemoresistance. 3D-QSAR analyses based on alignment independent descriptors were performed on a series of 81 Mer specific tyrosine kinase inhibitors. The fractional factorial design (FFD) and the enhanced replacement method (ERM) were applied and tested as variable selection algorithms for the selection of optimal subsets of molecular descriptors from a much greater pool of such regression variables. The data set was split into 65 molecules as the training set and 16 compounds as the test set. All descriptors were generated by using the GRid INdependent descriptors (GRIND) approach. After variable selection, GRIND were correlated with activity values (pIC50) by PLS regression. Of the two applied variable selection methods, ERM had a noticeable improvement on the statistical parameters of PLS model, and yielded a q (2) value of 0.77, an [Formula: see text] of 0.94, and a low RMSEP value of 0.25. The GRIND information contents influencing the affinity on Mer specific tyrosine kinase were also confirmed by docking studies. In a quantum calculation study, the energy difference between HOMO and LUMO (gap) implied the high interaction of the most active molecule in the active site of the protein. In addition, the molecular electrostatic potential energy at DFT level confirmed results obtained from the molecular docking. The identified key features obtained from the molecular modeling, enabled us to design novel kinase inhibitors.

  14. Combined therapeutic potential of nuclear receptors with receptor tyrosine kinase inhibitors in lung cancer

    International Nuclear Information System (INIS)

    Wairagu, Peninah M.; Park, Kwang Hwa; Kim, Jihye; Choi, Jong-Whan; Kim, Hyun-Won; Yeh, Byung-Il; Jung, Soon-Hee; Yong, Suk-Joong; Jeong, Yangsik

    2014-01-01

    Highlights: • The 48 NR genes and 48 biological anti-cancer targets are profiled in paired-cells. • Growth inhibition by NR ligands or TKIs is target receptor level-dependent. • T0901317 with gefitinib/PHA665752 shows additive growth inhibition in lung cells. - Abstract: Cancer heterogeneity is a big hurdle in achieving complete cancer treatment, which has led to the emergence of combinational therapy. In this study, we investigated the potential use of nuclear receptor (NR) ligands for combinational therapy with other anti-cancer drugs. We first profiled all 48 NRs and 48 biological anti-cancer targets in four pairs of lung cell lines, where each pair was obtained from the same patient. Two sets of cell lines were normal and the corresponding tumor cell lines while the other two sets consisted of primary versus metastatic tumor cell lines. Analysis of the expression profile revealed 11 NRs and 15 cancer targets from the two pairs of normal versus tumor cell lines, and 9 NRs and 9 cancer targets from the primary versus metastatic tumor cell lines had distinct expression patterns in each category. Finally, the evaluation of nuclear receptor ligand T0901317 for liver X receptor (LXR) demonstrated its combined therapeutic potential with tyrosine kinase inhibitors. The combined treatment of cMET inhibitor PHA665752 or EGFR inhibitor gefitinib with T0901317 showed additive growth inhibition in both H2073 and H1993 cells. Mechanistically, the combined treatment suppressed cell cycle progression by inhibiting cyclinD1 and cyclinB expression. Taken together, this study provides insight into the potential use of NR ligands in combined therapeutics with other biological anti-cancer drugs

  15. [Efficacy of icotinib for advanced non-small cell lung cancer patients with EGFR status identified].

    Science.gov (United States)

    Song, Zhengbo; Yu, Xinmin; Cai, Jufen; Shao, Lan; Lin, Baochai; He, Chunxiao; Zhang, Beibei; Zhang, Yiping

    2013-03-01

    As the first epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) in China, icotinib shows promising anticancer activity in vitro and vivo. The phase III clinical study (ICOGEN) showed that icotinib has a good efficacy and tolerability in Chinese patients with advanced non-small cell lung cancer (NSCLC) compared with gefitinib. This retrospective study aims to evaluate the efficacy and tolerability of icotinib monotherapy for advanced NSCLC patients with EGFR mutation and wild-type patients in our hospital. Patients with advanced NSCLC who were treated with icotinib in Zhejiang Cancer Hospital were retrospectively analyzed from August, 2011 to August, 2012. Survival was estimated using Kaplan-Meier analysis and Log-rank tests. The clinical data of 49 patients (13 with wild-type and 36 with EGFR mutation) with NSCLC were enrolled in the current study. The patients' overall objective response rate (ORR) was 58.3% and the disease control rate (DCR) in 36 EGFR mutation patients was 88.9%. The ORR was 7.7% and DCR was 53.8% in the wild-type patients. Median progression-free survival (PFS) with icotinib treatment in EGFR mutation patients was 9.5 months and 2.2 months in wild-type patients (Picotinib as first-line and 17 in further-line treatment. The PFS was 9.5 months in the first-line and 8.5 months for second-line or further-line patients (P=0.41). Median overall survival (OS) in EGFR mutation patients was not reached, but was 12.6 months in wild-type patients. Most of the drug-related adverse events were mild (grade I or II) and reversible with no grade IV toxicity. Icotinib monotherapy showed significant antitumor activity in advanced NSCLC EGFR mutation patients. The toxicity was well tolerated and acceptable.

  16. Bacterial single-stranded DNA-binding proteins are phosphorylated on tyrosine

    DEFF Research Database (Denmark)

    Mijakovic, Ivan; Petranovic, Dina; Macek, B

    2006-01-01

    for phosphotyrosine-containing proteins in Streptomyces griseus by immunoaffinity chromatography identified bacterial SSBs as a novel target of bacterial tyrosine kinases. Since genes encoding protein-tyrosine kinases (PTKs) have not been recognized in streptomycetes, and SSBs from Streptomyces coelicolor (Sc......SSB) and Bacillus subtilis (BsSSB) share 38.7% identity, we used a B.subtilis protein-tyrosine kinase YwqD to phosphorylate two cognate SSBs (BsSSB and YwpH) in vitro. We demonstrate that in vivo phosphorylation of B.subtilis SSB occurs on tyrosine residue 82, and this reaction is affected antagonistically...... by kinase YwqD and phosphatase YwqE. Phosphorylation of B.subtilis SSB increased binding almost 200-fold to single-stranded DNA in vitro. Tyrosine phosphorylation of B.subtilis, S.coelicolor and Escherichia coli SSBs occured while they were expressed in E.coli, indicating that tyrosine phosphorylation...

  17. Impact of active smoking on survival of patients with metastatic lung adenocarcinoma harboring an epidermal growth factor receptor (EGFR mutation

    Directory of Open Access Journals (Sweden)

    Bulent Erdogan

    2016-11-01

    Full Text Available Lung cancer in smokers and non-smokers demonstrates distinct genetic profiles, and cigarette smoking affects epidermal growth factor receptor (EGFR function and causes secondary EGFR tyrosine kinase resistance. We evaluated the effect of active smoking in patients with metastatic lung adenocarcinoma. A total of 132 metastatic lung adenocarcinoma patients, diagnosed between 2008 and 2013, with known EGFR mutation status, were evaluated retrospectively. Among these patients, 40 had an activating EGFR mutation. Patients who continued smoking during the treatment were defined as active smokers. Former smokers and never smokers were together defined as non-smokers. The outcomes of the treatment in relation to the EGFR mutation and smoking status were evaluated. The median follow-up time was 10.5 months. The overall response rate for the first-line therapy was significantly higher among the EGFR-mutant patients (p = 0.01, however, smoking status had no impact on the response rate (p = 0.1. The EGFR-mutant active smokers progressed earlier than the non-smokers (p < 0.01. The overall survival (OS of the non-smokers and patients treated with erlotinib was significantly longer (p = 0.02 and p = 0.01, respectively. Smoking status did not affect the OS in EGFR wild type tumors (p = 0.49 but EGFR-mutant non-smokers had a longer OS than the active smokers (p = 0.01.The active smokers treated with erlotinib had poorer survival than the non-smokers (p = 0.03. Multivariate analysis of EGFR-mutant patients showed that erlotinib treatment at any line and non-smoking were independent prognostic factors for the OS (p = 0.04 and p = 0.01, respectively. Smoking during treatment is a negative prognostic factor in metastatic lung adenocarcinoma with an EGFR mutation.

  18. TNK2 Tyrosine Kinase as a Novel Therapeutic Target in Triple-Negative Breast Cancer

    Science.gov (United States)

    2017-10-01

    Award Number: W81XWH-15-1-0311 TITLE: TNK2 Tyrosine Kinase as a Novel Therapeutic Target in Triple- Negative Breast Cancer PRINCIPAL...Distribution Unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT Triple-negative breast cancers (TNBCs) represent only 10%-15% of all breast cancers ; however... cancers (TNBC) represent 10-15% of all breast cancers . While significant advances have been made for targeted therapy of ER and HER2-positive breast

  19. Cost-Effectiveness Analysis of Afatinib versus Gefitinib for First-Line Treatment of Advanced EGFR-Mutated Advanced Non-Small Cell Lung Cancers.

    Science.gov (United States)

    Chouaid, Christos; Luciani, Laura; LeLay, Katell; Do, Pascal; Bennouna, Jaafar; Perol, Maurice; Moro-Sibilot, Denis; Vergnenègre, Alain; de Pouvourville, Gérard

    2017-10-01

    The irreversible ErbB family blocker afatinib and the reversible EGFR tyrosine kinase inhibitor gefitinib were compared in the multicenter, international, randomized, head-to-head phase 2b LUX-Lung 7 trial for first-line treatment of advanced EGFR mutation-positive NSCLCs. Afatinib and gefitinib costs and patients' outcomes in France were assessed. A partitioned survival model was designed to assess the cost-effectiveness of afatinib versus gefitinib for EGFR mutation-positive NSCLCs. Outcomes and safety were taken primarily from the LUX-Lung 7 trial. Resource use and utilities were derived from that trial, an expert-panel questionnaire, and published literature, limiting expenditures to direct costs. Incremental cost-effectiveness ratios (ICERs) were calculated over a 10-year time horizon for the entire population, and EGFR exon 19 deletion or exon 21 L858R mutation (L858R) subgroups. Deterministic and probabilistic sensitivity analyses were conducted. For all EGFR mutation-positive NSCLCs, the afatinib-versus-gefitinib ICER of was €45,211 per quality-adjusted life-year (QALY) (0.170 QALY gain for an incremental cost of €7697). ICERs for EGFR exon 19 deletion and L858R populations were €38,970 and €52,518, respectively. Afatinib had 100% probability to be cost-effective at a willingness-to-pay threshold of €70,000/QALY for patients with common EGFR mutations. First-line afatinib appears cost-effective compared with gefitinib for patients with EGFR mutation-positive NSCLCs. Copyright © 2017 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.

