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Sample records for activated egfr determines

  1. Response to the Dorsal Anterior Gradient of EGFR Signaling in Drosophila Oogenesis Is Prepatterned by Earlier Posterior EGFR Activation

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    Mariana Fregoso Lomas

    2013-08-01

    Full Text Available Spatially restricted epidermal growth factor receptor (EGFR activity plays a central role in patterning the follicular epithelium of the Drosophila ovary. In midoogenesis, localized EGFR activation is achieved by the graded dorsal anterior localization of its ligand, Gurken. Graded EGFR activity determines multiple dorsal anterior fates along the dorsal-ventral axis but cannot explain the sharp posterior limit of this domain. Here, we show that posterior follicle cells express the T-box transcription factors Midline and H15, which render cells unable to adopt a dorsal anterior fate in response to EGFR activation. The posterior expression of Midline and H15 is itself induced in early oogenesis by posteriorly localized EGFR signaling, defining a feedback loop in which early induction of Mid and H15 confers a molecular memory that fundamentally alters the outcome of later EGFR signaling. Spatial regulation of the EGFR pathway thus occurs both through localization of the ligand and through localized regulation of the cellular response.

  2. Frequent activation of EGFR in advanced chordomas

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    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

  3. Direct interaction between surface β1,4-galactosyltransferase 1 and epidermal growth factor receptor (EGFR) inhibits EGFR activation in hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Highlights: •β1,4GT1 interacts with EGFR both in vitro and in vivo. •β1,4GT1 co-localizes with EGFR on the cell surface. •β1,4GT1 inhibits 125I-EGF binding to EGFR. •β1,4GT1 inhibits EGF induced EGFR dimerization and phosphorylation. -- Abstract: Our previous studies showed that cell surface β1,4-galactosyltransferase 1 (β1,4GT1) negatively regulated cell survival through inhibition and modulation of the epidermal growth factor receptor (EGFR) signaling pathway in human hepatocellular carcinoma (HCC) SMMC-7721 cells. However, the underlying mechanism remains unclear. Here we demonstrated that β1,4-galactosyltransferase 1 (β1,4GT1) interacted with EGFR in vitro by GST pull-down analysis. Furthermore, we demonstrated that β1,4GT1 bound to EGFR in vivo by co-immunoprecipitation and determined the co-localization of β1,4GT1 and EGFR on the cell surface via confocal laser scanning microscopy analysis. Finally, using 125I-EGF binding experiments and Western blot analysis, we found that overexpression of β1,4GT1 inhibited 125I-EGF binding to EGFR, and consequently reduced the levels of EGFR dimerization and phosphorylation. In contrast, RNAi-mediated knockdown of β1,4GT1 increased the levels of EGFR dimerization and phosphorylation. These data suggest that cell surface β1,4GT1 interacts with EGFR and inhibits EGFR activation

  4. Determination of EGFR and KRAS mutational status in Greek non-small-cell lung cancer patients

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    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

    2015-01-01

    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

  5. Determination of EGFR and KRAS mutational status in Greek non-small-cell lung cancer patients

    OpenAIRE

    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

    2015-01-01

    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 clinicopat...

  6. Integrated Experimental and Model-based Analysis Reveals the Spatial Aspects of EGFR Activation Dynamics

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    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

  7. Trefoil factors: Tumor progression markers and mitogens via EGFR/MAPK activation in cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Kanuengnuch Kosriwong; Trevelyan R Menheniott; Andrew S Giraud; Patcharee Jearanaikoon; Banchob Sripa; Temduang Limpaiboon

    2011-01-01

    AIM: To investigate trefoil factor (TFF ) gene copy number, mRNA and protein expression as potential biomarkers in cholangiocarcinoma (CCA). METHODS: TFF mRNA levels, gene copy number and protein expression were determined respectively by quantitative reverse transcription polymerase chain reaction (PCR), quantitative PCR and immunohistochemistry in bile duct epithelium biopsies collected from individuals with CCA, precancerous bile duct dysplasia and from disease-free controls. The functional impact of recombinant human (rh)TFF2 peptide treatment on proliferation and epidermal growth factor receptor (EGFR)/mitogenactivated protein kinase (MAPK) signaling was assessed in the CCA cell line, KMBC, by viable cell counting and immunoblotting, respectively. RESULTS: TFF1 , TFF2 and TFF3 mRNA expression was significantly increased in CCA tissue compared to disease-free controls, and was unrelated to gene copy number. TFF1 immunoreactivity was strongly increased in both dysplasia and CCA, whereas TFF2 immunoreactivity was increased only in CCA compared to diseasefree controls. By contrast, TFF3 immunoreactivity was moderately decreased in dysplasia and further decreased in CCA. Kaplan-Meier analysis found no association of TFF mRNA, protein and copy number with age, gender, histological subtype, and patient survival time. Treatment of KMBC cells with rhTFF2 stimulated proliferation, triggered phosphorylation of EGFR and downstream extracellular signal related kinase (ERK), whereas co-incubation with the EGFR tyrosine kinase inhibitor, PD153035, blocked rhTFF2-dependent proliferation and EGFR/ERK responses. CONCLUSION: TFF mRNA/protein expression is indicative of CCA tumor progression, but not predictive for histological sub-type or survival time. TFF2 is mitogenic in CCA via EGFR/MAPK activation.

  8. Selective Antitumor Activity of Ibrutinib in EGFR-Mutant Non–Small Cell Lung Cancer Cells

    OpenAIRE

    Gao, Wen; Wang, Michael; Wang, Li; Lu, Haibo; Wu, Shuhong; Dai, Bingbing; Ou, Zhishuo; Zhang, Liang; Heymach, John V.; Gold, Kathryn A.; Minna, John ,; Roth, Jack A.; Hofstetter, Wayne L.; Swisher, Stephen G.; Fang, Bingliang

    2014-01-01

    Ibrutinib, which irreversibly inhibits Bruton tyrosine kinase, was evaluated for antitumor activity in a panel of non–small cell lung cancer (NSCLC) cell lines and found to selectively inhibit growth of NSCLC cells carrying mutations in the epidermal growth factor receptor (EGFR) gene, including T790M mutant and erlotinib-resistant H1975 cells. Ibrutinib induced dose-dependent inhibition of phosphor-EGFR at both Y1068 and Y1173 sites, suggesting ibrutinib functions as an EGFR inhibitor. Survi...

  9. Hypoxia activated EGFR signaling induces epithelial to mesenchymal transition (EMT.

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    Ashish Misra

    Full Text Available Metastasis is a multi-step process which requires the conversion of polarized epithelial cells to mesenchymal cells, Epithelial-Mesenchymal Transition (EMT. EMT is essential during embryonic morphogenesis and has been implicated in the progression of primary tumors towards metastasis. Hypoxia is known to induce EMT; however the molecular mechanism is still poorly understood. Using the A431 epithelial cancer cell line, we show that cells grown under hypoxic conditions migrated faster than cells grown under normal oxygen environment. Cells grown under hypoxia showed reduced adhesion to the extracellular matrix (ECM probably due to reduced number of Vinculin patches. Growth under hypoxic conditions also led to down regulation of E-cadherin and up regulation of vimentin expression. The increased motility of cells grown under hypoxia could be due to redistribution of Rac1 to the plasma membrane as opposed to increased expression of Rac1. EGF (Epidermal Growth Factor is a known inducer of EMT and growth of A431 cells in the absence of oxygen led to increased expression of EGFR (EGF Receptor. Treatment of A431 cells with EGF led to reduced cell adhesion to ECM, increased cell motility and other EMT characteristics. Furthermore, this transition was blocked by the monoclonal antibody Cetuximab. Cetuximab also blocked the hypoxia-induced EMT suggesting that cell growth under hypoxic conditions led to activation of EGFR signaling and induction of EMT phenotype.

  10. Lipopolysaccharide (LPS-Induced Biliary Epithelial Cell NRas Activation Requires Epidermal Growth Factor Receptor (EGFR.

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    Christy E Trussoni

    Full Text Available Cholangiocytes (biliary epithelial cells actively participate in microbe-induced proinflammatory responses in the liver and contribute to inflammatory and infectious cholangiopathies. We previously demonstrated that cholangiocyte TLR-dependent NRas activation contributes to proinflammatory/ proliferative responses. We test the hypothesis that LPS-induced activation of NRas requires the EGFR. SV40-transformed human cholangiocytes (H69 cells, or low passage normal human cholangiocytes (NHC, were treated with LPS in the presence or absence of EGFR or ADAM metallopeptidase domain 17 (TACE inhibitors. Ras activation assays, quantitative RT-PCR, and proliferation assays were performed in cells cultured with or without inhibitors or an siRNA to Grb2. Immunofluorescence for phospho-EGFR was performed on LPS-treated mouse samples and specimens from patients with primary sclerosing cholangitis, primary biliary cirrhosis, hepatitis C, and normal livers. LPS-treatment induced an association between the TLR/MyD88 and EGFR/Grb2 signaling apparatus, NRas activation, and EGFR phosphorylation. NRas activation was sensitive to EGFR and TACE inhibitors and correlated with EGFR phosphorylation. The TACE inhibitor and Grb2 depletion prevented LPS-induced IL6 expression (p<0.05 and proliferation (p<0.01. Additionally, cholangiocytes from LPS-treated mouse livers and human primary sclerosing cholangitis (PSC livers exhibited increased phospho-EGFR (p<0.01. Moreover, LPS-induced mouse cholangiocyte proliferation was inhibited by concurrent treatment with the EGFR inhibitor, Erlotinib. Our results suggest that EGFR is essential for LPS-induced, TLR4/MyD88-mediated NRas activation and induction of a robust proinflammatory cholangiocyte response. These findings have implications not only for revealing the signaling potential of TLRs, but also implicate EGFR as an integral component of cholangiocyte TLR-induced proinflammatory processes.

  11. Lipopolysaccharide (LPS)-Induced Biliary Epithelial Cell NRas Activation Requires Epidermal Growth Factor Receptor (EGFR).

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    Trussoni, Christy E; Tabibian, James H; Splinter, Patrick L; O'Hara, Steven P

    2015-01-01

    Cholangiocytes (biliary epithelial cells) actively participate in microbe-induced proinflammatory responses in the liver and contribute to inflammatory and infectious cholangiopathies. We previously demonstrated that cholangiocyte TLR-dependent NRas activation contributes to proinflammatory/ proliferative responses. We test the hypothesis that LPS-induced activation of NRas requires the EGFR. SV40-transformed human cholangiocytes (H69 cells), or low passage normal human cholangiocytes (NHC), were treated with LPS in the presence or absence of EGFR or ADAM metallopeptidase domain 17 (TACE) inhibitors. Ras activation assays, quantitative RT-PCR, and proliferation assays were performed in cells cultured with or without inhibitors or an siRNA to Grb2. Immunofluorescence for phospho-EGFR was performed on LPS-treated mouse samples and specimens from patients with primary sclerosing cholangitis, primary biliary cirrhosis, hepatitis C, and normal livers. LPS-treatment induced an association between the TLR/MyD88 and EGFR/Grb2 signaling apparatus, NRas activation, and EGFR phosphorylation. NRas activation was sensitive to EGFR and TACE inhibitors and correlated with EGFR phosphorylation. The TACE inhibitor and Grb2 depletion prevented LPS-induced IL6 expression (pphospho-EGFR (p<0.01). Moreover, LPS-induced mouse cholangiocyte proliferation was inhibited by concurrent treatment with the EGFR inhibitor, Erlotinib. Our results suggest that EGFR is essential for LPS-induced, TLR4/MyD88-mediated NRas activation and induction of a robust proinflammatory cholangiocyte response. These findings have implications not only for revealing the signaling potential of TLRs, but also implicate EGFR as an integral component of cholangiocyte TLR-induced proinflammatory processes.

  12. DUOX1 mediates persistent epithelial EGFR activation, mucous cell metaplasia, and airway remodeling during allergic asthma

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    Habibovic, Aida; Hristova, Milena; Heppner, David E.; Danyal, Karamatullah; Ather, Jennifer L.; Janssen-Heininger, Yvonne M.W.; Irvin, Charles G.; Poynter, Matthew E.; Lundblad, Lennart K.; Dixon, Anne E.; Geiszt, Miklos

    2016-01-01

    Chronic inflammation with mucous metaplasia and airway remodeling are hallmarks of allergic asthma, and these outcomes have been associated with enhanced expression and activation of EGFR signaling. Here, we demonstrate enhanced expression of EGFR ligands such as amphiregulin as well as constitutive EGFR activation in cultured nasal epithelial cells from asthmatic subjects compared with nonasthmatic controls and in lung tissues of mice during house dust mite–induced (HDM-induced) allergic inflammation. EGFR activation was associated with cysteine oxidation within EGFR and the nonreceptor tyrosine kinase Src, and both amphiregulin production and oxidative EGFR activation were diminished by pharmacologic or genetic inhibition of the epithelial NADPH oxidase dual oxidase 1 (DUOX1). DUOX1 deficiency also attenuated several EGFR-dependent features of HDM-induced allergic airway inflammation, including neutrophilic inflammation, type 2 cytokine production (IL-33, IL-13), mucous metaplasia, subepithelial fibrosis, and central airway resistance. Moreover, targeted inhibition of airway DUOX1 in mice with previously established HDM-induced allergic inflammation, by intratracheal administration of DUOX1-targeted siRNA or pharmacological NADPH oxidase inhibitors, reversed most of these outcomes. Our findings indicate an important function for DUOX1 in allergic inflammation related to persistent EGFR activation and suggest that DUOX1 targeting may represent an attractive strategy in asthma management.

  13. Expression of epidermal growth factor receptor (EGFR) and activated EGFR predict poor response to (chemo)radiation and survival in cervical cancer

    NARCIS (Netherlands)

    Noordhuis, M.G.; Eijsink, J.J.H.; ten Hoor, K.A.; Roossink, F.; Hollema, H.; Arts, H.J.G.; Pras, Elisabeth; Maduro, John; Reyners, A.K.L.; de Bock, G.H.; Wisman, G.B.A.; Schuuring, E.; van der Zee, A.G.J.

    2009-01-01

    PURPOSE: Activation of the epidermal growth factor receptor (EGFR) signaling pathway has been reported to induce resistance to (chemo)radiation in cancers, such as head and neck cancer, whereas EGFR-targeted agents in combination with (chemo)radiation seem to improve treatment efficacy. The aim of t

  14. TGFβ induces proHB-EGF shedding and EGFR transactivation through ADAM activation in gastric cancer cells

    International Nuclear Information System (INIS)

    Research highlights: → TGFβ induces EGFR transactivation through proHB-EGF shedding by activated ADAM members in gastric cancer cells. → TGFβ induces nuclear translocation of HB-EGF-CTF cleaved by ADAM members. → TGFβ enhances cell growth by EGFR transactivation and HB-EGF-CTF nuclear translocation and ADAM inhibitors block these effects. → Silencing of ADAM17 also blocks EGFR transactivation, HB-EGF-CTF nuclear translocation and cancer cell growth by TGFβ. → ADAM17 may play a crucial role in this TGFβ-HB-EGF signal transduction. -- Abstract: Background and aims: Transforming growth factor-beta (TGFβ) is known to potently inhibit cell growth. Loss of responsiveness to TGFβ inhibition on cell growth is a hallmark of many types of cancer, yet its mechanism is not fully understood. Membrane-anchored heparin-binding EGF-like growth factor (proHB-EGF) ectodomain is cleaved by a disintegrin and metalloproteinase (ADAM) members and is implicated in epidermal growth factor receptor (EGFR) transactivation. Recently, nuclear translocation of the C-terminal fragment (CTF) of pro-HB-EGF was found to induce cell growth. We investigated the association between TGFβ and HB-EGF signal transduction via ADAM activation. Materials and methods: The CCK-8 assay in two gastric cancer cell lines was used to determine the effect for cell growth by TGFβ. The effect of two ADAM inhibitors was also evaluated. Induction of EGFR phosphorylation by TGFβ was analyzed and the effect of the ADAM inhibitors was also examined. Nuclear translocation of HB-EGF-CTF by shedding through ADAM activated by TGFβ was also analyzed. EGFR transactivation, HB-EGF-CTF nuclear translocation, and cell growth were examined under the condition of ADAM17 knockdown. Result: TGFβ-induced EGFR phosphorylation of which ADAM inhibitors were able to inhibit. TGFβ induced shedding of proHB-EGF allowing HB-EGF-CTF to translocate to the nucleus. ADAM inhibitors blocked this nuclear translocation. TGF

  15. Integrated cell-based platform to study EGFR activation and transactivation.

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    Caruso, Marie-Elaine; Clément, Paule; Parent, Stéphane; Dupriez, Vincent; Bossé, Roger; Rouleau, Nathalie

    2013-09-01

    The epidermal growth factor receptor (EGFR) pathway is one of the most deregulated molecular pathways in human epithelial cancers. Many approved drugs were optimized to directly target EGFR but yielded only modest clinical improvement in cancer patients due to low efficacy and drug resistance. Transactivation of EGFR by other cell surface receptors such as G-protein-coupled receptors (GPCRs) was proposed to explain this lack of efficacy. Even if direct EGFR activation and transactivation by GPCR contribute to the activation of the same signaling pathways, they are often studied as independent events resulting in partial investigation of a drug's mechanism of action. We present a novel high-throughput approach that integrates interrogation of direct activation of EGFR and its transactivation via GPCR activation. Using distinct technology platforms, three readouts were used to measure (1) direct activation of GPCR via cyclic adenosine monophosphate (cAMP) detection, (2) direct activation of EGFR through the release of intracellular Ca(2+), and (3) EGFR transactivation by GPCR using the detection of p-extracellular-signal-regulated kinases 1/2 (p-ERK1/2). In addition to being simple, quick, and homogenous, our methods were shown to be more sensitive than those in current use. These enabling tools should improve the knowledge pertaining to GPCRs and receptor tyrosine kinases trans-regulation and facilitate the design of more potent and better targeted new therapeutic strategies.

  16. Dynamic recruitment of protein tyrosine phosphatase PTPD1 to EGF stimulation sites potentiates EGFR activation.

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    Pedro Roda-Navarro

    Full Text Available Balanced activity of protein tyrosine kinases and phosphatases (PTPs controls tyrosine phosphorylation levels and, consequently, is needed to prevent pathologies like cancer. Phosphatase activity is tightly regulated in space and time. Thus, in order to understand how phospho-tyrosine signalling is regulated, the intracellular dynamics of PTPs should be investigated. Here, we have studied the intracellular dynamics of PTPD1, a FERM (four-point-one, ezrin, radixin, moesin domain-containing PTP that is over expressed in cancer cells and potentiates EGFR signalling. Whereas PTPD1 was excluded from E-cadherin rich cell-cell adhesions in epithelial cell monolayers, it diffused from the cytoplasm to those membranes in contact with the extracellular medium. Localisation of PTPD1 at the plasma membrane was mediated by its FERM domain and enabled the formation of EGFR/PTPD1-containing signalling complexes that pre-existed at the plasma membrane before EGF stimulation. PTPD1 and EGFR transiently co-localised at EGF stimulation sites until the formation of macropinosomes containing active species of EGFR. Interference of PTPD1 expression caused a decrease in EGFR phosphorylated species at the periphery of the cell. Presented data suggest that the transient formation of dynamic PTPD1/EGFR signalling complexes strengthens EGF signalling by promoting the spatial propagation of EGFR phosphorylated species.

  17. Akt kinase-interacting protein1, a novel therapeutic target for lung cancer with EGFR-activating and gatekeeper mutations.

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    Yamada, T; Takeuchi, S; Fujita, N; Nakamura, A; Wang, W; Li, Q; Oda, M; Mitsudomi, T; Yatabe, Y; Sekido, Y; Yoshida, J; Higashiyama, M; Noguchi, M; Uehara, H; Nishioka, Y; Sone, S; Yano, S

    2013-09-12

    Despite initial dramatic response, epidermal growth factor receptor (EGFR) mutant lung cancer patients always acquire resistance to EGFR-tyrosine kinase inhibitors (TKIs). Gatekeeper T790M mutation in EGFR is the most prevalent genetic alteration underlying acquired resistance to EGFR-TKI, and EGFR mutant lung cancer cells are reported to be addictive to EGFR/Akt signaling even after acquired T790M mutation. Here, we focused on Akt kinase-interacting protein1 (Aki1), a scaffold protein of PI3K (phosphoinositide 3-kinase)/PDK1 (3-phosphoinositide-dependent protein kinase)/Akt that determines receptor signal selectivity for non-mutated EGFR, and assessed its role in EGFR mutant lung cancer with or without gatekeeper T790M mutation. Cell line-based assays showed that Aki1 constitutively associates with mutant EGFR in lung cancer cells with (H1975) or without (PC-9 and HCC827) T790M gatekeeper mutation. Silencing of Aki1 induced apoptosis of EGFR mutant lung cancer cells. Treatment with Aki1 siRNA dramatically inhibited growth of H1975 cells in a xenograft model. Moreover, silencing of Aki1 further potentiated growth inhibitory effect of new generation EGFR-TKIs against H1975 cells in vitro. Aki1 was frequently expressed in tumor cells of EGFR mutant lung cancer patients (53/56 cases), including those with acquired resistance to EGFR-TKI treatment (7/7 cases). Our data suggest that Aki1 may be a critical mediator of survival signaling from mutant EGFR to Akt, and may therefore be an ideal target for EGFR mutant lung cancer patients, especially those with acquired EGFR-TKI resistance due to EGFR T790M gatekeeper mutation. PMID:23045273

  18. Involvement of lipid rafts in adhesion-induced activation of Met and EGFR

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    Lu Ying-Che

    2011-10-01

    Full Text Available Abstract Background Cell adhesion has been shown to induce activation of certain growth factor receptors in a ligand-independent manner. However, the mechanism for such activation remains obscure. Methods Human epidermal carcinoma A431 cells were used as a model to examine the mechanism for adhesion-induced activation of hepatocyte growth factor receptor Met and epidermal growth factor receptor (EGFR. The cells were suspended and replated on culture dishes under various conditions. The phosphorylation of Met at Y1234/1235 and EGFR at Y1173 were used as indicators for their activation. The distribution of the receptors and lipid rafts on the plasma membrane were visualized by confocal fluorescent microscopy and total internal reflection microscopy. Results We demonstrate that Met and EGFR are constitutively activated in A431 cells, which confers proliferative and invasive potentials to the cells. The ligand-independent activation of Met and EGFR in A431 cells relies on cell adhesion to a substratum, but is independent of cell spreading, extracellular matrix proteins, and substratum stiffness. This adhesion-induced activation of Met and EGFR cannot be attributed to Src activation, production of reactive oxygen species, and the integrity of the cytoskeleton. In addition, we demonstrate that Met and EGFR are independently activated upon cell adhesion. However, partial depletion of Met and EGFR prevents their activation upon cell adhesion, suggesting that overexpression of the receptors is a prerequisite for their self-activation upon cell adhesion. Although Met and EGFR are largely distributed in 0.04% Triton-insoluble fractions (i.e. raft fraction, their activated forms are detected mainly in 0.04% Triton-soluble fractions (i.e. non-raft fraction. Upon cell adhesion, lipid rafts are accumulated at the cell surface close to the cell-substratum interface, while Met and EGFR are mostly excluded from the membrane enriched by lipid rafts. Conclusions

  19. Loss of EBP50 stimulates EGFR activity to induce EMT phenotypic features in biliary cancer cells.

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    Clapéron, A; Guedj, N; Mergey, M; Vignjevic, D; Desbois-Mouthon, C; Boissan, M; Saubaméa, B; Paradis, V; Housset, C; Fouassier, L

    2012-03-15

    Scaffold proteins form multiprotein complexes that are central to the regulation of intracellular signaling. The scaffold protein ezrin-radixin-moesin-binding phosphoprotein 50 (EBP50) is highly expressed at the plasma membrane of normal biliary epithelial cells and binds epidermal growth factor receptor (EGFR), a tyrosine kinase receptor with oncogenic properties. This study investigated EBP50-EGFR interplay in biliary cancer. We report that in a collection of 106 cholangiocarcinomas, EBP50 was delocalized to the cytoplasm of tumor cells in 66% of the cases. Ectopic expression of EBP50 was correlated with the presence of satellite nodules and with the expression of EGFR, which was at the plasma membrane, implying a loss of interaction with EBP50 in these cases. In vitro, loss of interaction between EBP50 and EGFR was mimicked by EBP50 depletion using a small interfering RNA approach in human biliary carcinoma cells co-expressing the two proteins at their plasma membrane, and in which interaction between EBP50 and EGFR was validated. EBP50 depletion caused an increase in EGFR expression at their surface, and a sustained activation of the receptor and of its downstream effectors (extracellular signal-regulated kinase 1/2, signal transducer and activator of transcription 3) in both basal and EGF-stimulated conditions. Cells lacking EBP50 showed epithelial-to-mesenchymal transition-associated features, including reduction in E-cadherin and cytokeratin-19 expression, induction of S100A4 and of the E-cadherin transcriptional repressor, Slug, and loss of cell polarity. Accordingly, depletion of EBP50 induced the disruption of adherens junctional complexes, the development of lamellipodia structures and the subsequent acquisition of motility properties. All these phenotypic changes were prevented upon inhibition of EGFR tyrosine kinase by gefitinib. These findings indicate that loss of EBP50 at the plasma membrane in tumor cells may contribute to biliary carcinogenesis

  20. EGFR inhibition evokes innate drug resistance in lung cancer cells by preventing Akt activity and thus inactivating Ets-1 function.

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    Phuchareon, Janyaporn; McCormick, Frank; Eisele, David W; Tetsu, Osamu

    2015-07-21

    Nonsmall cell lung cancer (NSCLC) is the leading cause of cancer death worldwide. About 14% of NSCLCs harbor mutations in epidermal growth factor receptor (EGFR). Despite remarkable progress in treatment with tyrosine kinase inhibitors (TKIs), only 5% of patients achieve tumor reduction >90%. The limited primary responses are attributed partly to drug resistance inherent in the tumor cells before therapy begins. Recent reports showed that activation of receptor tyrosine kinases (RTKs) is an important determinant of this innate drug resistance. In contrast, we demonstrate that EGFR inhibition promotes innate drug resistance despite blockade of RTK activity in NSCLC cells. EGFR TKIs decrease both the mitogen-activated protein kinase (MAPK) and Akt protein kinase pathways for a short time, after which the Ras/MAPK pathway becomes reactivated. Akt inhibition selectively blocks the transcriptional activation of Ets-1, which inhibits its target gene, dual specificity phosphatase 6 (DUSP6), a negative regulator specific for ERK1/2. As a result, ERK1/2 is activated. Furthermore, elevated c-Src stimulates Ras GTP-loading and activates Raf and MEK kinases. These observations suggest that not only ERK1/2 but also Akt activity is essential to maintain Ets-1 in an active state. Therefore, despite high levels of ERK1/2, Ets-1 target genes including DUSP6 and cyclins D1, D3, and E2 remain suppressed by Akt inhibition. Reduction of DUSP6 in combination with elevated c-Src renews activation of the Ras/MAPK pathway, which enhances cell survival by accelerating Bim protein turnover. Thus, EGFR TKIs evoke innate drug resistance by preventing Akt activity and inactivating Ets-1 function in NSCLC cells.

  1. Anti-epidermal growth factor receptor monoclonal antibody cetuximab inhibits EGFR/HER-2 heterodimerization and activation.