  20. Determination of EGFR and KRAS mutational status in Greek non-small-cell lung cancer patients.

    Science.gov (United States)

    Papadopoulou, Eirini; Tsoulos, Nikolaos; Tsirigoti, Angeliki; Apessos, Angela; Agiannitopoulos, Konstantinos; Metaxa-Mariatou, Vasiliki; Zarogoulidis, Konstantinos; Zarogoulidis, Pavlos; Kasarakis, Dimitrios; Kakolyris, Stylianos; Dahabreh, Jubrail; Vlastos, Fotis; Zoublios, Charalampos; Rapti, Aggeliki; Papageorgiou, Niki Georgatou; Veldekis, Dimitrios; Gaga, Mina; Aravantinos, Gerasimos; Karavasilis, Vasileios; Karagiannidis, Napoleon; Nasioulas, George

    2015-10-01

    It has been reported that certain patients with non-small-cell lung cancer (NSCLC) that harbor activating somatic mutations within the tyrosine kinase domain of the epidermal growth factor receptor ( EGFR ) gene may be effectively treated using targeted therapy. The use of EGFR inhibitors in patient therapy has been demonstrated to improve response and survival rates; therefore, it was suggested that clinical screening for EGFR mutations should be performed for all patients. Numerous clinicopathological factors have been associated with EGFR and Kirsten-rat sarcoma oncogene homolog (KRAS) mutational status including gender, smoking history and histology. In addition, it was reported that EGFR mutation frequency in NSCLC patients was ethnicity-dependent, with an incidence rate of ~30% in Asian populations and ~15% in Caucasian populations. However, limited data has been reported on intra-ethnic differences throughout Europe. The present study aimed to investigate the frequency and spectrum of EGFR mutations in 1,472 Greek NSCLC patients. In addition, KRAS mutation analysis was performed in patients with known smoking history in order to determine the correlation of type and mutation frequency with smoking. High-resolution melting curve (HRM) analysis followed by Sanger sequencing was used to identify mutations in exons 18-21 of the EGFR gene and in exon 2 of the KRAS gene. A sensitive next-generation sequencing (NGS) technology was also employed to classify samples with equivocal results. The use of sensitive mutation detection techniques in a large study population of Greek NSCLC patients in routine diagnostic practice revealed an overall EGFR mutation frequency of 15.83%. This mutation frequency was comparable to that previously reported in other European populations. Of note, there was a 99.8% concordance between the HRM method and Sanger sequencing. NGS was found to be the most sensitive method. In addition, female non-smokers demonstrated a high prevalence of

  1. Comparative evaluation of bone marrow cells morpho-functional activity in chronic myeloid leukemia patients treated with tyrosine kinase inhibitors of the first and second generation

    Directory of Open Access Journals (Sweden)

    I. O. Zhaleyko

    2014-07-01

    Full Text Available The efficiency of using the culture techniques of research for monitoring the patient’s response to the treatment by tyrosine kinase inhibitors of the first and second generation is shown. Thus, the functional activity of bone marrow cells in patients having the optimal treatment response to inhibitors of tyrosine kinases was significantly lower compared with patients with the acquired resistance to the drug, and patients who had CML diagnosed for first time. Furthermore, for patients with the optimal response to the nilotinib therapy, numbers of colonies in semi-solid agar in vitro was lower, than in patients with the optimal response to imatinib. When the leukaemic cell clone becomes resistant to tyrosine kinase inhibitors, the prevalence of early cells of granulocyte-macrophage hematopoietic stem cells is observed in CFU culture which can be an important prognostic factor for choosing the appropriate treatment strategy.

  2. Complement receptor-3 negatively regulates the phagocytosis of degenerated myelin through tyrosine kinase Syk and cofilin

    Directory of Open Access Journals (Sweden)

    Hadas Smadar

    2012-07-01

    Full Text Available Abstract Background Intact myelin, which normally surrounds axons, breaks down in Wallerian degeneration following axonal injury and during neurodegenerative diseases such as multiple sclerosis. Clearance of degenerated myelin by phagocytosis is essential since myelin impedes repair and exacerbates damage. CR3 (complement receptor-3 is a principal phagocytic receptor in myelin phagocytosis. We studied how tyrosine kinase Syk (spleen tyrosine kinase and cofilin control phagocytosis of degenerated myelin by CR3 in microglia and macrophages. Syk is a non-receptor tyrosine kinase that CR3 recruits to convey cellular functions. Cofilin is an actin-depolymerizing protein that controls F-actin (filamentous actin remodeling (i.e., disassembly and reassembly by shifting between active unphosphorylated and inactive phosphorylated states. Results Syk was continuously activated during prolonged phagocytosis. Phagocytosis increased when Syk activity and expression were reduced, suggesting that normally Syk down regulates CR3-mediated myelin phagocytosis. Levels of inactive p-cofilin (phosphorylated cofilin decreased transiently during prolonged phagocytosis. In contrast, p-cofilin levels decreased continuously when Syk activity and expression were continuously reduced, suggesting that normally Syk advances the inactive state of cofilin. Observations also revealed inverse relationships between levels of phagocytosis and levels of inactive p-cofilin, suggesting that active unphosphorylated cofilin advances phagocytosis. Active cofilin could advance phagocytosis by promoting F-actin remodeling, which supports the production of membrane protrusions (e.g., filopodia, which, as we also revealed, are instrumental in myelin phagocytosis. Conclusions CR3 both activates and downregulates myelin phagocytosis at the same time. Activation was previously documented. We presently demonstrate that downregulation is mediated through Syk, which advances the inactive

  3. Abelson tyrosine-protein kinase 2 regulates myoblast proliferation and controls muscle fiber length

    OpenAIRE

    Lee, Jennifer K; Hallock, Peter T; Burden, Steven J

    2017-01-01

    Muscle fiber length is nearly uniform within a muscle but widely different among different muscles. We show that Abelson tyrosine-protein kinase 2 (Abl2) has a key role in regulating myofiber length, as a loss of Abl2 leads to excessively long myofibers in the diaphragm, intercostal and levator auris muscles but not limb muscles. Increased myofiber length is caused by enhanced myoblast proliferation, expanding the pool of myoblasts and leading to increased myoblast fusion. Abl2 acts in myobla...

  4. Abelson tyrosine-protein kinase 2 Regulates Myoblast Proliferation and Controls Muscle Fiber Length

    OpenAIRE

    Burden, Steven; Lee, Jennifer

    2017-01-01

    Muscle fiber length is nearly uniform within a muscle but widely different among muscles. Here, we show that Abelson tyrosine-protein kinase 2 (Abl2) has a key role in regulating myofiber length, as a loss of Abl2 leads to excessively long myofibers in the diaphragm and other muscles. Increased myofiber length is caused by enhanced myoblast proliferation, expanding the pool of available myoblasts and leading to increased myoblast fusion. Abl2 acts in myoblasts, but expansion of the diaphragm ...

  5. The Fyn tyrosine kinase binds Irs-1 and forms a distinct signaling complex during insulin stimulation.

    Science.gov (United States)

    Sun, X J; Pons, S; Asano, T; Myers, M G; Glasheen, E; White, M F

    1996-05-03

    Irs-proteins link the receptors for insulin/IGF-1, growth hormones, and several interleukins and interferons to signaling proteins that contain Src homology-2 (SH2). To identify new Irs-1-binding proteins, we screened a mouse embryo expression library with recombinant [32P]Irs-1, which revealed a specific association between p59fyn and Irs-1. The SH2 domain in p59fyn bound to phosphorylated Tyr895 and Tyr1172, which are located in YXX(L/I) motifs. Mutation of p59fyn at the COOH-terminal tyrosine phosphorylation site (Tyr531) enhanced its binding to Irs-1 during insulin stimulation. Binding experiments with various SH2 protein revealed that Grb-2 was largely excluded from Irs-1 complexes containing p59fyn, whereas Grb-2 and p85 occurred in the same Irs-1 complex. By comparison with the insulin receptor, p59fyn kinase phosphorylated a unique cohort of tyrosine residues in Irs-1. These results outline a role for p59fyn or other related Src-kinases during insulin and cytokine signaling.

  6. JAK kinases are required for the bacterial RNA and poly I:C induced tyrosine phosphorylation of PKR

    OpenAIRE

    Bleiblo, Farag; Michael, Paul; Brabant, Danielle; Ramana, Chilakamarti V; Tai, TC; Saleh, Mazen; Parrillo, Joseph E; Kumar, Anand; Kumar, Aseem

    2012-01-01

    Discriminating the molecular patterns associated with RNA is central to innate immunity. The protein kinase PKR is a cytosolic sensor involved in the recognition of viral dsRNA and triggering interferon-induced signaling. Here, we identified bacterial RNA as a novel distinct pattern recognized by PKR. We show that the tyrosine phosphorylation of PKR induced by either bacterial RNA or poly I:C is impaired in mutant cells lacking TYK2, JAK1, or JAK2 kinases. PKR was found to be a direct substra...

  7. TAM receptor tyrosine kinase function and the immunopathology of liver disease.

    Science.gov (United States)

    Mukherjee, S K; Wilhelm, A; Antoniades, C G

    2016-06-01

    Tyro3, Axl, MERTK (TAM) receptor tyrosine kinases are implicated in the regulation of the innate immune response through clearance of apoptotic cellular debris and control of cytokine signaling cascades. As a result they are pivotal in regulating the inflammatory response to tissue injury. Within the liver, immune regulatory signaling is employed to prevent the overactivation of innate immunity in response to continual antigenic challenge from the gastrointestinal tract. In this review we appraise current understanding of the role of TAM receptor function in the regulation of both innate and adaptive immunity, with a focus on its impact upon hepatic inflammatory pathology. Copyright © 2016 the American Physiological Society.

  8. Muscle-Specific Tyrosine Kinase and Myasthenia Gravis Owing to Other Antibodies.

    Science.gov (United States)

    Rivner, Michael H; Pasnoor, Mamatha; Dimachkie, Mazen M; Barohn, Richard J; Mei, Lin

    2018-05-01

    Around 20% of patients with myasthenia gravis are acetylcholine receptor antibody negative; muscle-specific tyrosine kinase antibodies (MuSK) were identified as the cause of myasthenia gravis in 30% to 40% of these cases. Anti MuSK myasthenia gravis is associated with specific clinical phenotypes. One is a bulbar form with fewer ocular symptoms. Others show an isolated head drop or symptoms indistinguishable from acetylcholine receptor-positive myasthenia gravis. These patients usually respond well to immunosuppressive therapy, but not as well to cholinesterase inhibitors. Other antibodies associated with myasthenia gravis, including low-density lipoprotein receptor-related protein 4, are discussed. Copyright © 2018 Elsevier Inc. All rights reserved.

  9. Primary cilia and coordination of receptor tyrosine kinase (RTK) and transforming growth factor β (TGF-β) signaling

    DEFF Research Database (Denmark)

    Christensen, Søren Tvorup; Morthorst, Stine Kjær; Mogensen, Johanne Bay

    2017-01-01

    are at the root of a pleiotropic group of diseases and syndromic disorders called ciliopathies. In this review, we present an overview of primary cilia-mediated regulation of receptor tyrosine kinase (RTK) and transforming growth factor β (TGF-β) signaling. Further, we discuss how defects in the coordination...

  10. Tyrosine kinase inhibitor BIBF1120 ameliorates inflammation, angiogenesis and fibrosis in CCl4-induced liver fibrogenesis mouse model

    NARCIS (Netherlands)

    Öztürk Akcora, Büsra; Storm, Gerrit; Prakash, Jai; Bansal, Ruchi

    2017-01-01

    Hepatic fibrosis, a progressive chronic disease mainly caused by hepatitis viral infections, alcohol abuse or metabolic syndrome leading to liver dysfunction and is the growing cause of mortality worldwide. Tyrosine kinase inhibitor BIBF1120 (Nintedanib) has been evaluated in clinical trials for

  11. Tyrosine Kinase Receptor Landscape in Lung Cancer: Therapeutical Implications

    Directory of Open Access Journals (Sweden)

    A. Quintanal-Villalonga

    2016-01-01

    Full Text Available Lung cancer is a heterogeneous disease responsible for the most cases of cancer-related deaths. The majority of patients are clinically diagnosed at advanced stages, with a poor survival rate. For this reason, the identification of oncodrivers and novel biomarkers is decisive for the future clinical management of this pathology. The rise of high throughput technologies popularly referred to as “omics” has accelerated the discovery of new biomarkers and drivers for this pathology. Within them, tyrosine kinase receptors (TKRs have proven to be of importance as diagnostic, prognostic, and predictive tools and, due to their molecular nature, as therapeutic targets. Along this review, the role of TKRs in the different lung cancer histologies, research on improvement of anti-TKR therapy, and the current approaches to manage anti-TKR resistance will be discussed.

  12. Tyrosine phosphorylation in signal transduction

    International Nuclear Information System (INIS)

    Roberts, T.M.; Kaplan, D.; Morgan, W.; Keller, T.; Mamon, H.; Piwnica-Worms, H.; Druker, B.; Whitman, M.; Morrison, D.; Cohen, B.; Schaffhausen, B.; Cantley, L.; Rapp, U.

    1988-01-01

    Recent work has focused on the elucidation of the mechanisms by which membrane-bound tyrosine kinases transmit signals within the cell. To examine the role of tyrosine phosphorylation the authors have employed the following strategy. First, they have utilized antibodies to phosphotyrosine (anti-P.Tyr) to identify candidate substrates of various tyrosine kinases, such as pp60 c-src , the CSF- receptor, or the platelet-derived growth factor (PDGF) receptor. Second, they have attempted to characterize the biochemical properties of the putative substrates and to determine in what manner these properties are modified by phosphorylation on tyrosine residues. In this endeavor, they are recapitulating the classic biochemical analysis used to study the effect of kinases on metabolism. The final portion of our work consists of using modern molecular biological strategies to clone the genes or cDNAs for the substrates and overproduce the relevant proteins for studies in vitro in defined systems. This paper describes the first and second aspects of this strategy, the identification and characterization of novel substrate molecules

  13. Profiling EGFR activity in head and neck squamous cell carcinoma by using a novel layered membrane Western blot technology.