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    Patel, Dipa; Bassi, Rajiv; Hooper, Andrea; Prewett, Marie; Hicklin, Daniel J; Kang, Xiaoqiang

    2009-01-01

    Human carcinomas frequently express one or more members of the epidermal growth factor receptor family. Two family members, epidermal growth factor receptor (EGFR) and c-erbB2/neu (HER2), homodimerize or heterodimerize upon activation with ligand and trigger potent mechanisms of cellular proliferation, differentiation and migration. In this study, we examined the effect of the anti-EGFR monoclonal antibody Erbitux (cetuximab) on human tumor cells expressing both EGFR and HER2. Investigation of the effect of cetuximab on the activation of EGFR-EGFR, EGFR-HER2 and HER2-HER2 homodimers and heterodimers was conducted using the NCI-N87 human gastric carcinoma cell line. Treatment of NCI-N87 cells with cetuximab completely inhibited formation of EGFR-EGFR homodimers and EGFR-HER2 heterodimers. Activation of HER2-HER2 homodimers was not appreciably stimulated by exogenous ligand and was not inhibited by cetuximab treatment. Furthermore, cetuximab inhibited EGF-induced EGFR and HER2 phosphorylation in CAL27, NCI-H226 and NCI-N87 cells. The activation of downstream signaling molecules such as AKT, MAPK and STAT-3 were also inhibited by cetuximab in these cells. To examine the effect of cetuximab on the growth of tumors in vivo, athymic mice bearing established NCI-N87 or CAL27 xenografts were treated with cetuximab (1 mg, i.p., q3d). The growth of NCI-N87 and CAL27 tumors was significantly inhibited with cetuximab therapy compared to the control groups (p<0.0001 in both cases). In the CAL27 xenograft model, tumor growth inhibition by cetuximab treatment was similar to that by cetuximab and trastuzumab combination treatment. Immunohistological analysis of cetuximab-treated tumors showed a decrease in EGFR-HER2 signaling and reduced tumor cell proliferation. These results suggest that cetuximab may be useful in the treatment of carcinomas co-expressing EGFR and HER2. PMID:19082474

  2. K-RAS(V12) Induces Autocrine Production of EGFR Ligands and Mediates Radioresistance Through EGFR-Dependent Akt Signaling and Activation of DNA-PKcs

    International Nuclear Information System (INIS)

    Purpose: It is known that postirradiation survival of tumor cells presenting mutated K-RAS is mediated through autocrine activation of epidermal growth factor receptor (EGFR). In this study the molecular mechanism of radioresistance of cells overexpressing mutated K-RAS(V12) was investigated. Methods and Materials: Head-and-neck cancer cells (FaDu) presenting wild-type K-RAS were transfected with empty vector or vector expressing mutated K-RAS(V12). The effect of K-RAS(V12) on autocrine production of EGFR ligands, activation of EGFR downstream pathways, DNA damage repair, and postirradiation survival was analyzed. Results: Conditioned medium collected from K-RAS(V12)–transfected cells enhanced activation of the phosphatidylinositol-3-kinase–Akt pathway and increased postirradiation survival of wild-type K-RAS parental cells when compared with controls. These effects were reversed by amphiregulin (AREG)–neutralizing antibody. In addition, secretion of the EGFR ligands AREG and transforming growth factor α was significantly increased upon overexpression of K-RAS(V12). Expression of mutated K-RAS(V12) resulted in an increase in radiation-induced DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phosphorylation at S2056. This increase was accompanied by increased repair of DNA double-strand breaks. Abrogation of DNA-PKcs phosphorylation by serum depletion or AREG-neutralizing antibody underscored the role of autocrine production of EGFR ligands, namely, AREG, in regulating DNA-PKcs activation in K-RAS mutated cells. Conclusions: These data indicate that radioresistance of K-RAS mutated tumor cells is at least in part due to constitutive production of EGFR ligands, which mediate enhanced repair of DNA double-strand breaks through the EGFR–phosphatidylinositol-3-kinase–Akt cascade.

  3. Achievement of Cure with Gefitinib in Advanced Lung Adenocarcinoma Harboring an Activating EGFR Mutation: A Case Report

    Directory of Open Access Journals (Sweden)

    Taiji Kuwata

    2016-10-01

    Full Text Available Tyrosine kinase inhibitors (TKIs of the epidermal growth factor receptor (EGFR may achieve long-term survival in selected cases with advanced non-small cell lung cancer harboring activating mutations in the EGFR gene, but a cured case has not been reported yet. Here, we present the first case of EGFR-mutated lung adenocarcinoma cured with an EGFR-TKI, as the 75-year-old Japanese man has achieved complete response with gefitinib treatment and has survived without tumor 10 years after termination of gefitinib treatment.

  4. Proteomic profiling of patient-derived glioblastoma xenografts identifies a subset with activated EGFR: implications for drug development.

    Science.gov (United States)

    Brown, Kristine E; Chagoya, Gustavo; Kwatra, Shawn G; Yen, Timothy; Keir, Stephen T; Cooter, Mary; Hoadley, Katherine A; Rasheed, Ahmed; Lipp, Eric S; Mclendon, Roger; Ali-Osman, Francis; Bigner, Darell D; Sampson, John H; Kwatra, Madan M

    2015-06-01

    The development of drugs to inhibit glioblastoma (GBM) growth requires reliable pre-clinical models. To date, proteomic level validation of widely used patient-derived glioblastoma xenografts (PDGX) has not been performed. In the present study, we characterized 20 PDGX models according to subtype classification based on The Cancer Genome Atlas criteria, TP53, PTEN, IDH 1/2, and TERT promoter genetic analysis, EGFR amplification status, and examined their proteomic profiles against those of their parent tumors. The 20 PDGXs belonged to three of four The Cancer Genome Atlas subtypes: eight classical, eight mesenchymal, and four proneural; none neural. Amplification of EGFR gene was observed in 9 of 20 xenografts, and of these, 3 harbored the EGFRvIII mutation. We then performed proteomic profiling of PDGX, analyzing expression/activity of several proteins including EGFR. Levels of EGFR phosphorylated at Y1068 vary considerably between PDGX samples, and this pattern was also seen in primary GBM. Partitioning of 20 PDGX into high (n = 5) and low (n = 15) groups identified a panel of proteins associated with high EGFR activity. Thus, PDGX with high EGFR activity represent an excellent pre-clinical model to develop therapies for a subset of GBM patients whose tumors are characterized by high EGFR activity. Further, the proteins found to be associated with high EGFR activity can be monitored to assess the effectiveness of targeting EGFR. The development of drugs to inhibit glioblastoma (GBM) growth requires reliable pre-clinical models. We validated proteomic profiles using patient-derived glioblastoma xenografts (PDGX), characterizing 20 PDGX models according to subtype classification based on The Cancer Genome Atlas (TCGA) criteria, TP53, PTEN, IDH 1/2, and TERT promoter genetic analysis, EGFR amplification status, and examined their proteomic profiles against those of their parent tumors. Proteins found to be associated with high EGFR activity represent potential

  5. Activation of the epidermal growth factor receptor (EGFR) by a novel metalloprotease pathway.

    LENUS (Irish Health Repository)

    Bergin, David A

    2008-11-14

    Neutrophil Elastase (NE) is a pro-inflammatory protease present at higher than normal levels in the lung during inflammatory disease. NE regulates IL-8 production from airway epithelial cells and can activate both EGFR and TLR4. TACE\\/ADAM17 has been reported to trans-activate EGFR in response to NE. Here, using 16HBE14o-human bronchial epithelial cells we demonstrate a new mechanism by which NE regulates both of these events. A high molecular weight soluble metalloprotease activity detectable only in supernatants from NE-treated cells by gelatin and casein zymography was confirmed to be meprin alpha by Western immunoblotting. In vitro studies demonstrated the ability of NE to activate meprin alpha, which in turn could release soluble TGFalpha and induce IL-8 production from 16HBE14o- cells. These effects were abrogated by actinonin, a specific meprin inhibitor. NE-induced IL-8 expression was also inhibited by meprin alpha siRNA. Immunoprecipitation studies detected EGFR\\/TLR4 complexes in NE-stimulated cells overexpressing these receptors. Confocal studies confirmed colocalization of EGFR and TLR4 in 16HBE14o- cells stimulated with meprin alpha. NFkappaB was also activated via MyD88 in these cells by meprin alpha. In bronchoalveolar lavage fluid from NE knock-out mice infected intra-tracheally with Pseudomonas aeruginosa meprin alpha was significantly decreased compared with control mice, and was significantly increased and correlated with NE activity, in bronchoalveolar lavage fluid from individuals with cystic fibrosis but not healthy controls. The data describe a previously unidentified lung metalloprotease meprin alpha, and its role in NE-induced EGFR and TLR4 activation and IL-8 production.

  6. Potent Systemic Anticancer Activity of Adenovirally Expressed EGFR-Selective TRAIL Fusion Protein

    NARCIS (Netherlands)

    Bremer, Edwin; van Dam, Gooitzen M.; de Bruyn, Marco; van Riezen, Manon; Dijkstra, Marike; Kamps, Gera; Helfrich, Wijnand; Haisma, Hidde

    2008-01-01

    Previously, we demonstrated potent tumor cell-selective pro-apoptotic activity of scFv425:sTRAIL, a recombinant fusion protein comprised of EGFR-directed antibody fragment (scFv425) genetically fused to human soluble TNF-related apoptosis-inducing ligand (sTRAIL). Here, we report on the promising th

  7. Mutation specific functions of EGFR result in a mutation-specific downstream pathway activation

    NARCIS (Netherlands)

    L. Eraslan-Erdem (Lale); Y. Gao; N.K. Kloosterhof (Nanne); Y. Atlasi (Yaser); J.A.A. Demmers (Jeroen); A. Sacchetti (Andrea); J.M. Kros (Johan); P.A.E. Sillevis Smitt (Peter); J.G.J.V. Aerts (Joachim); P.J. French (Pim)

    2015-01-01

    markdownabstractBackground: Epidermal growth factor receptor (EGFR) is frequently mutated in various types of cancer. Although all oncogenic mutations are considered activating, different tumour types have different mutation spectra. It is possible that functional differences underlie this tumour-ty

  8. Effects of activated fibroblasts on phenotype modulation, EGFR signalling and cell cycle regulation in OSCC cells

    Energy Technology Data Exchange (ETDEWEB)

    Berndt, Alexander, E-mail: alexander.berndt@med.uni-jena.de [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Büttner, Robert, E-mail: Robert-Buettner@gmx.net [Institute of Biochemistry and Biophysics, Friedrich Schiller University Jena, 07740 Jena (Germany); Gühne, Stefanie, E-mail: stefanie_guehne@gmx.net [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Gleinig, Anna, E-mail: annagleinig@yahoo.com [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Richter, Petra, E-mail: P.Richter@med.uni-jena.de [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Chen, Yuan, E-mail: Yuan.Chen@med.uni-jena.de [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Franz, Marcus, E-mail: Marcus.Franz@med.uni-jena.de [Clinic of Internal Medicine I, Jena University Hospital, 07740 Jena (Germany); Liebmann, Claus, E-mail: Claus.Liebmann@uni-jena.de [Institute of Biochemistry and Biophysics, Friedrich Schiller University Jena, 07740 Jena (Germany)

    2014-04-01

    Crosstalk between carcinoma associated fibroblasts (CAFs) and oral squamous cell carcinoma (OSCC) cells is suggested to mediate phenotype transition of cancer cells as a prerequisite for tumour progression, to predict patients’ outcome, and to influence the efficacy of EGFR inhibitor therapies. Here we investigate the influence of activated fibroblasts as a model for CAFs on phenotype and EGFR signalling in OSCC cells in vitro. For this, immortalised hTERT-BJ1 fibroblasts were activated with TGFβ1 and PDGFAB to generate a myofibroblast or proliferative phenotype, respectively. Conditioned media (FCM{sub TGF}, FCM{sub PDGF}) were used to stimulate PE/CA-PJ15 OSCC cells. Results were compared to the effect of conditioned media of non-stimulated fibroblasts (FCM{sub B}). FCM{sub TGF} stimulation leads to an up-regulation of vimentin in the OSCC cells and an enhancement of invasive behaviour, indicating EMT-like effects. Similarly, FCM{sub TGF}≫FCM{sub PDGF} induced up-regulation of EGFR, but not of ErbB2/ErbB3. In addition, we detected an increase in basal activities of ERK, PI3K/Akt and Stat3 (FCM{sub TGF}>FCM{sub PDGF}) accompanied by protein interaction of vimentin with pERK. These effects are correlated with an increased proliferation. In summary, our results suggest that the activated myofibroblast phenotype provides soluble factors which are able to induce EMT-like phenomena and to increase EGFR signalling as well as cell proliferation in OSCC cells. Our results indicate a possible influence of activated myofibroblasts on EGFR-inhibitor therapy. Therefore, CAFs may serve as promising novel targets for combined therapy strategies. - Highlights: • A cell culture model for cancer associated fibroblasts is described. • The mutual interaction with OSCC cells leads to up-regulation of EGFR in tumour cells. • mCAF induces EGFR downstream signalling with increased proliferation in OSCC. • Erk activation is associated with protein interaction with vimentin

  9. Unravelling the pivotal role of Alix in MVB sorting and silencing of the activated EGFR.

    Science.gov (United States)

    Sun, Sheng; Zhou, Xi; Zhang, Wei; Gallick, Gary E; Kuang, Jian

    2015-03-15

    Endosomal sorting complex required for transport (ESCRT)-III-mediated membrane invagination and scission are a critical step in multivesicular body (MVB) sorting of ubiquitinated membrane receptors, and generally thought to be required for degradation of these receptors in lysosomes. The adaptor protein Alix is critically involved in multiple ESCRT-III-mediated, membrane-remodelling processes in mammalian cells. However, Alix knockdown does not inhibit degradation of the activated epidermal growth factor receptor (EGFR) in mammalian cell lines, leading to a widely held notion that Alix is not critically involved in MVB sorting of ubiquitinated membrane receptors in mammalian cells. In the present study, we demonstrate that, despite its non-essential role in degradation of the activated EGFR, Alix plays a critical role in its MVB sorting and silencing Epidermal growth factor (EGF) stimulation of mammalian cell lines induces Alix's interaction with the ubiquitinated EGFR via the Alix V domain, and increases Alix's association with membrane-bound charged multivesicular body protein 4 (CHMP4) via the Alix Bro1 domain. Under both continuous and pulse-chase EGF stimulation conditions, inhibition of Alix's interaction with membrane-bound CHMP4, inhibition of Alix dimerization through the V domain or Alix knockdown dramatically inhibits MVB sorting of the activated EGFR and promotes sustained activation of extracellular-signal regulated kinase (ERK)1/2. Under the continuous EGF stimulation conditions, these cell treatments also retard degradation of the activated EGFR. These findings indicate that Alix is critically involved in MVB sorting of ubiquitinated membrane receptors in mammalian cells.

  10. Synergistic anti-proliferative and pro-apoptotic activity of combined therapy with bortezomib, a proteasome inhibitor, with anti-epidermal growth factor receptor (EGFR) drugs in human cancer cells.

    Science.gov (United States)

    Cascone, Tina; Morelli, Maria Pia; Morgillo, Floriana; Kim, Woo-Young; Rodolico, Gabriella; Pepe, Stefano; Tortora, Giampaolo; Berrino, Liberato; Lee, Ho-Young; Heymach, John V; Ciardiello, Fortunato

    2008-09-01

    The proteasome plays a pivotal role in the turnover of regulatory transduction proteins induced by activated cell membrane growth factor receptors. The epidermal growth factor receptor (EGFR) pathway is crucial in the development and progression of human epithelial cancers. Proteasome inhibition may sensitize human cancer cell lines to EGFR inhibitors. We investigated the growth inhibitory and pro-apoptotic effects of the proteasome inhibitor bortezomib in combination with anti-EGFR drugs, such as gefitinib, vandetanib, and cetuximab in EGFR-expressing human cancer cell lines. Bortezomib determined dose-dependent growth inhibition in a nine cancer cell line panel (IC(50) values, range 6-42 nM). A significant synergistic growth inhibitory effect was observed with the combination of bortezomib and each EGFR inhibitor in all cell lines (combination index, CI, range 0.10-0.55), which was accompanied by a significant induction in apoptosis by the combined treatment with bortezomib, cetuximab and vandetanib. In HCT-116 colon cancer and A549 lung adenocarcinoma cells, bortezomib plus EGFR inhibitor treatment induced a more effective inhibition of EGFR-activated down-stream signals, including a marked suppression in activated, phosphorylated Akt (P-Akt). In contrast, overexpression of a constitutively active P-Akt protected A549 cells by cell growth inhibition and apoptosis following treatment with bortezomib and EGFR inhibitors. The combined treatment with bortezomib and EGFR inhibitors has a synergistic growth inhibitory and pro-apoptotic activity in different human cancer cells which possess a functional EGFR-dependent autocrine growth pathway through to a more efficient and sustained inhibition of Akt.

  11. Model-based analysis of HER activation in cells co-expressing EGFR, HER2 and HER3.

    Science.gov (United States)

    Shankaran, Harish; Zhang, Yi; Tan, Yunbing; Resat, Haluk

    2013-01-01

    The HER/ErbB family of receptor tyrosine kinases drives critical responses in normal physiology and cancer, and the expression levels of the various HER receptors are critical determinants of clinical outcomes. HER activation is driven by the formation of various dimer complexes between members of this receptor family. The HER dimer types can have differential effects on downstream signaling and phenotypic outcomes. We constructed an integrated mathematical model of HER activation, and trafficking to quantitatively link receptor expression levels to dimerization and activation. We parameterized the model with a comprehensive set of HER phosphorylation and abundance data collected in a panel of human mammary epithelial cells expressing varying levels of EGFR/HER1, HER2 and HER3. Although parameter estimation yielded multiple solutions, predictions for dimer phosphorylation were in agreement with each other. We validated the model using experiments where pertuzumab was used to block HER2 dimerization. We used the model to predict HER dimerization and activation patterns in a panel of human mammary epithelial cells lines with known HER expression levels in response to stimulations with ligands EGF and HRG. Simulations over the range of expression levels seen in various cell lines indicate that: i) EGFR phosphorylation is driven by HER1-HER1 and HER1-HER2 dimers, and not HER1-HER3 dimers, ii) HER1-HER2 and HER2-HER3 dimers both contribute significantly to HER2 activation with the EGFR expression level determining the relative importance of these species, and iii) the HER2-HER3 dimer is largely responsible for HER3 activation. The model can be used to predict phosphorylated dimer levels for any given HER expression profile. This information in turn can be used to quantify the potencies of the various HER dimers, and can potentially inform personalized therapeutic approaches. PMID:23990774

  12. Model-based analysis of HER activation in cells co-expressing EGFR, HER2 and HER3.

    Directory of Open Access Journals (Sweden)

    Harish Shankaran

    Full Text Available The HER/ErbB family of receptor tyrosine kinases drives critical responses in normal physiology and cancer, and the expression levels of the various HER receptors are critical determinants of clinical outcomes. HER activation is driven by the formation of various dimer complexes between members of this receptor family. The HER dimer types can have differential effects on downstream signaling and phenotypic outcomes. We constructed an integrated mathematical model of HER activation, and trafficking to quantitatively link receptor expression levels to dimerization and activation. We parameterized the model with a comprehensive set of HER phosphorylation and abundance data collected in a panel of human mammary epithelial cells expressing varying levels of EGFR/HER1, HER2 and HER3. Although parameter estimation yielded multiple solutions, predictions for dimer phosphorylation were in agreement with each other. We validated the model using experiments where pertuzumab was used to block HER2 dimerization. We used the model to predict HER dimerization and activation patterns in a panel of human mammary epithelial cells lines with known HER expression levels in response to stimulations with ligands EGF and HRG. Simulations over the range of expression levels seen in various cell lines indicate that: i EGFR phosphorylation is driven by HER1-HER1 and HER1-HER2 dimers, and not HER1-HER3 dimers, ii HER1-HER2 and HER2-HER3 dimers both contribute significantly to HER2 activation with the EGFR expression level determining the relative importance of these species, and iii the HER2-HER3 dimer is largely responsible for HER3 activation. The model can be used to predict phosphorylated dimer levels for any given HER expression profile. This information in turn can be used to quantify the potencies of the various HER dimers, and can potentially inform personalized therapeutic approaches.

  13. Model-based Analysis of HER Activation in Cells Co-Expressing EGFR, HER2 and HER3.

    Energy Technology Data Exchange (ETDEWEB)

    Shankaran, Harish; Zhang, Yi; Tan, Yunbing; Resat, Haluk

    2013-08-22

    The HER/ErbB family of receptor tyrosine kinases drive critical responses in normal physiology and cancer, and the expression levels of the various HER receptors are critical determinants of clinical outcomes. HER activation is driven by the formation of various dimer complexes between members of this receptor family. The HER dimer types can have differential effects on downstream signaling and phenotypic outcomes. We constructed an integrated mathematical model of HER activation and trafficking to quantitatively link receptor expression levels to dimerization and activation. We parameterized the model with a comprehensive set of HER phosphorylation and abundance data collected in a panel of human mammary epithelial cells expressing varying levels of EGFR, HER2 and HER3. Although parameter estimation yielded multiple solutions, predictions for dimer phosphorylation were in agreement with each other. We validated the model using experiments where pertuzumab was used to block HER2 dimerization. We used the model to predict HER dimerization and activation patterns in a panel of epithelial cells lines with known HER expression levels. Simulations over the range of expression levels seen in various cell lines indicate that: i) EGFR phosphorylation is driven by HER1/1 and HER1/2 dimers, and not HER1/3 dimers, ii) HER1/2 and HER2/3 dimers both contribute significantly to HER2 activation with the EGFR expression level determining the relative importance of these species, and iii) the HER2/3 dimer is largely responsible for HER3 activation. The model can be used to predict phosphorylated dimer levels for any given HER expression profile. This information in turn can be used to quantify the potencies of the various HER dimers, and can potentially inform personalized therapeutic approaches.

  14. Alpha6beta4 integrin crosslinking induces EGFR clustering and promotes EGF-mediated Rho activation in breast cancer

    Directory of Open Access Journals (Sweden)

    Woodward Wendy A

    2009-05-01

    Full Text Available Abstract Background The α6β4 integrin is overexpressed in the basal subtype of breast cancer and plays an important role in tumor cell motility and invasion. EGFR is also overexpressed in the basal subtype of breast cancer, and crosstalk between α6β4 integrin and EGFR appears to be important in tumor progression. Methods We evaluated the effects of α6β4 crosslinking on the distribution and function of EGFR in breast carcinoma cell line MDA-MB-231. Receptor distribution was evaluated by fluorescence microscopy and multispectral imaging flow cytometry, and ligand-mediated EGFR signaling was evaluated using Western blots and a Rho pull-down assay. Results Antibody-mediated crosslinking of α6β4 integrin was sufficient to induce cell-surface clustering of not only α6β4 but also EGFR in nonadherent cells. The induced clustering of EGFR was observed minimally after 5 min of integrin crosslinking but was more prominent after 15 min. EGFR clustering had minimal effect on the phosphorylation of Akt or Erk1,2 in response to EGF in suspended cells or in response to HB-EGF in adherent cells. However, EGFR clustering induced by crosslinking α6β4 had a marked effect on Rho activation in response to EGF. Conclusion Crosslinking α6β4 integrin in breast carcinoma cells induces EGFR clustering and preferentially promotes Rho activation in response to EGF. We hypothesize that this integrin-EGFR crosstalk may facilitate tumor cell cytoskeletal rearrangements important for tumor progression.

  15. EGFR is dispensable for c-Met-mediated proliferation and survival activities in mouse adult liver oval cells.

    Science.gov (United States)

    Martínez-Palacián, A; del Castillo, G; Herrera, B; Fernández, M; Roncero, C; Fabregat, I; Sánchez, A

    2012-02-01

    Liver progenitor cells rise as potential critical players in hepatic regeneration but also carcinogenesis. It is therefore mandatory to define the signals controlling their activation and expansion. Recently, by using a novel in vitro model of oval cell lines expressing a mutant tyrosine kinase-inactive form of c-Met we demonstrated that autocrine c-Met signalling plays an essential role in promoting oval cell survival. Here, we investigated the significance of the epidermal growth factor receptor (EGFR) signalling in oval cell proliferation and survival, as well as a potential functional crosstalk between the c-Met and the EGFR pathways. We found an autocrine activation of the EGFR-triggered pathway in Met(flx/flx) and Met(-/-) oval cells as judged by constitutive expression of the EGFR ligands, transforming growth factor-alpha (TGF-α) and heparin-binding EGF like growth factor (HB-EGF), and activation of EGFR. On the other hand, treatment with AG1478, a specific inhibitor of EGFR, effectively blocked endogenous and EGF-induced proliferation, while increased serum withdrawal and transforming growth factor-beta (TGF-β)-induced apoptosis. These results suggest that constitutively activated EGFR might promote oval cell proliferation and survival. We found that hepatocyte growth factor (HGF) does not transactivate EGFR nor EGF transactivates c-Met. Furthermore, treatment with AG1478 or EGFR gene silencing did not interfere with HGF-mediated activation of target signals, such as protein kinase B (AKT/PKB), and extracellular signal-regulated kinases 1/2 (ERK 1/2), nor did it have any effect on HGF-induced proliferative and antiapoptotic activities in Met(flx/flx) cells, showing that HGF does not require EGFR activation to mediate such responses. EGF induced proliferation and survival equally in Met(flx/flx) and Met(-/-) oval cells, proving that EGFR signalling does not depend on c-Met tyrosine kinase activity. Together, our results provide strong evidence that in

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

    International Nuclear Information System (INIS)

    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. 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). 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. 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

  17. NF-{kappa}B signaling is activated and confers resistance to apoptosis in three-dimensionally cultured EGFR-mutant lung adenocarcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Sakuma, Yuji, E-mail: ysakuma@gancen.asahi.yokohama.jp [Molecular Pathology and Genetics Division, Kanagawa Cancer Center Research Institute, Yokohama (Japan); Yamazaki, Yukiko; Nakamura, Yoshiyasu; Yoshihara, Mitsuyo; Matsukuma, Shoichi; Koizume, Shiro; Miyagi, Yohei [Molecular Pathology and Genetics Division, Kanagawa Cancer Center Research Institute, Yokohama (Japan)

    2012-07-13

    Highlights: Black-Right-Pointing-Pointer EGFR-mutant cells in 3D culture resist EGFR inhibition compared with suspended cells. Black-Right-Pointing-Pointer Degradation of I{kappa}B and activation of NF-{kappa}B are observed in 3D-cultured cells. Black-Right-Pointing-Pointer Inhibiting NF-{kappa}B 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 I{kappa}B{alpha}, the inhibitor of nuclear factor (NF)-{kappa}B, was observed only in the 3D-cultured cells, leading to an increase in the activation of NF-{kappa}B. Moreover, the inhibition of NF-{kappa}B with pharmacological inhibitors enhanced EGFR TKI-induced apoptosis in 3D-cultured EGFR-mutant cells. These results suggest that inhibition of NF-{kappa}B signaling would render ECM-adherent EGFR-mutant lung adenocarcinoma cells in vivo more susceptible to EGFR TKI-induced cell death.

  18. Exocyst Sec10 protects renal tubule cells from injury by EGFR/MAPK activation and effects on endocytosis.

    Science.gov (United States)

    Fogelgren, Ben; Zuo, Xiaofeng; Buonato, Janine M; Vasilyev, Aleksandr; Baek, Jeong-In; Choi, Soo Young; Chacon-Heszele, Maria F; Palmyre, Aurélien; Polgar, Noemi; Drummond, Iain; Park, Kwon Moo; Lazzara, Matthew J; Lipschutz, Joshua H

    2014-12-15

    Acute kidney injury is common and has a high mortality rate, and no effective treatment exists other than supportive care. Using cell culture models, we previously demonstrated that exocyst Sec10 overexpression reduced damage to renal tubule cells and speeded recovery and that the protective effect was mediated by higher basal levels of mitogen-activated protein kinase (MAPK) signaling. The exocyst, a highly-conserved eight-protein complex, is known for regulating protein trafficking. Here we show that the exocyst biochemically interacts with the epidermal growth factor receptor (EGFR), which is upstream of MAPK, and Sec10-overexpressing cells express greater levels of phosphorylated (active) ERK, the final step in the MAPK pathway, in response to EGF stimulation. EGFR endocytosis, which has been linked to activation of the MAPK pathway, increases in Sec10-overexpressing cells, and gefitinib, a specific EGFR inhibitor, and Dynasore, a dynamin inhibitor, both reduce EGFR endocytosis. In turn, inhibition of the MAPK pathway reduces ligand-mediated EGFR endocytosis, suggesting a potential feedback of elevated ERK activity on EGFR endocytosis. Gefitinib also decreases MAPK signaling in Sec10-overexpressing cells to levels seen in control cells and, demonstrating a causal role for EGFR, reverses the protective effect of Sec10 overexpression following cell injury in vitro. Finally, using an in vivo zebrafish model of acute kidney injury, morpholino-induced knockdown of sec10 increases renal tubule cell susceptibility to injury. Taken together, these results suggest that the exocyst, acting through EGFR, endocytosis, and the MAPK pathway is a candidate therapeutic target for acute kidney injury.

  19. ZD6474, an inhibitor of VEGFR and EGFR tyrosine kinase activity in combination with radiotherapy

    International Nuclear Information System (INIS)

    Radiation enhances both epithelial growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF) expression, which are a part of key pathways for tumor progression. Some tumors may not respond well to EGFR inhibitors alone or may develop resistance to EGFR inhibitors. Therefore, drug therapy targeted to VEGF receptors and EGFRs, when combined with radiotherapy (RT), may improve tumor control and provide wider applicability. This article focuses on ZD6474, an inhibitor of EGFR and VEGF receptor signaling in combination with RT. We discuss preclinical and clinical studies with RT and inhibitors of VEGF or EGFR signaling first. We then address issues associated with ZD6474 pharmacokinetic dosing, and scheduling when combined with RT. We also discuss ZD6474 in the context of anti-EGFR therapy resistance. Dual inhibition of EGFR and VEGF receptor signaling pathways shows promise in enhancing RT efficacy

  20. Increased concentrations of growth factors and activation of the EGFR system in breast cancer

    DEFF Research Database (Denmark)

    Aalund Olsen, Dorte; Bechmann, Troels; Østergaard, Birthe;

    2012-01-01

    In this study the total and phosphorylated amount of epidermal growth factor receptor 1 (EGFR) and 2 (HER2) were measured together with EGFR ligands in tissue samples of breast cancer patients in order to investigate interrelations and possible prognostic values.......In this study the total and phosphorylated amount of epidermal growth factor receptor 1 (EGFR) and 2 (HER2) were measured together with EGFR ligands in tissue samples of breast cancer patients in order to investigate interrelations and possible prognostic values....

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

    Directory of Open Access Journals (Sweden)

    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.