    Science.gov (United States)

    Patel, Vyomesh; Ramesh, Arun; Traicoff, June L; Baibakov, Galina; Emmert-Buck, Michael R; Gutkind, J Silvio; Knezevic, Vladimir

    2005-05-01

    Given the role of epidermal growth factor receptor (EGFR) in head and neck squamous cell carcinomas (HNSCC), several rational approaches have now been utilized to abrogate tyrosine kinase activity and its disengagement from downstream signal transducers. Monitoring the activity of these molecules could potentially be useful to determine not only drug efficacy but also to identify HNSCC patients most likely to benefit from this type of therapy. In this study we have used a novel high throughput multi-layered Western blotting (MLWestern) method that allows the detection of multiple proteins from a single experiment in order to characterize key components in the EGFR signaling pathway in HNSCC cells. Total and activated forms of EGFR and the downstream effectors, Erk and Akt were readily detected in HNSCC cells, where in the control cells (HaCaT) these proteins could only be detected in EGF stimulated cells. Results from conventional Western blot and MLWestern were comparable. Clustering analysis of protein expression revealed similarities in cellular response between some of the cell lines indicative of similarities in their biological response. The data indicate that MLWestern can be potentially applied to identify molecular targets that could be used for rational therapeutic intervention strategies.

  14. Promiscuity and selectivity of small-molecule inhibitors across TAM receptor tyrosine kinases in pediatric leukemia.

    Science.gov (United States)

    Liu, Mao-Hua; Chen, Shi-Bing; Yu, Juan; Liu, Cheng-Jun; Zhang, Xiao-Jing

    2017-08-01

    The TAM receptor tyrosine kinase family member Mer has been recognized as an attractive therapeutic target for pediatric leukemia. Beside Mer the family contains other two kinases, namely, Tyro3 and Axl, which are highly homologues with Mer and thus most existing small-molecule inhibitors show moderate or high promiscuity across the three kinases. Here, the structural basis and energetic property of selective binding of small-molecule inhibitors to the three kinases were investigated at molecular level. It is found that the selectivity is primarily determined by the size, shape and configuration of kinase's ATP-binding site; the Mer and Axl possess a small, closed active pocket as compared to the bulky, open pocket of Tyro3. The location and conformation of active-site residues of Mer and Axl are highly consistent, suggesting that small-molecule inhibitors generally have a low Mer-over-Axl selectivity and a high Mer-over-Tyro3 selectivity. We demonstrated that the difference in ATP binding potency to the three kinases is also responsible for inhibitor selectivity. We also found that the long-range interactions and allosteric effect arising from rest of the kinase's active site can indirectly influence inhibitor binding and selectivity. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Suppression of adhesion-induced protein tyrosine phosphorylation decreases invasive and metastatic potentials of B16-BL6 melanoma cells by protein tyrosine kinase inhibitor genistein.

    Science.gov (United States)

    Yan, C; Han, R

    1997-01-01

    Protein tyrosine kinase (PTK) appears to be involved in the activation of signaling during cell attachment to and spreading on extracellular matrix (ECM) in the metastatic cascade. To verify the assumption that PTK inhibitors might impair ECM signaling and prevent cancer metastasis, the highly metastatic B16-BL6 mouse melanoma cells were exposed to the PTK inhibitor genistein for 3 days. The ability of the cells to invade through reconstituted basement membrane (Matrigel) and to establish experimental pulmonary metastatic foci in C57BL/6 mice decreased after genistein exposure. The genistein-treated cells were also prevented from attaching to Matrigel and spread extremely poorly on the ECM substratum. Immunoblot analysis showed that tyrosine phosphorylation of a 125-kD protein in response to cell spreading on Matrigel was suppressed in the genistein-treated cells. Adhesion-induced protein tyrosine phosphorylation represents the earlier and specific event in the activation of ECM signaling, so this result implied ECM signaling was impaired in the treated cells. With immunofluorescence microscopy, the adhesion-induced tyrosine phosphorylated proteins were located at the pericytoplasms of well-spread cells, but not at the periphery of poorly spread genistein-treated cells. Therefore, this paper suggests that genistein might impair ECM signaling and subsequently prevent cancer cells from spreading well and invading or establishing metastasis through the suppression of adhesion-induced protein tyrosine phosphorylation. PTKs and adhesion-induced protein tyrosine phosphorylation might play a role in the control of invasion and metastasis.

  16. p56Lck and p59Fyn Regulate CD28 Binding to Phosphatidylinositol 3-Kinase, Growth Factor Receptor-Bound Protein GRB-2, and T Cell-Specific Protein-Tyrosine Kinase ITK: Implications for T-Cell Costimulation

    Science.gov (United States)

    Raab, Monika; Cai, Yun-Cai; Bunnell, Stephen C.; Heyeck, Stephanie D.; Berg, Leslie J.; Rudd, Christopher E.

    1995-09-01

    T-cell activation requires cooperative signals generated by the T-cell antigen receptor ξ-chain complex (TCRξ-CD3) and the costimulatory antigen CD28. CD28 interacts with three intracellular proteins-phosphatidylinositol 3-kinase (PI 3-kinase), T cell-specific protein-tyrosine kinase ITK (formerly TSK or EMT), and the complex between growth factor receptor-bound protein 2 and son of sevenless guanine nucleotide exchange protein (GRB-2-SOS). PI 3-kinase and GRB-2 bind to the CD28 phosphotyrosine-based Tyr-Met-Asn-Met motif by means of intrinsic Src-homology 2 (SH2) domains. The requirement for tyrosine phosphorylation of the Tyr-Met-Asn-Met motif for SH2 domain binding implicates an intervening protein-tyrosine kinase in the recruitment of PI 3-kinase and GRB-2 by CD28. Candidate kinases include p56Lck, p59Fyn, ξ-chain-associated 70-kDa protein (ZAP-70), and ITK. In this study, we demonstrate in coexpression studies that p56Lck and p59Fyn phosphorylate CD28 primarily at Tyr-191 of the Tyr-Met-Asn-Met motif, inducing a 3- to 8-fold increase in p85 (subunit of PI 3-kinase) and GRB-2 SH2 binding to CD28. Phosphatase digestion of CD28 eliminated binding. In contrast to Src kinases, ZAP-70 and ITK failed to induce these events. Further, ITK binding to CD28 was dependent on the presence of p56Lck and is thus likely to act downstream of p56Lck/p59Fyn in a signaling cascade. p56Lck is therefore likely to be a central switch in T-cell activation, with the dual function of regulating CD28-mediated costimulation as well as TCR-CD3-CD4 signaling.

  17. Epidermal growth factor receptor activation in glioblastoma through novel missense mutations in the extracellular domain.

    Directory of Open Access Journals (Sweden)

    Jeffrey C Lee

    2006-12-01

    Full Text Available Protein tyrosine kinases are important regulators of cellular homeostasis with tightly controlled catalytic activity. Mutations in kinase-encoding genes can relieve the autoinhibitory constraints on kinase activity, can promote malignant transformation, and appear to be a major determinant of response to kinase inhibitor therapy. Missense mutations in the EGFR kinase domain, for example, have recently been identified in patients who showed clinical responses to EGFR kinase inhibitor therapy.Encouraged by the promising clinical activity of epidermal growth factor receptor (EGFR kinase inhibitors in treating glioblastoma in humans, we have sequenced the complete EGFR coding sequence in glioma tumor samples and cell lines. We identified novel missense mutations in the extracellular domain of EGFR in 13.6% (18/132 of glioblastomas and 12.5% (1/8 of glioblastoma cell lines. These EGFR mutations were associated with increased EGFR gene dosage and conferred anchorage-independent growth and tumorigenicity to NIH-3T3 cells. Cells transformed by expression of these EGFR mutants were sensitive to small-molecule EGFR kinase inhibitors.Our results suggest extracellular missense mutations as a novel mechanism for oncogenic EGFR activation and may help identify patients who can benefit from EGFR kinase inhibitors for treatment of glioblastoma.

  18. Role of protein kinase C and epidermal growth factor receptor signalling in growth stimulation by neurotensin in colon carcinoma cells

    Directory of Open Access Journals (Sweden)

    Dajani Olav

    2011-10-01

    Full Text Available Abstract Background Neurotensin has been found to promote colon carcinogenesis in rats and mice, and proliferation of human colon carcinoma cell lines, but the mechanisms involved are not clear. We have examined signalling pathways activated by neurotensin in colorectal and pancreatic carcinoma cells. Methods Colon carcinoma cell lines HCT116 and HT29 and pancreatic adenocarcinoma cell line Panc-1 were cultured and stimulated with neurotensin or epidermal growth factor (EGF. DNA synthesis was determined by incorporation of radiolabelled thymidine into DNA. Levels and phosphorylation of proteins in signalling pathways were assessed by Western blotting. Results Neurotensin stimulated the phosphorylation of both extracellular signal-regulated kinase (ERK and Akt in all three cell lines, but apparently did so through different pathways. In Panc-1 cells, neurotensin-induced phosphorylation of ERK, but not Akt, was dependent on protein kinase C (PKC, whereas an inhibitor of the β-isoform of phosphoinositide 3-kinase (PI3K, TGX221, abolished neurotensin-induced Akt phosphorylation in these cells, and there was no evidence of EGF receptor (EGFR transactivation. In HT29 cells, in contrast, the EGFR tyrosine kinase inhibitor gefitinib blocked neurotensin-stimulated phosphorylation of both ERK and Akt, indicating transactivation of EGFR, independently of PKC. In HCT116 cells, neurotensin induced both a PKC-dependent phosphorylation of ERK and a metalloproteinase-mediated transactivation of EGFR that was associated with a gefitinib-sensitive phosphorylation of the downstream adaptor protein Shc. The activation of Akt was also inhibited by gefitinib, but only partly, suggesting a mechanism in addition to EGFR transactivation. Inhibition of PKC blocked neurotensin-induced DNA synthesis in HCT116 cells. Conclusions While acting predominantly through PKC in Panc-1 cells and via EGFR transactivation in HT29 cells, neurotensin used both these pathways in HCT116

  19. Role of protein kinase C and epidermal growth factor receptor signalling in growth stimulation by neurotensin in colon carcinoma cells

    International Nuclear Information System (INIS)

    Müller, Kristin M; Tveteraas, Ingun H; Aasrum, Monica; Ødegård, John; Dawood, Mona; Dajani, Olav; Christoffersen, Thoralf; Sandnes, Dagny L

    2011-01-01

    Neurotensin has been found to promote colon carcinogenesis in rats and mice, and proliferation of human colon carcinoma cell lines, but the mechanisms involved are not clear. We have examined signalling pathways activated by neurotensin in colorectal and pancreatic carcinoma cells. Colon carcinoma cell lines HCT116 and HT29 and pancreatic adenocarcinoma cell line Panc-1 were cultured and stimulated with neurotensin or epidermal growth factor (EGF). DNA synthesis was determined by incorporation of radiolabelled thymidine into DNA. Levels and phosphorylation of proteins in signalling pathways were assessed by Western blotting. Neurotensin stimulated the phosphorylation of both extracellular signal-regulated kinase (ERK) and Akt in all three cell lines, but apparently did so through different pathways. In Panc-1 cells, neurotensin-induced phosphorylation of ERK, but not Akt, was dependent on protein kinase C (PKC), whereas an inhibitor of the β-isoform of phosphoinositide 3-kinase (PI3K), TGX221, abolished neurotensin-induced Akt phosphorylation in these cells, and there was no evidence of EGF receptor (EGFR) transactivation. In HT29 cells, in contrast, the EGFR tyrosine kinase inhibitor gefitinib blocked neurotensin-stimulated phosphorylation of both ERK and Akt, indicating transactivation of EGFR, independently of PKC. In HCT116 cells, neurotensin induced both a PKC-dependent phosphorylation of ERK and a metalloproteinase-mediated transactivation of EGFR that was associated with a gefitinib-sensitive phosphorylation of the downstream adaptor protein Shc. The activation of Akt was also inhibited by gefitinib, but only partly, suggesting a mechanism in addition to EGFR transactivation. Inhibition of PKC blocked neurotensin-induced DNA synthesis in HCT116 cells. While acting predominantly through PKC in Panc-1 cells and via EGFR transactivation in HT29 cells, neurotensin used both these pathways in HCT116 cells. In these cells, neurotensin-induced activation of ERK

  20. Rapid phospho-turnover by receptor tyrosine kinases impacts downstream signaling and drug binding.

    Science.gov (United States)

    Kleiman, Laura B; Maiwald, Thomas; Conzelmann, Holger; Lauffenburger, Douglas A; Sorger, Peter K

    2011-09-02

    Epidermal growth factor receptors (ErbB1-4) are oncogenic receptor tyrosine kinases (RTKs) that regulate diverse cellular processes. In this study, we combine measurement and mathematical modeling to quantify phospho-turnover at ErbB receptors in human cells and to determine the consequences for signaling and drug binding. We find that phosphotyrosine residues on ErbB1 have half-lives of a few seconds and therefore turn over 100-1000 times in the course of a typical immediate-early response to ligand. Rapid phospho-turnover is also observed for EGF-activated ErbB2 and ErbB3, unrelated RTKs, and multiple intracellular adaptor proteins and signaling kinases. Thus, the complexes formed on the cytoplasmic tail of active receptors and the downstream signaling kinases they control are highly dynamic and antagonized by potent phosphatases. We develop a kinetic scheme for binding of anti-ErbB1 drugs to receptors and show that rapid phospho-turnover significantly impacts their mechanisms of action. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Rapid Phospho-Turnover by Receptor Tyrosine Kinases Impacts Downstream Signaling and Drug Binding

    OpenAIRE

    Kleiman, Laura B.; Maiwald, Thomas; Conzelmann, Holger; Lauffenburger, Douglas A.; Sorger, Peter K.