  2. TWIST1 a new determinant of epithelial to mesenchymal transition in EGFR mutated lung adenocarcinoma.

    Directory of Open Access Journals (Sweden)

    Karine Pallier

    Full Text Available Metastasis is a multistep process and the main cause of mortality in lung cancer patients. We previously showed that EGFR mutations were associated with a copy number gain at a locus encompassing the TWIST1 gene on chromosome 7. TWIST1 is a highly conserved developmental gene involved in embryogenesis that may be reactivated in cancers promoting both malignant conversion and cancer progression through an epithelial to mesenchymal transition (EMT. The aim of this study was to investigate the possible implication of TWIST1 reactivation on the acquisition of a mesenchymal phenotype in EGFR mutated lung cancer. We studied a series of consecutive lung adenocarcinoma from Caucasian non-smokers for which surgical frozen samples were available (n = 33 and showed that TWIST1 expression was linked to EGFR mutations (P<0.001, to low CDH1 expression (P<0.05 and low disease free survival (P = 0.044. To validate that TWIST1 is a driver of EMT in EGFR mutated lung cancer, we used five human lung cancer cell lines and demonstrated that EMT and the associated cell mobility were dependent upon TWIST1 expression in cells with EGFR mutation. Moreover a decrease of EGFR pathway stimulation through EGF retrieval or an inhibition of TWIST1 expression by small RNA technology reversed the phenomenon. Collectively, our in vivo and in vitro findings support that TWIST1 collaborates with the EGF pathway in promoting EMT in EGFR mutated lung adenocarcinoma and that large series of EGFR mutated lung cancer patients are needed to further define the prognostic role of TWIST1 reactivation in this subgroup.

  3. CRIPTO1 expression in EGFR-mutant NSCLC elicits intrinsic EGFR-inhibitor resistance

    OpenAIRE

    Park, Kang-Seo; Raffeld, Mark; Moon, Yong Wha; Xi, Liqiang; Bianco, Caterina; Van Pham, Trung; Lee, Liam C.; Mitsudomi, Tetsuya; Yatabe, Yasushi; Okamoto, Isamu; Subramaniam, Deepa; Mok, Tony; Rosell, Rafael; Luo, Ji; Salomon, David S.

    2014-01-01

    The majority of non–small cell lung cancer (NSCLC) patients harbor EGFR-activating mutations that can be therapeutically targeted by EGFR tyrosine kinase inhibitors (EGFR-TKI), such as erlotinib and gefitinib. Unfortunately, a subset of patients with EGFR mutations are refractory to EGFR-TKIs. Resistance to EGFR inhibitors reportedly involves SRC activation and induction of epithelial-to-mesenchymal transition (EMT). Here, we have demonstrated that overexpression of CRIPTO1, an EGF-CFC protei...

  4. 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.

  5. Synthesis of new 4-anilinoquinazoline analogues and evaluation of their EGFR inhibitor activity.

    Science.gov (United States)

    Wang, Zheng; WANG, Cui-Ling; Li, Jun-lin; Zhang, Ning; Sun, Yan-ni; Liu, Zhu-lan; Tang, Zhi-shu; Liu, Jian-li

    2015-12-01

    Thirteen of 4-anilinoquinazoline derivatives with imine groups at position 6 of quinazoline ring were synthesized and their antitumor activities were evaluated by MTT assay and Western blotting analysis. Among these compounds, 13a-131 were reported first time. The MTT assay was carried out on three human cancer cell lines (A549, HepG2 and SMMC7721) with EGFR highly expressed. Among the tested compounds, 13i and 13j exhibited notable inhibition potency and their IC50 values on three cell lines were equivalent to or less than those of gefitinib. Compound 14, without imine group substituted, displayed excellent inhibitor potency only on A549 cell line. Compounds 14 and 13j were chosen to perform Western blotting analysis on A549. The results showed that both of the compounds could inhibit the expression level of phosphorylated EGFR remarkably. It was concluded that the inhibitor potency of compound 14 was almost equivalent to that of gefitinib and the inhibitor potency of 13j was better than that of gefitinib. PMID:27169285

  6. Deoxycholic acid mediates non-canonical EGFR-MAPK activation through the induction of calcium signaling in colon cancer cells.

    Science.gov (United States)

    Centuori, Sara M; Gomes, Cecil J; Trujillo, Jesse; Borg, Jamie; Brownlee, Joshua; Putnam, Charles W; Martinez, Jesse D

    2016-07-01

    Obesity and a western diet have been linked to high levels of bile acids and the development of colon cancer. Specifically, increased levels of the bile acid deoxycholic acid (DCA), an established tumor promoter, has been shown to correlate with increased development of colorectal adenomas and progression to carcinoma. Herein we investigate the mechanism by which DCA leads to EGFR-MAPK activation, a candidate mechanism by which DCA may promote colorectal tumorigenesis. DCA treated colon cancer cells exhibited strong and prolonged activation of ERK1/2 when compared to EGF treatment alone. We also showed that DCA treatment prevents EGFR degradation as opposed to the canonical EGFR recycling observed with EGF treatment. Moreover, the combination of DCA and EGF treatment displayed synergistic activity, suggesting DCA activates MAPK signaling in a non-canonical manner. Further evaluation showed that DCA treatment increased intracellular calcium levels and CAMKII phosphorylation, and that blocking calcium with BAPTA-AM abrogated MAPK activation induced by DCA, but not by EGF. Finally we showed that DCA-induced CAMKII leads to MAPK activation through the recruitment of c-Src. Taken together, we demonstrated that DCA regulates MAPK activation through calcium signaling, an alternative mechanism not previously recognized in human colon cancer cells. Importantly, this mechanism allows for EGFR to escape degradation and thus achieve a constitutively active state, which may explain its tumor promoting effects.

  7. Novel matrine derivative MD-1 attenuates hepatic fibrosis by inhibiting EGFR activation of hepatic stellate cells.

    Science.gov (United States)

    Feng, Yi; Ying, Hai-Yan; Qu, Ying; Cai, Xiao-Bo; Xu, Ming-Yi; Lu, Lun-Gen

    2016-09-01

    Matrine (MT), the effective component of Sophora flavescens Ait, has been shown to have anti-inflammation, immune-suppressive, anti-tumor, and anti-hepatic fibrosis activities. However, the pharmacological effects of MT still need to be strengthened due to its relatively low efficacy and short half-life. In the present study, we report a more effective thio derivative of MT, MD-1, and its inhibitory effects on the activation of hepatic stellate cells (HSCs) in both cell culture and animal models. Cytological experiments showed that MD-1 can inhibit the proliferation of HSC-T6 cells with a half-maximal inhibitory concentration (IC50) of 62 μmol/L. In addition, MD-1 more strongly inhibits the migration of HSC-T6 cells compared to MT and can more effectively induce G0/G1 arrest and apoptosis. Investigating the biological mechanisms underlying anti-hepatic fibrosis in the presence of MD-1, we found that MD-1 can bind the epidermal growth factor receptor (EGFR) on the surface of HSC-T6 cells, which can further inhibit the phosphorylation of EGFR and its downstream protein kinase B (Akt), resulting in decreased expression of cyclin D1 and eventual inhibition of the activation of HSC-T6 cells. Furthermore, in rats with dimethylnitrosamine (DMN)-induced hepatic fibrosis, MD-1 slowed the development and progression of hepatic fibrosis, protecting hepatic parenchymal cells and improving hepatic functions. Therefore, MD-1 is a potential drug for anti-hepatic fibrosis.

  8. Regulation of EGF-induced ERK/MAPK Activation and EGFR Internalization by G Protein-coupled Receptor Kinase 2

    Institute of Scientific and Technical Information of China (English)

    Jingxia GAO; Jiali LI; Lan MA

    2005-01-01

    G protein-coupled receptor kinases (GRKs) mediate agonist-induced phosphorylation and desensitization of various G protein-coupled receptors (GPCRs). We investigate the role of GRK2 on epidermal growth factor (EGF) receptor signaling, including EGF-induced extracellular signal-regulated kinase and mitogen-activated protein kinase (ERK/MAPK) activation and EGFR internalization. Immunoprecipitation and immunofluorescence experiments show that EGF stimulates GRK2 binding to EGFR complex and GRK2 translocating from cytoplasm to the plasma membrane in human embryonic kidney 293 cells. Western blotting assay shows that EGF-induced ERK/MAPK phosphorylation increases 1.9-fold, 1.1-fold and 1.5-fold (P<0.05) at time point 30, 60 and 120 min, respectively when the cells were transfected with GRK2,suggesting the regulatory role of GRK2 on EGF-induced ERK/MAPK activation. Flow cytometry experiments show that GRK2 overexpression has no effect on EGF-induced EGFR internalization, however, it increases agonist-induced G protein-coupled δ opioid receptor internalization by approximately 40% (P<0.01). Overall,these data suggest that GRK2 has a regulatory role in EGF-induced ERK/MAPK activation, and that the mechanisms underlying the modulatory role of GRK2 in EGFR and GPCR signaling pathways are somewhat different at least in receptor internalization.

  9. Design, synthesis and molecular docking of α,β-unsaturated cyclohexanone analogous of curcumin as potent EGFR inhibitors with antiproliferative activity.

    Science.gov (United States)

    Xu, Yun-Yun; Cao, Yi; Ma, Hailkuo; Li, Huan-Qiu; Ao, Gui-Zhen

    2013-01-15

    A type of novel α,β-unsaturated cyclohexanone analogous, which designed based on the curcumin core structure, have been discovered as potential EGFR inhibitors. These compounds exhibit potent antiproliferative activity in two human tumor cell lines (Hep G2 and B16-F10). Among them, compounds I(3) and I(12) displayed the most potent EGFR inhibitory activity (IC(50) = 0.43 μM and 1.54 μM, respectively). Molecular docking of I(12) into EGFR TK active site was also performed. This inhibitor nicely fitting the active site might well explain its excellent inhibitory activity.

  10. 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

  11. In vivo and in vitro antitumor activity of oxaliplatin in combination with cetuximab in human colorectal tumor cell lines expressing different level of EGFR.

    Science.gov (United States)

    Balin-Gauthier, Diane; Delord, Jean-Pierre; Rochaix, Philippe; Mallard, Valérie; Thomas, Fabienne; Hennebelle, Isabelle; Bugat, Roland; Canal, Pierre; Allal, Cuider

    2006-06-01

    This study aimed to assess the effect of cetuximab (C225, Erbitux, a chimeric anti-epidermal growth factor receptor (EGFR) monoclonal antibody) in combination with oxaliplatin in vitro and in vivo on four colon cancer cell lines (HCT-8; HT-29, SW620, HCT-116) expressing different levels of EGFR. In vitro, cetuximab combined with oxaliplatin significantly decreased the IC50 values of oxaliplatin in HCT-8 (EGF-R moderate) and HT-29 (EGF-R weak) cell lines, while SW620 (EGF-R negative) and HCT-116 (EGFR strong) cell lines remained unresponsive. This combination was synergistic in HCT-8 and HT-29 cell lines while cetuximab induced no major modification of the IC50 of oxaliplatin in HCT-116 or SW620 cell lines. We then determined the effect of cetuximab on the EGF-induced EGFR phosphorylation and we highlight a correlation between the basal level of phospho-EGFR and the response to the combination. In vivo, the combination of cetuximab plus oxaliplatin significantly inhibited tumor growth of HCT-8 and HT-29 (tumor delay or Td = 21.6+/-2.9 and 18.0+/-2.9 days respectively, synergistic effect) compared to either oxaliplatin (Td=12.6+/-2.3 and 14.4+/-3.2 days respectively) or cetuximab (Td=13.4+/-2.9 and 14.5+/-2.4 days, respectively) alone in xenograft models. The combination had no effect on HCT-116 and SW-620 cell lines. The observed responses are strictly dependent on the cell type, and are not correlated with the level of EGFR expression but related to the basal level of phospho-EGFR. This study provides promising preclinical results for a possible clinical investigation of the combination of oxaliplatin plus cetuximab in chemorefractory colorectal tumors.

  12. EGF‑stimulated AKT activation is mediated by EGFR recycling via an early endocytic pathway in a gefitinib‑resistant human lung cancer cell line.

    Science.gov (United States)

    Nishimura, Yukio; Takiguchi, Soichi; Ito, Shigeru; Itoh, Kazuyuki

    2015-04-01

    The receptor tyrosine kinase epidermal growth factor receptor (EGFR) and its ligand epidermal growth factor (EGF) are known to play important roles in malignant tumor cells, and the EGFR signaling pathway is one of the most important targets in various tumors, including non-small cell lung cancer (NSCLC). We reported recently that an aberration in certain steps of EGF-stimulated phosphorylated epidermal growth factor receptor (pEGFR) endocytic trafficking from the early endosomes to the late endosomes occurs in the gefitinib-resistant NSCLC cells, in which large amounts of sorting nexin 1 (SNX1) are colocalized with EGFR in the aggregated early endosomes where the internalized pEGFR is also accumulated of these cells. To further investigate the role of SNX1 in EGF‑stimulated pEGFR endocytosis, followed by downstream signaling leading to the activation of phosphatidylinositol 3-kinase (PI3K)--the serine/threonine kinase AKT pathway, we examined the effect of depletion of SNX1 knock-down expression by siRNA and an inhibition of targeting membrane recycling using monensin. Using immunofluorescence, we observed an efficient endocytic transport of pEGFR from early endosomes to late endosomes/lysosomes after EGF-stimulation in the cells transfected with siRNA‑SNX1, whereas the delayed endocytic delivery of pEGFR was evident in the siRNA-control-transfected cells. Furthermore, a large amount of endocytosed pEGFR was accumulated in the presence of monensin in the early endosomes of the SNX1 knock-down cells. In western blot analysis, EGF stimulation of both control and cells transfected with siRNA-SNX1 resulted in rapid phosphorylation of EGFR and enhanced AKT phosphorylation. Monensin-dependent inhibition of AKT phosphorylation was stronger in SNX1 knock-down cells than in controls. In contrast, however, monensin had no effect on AKT phosphorylation triggered by activation of the MET receptor tyrosine kinase. Collectively, we suggest that EGF-stimulated recycling of

  13. Conversion from the "oncogene addiction" to "drug addiction" by intensive inhibition of the EGFR and MET in lung cancer with activating EGFR mutation.

    Science.gov (United States)

    Suda, Kenichi; Tomizawa, Kenji; Osada, Hirotaka; Maehara, Yoshihiko; Yatabe, Yasushi; Sekido, Yoshitaka; Mitsudomi, Tetsuya

    2012-06-01

    Emergence of acquired resistance is virtually inevitable in patients with a mutation in the epidermal growth factor receptor gene (EGFR) treated with EGFR tyrosine kinase inhibitors (TKIs). Several novel TKIs that may prevent or overcome the resistance mechanisms are now under clinical development. However, it is unknown how tumor cells will respond to intensive treatment using these novel TKIs. We previously established HCC827EPR cells, which are T790M positive, through combined treatment with erlotinib and a MET-TKI from erlotinib-hypersensitive HCC827 cells. In this study, we treated HCC827EPR cells sequentially with an irreversible EGFR-TKI, CL-387,785, to establish resistant cells (HCC827CLR), and we analyzed the mechanisms responsible for resistance. In HCC827CLR cells, PTEN expression was downregulated and Akt phosphorylation persisted in the presence of CL-387,785. Akt inhibition restored CL-387,785 sensitivity. In addition, withdrawal of CL-387,785 reduced cell viability in HCC827CLR cells, indicating that these cells were "addicted" to CL-387,785. HCC827CLR cells overexpressed the EGFR, and inhibition of the EGFR or MEK-ERK was needed to maintain cell proliferation. Increased senescence was observed in HCC827CLR cells in the drug-free condition. Through long-term culture of HCC827CLR cells without CL-387,785, we established HCC827-CL-387,785-independent cells, which exhibited decreased EGFR expression and a mesenchymal phenotype. In conclusion, PTEN downregulation is a newly identified mechanism underlying the acquired resistance to irreversible EGFR-TKIs after acquisition of T790M against erlotinib. This series of experiments highlights the flexibility of cancer cells that have adapted to environmental stresses induced by intensive treatment with TKIs. PMID:22133747

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

    Science.gov (United States)

    Jackson, Nicole M; Ceresa, Brian P

    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. PMID:27381222

  15. The use of EGFR exon 19 and 21 unlabeled DNA probes to screen for activating mutations in non-small cell lung cancer.

    Science.gov (United States)

    Willmore-Payne, Carlynn; Holden, Joseph A; Wittwer, Carl T; Layfield, Lester J

    2008-07-01

    Activating mutations in epidermal growth factor receptor-1 (EGFR) are found in 10-15% of Caucasian patients with non-small cell lung carcinoma (NSCLC). Approximately 90% of the mutations are deletions of several amino acids in exon 19 or point mutations in exon 21. Some studies suggest that these mutations identify patients that might benefit from targeted EGFR inhibitor therapy. DNA melting analysis of polymerase chain reaction products can screen for these mutations to identify this patient population. However, amplicon DNA melting analysis, although easily capable of detecting heterozygous mutations by heterodimer formation, becomes more difficult if mutations are homozygous or if the mutant allele is selectively amplified over wild type. Amplification of EGFR is common in NSCLC and this could compromise mutation detection by amplicon melting analysis. To overcome this potential limitation, we developed unlabeled, single-stranded DNA probes, complimentary to EGFR exon 19 and exon 21 where the common activating mutations occur. The unlabeled probes are incorporated into a standard polymerase chain reaction during the amplification of EGFR exons 19 and 21. The probe melting peak is easily distinguished from the amplicon melting peak, and probe melting is altered if mutations are present. This allows for easy identification of activating mutations even in homozygous or amplified states and is useful in the screening of NSCLC for the common EGFR activating mutations. PMID:19137110

  16. Docetaxel for non small cell lung cancer harboring the activated EGFR mutation with T790M at initial presentation

    Directory of Open Access Journals (Sweden)

    Yamane H

    2013-03-01

    Full Text Available Hiromichi Yamane,1 Nobuaki Ochi,1 Masayuki Yasugi,2 Takayuki Tabayashi,1 Tomoko Yamagishi,1 Yasumasa Monobe,3 Akiko Hisamoto,4 Katsuyuki Kiura,4 Nagio Takigawa1 1Department of General Internal Medicine 4, Kawasaki Medical School, Okayama, Japan; 2Department of Respiratory Medicine, National Hospital Organization Fukuyama Medical Center, Fukuyama, Japan; 3Department of Pathology, Kawasaki Medical School Kawasaki Hospital, Okayama, Japan; 4Department of Hematology, Oncology, and Respiratory Medicine, Okayama University Graduate School of Medicine, Okayama, Japan Abstract: A 72-year-old woman was referred to our hospital with Stage IV non-small-cell lung cancer (NSCLC. Chest computed tomography revealed a mass in the upper lobe of the right lung, with pleural effusion. Cytologic examination identified adenocarcinoma cells in the right pleural effusion. Furthermore, both a deletion mutation in exon 19 and a threonine–methionine substitution mutation at position 790 in exon 20 (T790M were detected in the epidermal growth factor receptors (EGFR in the malignant cells. As systemic chemotherapy consisting of carboplatin and pemetrexed or erlotinib proved ineffective, docetaxel monotherapy was initiated as a third-line treatment. Following salvage chemotherapy, her Eastern Cooperative Oncology Group performance status improved from 3 to 1, with tumor regression over 5 months. To the best of our knowledge, this is the first report of successful docetaxel treatment for a patient with NSCLC harboring the T790M EGFR-activating mutation identified before treatment with EGFR tyrosine kinase inhibitors. Keywords: non-small-cell lung cancer, EGFR mutation, pretreatment mutation, T790M, docetaxel

  17. Up-regulation of fatty acid synthase induced by EGFR/ERK activation promotes tumor growth in pancreatic cancer

    Energy Technology Data Exchange (ETDEWEB)

    Bian, Yong, E-mail: drbiany@126.com [Department of Science and Technology, Nanjing University of Chinese Medicine, 210023 (China); Yu, Yun [College of Pharmacy, Nanjing University of Chinese Medicine, 210023 (China); Wang, Shanshan; Li, Lin [Department of Science and Technology, Nanjing University of Chinese Medicine, 210023 (China)

    2015-08-07

    Lipid metabolism is dysregulated in many human diseases including atherosclerosis, type 2 diabetes and cancers. Fatty acid synthase (FASN), a key lipogenic enzyme involved in de novo lipid biosynthesis, is significantly upregulated in multiple types of human cancers and associates with tumor progression. However, limited data is available to understand underlying biological functions and clinical significance of overexpressed FASN in pancreatic ductal adenocarcinoma (PDAC). Here, upregulated FASN was more frequently observed in PDAC tissues compared with normal pancreas in a tissue microarray. Kaplan–Meier survival analysis revealed that high expression level of FASN resulted in a significantly poor prognosis of PDAC patients. Knockdown or inhibition of endogenous FASN decreased cell proliferation and increased cell apoptosis in HPAC and AsPC-1 cells. Furthermore, we demonstrated that EGFR/ERK signaling accounts for elevated FASN expression in PDAC as ascertained by performing siRNA assays and using specific pharmacological inhibitors. Collectively, our results indicate that FASN exhibits important roles in tumor growth and EGFR/ERK pathway is responsible for upregulated expression of FASN in PDAC. - Highlights: • Increased expression of FASN indicates a poor prognosis in PDAC. • Elevated FASN favors tumor growth in PDAC in vitro. • Activation of EGFR signaling contributes to elevated FASN expression.

  18. Up-regulation of fatty acid synthase induced by EGFR/ERK activation promotes tumor growth in pancreatic cancer

    International Nuclear Information System (INIS)

    Lipid metabolism is dysregulated in many human diseases including atherosclerosis, type 2 diabetes and cancers. Fatty acid synthase (FASN), a key lipogenic enzyme involved in de novo lipid biosynthesis, is significantly upregulated in multiple types of human cancers and associates with tumor progression. However, limited data is available to understand underlying biological functions and clinical significance of overexpressed FASN in pancreatic ductal adenocarcinoma (PDAC). Here, upregulated FASN was more frequently observed in PDAC tissues compared with normal pancreas in a tissue microarray. Kaplan–Meier survival analysis revealed that high expression level of FASN resulted in a significantly poor prognosis of PDAC patients. Knockdown or inhibition of endogenous FASN decreased cell proliferation and increased cell apoptosis in HPAC and AsPC-1 cells. Furthermore, we demonstrated that EGFR/ERK signaling accounts for elevated FASN expression in PDAC as ascertained by performing siRNA assays and using specific pharmacological inhibitors. Collectively, our results indicate that FASN exhibits important roles in tumor growth and EGFR/ERK pathway is responsible for upregulated expression of FASN in PDAC. - Highlights: • Increased expression of FASN indicates a poor prognosis in PDAC. • Elevated FASN favors tumor growth in PDAC in vitro. • Activation of EGFR signaling contributes to elevated FASN expression

  19. The Use of EGFR Exon 19 and 21 Unlabeled DNA Probes to Screen for Activating Mutations in Non–Small Cell Lung Cancer

    OpenAIRE

    Willmore-Payne, Carlynn; Holden, Joseph A.; Wittwer, Carl T.; Layfield, Lester J.

    2008-01-01

    Activating mutations in epidermal growth factor receptor-1 (EGFR) are found in 10–15% of Caucasian patients with non–small cell lung carcinoma (NSCLC). Approximately 90% of the mutations are deletions of several amino acids in exon 19 or point mutations in exon 21. Some studies suggest that these mutations identify patients that might benefit from targeted EGFR inhibitor therapy. DNA melting analysis of polymerase chain reaction products can screen for these mutations to identify this patient...

  20. Akt kinase-interacting protein1, a novel therapeutic target for lung cancer with EGFR-activating and gatekeeper mutations

    OpenAIRE

    Yamada, Tadaaki; Takeuchi, Shinji; Fujita, Naoya; Nakamura, Akito; Wang, Wei; Li, Qi; Oda, Makoto; Mitsudomi, Tetsuya; Yatabe, Yasushi; Sekido, Yoshitaka; Yoshida, Junji; Higashiyama, Masahiko; Noguchi, Masayuki; Uehara, Hisanori; Nishioka, Yasuhiko

    2013-01-01

    Despite initial dramatic response, epidermal growth factor receptor (EGFR) mutant lung cancer patients always acquire resistance to EGFR-tyrosine kinase inhibitors (TKIs). Gatekeeper T790M mutation in EGFR is the most prevalent genetic alteration underlying acquired resistance to EGFR-TKI, and EGFR mutant lung cancer cells are reported to be addictive to EGFR/Akt signaling even after acquired T790M mutation. Here, we focused on Akt kinase-interacting protein1 (Aki1), a scaffold protein of PI3...

  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 fractio...

  2. Lapatinib, a dual EGFR and HER2 tyrosine kinase inhibitor, downregulates thymidylate synthase by inhibiting the nuclear translocation of EGFR and HER2.

    Directory of Open Access Journals (Sweden)

    Hwang-Phill Kim

    Full Text Available BACKGROUND: Epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI has been shown to exert a synergistic antitumor effect when combined with fluoropyrimidine. This synergy may be attributable to the downregulation of thymidylate synthase (TS, which is frequently overexpressed in fluoropyrimidine-resistant cancer cells. However, the molecular mechanism underlying the downregulation of TS has yet to be clearly elucidated. METHODOLOGY AND PRINCIPAL FINDINGS: In this study, we demonstrate that lapatinib, a dual TKI of EGFR and HER2 downregulates TS via inhibition of the nuclear translocation of EGFR and HER2. From our cDNA microarray experiments, we determined that a variety of nucleotide synthesis-related genes, including TS, were downregulated with lapatinib, and this was apparent in HER2-amplified cells. Targeted and pharmacologic inhibition assays confirmed that the dual inhibition of EGFR and HER2 is required for the more effective reduction of TS as compared to what was observed with gefitinib or trasutuzumab alone. Additionally, we determined that co-transfected EGFR and HER2 activate the TS gene promoter more profoundly than do either EGFR or HER2 alone. The translocation of EGFR and HER2 into the nucleus and the subsequent activation of the TS promoter were inhibited by lapatinib. CONCLUSIONS AND SIGNIFICANCE: These results demonstrate that lapatinib inhibits the nuclear translocation of EGFR and HER2 and downregulates TS, thus sensitizing cancer cells to fluoropyrimidine.

  3. EGFR trans-activation by urotensin II receptor is mediated by β-arrestin recruitment and confers cardioprotection in pressure overload-induced cardiac hypertrophy.

    Science.gov (United States)

    Esposito, Giovanni; Perrino, Cinzia; Cannavo, Alessandro; Schiattarella, Gabriele G; Borgia, Francesco; Sannino, Anna; Pironti, Gianluigi; Gargiulo, Giuseppe; Di Serafino, Luigi; Franzone, Anna; Scudiero, Laura; Grieco, Paolo; Indolfi, Ciro; Chiariello, Massimo

    2011-06-01

    Urotensin II (UTII) and its seven trans-membrane receptor (UTR) are up-regulated in the heart under pathological conditions. Previous in vitro studies have shown that UTII trans-activates the epidermal growth factor receptor (EGFR), however, the role of such novel signalling pathway stimulated by UTII is currently unknown. In this study, we hypothesized that EGFR trans-activation by UTII might exert a protective effect in the overloaded heart. To test this hypothesis, we induced cardiac hypertrophy by transverse aortic constriction (TAC) in wild-type mice, and tested the effects of the UTII antagonist Urantide (UR) on cardiac function, structure, and EGFR trans-activation. After 7 days of pressure overload, UR treatment induced a rapid and significant impairment of cardiac function compared to vehicle. In UR-treated TAC mice, cardiac dysfunction was associated with reduced phosphorylation levels of the EGFR and extracellular-regulated kinase (ERK), increased apoptotic cell death and fibrosis. In vitro UTR stimulation induced membrane translocation of β-arrestin 1/2, EGFR phosphorylation/internalization, and ERK activation in HEK293 cells. Furthermore, UTII administration lowered apoptotic cell death induced by serum deprivation, as shown by reduced TUNEL/Annexin V staining and caspase 3 activation. Interestingly, UTII-mediated EGFR trans-activation could be prevented by UR treatment or knockdown of β-arrestin 1/2. Our data show, for the first time in vivo, a new UTR signalling pathway which is mediated by EGFR trans-activation, dependent by β-arrestin 1/2, promoting cell survival and cardioprotection.

  4. 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.