    2011-01-01

    Epidermal growth factor receptors (ErbB1–4) are oncogenic receptor tyrosine kinases (RTKs) that regulate diverse cellular processes. In this study, we combine measurement and mathematical modeling to quantify phospho-turnover at ErbB receptors in human cells and to determine the consequences for signaling and drug binding. We find that phosphotyrosine residues on ErbB1 have half-lives of a few seconds and therefore turn over 100–1000 times in the course of a typical immediate-early response t...

  2. E3B1, a human homologue of the mouse gene product Abi-1, sensitizes activation of Rap1 in response to epidermal growth factor

    International Nuclear Information System (INIS)

    Jenei, Veronika; Andersson, Tommy; Jakus, Judit; Dib, Karim

    2005-01-01

    E3B1, a human homologue of the mouse gene product Abi-1, has been implicated in growth-factor-mediated regulation of the small GTPases p21 Ras and Rac. E3b1 is a regulator of Rac because it can form a complex with Sos-1 and eps8, and such a Sos-1-e3B1-eps8 complex serves as a guanine nucleotide exchange factor for Rac. In the present study, we found that overexpression of e3B1 in NIH3T3/EGFR cells sensitized EGF-induced activation of Rac1, whereas it had no impact on EGF-induced activation of p21 Ras . Remarkably, we found that EGF-induced activation of the p21 Ras -related GTPase Rap1 was also sensitized in NIH3T3/EGFR-e3B1 cells. Thus, in NIH3T3/EGFR-e3B1 cells, maximal EGF-induced activation of Rap1 occurs with a dose of EGF much lower than in NIH3T3/EGFR cells. We also report that overexpression of e3B1 in NIH3T3/EGFR cells renders EGF-induced activation of Rap1 completely dependent on Src tyrosine kinases but not on c-Abl. However, EGF-induced tyrosine phosphorylation of the Rap GEF C3G occurred regardless of whether e3B1 was overexpressed or not, and this did not involve Src tyrosine kinases. Accordingly, we propose that overexpression of e3B1 in NIH3T3/EGFR cells leads to mobilization of Src tyrosine kinases that participate in EGF-induced activation of Rap1 and inhibition of cell proliferation

  3. X-linked agammaglobulinemia - first case with Bruton tyrosine kinase mutation from Pakistan.

    Science.gov (United States)

    Zaidi, Samreen Kulsom; Qureshi, Sonia; Qamar, Farah Naz

    2017-03-01

    X-linked agammaglobulinemia (XLA) is a primary immunodeficiency with more than 600 mutations in Bruton tyrosine kinase (Bkt) gene which are responsible for early-onset agammaglobulinemia and repeated infections. Herein we present a case of a 3-year-old boy with history of repeated diarrhoea and an episode of meningoencephalitis with hemiplegia. The workup showed extremely low levels of immunoglobulin with low CD+19 cells. Genetic analysis showed Btk mutation 18 c.1883delCp.T628fs. To the best of our knowledge this is the first report of a case of XLA confirmed by molecular technique from Pakistan.

  4. Cell adhesion and EGFR activation regulate EphA2 expression in cancer

    DEFF Research Database (Denmark)

    Larsen, Alice Bjerregaard; Stockhausen, Marie-Thérése; Poulsen, Hans Skovgaard

    2010-01-01

    largely unknown. Here we show that the expression of EphA2 in in vitro cultured cells, is restricted to cells growing adherently and that adhesion-induced EphA2 expression is dependent upon activation of the epidermal growth factor receptor (EGFR), mitogen activated protein kinase kinase (MEK) and Src...... family kinases (SRC). Moreover, the results show that adhesion-induced EGFR activation and EphA2 expression is affected by interactions with extracellular matrix (ECM) proteins working as integrin ligands. Stimulation with the EphA2 ligand, ephrinA1 inhibited ERK phosphorylation and cancer cell viability....... These effects were however abolished by activation of the EGF-receptor ligand system favoring Ras/MAPK signaling and cell proliferation. Based on our results, we propose a regulatory mechanism where cell adhesion induces EGFR kinase activation and EphA2 expression; and where the effect of ephrinA1 mediated...

  5. Phosphorylation of SLP-76 by the ZAP-70 protein-tyrosine kinase is required for T-cell receptor function.

    Science.gov (United States)

    Bubeck Wardenburg, J; Fu, C; Jackman, J K; Flotow, H; Wilkinson, S E; Williams, D H; Johnson, R; Kong, G; Chan, A C; Findell, P R

    1996-08-16

    Two families of tyrosine kinases, the Src and Syk families, are required for T-cell receptor activation. While the Src kinases are responsible for phosphorylation of receptor-encoded signaling motifs and for up-regulation of ZAP-70 activity, the downstream substrates of ZAP-70 are unknown. Evidence is presented herein that the Src homology 2 (SH2) domain-containing leukocyte protein of 76 kDa (SLP-76) is a substrate of ZAP-70. Phosphorylation of SLP-76 is diminished in T cells that express a catalytically inactive ZAP-70. Moreover, SLP-76 is preferentially phosphorylated by ZAP-70 in vitro and in heterologous cellular systems. In T cells, overexpression of wild-type SLP-76 results in a hyperactive receptor, while expression of a SLP-76 molecule that is unable to be tyrosine-phosphorylated attenuates receptor function. In addition, the SH2 domain of SLP-76 is required for T-cell receptor function, although its role is independent of the ability of SLP-76 to undergo tyrosine phosphorylation. As SLP-76 interacts with both Grb2 and phospholipase C-gamma1, these data indicate that phosphorylation of SLP-76 by ZAP-70 provides an important functional link between the T-cell receptor and activation of ras and calcium pathways.

  6. Molecular mechanism of 17-allylamino-17-demethoxygeldanamycin (17-AAG)-induced AXL receptor tyrosine kinase degradation.

    Science.gov (United States)

    Krishnamoorthy, Gnana Prakasam; Guida, Teresa; Alfano, Luigi; Avilla, Elvira; Santoro, Massimo; Carlomagno, Francesca; Melillo, Rosa Marina

    2013-06-14

    The receptor tyrosine kinase AXL is overexpressed in many cancer types including thyroid carcinomas and has well established roles in tumor formation and progression. Proper folding, maturation, and activity of several oncogenic receptor tyrosine kinases require HSP90 chaperoning. HSP90 inhibition by the antibiotic geldanamycin or its derivative 17-allylamino-17-demethoxygeldanamycin (17-AAG) causes destabilization of its client proteins. Here we show that AXL is a novel client protein of HSP90. 17-AAG induced a time- and dose-dependent down-regulation of endogenous or ectopically expressed AXL protein, thereby inhibiting AXL-mediated signaling and biological activity. 17-AAG-induced AXL down-regulation specifically affected fully glycosylated mature receptor present on cell membrane. By using biotin and [(35)S]methionine labeling, we showed that 17-AAG caused depletion of membrane-localized AXL by mediating its degradation in the intracellular compartment, thus restricting its exposure on the cell surface. 17-AAG induced AXL polyubiquitination and subsequent proteasomal degradation; under basal conditions, AXL co-immunoprecipitated with HSP90. Upon 17-AAG treatment, AXL associated with the co-chaperone HSP70 and the ubiquitin E3 ligase carboxyl terminus of HSC70-interacting protein (CHIP). Overexpression of CHIP, but not of the inactive mutant CHIP K30A, induced accumulation of AXL polyubiquitinated species upon 17-AAG treatment. The sensitivity of AXL to 17-AAG required its intracellular domain because an AXL intracellular domain-deleted mutant was insensitive to the compound. Active AXL and kinase-dead AXL were similarly sensitive to 17-AAG, implying that 17-AAG sensitivity does not require receptor phosphorylation. Overall our data elucidate the molecular basis of AXL down-regulation by HSP90 inhibitors and suggest that HSP90 inhibition in anticancer therapy can exert its effect through inhibition of multiple kinases including AXL.

  7. Molecular Mechanism of 17-Allylamino-17-demethoxygeldanamycin (17-AAG)-induced AXL Receptor Tyrosine Kinase Degradation*

    Science.gov (United States)

    Krishnamoorthy, Gnana Prakasam; Guida, Teresa; Alfano, Luigi; Avilla, Elvira; Santoro, Massimo; Carlomagno, Francesca; Melillo, Rosa Marina

    2013-01-01

    The receptor tyrosine kinase AXL is overexpressed in many cancer types including thyroid carcinomas and has well established roles in tumor formation and progression. Proper folding, maturation, and activity of several oncogenic receptor tyrosine kinases require HSP90 chaperoning. HSP90 inhibition by the antibiotic geldanamycin or its derivative 17-allylamino-17-demethoxygeldanamycin (17-AAG) causes destabilization of its client proteins. Here we show that AXL is a novel client protein of HSP90. 17-AAG induced a time- and dose-dependent down-regulation of endogenous or ectopically expressed AXL protein, thereby inhibiting AXL-mediated signaling and biological activity. 17-AAG-induced AXL down-regulation specifically affected fully glycosylated mature receptor present on cell membrane. By using biotin and [35S]methionine labeling, we showed that 17-AAG caused depletion of membrane-localized AXL by mediating its degradation in the intracellular compartment, thus restricting its exposure on the cell surface. 17-AAG induced AXL polyubiquitination and subsequent proteasomal degradation; under basal conditions, AXL co-immunoprecipitated with HSP90. Upon 17-AAG treatment, AXL associated with the co-chaperone HSP70 and the ubiquitin E3 ligase carboxyl terminus of HSC70-interacting protein (CHIP). Overexpression of CHIP, but not of the inactive mutant CHIP K30A, induced accumulation of AXL polyubiquitinated species upon 17-AAG treatment. The sensitivity of AXL to 17-AAG required its intracellular domain because an AXL intracellular domain-deleted mutant was insensitive to the compound. Active AXL and kinase-dead AXL were similarly sensitive to 17-AAG, implying that 17-AAG sensitivity does not require receptor phosphorylation. Overall our data elucidate the molecular basis of AXL down-regulation by HSP90 inhibitors and suggest that HSP90 inhibition in anticancer therapy can exert its effect through inhibition of multiple kinases including AXL. PMID:23629654

  8. Research Progress of the Resistance Mechanism of Non-small Cell Lung Cancer 
to EGFR-TKIs

    Directory of Open Access Journals (Sweden)

    Huihui LIU

    2013-10-01

    Full Text Available Nowadays, lung cancer is the malignant tumor of the highest morbidity and mortality over the world, and non-small cell lung cancer (NSCLC makes up about 80%. There is a great many NSCLC patients have been in advanced stage when diagnosed. As a result, people pay more attention to curing advanced NSCLC. The standard treatment to advanced NSCLC is platinum-based combined chemotherapy. However, chemotherapy drugs usually have limited effects on improving the survival of the patients. Then exploring new therapies is extremely urgent to us. Now, molecular targeted therapy has been the most promising research area for the treatment of NSCLC with researches going deep into pathogenesis and biological behavior of lung cancer. Epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs have achieved a great success in the treatment of advanced NSCLC. Their representatives are erlotinib and gefitinib. The two drugs have been widely used to treat advanced NSCLCs worldwide, especially for the patients with EGFR activating mutations. However, after a period of treatment (median time is 6 to 12 months, most patients will develop drug resistance to EGFR-TKIs. Intense research in these NSCLCs has identified two major mechanisms of resistance to TKIs: primary and acquired resistances. The research about resistance mechanism of NSCLC to EGFR-TKIs is a hot one because of their excellent effects on improving overall and progression-free survival. The aim of this article was to summarize the development of the resistance mechanisms.