  5. CRIPTO1 expression in EGFR-mutant NSCLC elicits intrinsic EGFR-inhibitor resistance.

    Science.gov (United States)

    Park, Kang-Seo; Raffeld, Mark; Moon, Yong Wha; Xi, Liqiang; Bianco, Caterina; Pham, Trung; Lee, Liam C; Mitsudomi, Tetsuya; Yatabe, Yasushi; Okamoto, Isamu; Subramaniam, Deepa; Mok, Tony; Rosell, Rafael; Luo, Ji; Salomon, David S; Wang, Yisong; Giaccone, Giuseppe

    2014-07-01

    The majority of non-small cell lung cancer (NSCLC) patients harbor EGFR-activating mutations that can be therapeutically targeted by EGFR tyrosine kinase inhibitors (EGFR-TKI), such as erlotinib and gefitinib. Unfortunately, a subset of patients with EGFR mutations are refractory to EGFR-TKIs. Resistance to EGFR inhibitors reportedly involves SRC activation and induction of epithelial-to-mesenchymal transition (EMT). Here, we have demonstrated that overexpression of CRIPTO1, an EGF-CFC protein family member, renders EGFR-TKI-sensitive and EGFR-mutated NSCLC cells resistant to erlotinib in culture and in murine xenograft models. Furthermore, tumors from NSCLC patients with EGFR-activating mutations that were intrinsically resistant to EGFR-TKIs expressed higher levels of CRIPTO1 compared with tumors from patients that were sensitive to EGFR-TKIs. Primary NSCLC cells derived from a patient with EGFR-mutated NSCLC that was intrinsically erlotinib resistant were CRIPTO1 positive, but gained erlotinib sensitivity upon loss of CRIPTO1 expression during culture. CRIPTO1 activated SRC and ZEB1 to promote EMT via microRNA-205 (miR-205) downregulation. While miR-205 depletion induced erlotinib resistance, miR-205 overexpression inhibited CRIPTO1-dependent ZEB1 and SRC activation, restoring erlotinib sensitivity. CRIPTO1-induced erlotinib resistance was directly mediated through SRC but not ZEB1; therefore, cotargeting EGFR and SRC synergistically attenuated growth of erlotinib-resistant, CRIPTO1-positive, EGFR-mutated NSCLC cells in vitro and in vivo, suggesting that this combination may overcome intrinsic EGFR-inhibitor resistance in patients with CRIPTO1-positive, EGFR-mutated NSCLC. PMID:24911146

  6. ADAM17-siRNA inhibits MCF-7 breast cancer through EGFR-PI3K-AKT activation.

    Science.gov (United States)

    Meng, Xiangchao; Hu, Baoshan; Hossain, Mohammad Monir; Chen, Guofu; Sun, Ying; Zhang, Xuepeng

    2016-08-01

    A disintegrin and metalloproteinase-17 (ADAM17) can cut and release a wide variety of epidermal growth factor receptor (EGFR) ligands to promote survival, invasion and proliferation of cancer cell, and therefore, is considered to be a potential therapeutic target for cancer. The main goal of the present study was to observe the effects of ADAM17 small interfering RNA (ADAM17-siRNA) on human MCF-7 breast cancer and investigate its activation pathway. In vitro, MCF-7 cells were divided into ADAM17-siRNA groups, nonsense siRNA groups, AG1478 (selective EGFR blocker) groups, LY294002 [phosphatidylinositol 3-kinase (PI3K) phosphorylation inhibitor] groups, PD0325901 [mitogen extracellular kinase (MEK) inhibitor] groups and control groups. In vivo, MCF-7 cells were implanted subcutaneously into nude mice and then these mice were randomly divided into ADAM17-siRNA groups, vector groups and control groups. Our data showed that compared with the control groups, ADAM17-siRNA, AG1478 and LY294002 could inhibit the migration and proliferation of MCF-7 cells, but PD0325901 and nonsense siRNA did not show this effect. Except that specific ADAM17-siRNA could inhibit the expression of ADAM17 mRNA, others did not change it. Western blot analysis further confirmed that EGFR-PI3K-AKT signaling pathway is involved in ADAM17-siRNA inhibiting migration and proliferation of MCF-7 cells. Similarly to the former, the growth of MCF-7 breast cancer in nude mice was significantly inhibited by ADAM17-siRNA. Compared with the control group and the vector group, the tumor volume was smaller in the ADAM17-siRNA group, the tissues developed large areas of necrosis, immunohistochemistry showed low expressions of ADAM17 and Ki-67 and western blot analysis proved that the expression of ADAM17 protein in the tissue was also reduced. The present study suggests that ADAM17-siRNA inhibits MCF-7 breast cancer and is activated through the EGFR-PI3K-AKT signaling pathway. PMID:27221510

  7. Nuclear epidermal growth factor receptor (EGFR) interacts with signal transducer and activator of transcription 5 (STAT5) in activating Aurora-A gene expression

    OpenAIRE

    Hung, Liang-Yi; Tseng, Joseph T.; Lee, Yi-Chao; Xia, Weiya; Wang, Ying-Nai; Wu, Min-Li; Chuang, Yu-Hsuan; Lai, Chein-Hsien; Chang, Wen-Chang

    2008-01-01

    Loss of the maintenance of genetic material is a critical step leading to tumorigenesis. It was reported that overexpression of Aurora-A and the constitutive activation of the epidermal growth factor (EGF) receptor (EGFR) are implicated in chromosome instability. In this study, we examined that when cells treated with EGF result in centrosome amplification and microtubule disorder, which are critical for genetic instability. Interestingly, the expression of Aurora-A was also increased by EGF ...

  8. The scaffolding protein NHERF1 sensitizes EGFR-dependent tumor growth, motility and invadopodia function to gefitinib treatment in breast cancer cells.

    Science.gov (United States)

    Bellizzi, Antonia; Greco, Maria Raffaella; Rubino, Rosa; Paradiso, Angelo; Forciniti, Stefania; Zeeberg, Katrine; Cardone, Rosa Angela; Reshkin, Stephan Joel

    2015-03-01

    Triple negative breast cancer (TNBC) patients cannot be treated with endocrine therapy or targeted therapies due to lack of related receptors. These patients overexpress the epidermal growth factor receptor (EGFR), but are resistant to tyrosine kinase inhibitors (TKIs) and anti-EGFR therapies. Mechanisms suggested for resistance to TKIs include EGFR independence, mutations and alterations in EGFR and in its downstream signalling pathways. Ligand-induced endocytosis and degradation of EGFR play important roles in the downregulation of the EGFR signal suggesting that its activity could be regulated by targeting its trafficking. Evidence in normal cells showing that the scaffolding protein Na+/H+ exchanger regulatory factor 1 (NHERF1) can associate with EGFR to regulate its trafficking, led us to hypothesize that NHERF1 expression levels could regulate EGFR trafficking and functional expression in TNBC cells and, in this way, modulate its role in progression and response to treatment. We investigated the subcellular localization of NHERF1 and its interaction with EGFR in a metastatic basal like TNBC cell model, MDA-MB‑231, and the role of forced NHERF1 overexpression and/or stimulation with EGF on the sensitivity to EGFR specific TKI treatment with gefitinib. Stimulation with EGF induces an interaction of NHERF1 with EGFR to regulate its localization, degradation and function. NHERF1 overexpression is sufficient to drive its interaction with EGFR in non-stimulated conditions, inhibits EGFR degradation and increases its retention time in the plasma membrane. Importantly, NHERF1 overexpression strongly sensitized the cell to the pharmacological inhibition by gefitinib of EGFR-driven growth, motility and invadopodia-dependent ECM proteolysis. The further determination of how the NHERF1‑EGFR interaction is regulated may improve our understanding of TNBC resistance to the action of existing anticancer drugs.

  9. TGF-β and Hypoxia/Reoxygenation Promote Radioresistance of A549 Lung Cancer Cells through Activation of Nrf2 and EGFR

    Directory of Open Access Journals (Sweden)

    Sae-lo-oom Lee

    2016-01-01

    Full Text Available Although many studies have examined the roles of hypoxia and transforming growth factor- (TGF- β separately in the tumor microenvironment, the effects of simultaneous treatment with hypoxia/reoxygenation and TGF-β on tumor malignancy are unclear. Here, we investigated the effects of redox signaling and oncogenes on cell proliferation and radioresistance in A549 human lung cancer cells in the presence of TGF-β under hypoxia/reoxygenation conditions. Combined treatment with TGF-β and hypoxia activated epidermal growth factor receptor (EGFR and nuclear factor (erythroid-derived 2-like 2 (Nrf2, a redox-sensitive transcription factor. Interestingly, Nrf2 knockdown suppressed the effects of combined treatment on EGFR phosphorylation. In addition, blockade of EGFR signaling also suppressed induction of Nrf2 following combined treatment with hypoxia and TGF-β, indicating that the combined treatment induced positive crosstalk between Nrf2 and EGFR. TGF-β and hypoxia/reoxygenation increased the accumulation of reactive oxygen species (ROS, while treatment with N-acetyl-L-cysteine abolished the activation of Nrf2 and EGFR. Treatment with TGF-β under hypoxic conditions increased the proliferation of A549 cells compared with that after vehicle treatment. Moreover, cells treated with the combined treatment exhibited resistance to ionizing radiation (IR, and knockdown of Nrf2 increased IR-induced cell death under these conditions. Thus, taken together, our findings suggested that TGF-β and hypoxia/reoxygenation promoted tumor progression and radioresistance of A549 cells through ROS-mediated activation of Nrf2 and EGFR.

  10. EGFR mediates astragaloside IV-induced Nrf2 activation to protect cortical neurons against in vitro ischemia/reperfusion damages

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Da-min [Department of Anesthesiology, Affiliated Yixing People' s Hospital, Jiangsu University, Yixing (China); Lu, Pei-Hua, E-mail: lphty1_1@163.com [Department of Medical Oncology, Wuxi People' s Hospital Affiliated to Nanjing Medical University, Wuxi (China); Zhang, Ke; Wang, Xiang [Department of Anesthesiology, Affiliated Yixing People' s Hospital, Jiangsu University, Yixing (China); Sun, Min [Department of General Surgery, Affiliated Yixing People' s Hospital, Jiangsu University, Yixing (China); Chen, Guo-Qian [Department of Clinical Laboratory, Wuxi People' s Hospital Affiliated to Nanjing Medical University, Wuxi (China); Wang, Qiong, E-mail: WangQiongprof1@126.com [Department of Clinical Laboratory, Wuxi People' s Hospital Affiliated to Nanjing Medical University, Wuxi (China)

    2015-02-13

    In this study, we tested the potential role of astragaloside IV (AS-IV) against oxygen and glucose deprivation/re-oxygenation (OGD/R)-induced damages in murine cortical neurons, and studied the associated signaling mechanisms. AS-IV exerted significant neuroprotective effects against OGD/R by reducing reactive oxygen species (ROS) accumulation, thereby attenuating oxidative stress and neuronal cell death. We found that AS-IV treatment in cortical neurons resulted in NF-E2-related factor 2 (Nrf2) signaling activation, evidenced by Nrf2 Ser-40 phosphorylation, and its nuclear localization, as well as transcription of antioxidant-responsive element (ARE)-regulated genes: heme oxygenase-1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO-1) and sulphiredoxin 1 (SRXN-1). Knockdown of Nrf2 through lentiviral shRNAs prevented AS-IV-induced ARE genes transcription, and abolished its anti-oxidant and neuroprotective activities. Further, we discovered that AS-IV stimulated heparin-binding-epidermal growth factor (HB-EGF) release to trans-activate epidermal growth factor receptor (EGFR) in cortical neurons. Blockage or silencing EGFR prevented Nrf2 activation by AS-IV, thus inhibiting AS-IV-mediated anti-oxidant and neuroprotective activities against OGD/R. In summary, AS-IV protects cortical neurons against OGD/R damages through activating of EGFR-Nrf2 signaling. - Highlights: • Pre-treatment of astragaloside IV (AS-IV) protects murine cortical neurons from OGD/R. • AS-IV activates Nrf2-ARE signaling in murine cortical neurons. • Nrf2 is required for AS-IV-mediated anti-oxidant and neuroprotective activities. • AS-IV stimulates HB-EGF release to trans-activate EGFR in murine cortical neurons. • EGFR mediates AS-IV-induced Nrf2 activation and neuroprotection against OGD/R.

  11. Oligomerization of epidermal growth factor receptors (EGFR) on A431 cells studied by time-resolved fluorescence imaging microscopy: a stereochemical model for tyrosine kinase receptor activation

    NARCIS (Netherlands)

    Th.W.J. Gadella; T.M. Jovin

    1995-01-01

    The aggregation states of the epidermal growth factor receptor (EGFR) on single A431 human epidermoid carcinoma cells were assessed with two new techniques for determining fluorescence resonance en- ergy transfer: donor photobleaching fluorescence reso- nance energy transfer (pbFRET) microscopy and

  12. CXCL12/CXCR4 Axis Activation Mediates Prostate Myofibroblast Phenoconversion through Non-Canonical EGFR/MEK/ERK Signaling

    Science.gov (United States)

    Rodríguez-Nieves, José A.; Patalano, Susan C.; Almanza, Diego; Gharaee-Kermani, Mehrnaz; Macoska, Jill A.

    2016-01-01

    Benign prostate hyperplasia (BPH), an enlargement of the prostate common in aging in men, is associated with urinary voiding dysfunction manifest as Lower Urinary Tract Symptoms (LUTS). Although inflammation and abnormal smooth muscle contractions are known to play key roles in the development of LUTS, tissue fibrosis may also be an important and previously unrecognized contributing factor. Tissue fibrosis arises from the unregulated differentiation of fibroblasts or other precursor cell types into myofibroblasts, which is usually accomplished by activation of the TGFβ/TGFβR axis. Previously we reported that the CXC-type chemokines, CXCL5, CXCL8 and CXCL12, which are up-regulated in the aging in the prostate, can drive this differentiation process as well in the absence of TGFβ. Based on this data we sought to elucidate the molecular mechanisms employed by CXCL12, and its receptor CXCR4, during prostate myofibroblast phenoconversion. The results of these studies suggest that CXCL12/CXCR4-mediated signaling events in prostate myofibroblast phenoconversion may proceed through non-canonical pathways that do not depend on TGFβ/TGFβR axis activation or Smad signaling. Here we report that CXCL12/CXCR4 axis activation promotes signaling through the EGFR and downstream MEK/ERK and PI3K/Akt pathways during myofibroblast phenoconversion, but not through TGFβ/TGFβR and downstream Smad signaling, in prostate fibroblasts undergoing myofibroblast phenoconversion. We document that EGFR transactivation is required for CXCL12-mediated signaling and expression of genes associate with myofibroblast phenoconversion (α-SMA, COL1a1). Our study successfully identified TGFβ/TGFβR-independent molecular mechanisms that promote CXCL12/CXCR4-induced myofibroblast phenoconversion. This information may be crucial for the development of novel therapies and potential biomarkers for prostatic fibrosis. PMID:27434301

  13. Markers for EGFR pathway activation as predictor of outcome in metastatic colorectal cancer patients treated with or without cetuximab.

    NARCIS (Netherlands)

    Tol, J.; Dijkstra, J.R.; Klomp, M.; Teerenstra, S.; Dommerholt, M.; Vink-Borger, M.E.; Cleef, P.H. van; Krieken, J.H.J.M. van; Punt, C.J.A.; Nagtegaal, I.D.

    2010-01-01

    BACKGROUND: Anti-EGFR monoclonal antibodies in metastatic colorectal cancer (mCRC) treatment are only effective in patients with KRAS wild type tumours. Here we assess the predictive value of other potential relevant markers involved in the epidermal growth factor receptor (EGFR) signalling pathways

  14. T11TS inhibits Angiopoietin-1/Tie-2 signaling, EGFR activation and Raf/MEK/ERK pathway in brain endothelial cells restraining angiogenesis in glioma model.