  9. PTP1B Inhibition Causes Rac1 Activation by Enhancing Receptor Tyrosine Kinase Signaling

    Directory of Open Access Journals (Sweden)

    Ayako Tsuchiya

    2014-04-01

    Full Text Available Background/Aims: The present study investigated the signaling pathway underlying Rac1 activation induced by the linoleic acid derivative 8-[2-(2-pentyl-cyclopropylmethyl-cyclopropyl]-octanoic acid (DCP-LA. Methods: Activity of protein tyrosine phosphatase 1B (PTP1B was assayed under cell-free conditions. Western blot was carried out to quantify phosphorylation of insulin receptor substrate-1 (IRS-1 and Akt in PC-12 cells. Rac1 activity was monitored in the föerster resonance energy transfer (FRET analysis using living and fixed PC-12 cells. Results: DCP-LA markedly suppressed PTP1B activity in a concentration (100 pM-100 µM-dependent manner. In the DCP-LA binding assay, fluorescein-conjugated DCP-LA produced a single fluorescent signal band at 60 kDa, corresponding to the molecule of PTP1B, and the signal was attenuated or abolished by co-treatment or pretreatment with non-conjugated DCP-LA. DCP-LA significantly enhanced nerve growth factor (NGF-stimulated phosphorylation of IRS-1 at Tyr1222 and Akt1/2 at Thr308/309 and Ser473/474 in PC-12 cells. In the FRET analysis, DCP-LA significantly enhanced NGF-stimulated Rac1 activation, which is abrogated by the phosphatidylinositol 3 kinase (PI3K inhibitor wortmannin, the 3-phosphoinositide-dependent protein kinase-1 (PDK1 inhibitor BX912, or the Akt inhibitor MK2206. Conclusion: The results of the present study show that DCP-LA-induced PTP1B inhibition, possibly through its direct binding, causes Rac1 activation by enhancing a pathway along a receptor tyrosine kinase (RTK/IRS-1/PI3K/Akt/Rac1 axis.

  10. Role of tyrosine phosphatase inhibitors in cancer treatment with emphasis on SH2 domain-containing tyrosine phosphatases (SHPs)

    NARCIS (Netherlands)

    Irandoust, Mahban; van den Berg, Timo K.; Kaspers, Gertjan J. L.; Cloos, Jacqueline

    2009-01-01

    Protein tyrosine phosphorylation is one of the key mechanisms involved in signal transduction pathways. This modification is regulated by concerted action of protein tyrosine phosphatases and protein tyrosine kinases. Deregulation of either of these key regulators lead to abnormal cellular

  11. Crystal structure of the EphA4 protein tyrosine kinase domain in the apo-and dasantinib-bound state

    NARCIS (Netherlands)

    Farenc, C; Celie, C; Tensen, P.H.N; de Esch, I.J.P.; Siegal, C.P.

    2011-01-01

    The Eph family of receptor tyrosine kinases regulates diverse cellular processes while the over-expression of a member of this family, EphA4, has been reported in a variety of malignant carcinomas. To gain insight into molecular mechanisms and to facilitate structure-based inhibitor design, we

  12. 1-Methoxy-agroclavine from Penicillium sp. WC75209, a novel inhibitor of the Lck tyrosine kinase.

    Science.gov (United States)

    Padmanabha, R; Shu, Y Z; Cook, L S; Veitch, J A; Donovan, M; Lowe, S; Huang, S; Pirnik, D; Manly, S P

    1998-03-17

    A high-throughput screen was developed and implemented to identify inhibitors of the Lck tyrosine kinase. This report describes the identification of a specific inhibitor of this enzyme from the solid fermentation culture of the Penicillium sp., WC75209. The active compound was isolated and structurally characterized as 1-methoxy-5R, 10S-agroclavine, a new member of the ergot alkaloid family.

  13. Identification of SH2-Bbeta as a substrate of the tyrosine kinase JAK2 involved in growth hormone signaling.

    OpenAIRE

    Rui, L; Mathews, L S; Hotta, K; Gustafson, T A; Carter-Su, C

    1997-01-01

    Activation of the tyrosine kinase JAK2 is an essential step in cellular signaling by growth hormone (GH) and multiple other hormones and cytokines. Murine JAK2 has a total of 49 tyrosines which, if phosphorylated, could serve as docking sites for Src homology 2 (SH2) or phosphotyrosine binding domain-containing signaling molecules. Using a yeast two-hybrid screen of a rat adipocyte cDNA library, we identified a splicing variant of the SH2 domain-containing protein SH2-B, designated SH2-Bbeta,...

  14. Acquired EGFR L718V mutation mediates resistance to osimertinib in non-small cell lung cancer but retains sensitivity to afatinib.

    Science.gov (United States)

    Liu, Yutao; Li, Yan; Ou, Qiuxiang; Wu, Xue; Wang, Xiaonan; Shao, Yang W; Ying, Jianming

    2018-04-01

    Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are promising targeted therapies for EGFR-mutated non-small-cell lung cancer (NSCLC) patients. However, acquired resistance inevitably develops. Comprehensive and dynamic companion genomic diagnosis can gain insights into underlying resistance mechanisms, thereby help oncologists and patients to make informed decision on the potential benefit of the treatment. A 67-year-old male who was initially diagnosed of EGFR L858R-mediated NSCLC received multiple lines of chemotherapy and EGFR TKI therapies after surgery. The EGFR mutational status of individual metastatic lesion was determined by genetic testing of the tumor tissue biopsies using next generation sequencing (NGS) throughout the patient's clinical course. An acquired potentially drug-resistant EGFR mutation was functionally validated in vitro and its sensitivity to different EGFR TKIs was assessed simultaneously. We have identified distinct resistance mechanisms to EGFR blockade in different metastatic lung lesions. Acquired EGFR T790M was first detected that leads to the resistance to the gefitinib treatment. Consequently, osimertinib was administrated and the response lasted until disease progressed. We identified a newly acquired EGFR L718V mutation in one lesion in conjunction with L858R, but not T790M, which showed stable disease on the following erlotinib treatment, while EGFR C797S together with L858R/T790M was detected in the other lesion that continuously progressed. In vitro functional studies demonstrated that EGFR-L858R/L718V confers resistance to osimertinib, but retains sensitivity to the second generation TKI afatinib. We reported that distinct resistance mechanisms could arise in different metastases within the same patient in response to EGFR blockade. We also demonstrated in vitro that EGFR L718V mutation mediates resistance to osimertinib, but retains sensitivity to afatinib. We evidenced that dynamic companion genomic

  15. Effects of Src Kinase Inhibition on Expression of Protein Tyrosine Phosphatase 1B after Brain Hypoxia in a Piglet Animal Model

    Directory of Open Access Journals (Sweden)

    Dimitrios Angelis

    2017-01-01

    Full Text Available Background. Protein tyrosine phosphatases (PTPs in conjunction with protein tyrosine kinases (PTKs regulate cellular processes by posttranslational modifications of signal transduction proteins. PTP nonreceptor type 1B (PTP-1B is an enzyme of the PTP family. We have previously shown that hypoxia induces an increase in activation of a class of nonreceptor PTK, the Src kinases. In the present study, we investigated the changes that occur in the expression of PTP-1B in the cytosolic component of the brain of newborn piglets acutely after hypoxia as well as long term for up to 2 weeks. Methods. Newborn piglets were divided into groups: normoxia, hypoxia, hypoxia followed by 1 day and 15 days in FiO2 0.21, and hypoxia pretreated with Src kinase inhibitor PP2, prior to hypoxia followed by 1 day and 15 days. Hypoxia was achieved by providing 7% FiO2 for 1 hour and PTP-1B expression was measured via immunoblotting. Results. PTP-1B increased posthypoxia by about 30% and persisted for 2 weeks while Src kinase inhibition attenuated the expected PTP-1B-increased expression. Conclusions. Our study suggests that Src kinase mediates a hypoxia-induced increased PTP-1B expression.

  16. Allosteric inhibition of SHP2 phosphatase inhibits cancers driven by receptor tyrosine kinases

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ying-Nan P.; LaMarche, Matthew J.; Chan, Ho Man; Fekkes, Peter; Garcia-Fortanet, Jorge; Acker, Michael G.; Antonakos, Brandon; Chen, Christine Hiu-Tung; Chen, Zhouliang; Cooke, Vesselina G.; Dobson, Jason R.; Deng, Zhan; Fei, Feng; Firestone, Brant; Fodor, Michelle; Fridrich, Cary; Gao, Hui; Grunenfelder, Denise; Hao, Huai-Xiang; Jacob, Jaison; Ho, Samuel; Hsiao, Kathy; Kang, Zhao B.; Karki, Rajesh; Kato, Mitsunori; Larrow, Jay; La Bonte, Laura R.; Lenoir, Francois; Liu, Gang; Liu, Shumei; Majumdar, Dyuti; Meyer, Matthew J.; Palermo, Mark; Perez, Lawrence; Pu, Minying; Price, Edmund; Quinn, Christopher; Shakya, Subarna; Shultz, Michael D.; Slisz, Joanna; Venkatesan, Kavitha; Wang, Ping; Warmuth, Markus; Williams, Sarah; Yang, Guizhi; Yuan, Jing; Zhang, Ji-Hu; Zhu, Ping; Ramsey, Timothy; Keen, Nicholas J.; Sellers, William R.; Stams, Travis; Fortin , Pascal D. (Novartis)

    2016-06-29

    The non-receptor protein tyrosine phosphatase SHP2, encoded by PTPN11, has an important role in signal transduction downstream of growth factor receptor signalling and was the first reported oncogenic tyrosine phosphatase1. Activating mutations of SHP2 have been associated with developmental pathologies such as Noonan syndrome and are found in multiple cancer types, including leukaemia, lung and breast cancer and neuroblastoma1, 2, 3, 4, 5. SHP2 is ubiquitously expressed and regulates cell survival and proliferation primarily through activation of the RAS–ERK signalling pathway2, 3. It is also a key mediator of the programmed cell death 1 (PD-1) and B- and T-lymphocyte attenuator (BTLA) immune checkpoint pathways6, 7. Reduction of SHP2 activity suppresses tumour cell growth and is a potential target of cancer therapy8, 9. Here we report the discovery of a highly potent (IC50 = 0.071 μM), selective and orally bioavailable small-molecule SHP2 inhibitor, SHP099, that stabilizes SHP2 in an auto-inhibited conformation. SHP099 concurrently binds to the interface of the N-terminal SH2, C-terminal SH2, and protein tyrosine phosphatase domains, thus inhibiting SHP2 activity through an allosteric mechanism. SHP099 suppresses RAS–ERK signalling to inhibit the proliferation of receptor-tyrosine-kinase-driven human cancer cells in vitro and is efficacious in mouse tumour xenograft models. Together, these data demonstrate that pharmacological inhibition of SHP2 is a valid therapeutic approach for the treatment of cancers.

  17. Regulation of Src family kinases involved in T cell receptor signaling by protein-tyrosine phosphatase CD148

    Czech Academy of Sciences Publication Activity Database

    Štěpánek, Ondřej; Kalina, T.; Dráber, Peter; Skopcová, Tereza; Svojgr, K.; Angelisová, Pavla; Hořejší, Václav; Weiss, A.; Brdička, Tomáš

    2011-01-01

    Roč. 286, č. 25 (2011), s. 22101-22112 ISSN 0021-9258 R&D Projects: GA MŠk 2B06064; GA MŠk 1M0506 Institutional research plan: CEZ:AV0Z50520514 Keywords : CD148 * tyrosine phosphatase * Src family kinases Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.773, year: 2011

  18. Tyrosine kinase inhibitors induced immune thrombocytopenia in chronic myeloid leukemia?

    Directory of Open Access Journals (Sweden)

    Avital F. Barak

    2011-12-01

    Full Text Available The outcome and quality of life of chronic myeloid leukemia (CML patients has remarkably changed with the treatment of tyrosine kinase inhibitors (TKIs. Currently, hematopoietic stem cell transplantation (HSCT is considered mainly as a third line salvage therapy in cases of TKIs resistance or intolerance. Here we describe a patient with chronic phase CML who developed both resistance and late occurrence of s severe thrombocytopenia on first and second generation TKIs and eventually underwent HSCT. Although the mechanism of the myelosuppression is not fully understood, we showed for the first time the development of dose dependent platelet antibodies in the presence of TKIs, suggesting the possibility of TKIs induced thrombocytopenia. Our case emphasizes that late development of severe myelosuppression during imatinib treatment is probably an important indication for consideration of early HSCT.

  19. A bipolar clamp mechanism for activation of Jak-family protein tyrosine kinases.

    Directory of Open Access Journals (Sweden)

    Dipak Barua

    2009-04-01

    Full Text Available Most cell surface receptors for growth factors and cytokines dimerize in order to mediate signal transduction. For many such receptors, the Janus kinase (Jak family of non-receptor protein tyrosine kinases are recruited in pairs and juxtaposed by dimerized receptor complexes in order to activate one another by trans-phosphorylation. An alternative mechanism for Jak trans-phosphorylation has been proposed in which the phosphorylated kinase interacts with the Src homology 2 (SH2 domain of SH2-B, a unique adaptor protein with the capacity to homo-dimerize. Building on a rule-based kinetic modeling approach that considers the concerted nature and combinatorial complexity of modular protein domain interactions, we examine these mechanisms in detail, focusing on the growth hormone (GH receptor/Jak2/SH2-Bbeta system. The modeling results suggest that, whereas Jak2-(SH2-Bbeta(2-Jak2 heterotetramers are scarcely expected to affect Jak2 phosphorylation, SH2-Bbeta and dimerized receptors synergistically promote Jak2 trans-activation in the context of intracellular signaling. Analysis of the results revealed a unique mechanism whereby SH2-B and receptor dimers constitute a bipolar 'clamp' that stabilizes the active configuration of two Jak2 molecules in the same macro-complex.

  20. [Literature review and presentation of our own research results regarding the effects on bone of tyrosine kinase inhibitors imatinib and nilotinib used in the treatment of oncohematological diseases].

    Science.gov (United States)

    Kirschner, Gyöngyi; Balla, Bernadett; Kósa, János; Horváth, Péter; Kövesdi, Andrea; Lakatos, Gergely; Takács, István; Nagy, Zsolt; Tóbiás, Bálint; Árvai, Kristóf; Lakatos, Péter

    2016-09-01

    Tyrosine kinase inhibitors are widely used for treatment of certain oncohematological diseases. Several clinical studies have confirmed that specific BCR-ABL tyrosine kinase inhibitors alter the physiological process of bone tissue in a complex and unclearly identified manner. Since these treatments are being given to more and more patients, and the therapy takes decades or lasts even lifelong, it is justifiable to obtain more detailed knowledge of the molecular background of these mechanisms. In this article the authors summarize preliminary research results and human clinical observations on imatinib and nilotinib which are related to bone metabolism, and present the results of their own experiments in in vitro osteoblast cultures. Based on the presented results, the effects of imatinib and nilotinib on bone cells depend on the concentration of imatinib and nilotinib, the maturation stage of the cells and the distribution ratio of receptor tyrosine kinase signaling pathways. In this study the authors firstly prepared a stop-gap, comprehensive review in the Hungarian literature, regarding the effects of tyrosine kinase inhibitors on bone metabolism. In addition they firstly performed whole transcriptome analysis on osteoblasts in order to obtain a better understanding of the cellular molecular mechanisms. Orv. Hetil., 2016, 157(36), 1429-1437.

  1. Ibrutinib (PCI-32765), the first BTK (Bruton's tyrosine kinase) inhibitor in clinical trials.

    Science.gov (United States)

    Brown, Jennifer R

    2013-03-01

    Ibrutinib is a potent covalent kinase inhibitor that targets BTK. BTK, or Bruton's tyrosine kinase, is an obvious target for therapy of B cell diseases because inactivating mutations lead to B cell aplasia in humans and the disease X-linked agammaglobulinemia. Ibrutinib has modest cytotoxicity against CLL cells in vitro but also blocks trophic stimuli from the microenvironment. As with other inhibitors of the BCR pathway, ibrutinib causes rapid nodal reduction and response associated with rapid increase in lymphocytosis, which then returns to baseline over time. The ORR of ibrutinib in relapsed refractory CLL is 67 % with PFS 88 % at 15 months. In a cohort of untreated patients 65 years and over, the estimated 15 month PFS is 96 %. Registration trials have been initiated, and the difficult task that remains is to determine where in the course of CLL therapy this drug will have the greatest impact and benefit for patients.

  2. Integrated Experimental and Model-based Analysis Reveals the Spatial Aspects of EGFR Activation Dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Shankaran, Harish; Zhang, Yi; Chrisler, William B.; Ewald, Jonathan A.; Wiley, H. S.; Resat, Haluk

    2012-10-02

    The epidermal growth factor receptor (EGFR) belongs to the ErbB family of receptor tyrosine kinases, and controls a diverse set of cellular responses relevant to development and tumorigenesis. ErbB activation is a complex process involving receptor-ligand binding, receptor dimerization, phosphorylation, and trafficking (internalization, recycling and degradation), which together dictate the spatio-temporal distribution of active receptors within the cell. The ability to predict this distribution, and elucidation of the factors regulating it, would help to establish a mechanistic link between ErbB expression levels and the cellular response. Towards this end, we constructed mathematical models for deconvolving the contributions of receptor dimerization and phosphorylation to EGFR activation, and to examine the dependence of these processes on sub-cellular location. We collected experimental datasets for EGFR activation dynamics in human mammary epithelial cells, with the specific goal of model parameterization, and used the data to estimate parameters for several alternate models. Model-based analysis indicated that: 1) signal termination via receptor dephosphorylation in late endosomes, prior to degradation, is an important component of the response, 2) less than 40% of the receptors in the cell are phosphorylated at any given time, even at saturating ligand doses, and 3) receptor dephosphorylation rates at the cell surface and early endosomes are comparable. We validated the last finding by measuring EGFR dephosphorylation rates at various times following ligand addition both in whole cells, and in endosomes using ELISAs and fluorescent imaging. Overall, our results provide important information on how EGFR phosphorylation levels are regulated within cells. Further, the mathematical model described here can be extended to determine receptor dimer abundances in cells co-expressing various levels of ErbB receptors. This study demonstrates that an iterative cycle of

  3. Focal Adhesion Kinase Inhibitors in Combination with Erlotinib Demonstrate Enhanced Anti-Tumor Activity in Non-Small Cell Lung Cancer.

    Directory of Open Access Journals (Sweden)

    Grant A Howe

    Full Text Available Blockade of epidermal growth factor receptor (EGFR activity has been a primary therapeutic target for non-small cell lung cancers (NSCLC. As patients with wild-type EGFR have demonstrated only modest benefit from EGFR tyrosine kinase inhibitors (TKIs, there is a need for additional therapeutic approaches in patients with wild-type EGFR. As a key component of downstream integrin signalling and known receptor cross-talk with EGFR, we hypothesized that targeting focal adhesion kinase (FAK activity, which has also been shown to correlate with aggressive stage in NSCLC, would lead to enhanced activity of EGFR TKIs. As such, EGFR TKI-resistant NSCLC cells (A549, H1299, H1975 were treated with the EGFR TKI erlotinib and FAK inhibitors (PF-573,228 or PF-562,271 both as single agents and in combination. We determined cell viability, apoptosis and 3-dimensional growth in vitro and assessed tumor growth in vivo. Treatment of EGFR TKI-resistant NSCLC cells with FAK inhibitor alone effectively inhibited cell viability in all cell lines tested; however, its use in combination with the EGFR TKI erlotinib was more effective at reducing cell viability than either treatment alone when tested in both 2- and 3-dimensional assays in vitro, with enhanced benefit seen in A549 cells. This increased efficacy may be due in part to the observed inhibition of Akt phosphorylation when the drugs were used in combination, where again A549 cells demonstrated the most inhibition following treatment with the drug combination. Combining erlotinib with FAK inhibitor was also potent in vivo as evidenced by reduced tumor growth in the A549 mouse xenograft model. We further ascertained that the enhanced sensitivity was irrespective of the LKB1 mutational status. In summary, we demonstrate the effectiveness of combining erlotinib and FAK inhibitors for use in known EGFR wild-type, EGFR TKI resistant cells, with the potential that a subset of cell types, which includes A549, could be

  4. Structural coupling of SH2-kinase domains links Fes and Abl substrate recognition and kinase activation.

    Science.gov (United States)

    Filippakopoulos, Panagis; Kofler, Michael; Hantschel, Oliver; Gish, Gerald D; Grebien, Florian; Salah, Eidarus; Neudecker, Philipp; Kay, Lewis E; Turk, Benjamin E; Superti-Furga, Giulio; Pawson, Tony; Knapp, Stefan

    2008-09-05

    The SH2 domain of cytoplasmic tyrosine kinases can enhance catalytic activity and substrate recognition, but the molecular mechanisms by which this is achieved are poorly understood. We have solved the structure of the prototypic SH2-kinase unit of the human Fes tyrosine kinase, which appears specialized for positive signaling. In its active conformation, the SH2 domain tightly interacts with the kinase N-terminal lobe and positions the kinase alphaC helix in an active configuration through essential packing and electrostatic interactions. This interaction is stabilized by ligand binding to the SH2 domain. Our data indicate that Fes kinase activation is closely coupled to substrate recognition through cooperative SH2-kinase-substrate interactions. Similarly, we find that the SH2 domain of the active Abl kinase stimulates catalytic activity and substrate phosphorylation through a distinct SH2-kinase interface. Thus, the SH2 and catalytic domains of active Fes and Abl pro-oncogenic kinases form integrated structures essential for effective tyrosine kinase signaling.

  5. 2D QSAR studies of the inhibitory activity of a series of substituted purine derivatives against c-Src tyrosine kinase

    Directory of Open Access Journals (Sweden)

    Mukesh C. Sharma

    2016-07-01

    Full Text Available A series of 34 substituted purine analogues derivatives were subjected to quantitative structure-activity relationship analyses as inhibitors of c-Src tyrosine kinase. Partial least squares regression was applied to derive QSAR models, which were further validated for statistical significance by internal and external validation. The best QSAR model developed had a good predictive correlation coefficient (r2 of 0.8319, a significant cross-validated correlation coefficient (q2 of 0.7550, and an r2 for the external test set (pred_r2 of 0.7983. It was developed from the PLS method with descriptors including the SsCH3E-index, H-Donor Count, T_2_Cl_3, and negative correlation with SsOHcount. The current study provides better insight into the future design of more potent c-Src tyrosine kinase inhibitors prior to synthesis.

  6. Requirement of ERα and basal activities of EGFR and Src kinase in Cd-induced activation of MAPK/ERK pathway in human breast cancer MCF-7 cells

    Energy Technology Data Exchange (ETDEWEB)

    Song, Xiulong, E-mail: songxiulong@hotmail.com; Wei, Zhengxi; Shaikh, Zahir A., E-mail: zshaikh@uri.edu

    2015-08-15

    Cadmium (Cd) is a common environmental toxicant and an established carcinogen. Epidemiological studies implicate Cd with human breast cancer. Low micromolar concentrations of Cd promote proliferation of human breast cancer cells in vitro. The growth promotion of breast cancer cells is associated with the activation of MAPK/ERK pathway. This study explores the mechanism of Cd-induced activation of MAPK/ERK pathway. Specifically, the role of cell surface receptors ERα, EGFR, and Src kinase was evaluated in human breast cancer MCF-7 cells treated with 1–3 μM Cd. The activation of ERK was studied using a serum response element (SRE) luciferase reporter assay. Receptor phosphorylation was detected by Western blot analyses. Cd treatment increased both the SRE reporter activity and ERK1/2 phosphorylation in a concentration-dependent manner. Cd treatment had no effect on reactive oxygen species (ROS) generation. Also, blocking the entry of Cd into the cells with manganese did not diminish Cd-induced activation of MAPK/ERK. These results suggest that the effect of Cd was likely not caused by intracellular ROS generation, but through interaction with the membrane receptors. While Cd did not appear to activate either EGFR or Src kinase, their inhibition completely blocked the Cd-induced activation of ERK as well as cell proliferation. Similarly, silencing ERα with siRNA or use of ERα antagonist blocked the effects of Cd. Based on these results, it is concluded that not only ERα, but also basal activities of EGFR and Src kinase are essential for Cd-induced signal transduction and activation of MAPK/ERK pathway for breast cancer cell proliferation. - Highlights: • Low micromolar concentrations of Cd rapidly activate ERK1/2 in MCF-7 cells. • Signal transduction and resulting cell proliferation require EGFR, ERα, and Src. • These findings implicate Cd in promotion of breast cancer.

  7. Complete response in gallbladder cancer to erlotinib plus gemcitabine does not require mutation of the epidermal growth factor receptor gene: a case report

    Directory of Open Access Journals (Sweden)

    Lincer Robert

    2010-10-01

    Full Text Available Abstract Background Gallbladder cancer typically follows an aggressive course, with chemotherapy the standard of care for advanced disease; complete remissions are rarely encountered. The epidermal growth factor receptor (EGFR is a promising therapeutic target but the activity of single agent oral EGFR tyrosine kinase inhibitors is low. There have been no previous reports of chemotherapy plus an EGFR-tyrosine kinase inhibitor (TKI to treat gallbladder cancer or correlations of response with the mutation status of the tyrosine kinase domain of the EGFR gene. Case presentation A 67 year old man with metastatic gallbladder cancer involving the liver and abdominal lymph nodes was treated with gemcitabine (1000 mg/m2 on day 1 and 8 every 21 days as well as daily erlotinib (100 mg. After four cycles of therapy, the CA 19-9 normalized and a PET/CT showed a complete remission; this response was maintained by the end of 12 cycles of therapy. Gemcitabine was then discontinued and single agent erlotinib was continued as maintenance therapy. The disease remains in good control 18 months after initiation of therapy, including 6 months on maintenance erlotinib. The only grade 3 toxicity was a typical EGFR-related skin rash. Because of the remarkable response to erlotinib plus gemcitabine, we performed tumor genotyping of the EGFR gene for response predicting mutations in exons 18, 19 and 21. This disclosed the wild-type genotype with no mutations found. Conclusion This case report demonstrates a patient with stage IV gallbladder cancer who experienced a rarely encountered complete, prolonged response after treatment with an oral EGFR-TKI plus chemotherapy. This response occurred in the absence of an EGFR gene mutation. These observations should inform the design of clinical trials using EGFR-TKIs to treat gallbladder and other biliary tract cancers; such trials should not select patients based on EGFR mutation status.

  8. Complete response in gallbladder cancer to erlotinib plus gemcitabine does not require mutation of the epidermal growth factor receptor gene: a case report

    International Nuclear Information System (INIS)

    Mody, Kabir; Strauss, Edward; Lincer, Robert; Frank, Richard C

    2010-01-01

    Gallbladder cancer typically follows an aggressive course, with chemotherapy the standard of care for advanced disease; complete remissions are rarely encountered. The epidermal growth factor receptor (EGFR) is a promising therapeutic target but the activity of single agent oral EGFR tyrosine kinase inhibitors is low. There have been no previous reports of chemotherapy plus an EGFR-tyrosine kinase inhibitor (TKI) to treat gallbladder cancer or correlations of response with the mutation status of the tyrosine kinase domain of the EGFR gene. A 67 year old man with metastatic gallbladder cancer involving the liver and abdominal lymph nodes was treated with gemcitabine (1000 mg/m2) on day 1 and 8 every 21 days as well as daily erlotinib (100 mg). After four cycles of therapy, the CA 19-9 normalized and a PET/CT showed a complete remission; this response was maintained by the end of 12 cycles of therapy. Gemcitabine was then discontinued and single agent erlotinib was continued as maintenance therapy. The disease remains in good control 18 months after initiation of therapy, including 6 months on maintenance erlotinib. The only grade 3 toxicity was a typical EGFR-related skin rash. Because of the remarkable response to erlotinib plus gemcitabine, we performed tumor genotyping of the EGFR gene for response predicting mutations in exons 18, 19 and 21. This disclosed the wild-type genotype with no mutations found. This case report demonstrates a patient with stage IV gallbladder cancer who experienced a rarely encountered complete, prolonged response after treatment with an oral EGFR-TKI plus chemotherapy. This response occurred in the absence of an EGFR gene mutation. These observations should inform the design of clinical trials using EGFR-TKIs to treat gallbladder and other biliary tract cancers; such trials should not select patients based on EGFR mutation status

  9. Complete response in gallbladder cancer to erlotinib plus gemcitabine does not require mutation of the epidermal growth factor receptor gene: a case report.

    Science.gov (United States)

    Mody, Kabir; Strauss, Edward; Lincer, Robert; Frank, Richard C

    2010-10-20

    Gallbladder cancer typically follows an aggressive course, with chemotherapy the standard of care for advanced disease; complete remissions are rarely encountered. The epidermal growth factor receptor (EGFR) is a promising therapeutic target but the activity of single agent oral EGFR tyrosine kinase inhibitors is low. There have been no previous reports of chemotherapy plus an EGFR-tyrosine kinase inhibitor (TKI) to treat gallbladder cancer or correlations of response with the mutation status of the tyrosine kinase domain of the EGFR gene. A 67 year old man with metastatic gallbladder cancer involving the liver and abdominal lymph nodes was treated with gemcitabine (1000 mg/m2) on day 1 and 8 every 21 days as well as daily erlotinib (100 mg). After four cycles of therapy, the CA 19-9 normalized and a PET/CT showed a complete remission; this response was maintained by the end of 12 cycles of therapy. Gemcitabine was then discontinued and single agent erlotinib was continued as maintenance therapy. The disease remains in good control 18 months after initiation of therapy, including 6 months on maintenance erlotinib. The only grade 3 toxicity was a typical EGFR-related skin rash. Because of the remarkable response to erlotinib plus gemcitabine, we performed tumor genotyping of the EGFR gene for response predicting mutations in exons 18, 19 and 21. This disclosed the wild-type genotype with no mutations found. This case report demonstrates a patient with stage IV gallbladder cancer who experienced a rarely encountered complete, prolonged response after treatment with an oral EGFR-TKI plus chemotherapy. This response occurred in the absence of an EGFR gene mutation. These observations should inform the design of clinical trials using EGFR-TKIs to treat gallbladder and other biliary tract cancers; such trials should not select patients based on EGFR mutation status.

  10. Comparison of EGFR and KRAS Status between Primary Non-small Cell Lung Cancer and Corresponding Metastases: A Systematic Review and meta-analysis

    Directory of Open Access Journals (Sweden)

    Chengbo HAN

    2010-09-01

    Full Text Available Background and objective Epidermal growth factor receptor (EGFR and KRAS status were particularly critical for the choice of first-line targeted therapy of non-small cell lung cancer (NSCLC, while the primary tumor and metastases might be different in the EGFR and KRAS gene status. The aim of this pooled analysis is to compare EGFR and KRAS status in matching primary NSCLC and metastases and further to guide clinical practice. Methods Systematic computerized searches of the Pubmed and Medline databases (up to May 10, 2010 meeting specified search criteria were performed, followed by a further screening according to inclusive and exclusive criteria. Results Fourteen articles were selected into the final meta-analysis with paired primary and metastatic cases of 598. Expression level of EGFR protein and mutation frequency of KRAS gene in primary tumors were higher than that in metastases, relative risk (RR=1.13 (95%CI: 0.98-1.31, P=0.09 and RR=1.39 (95%CI: 0.95-2.03, P=0.09, respectively. EGFR gene copy number in metastases was higher than that in primary tumor, RR=0.74 (95%CI: 0.53-1.02, P=0.06. There was no statistically significant difference of EGFR mutation frequency in primary tumors and metastases (P=0.31. The discordant rate in primary and metastases was 17.09% for EGFR mutation, 27.07% for EGFR amplification, 27.84% for EGFR protein expression and 25.91% for KRAS mutation. Conclusion The systematic analysis showed that the EGFR mutation status in primary lung cancer and corresponding metastases was more stable than KRAS gene. KRAS mutation in primary lung cancerous foci seems to better reflect systemically cancerous genetic characteristics of KRAS gene. Determination of KRAS gene status based merely on metastatic foci might lead to more resistant selections of EGFR tyrosine kinase inhibitor (TKI therapy. Combined detection of EGFR and KRAS mutation from primary NSCLC foci might serve as a better predictive biomarker for anti-EGFR targeted

  11. MHC class I ligation of human T cells activates the ZAP70 and p56lck tyrosine kinases, leads to an alternative phenotype of the TCR/CD3 zeta-chain, and induces apoptosis

    DEFF Research Database (Denmark)

    Skov, S; Bregenholt, S; Claesson, Mogens Helweg

    1997-01-01

    Cross-linking of MHC class I (MHC-I) molecules on human T cells induces signal-transduction events, including activation of tyrosine kinases, tyrosine phosphorylation of phospholipase C-gamma 1, and elevation of the intracellular free calcium concentration. In this study, we demonstrate...... that the ZAP70 tyrosine kinase is tyrosine phosphorylated in Jurkat T cells and in purified peripheral T cells after MHC-I ligation. The tyrosine-phosphorylated ZAP70 kinase exhibits a particular phenotype with low affinities for proteins at 21, 40, 60, and 120 kDa, proteins normally co-precipitated with ZAP70...... after TCR/CD3 stimulation. The phosphorylation of ZAP70 after MHC-I ligation was dependent on TCR/CD3 surface expression. One of the natural substrates for ZAP70 is the zeta-chain dimer of the TCR/CD3 complex. MHC-I cross-linking induces a phosphorylated zeta-protein that migrates as a dimer at 42 k...

  12. Interleukins 2, 4, 7, and 15 stimulate tyrosine phosphorylation of insulin receptor substrates 1 and 2 in T cells. Potential role of JAK kinases.

    Science.gov (United States)

    Johnston, J A; Wang, L M; Hanson, E P; Sun, X J; White, M F; Oakes, S A; Pierce, J H; O'Shea, J J

    1995-12-01

    The signaling molecules insulin receptor substrate (IRS)-1 and the newly described IRS-2 (4PS) molecule are major insulin and interleukin 4 (IL-4)-dependent phosphoproteins. We report here that IL-2, IL-7, and IL-15, as well as IL-4, rapidly stimulate the tyrosine phosphorylation of IRS-1 and IRS-2 in human peripheral blood T cells, NK cells, and in lymphoid cell lines. In addition, we show that the Janus kinases, JAK1 and JAK3, associate with IRS-1 and IRS-2 in T cells. Coexpression studies demonstrate that these kinases can tyrosine-phosphorylate IRS-2, suggesting a possible mechanism by which cytokine receptors may induce the tyrosine phosphorylation of IRS-1 and IRS-2. We further demonstrate that the p85 subunit of phosphoinositol 3-kinase associates with IRS-1 in response to IL-2 and IL-4 in T cells. Therefore, these data indicate that IRS-1 and IRS-2 may have important roles in T lymphocyte activation not only in response to IL-4, but also in response to IL-2, IL-7, and IL-15.

  13. syk kinase activation by a src kinase-initiated activation loop phosphorylation chain reaction

    Science.gov (United States)

    El-Hillal, O.; Kurosaki, T.; Yamamura, H.; Kinet, J.-P.; Scharenberg, A. M.

    1997-01-01

    Activation of the syk tyrosine kinase occurs almost immediately following engagement of many types of antigen receptors, including Fc receptors, but the mechanism through which syk is activated is currently unclear. Here we demonstrate that Fc receptor-induced syk activation occurs as the result of phosphorylation of the syk activation loop by both src family kinases and other molecules of activated syk, suggesting that syk activation occurs as the result of a src kinase-initiated activation loop phosphorylation chain reaction. This type of activation mechanism predicts that syk activation would exhibit exponential kinetics, providing a potential explanation for its rapid and robust activation by even weak antigen receptor stimuli. We propose that a similar mechanism may be responsible for generating rapid activation of other cytoplasmic tyrosine kinases, such as those of the Bruton tyrosine kinase/tec family, as well. PMID:9050880

  14. Design, Synthesis, and X-ray Crystal Structures of 2,4-Diaminofuro[2,3-d]pyrimidines as Multireceptor Tyrosine Kinase and Dihydrofolate Reductase Inhibitors

    Science.gov (United States)

    Gangjee, Aleem; Li, Wei; Lin, Lu; Zeng, Yibin; Ihnat, Michael; Warnke, Linda A.; Green, Dixy W.; Cody, Vivian; Pace, Jim; Queener, Sherry F.

    2009-01-01

    To optimize dual receptor tyrosine kinase (RTK) and dihydrofolate reductase (DHFR) inhibition, the E- and Z-isomers of 5-[2-(2-methoxyphenyl)prop-1-en-1-yl]furo[2,3-d]pyrimidine-2,4-diamines (1a and 1b) were separated by HPLC and the X-ray crystal structures (2.0 Å and 1.4 Å respectively) with mouse DHFR and NADPH as well as 1b with human DHFR (1.5 Å) were determined. The E- and Z-isomers adopt different binding modes when bound to mouse DHFR. A series of 2,4-diaminofuro[2,3-d]pyrimidines 2–13 were designed and synthesized using the X-ray crystal structures of 1a and 1b with DHFR to increase their DHFR inhibitory activity. Wittig reactions of appropriate 2-methoxyphenyl ketones with 2,4-diamino-6-chloromethyl furo[2,3-d]pyrimidine afforded the C8–C9 unsaturated compounds 2–7 and catalytic reduction gave the saturated 8–13. Homologation of the C9-methyl analog maintains DHFR inhibitory activity. In addition, inhibition of EGFR and PDGFR-β were discovered for saturated C9-homologated analogs 9 and 10 that were absent in the saturated C9-methyl analogs. PMID:19748785

  15. EGFR-expression in primary urinary bladder cancer and corresponding metastases and the relation to HER2-expression. On the possibility to target these receptors with radionuclides

    International Nuclear Information System (INIS)

    Carlsson, Jörgen; Wester, Kenneth; De La Torre, Manuel; Malmström, Per-Uno; Gårdmark, Truls

    2015-01-01

    There is limited effect of tyrosine kinase inhibitors or “naked” antibodies binding EGFR or HER2 for therapy of metastasized urinary bladder cancer and these methods are therefore not routinely used. Targeting radio-nuclides to the extracellular domain of the receptors is potentially a better possibility. EGFR- and HER2-expression was analyzed for primary tumors and corresponding metastases from 72 patients using immunohistochemistry and the internationally recommended HercepTest. Intracellular mutations were not analyzed since only the receptors were considered as targets and intracellular abnormalities should have minor effect on radiation dose. EGFR was positive in 71% of the primary tumors and 69% of corresponding metastases. Local and distant metastases were EGFR-positive in 75% and 66% of the cases, respectively. The expression frequency of HER2 in related lesions was slightly higher (data from previous study). The EGFR-positive tumors expressed EGFR in metastases in 86% of the cases. The co-expression of EGFR and HER2 was 57% for tumors and 53% for metastases. Only 3% and 10% of the lesions were negative for both receptors in tumors and metastases, respectively. Thus, targeting these receptors with radionuclides might be applied for most patients. At least one of the EGFR- or HER2-receptors was present in most cases and co-expressed in more than half the cases. It is therefore interesting to deliver radionuclides for whole-body receptor-analysis, dosimetry and therapy. This can hopefully compensate for resistance to other therapies and more patients can hopefully be treated with curative instead of palliative intention

  16. Cell adhesion and EGFR activation regulate EphA2 expression in cancer

    DEFF Research Database (Denmark)

    Larsen, Alice Bjerregaard; Stockhausen, Marie-Thérése; Poulsen, Hans Skovgaard

    2010-01-01

    family kinases (SRC). Moreover, the results show that adhesion-induced EGFR activation and EphA2 expression is affected by interactions with extracellular matrix (ECM) proteins working as integrin ligands. Stimulation with the EphA2 ligand, ephrinA1 inhibited ERK phosphorylation and cancer cell viability...... largely unknown. Here we show that the expression of EphA2 in in vitro cultured cells, is restricted to cells growing adherently and that adhesion-induced EphA2 expression is dependent upon activation of the epidermal growth factor receptor (EGFR), mitogen activated protein kinase kinase (MEK) and Src...

  17. Molecular Epidemiology of EGFR Mutations in Asian Patients with Advanced Non-Small-Cell Lung Cancer of Adenocarcinoma Histology – Mainland China Subset Analysis of the PIONEER study

    Science.gov (United States)

    Shi, Yuankai; Li, Junling; Zhang, Shucai; Wang, Mengzhao; Yang, Shujun; Li, Ning; Wu, Gang; Liu, Wei; Liao, Guoqing; Cai, Kaican; Chen, Liang’an; Zheng, Meizhen; Yu, Ping; Wang, Xiuwen; Liu, Yunpeng; Guo, Qisen; Nie, Ligong; Liu, Jiwei; Han, Xiaohong

    2015-01-01

    Epidermal growth factor receptor (EGFR) mutations are the strongest response predictors to EGFR tyrosine kinase inhibitors (TKI) therapy, but knowledge of the EGFR mutation frequency on lung adenocarcinoma is still limited to retrospective studies. The PIONEER study (NCT01185314) is a prospective molecular epidemiology study in Asian patients with newly diagnosed advanced lung adenocarcinoma, aiming to prospectively analyze EGFR mutation status in IIIB/IV treatment-naïve lung adenocarcinomas in Asia. We report the mainland China subset results. Eligible patients (≥20 yrs old, IIIB/IV adenocarcinoma and treatment-naïve) were registered in 17 hospitals in mainland China. EGFR was tested for mutations by amplification refractory mutation system using biopsy samples. Demographic and clinical characteristics were collected for subgroup analyses. A total of 747 patients were registered. Successful EGFR mutation analysis was performed in 741, with an overall mutation rate of 50.2%. The EGFR active mutation rate is 48.0% (with 1.3% of combined active and resistance mutations). Tobacco use (>30 pack-year vs. 0–10 pack-year, OR 0.27, 95%CI: 0.17–0.42) and regional lymph nodes involvement (N3 vs. N0, OR 0.47, 95%CI: 0.29–0.76) were independent predictors of EGFR mutation in multivariate analysis. However, even in regular smokers, the EGFR mutation frequency was 35.3%. The EGFR mutation frequency was similar between diverse biopsy sites and techniques. The overall EGFR mutation frequency of the mainland China subset was 50.2%, independently associated with the intensity of tobacco use and regional lymph nodes involvement. The relatively high frequency of EGFR mutations in the mainland China subset suggest that any effort to obtain tissue sample for EGFR mutation testing should be encouraged. PMID:26599344

  18. Tyrosine phosphorylation in T cells is regulated by phosphatase activity: studies with phenylarsine oxide.

    OpenAIRE

    Garcia-Morales, P; Minami, Y; Luong, E; Klausner, R D; Samelson, L E

    1990-01-01

    Activation of T cells induces rapid tyrosine phosphorylation on the T-cell receptor zeta chain and other substrates. These phosphorylations can be regulated by a number of protein-tyrosine kinases (ATP: protein-tyrosine O-phosphotransferase, EC 2.7.1.112) and protein-tyrosine-phosphatases (protein-tyrosine-phosphate phosphohydrolase, EC 3.1.3.48). In this study, we demonstrate that phenylarsine oxide can inhibit tyrosine phosphatases while leaving tyrosine kinase function intact. We use this ...

  19. Nuclear localization of lymphocyte-specific protein tyrosine kinase (Lck) and its role in regulating LIM domain only 2 (Lmo2) gene

    Energy Technology Data Exchange (ETDEWEB)

    Venkitachalam, Srividya; Chueh, Fu-Yu [Department of Microbiology and Immunology, H. M. Bligh Cancer Research Laboratories, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL 60064 (United States); Yu, Chao-Lan, E-mail: chaolan.yu@rosalindfranklin.edu [Department of Microbiology and Immunology, H. M. Bligh Cancer Research Laboratories, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL 60064 (United States)

    2012-01-20

    Highlights: Black-Right-Pointing-Pointer Lmo2 expression is elevated in Lck-transformed cells. Black-Right-Pointing-Pointer Both endogenous and exogenous Lck localize in the nucleus. Black-Right-Pointing-Pointer Nuclear Lck is active in Lck-transformed cells. Black-Right-Pointing-Pointer Lck binds to the promoter region of Lmo2 gene in vivo. Black-Right-Pointing-Pointer In contrast to JAK2, Lck does not increase histone H3 phosphorylation on Tyr 41. -- Abstract: LIM domain only protein 2 (Lmo2) is a transcription factor that plays a critical role in the development of T-acute lymphoblastic leukemia (T-ALL). A previous report established a link between Lmo2 expression and the nuclear presence of oncogenic Janus kinase 2 (JAK2), a non-receptor protein tyrosine kinase. The oncogenic JAK2 kinase phosphorylates histone H3 on Tyr 41 that leads to the relief of Lmo2 promoter repression and subsequent gene expression. Similar to JAK2, constitutive activation of lymphocyte-specific protein tyrosine kinase (Lck) has been implicated in lymphoid malignancies. However, it is not known whether oncogenic Lck regulates Lmo2 expression through a similar mechanism. We show here that Lmo2 expression is significantly elevated in T cell leukemia LSTRA overexpressing active Lck kinase and in HEK 293 cells expressing oncogenic Y505FLck kinase. Nuclear localization of active Lck kinase was confirmed in both Lck-transformed cells by subcellular fractionation and immunofluorescence microscopy. More importantly, in contrast to oncogenic JAK2, oncogenic Lck kinase does not result in significant increase in histone H3 phosphorylation on Tyr 41. Instead, chromatin immunoprecipitation experiment shows that oncogenic Y505FLck kinase binds to the Lmo2 promoter in vivo. This result raises the possibility that oncogenic Lck may activate Lmo2 promoter through direct interaction.

  20. Platelet-activating factor stimulation of tyrosine kinase and its relationship to phospholipase C in rabbit platelets: Studies with genistein and monoclonal antibody to phosphotyrosine

    International Nuclear Information System (INIS)

    Dhar, A.; Paul, A.K.; Shukla, S.D.

    1990-01-01

    Platelet-activating factor (PAF) is a proinflammatory lipid that has platelet-stimulating property. PAF receptor-coupled activation of phosphoinositide-specific phospholipase C (PLC) and phosphorylation of several proteins has already been established in our laboratory. To investigate further the molecular mechanism and relationship between activation of PLC and protein phosphorylation, we have used Genistein (a putative inhibitor of tyrosine-specific protein kinases), phosphotyrosine antibody, and phosphoamino acid analysis to probe the involvement of tyrosine kinase in this process. Washed rabbit platelets were loaded with myo-[2-3H]inositol and challenged with PAF (100 nM) after pretreatment with Genistein. PLC-mediated production of radioactive inositol monophosphate, inositol diphosphate, and inositol triphosphate was monitored. PAF alone caused stimulation of PLC activity [( 3H]inositol triphosphate production), whereas pretreatment with Genistein (0.5 mM) diminished PAF-stimulated PLC activity to basal level. Genistein also blocked PAF-stimulated platelet aggregation at this dose. In contrast to Genistein, staurosporine which inhibits protein kinase C, potentiated PAF-stimulated [3H]inositol triphosphate production. Genistein substantially inhibited the combined effects of staurosporine and PAF on inositol triphosphate production. Genistein also reduced PAF-induced phosphorylation of Mr 20,000 and 50,000 proteins. Phorbol 12-myristate 13-acetate-induced Mr 40,000 protein phosphorylation was also affected by Genistein. The above results suggested that Genistein inhibited tyrosine kinase at an early stage of signal transduction by inhibiting PLC. This, in turn, decreased the activation of protein kinase C and, therefore, caused a reduction in Mr 40,000 protein phosphorylation