    Science.gov (United States)

    Bhattacharya, Debanjan; Chaudhuri, Suhnrita; Singh, Manoj Kumar; Chaudhuri, Swapna

    2015-06-01

    Malignant gliomas represent one of the most aggressive and hypervascular primary brain tumors. Angiopoietin-1, the peptide growth factor activates endothelial Tie-2 receptor promoting vessel maturation and vascular stabilization steps of angiogenesis in glioma. Epidermal growth factor receptor (EGFR) and Tie-2 receptor on endothelial cells once activated transmits signals through downstream Raf/MEK/ERK pathway promoting endothelial cell proliferation and migration which are essential for angiogenesis induction. The in vivo effect of sheep erythrocyte membrane glycopeptide T11-target structure (T11TS) on angiopoietin-1/Tie-2 axis, EGFR signaling and Raf/MEK/ERK pathway in glioma associated endothelial cells has not been investigated previously. The present study performed with rodent glioma model aims to investigate the effect of T11TS treatment on angiopoietin-1/Tie-2 signaling, EGFR activity and Raf/MEK/ERK pathway in glioma associated endothelial cells within glioma milieu. T11TS administration in rodent glioma model inhibited angiopoietin-1 expression and attenuated Tie-2 expression and activation in glioma associated brain endothelial cells. T11TS treatment also downregulated total and phosphorylated EGFR expression in glioma associated endothelial cells. Additionally T11TS treatment inhibited Raf-1 expression, MEK-1 and ERK-1/2 expression and phosphorylation in glioma associated brain endothelial cells. Thus T11TS therapy remarkably inhibits endothelial angiopoietin-1/Tie-2 signaling associated with vessel maturation and simultaneously antagonizes endothelial cell proliferation signaling by blocking EGFR activation and components of Raf/MEK/ERK pathway. Collectively, the findings demonstrate a multi-targeted anti-angiogenic activity of T11TS which augments the potential for clinical translation of T11TS as an effective angiogenesis inhibitor for glioma treatment.

  15. EBP50 inhibits EGF-induced breast cancer cell proliferation by blocking EGFR phosphorylation.

    Science.gov (United States)

    Yao, Wenfang; Feng, Duiping; Bian, Weihua; Yang, Longyan; Li, Yang; Yang, Zhiyu; Xiong, Ying; Zheng, Junfang; Zhai, Renyou; He, Junqi

    2012-11-01

    Ezrin-radixin-moesin-binding phosphoprotein-50 (EBP50) suppresses breast cancer cell proliferation, potentially through its regulatory effect on epidermal growth factor receptor (EGFR) signaling, although the mechanism by which this occurs remains unknown. Thus in our studies, we aimed to determine the effect of EBP50 expression on EGF-induced cell proliferation and activation of EGFR signaling in the breast cancer cell lines, MDA-MB-231 and MCF-7. In MDA-MB-231 cells, which express low levels of EBP50, EBP50 overexpression inhibited EGF-induced cell proliferation, ERK1/2 and AKT phosphorylation. In MCF-7 cells, which express high levels of EBP50, EBP50 knockdown promoted EGF-induced cell proliferation, ERK1/2 and AKT phosphorylation. Knockdown of EBP50 in EBP50-overexpressed MDA-MB-231 cells abrogated the inhibitory effect of EBP50 on EGF-stimulated ERK1/2 phosphorylation and restoration of EBP50 expression in EBP50-knockdown MCF-7 cells rescued the inhibition of EBP50 on EGF-stimulated ERK1/2 phosphorylation, further confirming that the activation of EGF-induced downstream molecules could be specifically inhibited by EBP50 expression. Since EGFR signaling was triggered by EGF ligands via EGFR phosphorylation, we further detected the phosphorylation status of EGFR in the presence or absence of EBP50 expression. Overexpression of EBP50 in MDA-MB-231 cells inhibited EGF-stimulated EGFR phosphorylation, whereas knockdown of EBP50 in MCF-7 cells enhanced EGF-stimulated EGFR phosphorylation. Meanwhile, total expression levels of EGFR were unaffected during EGF stimulation. Taken together, our data shows that EBP50 can suppress EGF-induced proliferation of breast cancer cells by inhibiting EGFR phosphorylation and blocking EGFR downstream signaling in breast cancer cells. These results provide further insight into the molecular mechanism by which EBP50 regulates the development and progression of breast cancer.

  16. 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...... that EphA2 expression in in vitro cultured cells is restricted to cells growing adherently. EphA2 and EGFR are both cell surface trans-membrane proteins and we have found that the two receptors co-localize on human cancer cells. EGFR and EphA2 co-immunoprecipitates and their interactions affects receptor...

  17. Targeting SHP2 for EGFR inhibitor resistant non-small cell lung carcinoma

    International Nuclear Information System (INIS)

    Highlights: •SHP2 is required for EGFR inhibitor resistant NSCLC H1975 cell proliferation. •SHP2 inhibitor blocks EGF-stimulated ERK1/2 activation and proliferation. •SHP2 inhibitor exhibits marked anti-tumor activity in H1975 xenograft mice. •SHP2 inhibitor synergizes with PI3K inhibitor in suppressing cell growth. •Targeting SHP2 represents a novel strategy for EGFR inhibitor resistant NSCLCs. -- Abstract: Targeted therapy with inhibitors of epidermal growth factor receptor (EGFR) has produced a noticeable benefit to non-small cell lung cancer (NSCLC) patients whose tumors carry activating mutations (e.g. L858R) in EGFR. Unfortunately, these patients develop drug resistance after treatment, due to acquired secondary gatekeeper mutations in EGFR (e.g. T790M). Given the critical role of SHP2 in growth factor receptor signaling, we sought to determine whether targeting SHP2 could have therapeutic value for EGFR inhibitor resistant NSCLC. We show that SHP2 is required for EGF-stimulated ERK1/2 phosphorylation and proliferation in EGFR inhibitor resistant NSCLC cell line H1975, which harbors the EGFR T790M/L858R double-mutant. We demonstrate that treatment of H1975 cells with II-B08, a specific SHP2 inhibitor, phenocopies the observed growth inhibition and reduced ERK1/2 activation seen in cells treated with SHP2 siRNA. Importantly, we also find that II-B08 exhibits marked anti-tumor activity in H1975 xenograft mice. Finally, we observe that combined inhibition of SHP2 and PI3K impairs both the ERK1/2 and PI3K/AKT signaling axes and produces significantly greater effects on repressing H1975 cell growth than inhibition of either protein individually. Collectively, these results suggest that targeting SHP2 may represent an effective strategy for treatment of EGFR inhibitor resistant NSCLCs

  18. Targeting SHP2 for EGFR inhibitor resistant non-small cell lung carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Jie; Zeng, Li-Fan; Shen, Weihua [Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (United States); Turchi, John J. [Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (United States); Department of Medicine, Indiana University School of Medicine, Indianapolis (United States); Zhang, Zhong-Yin, E-mail: zyzhang@iu.edu [Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (United States)

    2013-10-04

    Highlights: •SHP2 is required for EGFR inhibitor resistant NSCLC H1975 cell proliferation. •SHP2 inhibitor blocks EGF-stimulated ERK1/2 activation and proliferation. •SHP2 inhibitor exhibits marked anti-tumor activity in H1975 xenograft mice. •SHP2 inhibitor synergizes with PI3K inhibitor in suppressing cell growth. •Targeting SHP2 represents a novel strategy for EGFR inhibitor resistant NSCLCs. -- Abstract: Targeted therapy with inhibitors of epidermal growth factor receptor (EGFR) has produced a noticeable benefit to non-small cell lung cancer (NSCLC) patients whose tumors carry activating mutations (e.g. L858R) in EGFR. Unfortunately, these patients develop drug resistance after treatment, due to acquired secondary gatekeeper mutations in EGFR (e.g. T790M). Given the critical role of SHP2 in growth factor receptor signaling, we sought to determine whether targeting SHP2 could have therapeutic value for EGFR inhibitor resistant NSCLC. We show that SHP2 is required for EGF-stimulated ERK1/2 phosphorylation and proliferation in EGFR inhibitor resistant NSCLC cell line H1975, which harbors the EGFR T790M/L858R double-mutant. We demonstrate that treatment of H1975 cells with II-B08, a specific SHP2 inhibitor, phenocopies the observed growth inhibition and reduced ERK1/2 activation seen in cells treated with SHP2 siRNA. Importantly, we also find that II-B08 exhibits marked anti-tumor activity in H1975 xenograft mice. Finally, we observe that combined inhibition of SHP2 and PI3K impairs both the ERK1/2 and PI3K/AKT signaling axes and produces significantly greater effects on repressing H1975 cell growth than inhibition of either protein individually. Collectively, these results suggest that targeting SHP2 may represent an effective strategy for treatment of EGFR inhibitor resistant NSCLCs.

  19. Synergistic antitumor effect between gefitinib and fractionated irradiation in anaplastic oligodendrogliomas cannot be predicted by the Egfr signaling activity.

    Directory of Open Access Journals (Sweden)

    Sophie Pinel

    Full Text Available In high-grade gliomas, the identification of patients that could benefit from EGFR inhibitors remains a challenge, hindering the use of these agents. Using xenografts models, we evaluated the antitumor effect of the combined treatment "gefitinib + radiotherapy" and aimed to identify the profile of responsive tumors. Expression of phosphorylated proteins involved in the EGFR-dependent signaling pathways was analyzed in 10 glioma models. We focused on three models of anaplastic oligodendrogliomas (TCG2, TCG3 and TCG4 harboring high levels of phospho-EGFR, phospho-AKT and phospho-MEK1. They were treated with gefitinib (GEF 75 mg/kg/day x 5 days/week, for 2 weeks and/or fractionated radiotherapy (RT: 5x2Gy/week for 2 weeks. Our results showed that GEF and/or RT induced significant tumor growth delays. However, only the TCG3 xenografts were highly responsive to the combination GEF+RT, with ∼50% of tumor cure. Phosphoproteins analysis five days after treatment onset demonstrated in TCG3 xenografts, but not in TCG2 model, that the EGFR-dependent pathways were inhibited after GEF treatment. Moreover, TCG3-bearing mice receiving GEF monotherapy exhibited a transient beneficial therapeutic response, rapidly followed by tumor regrowth, along with a major vascular remodeling. Taken together, our data evoked an "EGFR-addictive" behavior for TCG3 tumors. This study confirms that combination of gefitinib with fractionated irradiation could be a potent therapeutic strategy for anaplastic oligodendrogliomas harboring EGFR abnormalities but this treatment seems mainly beneficial for "EGFR-addictive" tumors. Unfortunately, neither the usual molecular markers (EGFR amplification, PTEN loss nor the basal overexpression of phosphoproteins were useful to distinguish this responsive tumor. Evaluating the impact of TKIs on the EGFR-dependent pathways during the treatment might be more relevant, and requires further validation.

  20. Renal cortical hexokinase and pentose phosphate pathway activation through the EGFR/Akt signaling pathway in endotoxin-induced acute kidney injury.

    Science.gov (United States)

    Smith, Joshua A; Stallons, L Jay; Schnellmann, Rick G

    2014-08-15

    While disruption of energy production is an important contributor to renal injury, metabolic alterations in sepsis-induced AKI remain understudied. We assessed changes in renal cortical glycolytic metabolism in a mouse model of sepsis-induced AKI. A specific and rapid increase in hexokinase (HK) activity (∼2-fold) was observed 3 h after LPS exposure and maintained up to 18 h, in association with a decline in renal function as measured by blood urea nitrogen (BUN). LPS-induced HK activation occurred independently of HK isoform expression or mitochondrial localization. No other changes in glycolytic enzymes were observed. LPS-mediated HK activation was not sufficient to increase glycolytic flux as indicated by reduced or unchanged pyruvate and lactate levels in the renal cortex. LPS-induced HK activation was associated with increased glucose-6-phosphate dehydrogenase activity but not glycogen production. Mechanistically, LPS-induced HK activation was attenuated by pharmacological inhibitors of the EGF receptor (EGFR) and Akt, indicating that EGFR/phosphatidylinositol 3-kinase/Akt signaling is responsible. Our findings reveal LPS rapidly increases renal cortical HK activity in an EGFR- and Akt-dependent manner and that HK activation is linked to increased pentose phosphate pathway activity.

  1. Multi-determinants analysis of molecular alterations for predicting clinical benefit to EGFR-targeted monoclonal antibodies in colorectal cancer.

    Directory of Open Access Journals (Sweden)

    Andrea Sartore-Bianchi

    Full Text Available BACKGROUND: KRAS mutations occur in 35-45% of metastatic colorectal cancers (mCRC and preclude responsiveness to EGFR-targeted therapy with cetuximab or panitumumab. However, less than 20% patients displaying wild-type KRAS tumors achieve objective response. Alterations in other effectors downstream of the EGFR, such as BRAF, and deregulation of the PIK3CA/PTEN pathway have independently been found to give rise to resistance. We present a comprehensive analysis of KRAS, BRAF, PIK3CA mutations, and PTEN expression in mCRC patients treated with cetuximab or panitumumab, with the aim of clarifying the relative contribution of these molecular alterations to resistance. METHODOLOGY/PRINCIPAL FINDINGS: We retrospectively analyzed objective tumor response, progression-free (PFS and overall survival (OS together with the mutational status of KRAS, BRAF, PIK3CA and expression of PTEN in 132 tumors from cetuximab or panitumumab treated mCRC patients. Among the 106 non-responsive patients, 74 (70% had tumors with at least one molecular alteration in the four markers. The probability of response was 51% (22/43 among patients with no alterations, 4% (2/47 among patients with 1 alteration, and 0% (0/24 for patients with > or =2 alterations (p or =2 molecular alteration(s (p<0.001. CONCLUSIONS/SIGNIFICANCE: When expression of PTEN and mutations of KRAS, BRAF and PIK3CA are concomitantly ascertained, up to 70% of mCRC patients unlikely to respond to anti-EGFR therapies can be identified. We propose to define as 'quadruple negative', the CRCs lacking alterations in KRAS, BRAF, PTEN and PIK3CA. Comprehensive molecular dissection of the EGFR signaling pathways should be considered to select mCRC patients for cetuximab- or panitumumab-based therapies.

  2. Molecular docking based screening of novel designed chalcone series of compounds for their anti-cancer activity targeting EGFR kinase domain

    Science.gov (United States)

    Rao, Chennu Maruthi Malya Prasada; Yejella, Rajendra Prasad; Rehman, Rehman Shaik Abdul; Basha, Syed Hussain

    2015-01-01

    Epidermal growth factor receptors (EGFR) are critical for the growth of many tumors and expressed at high levels in about one third of epithelial cancers. Hence, blockade of the binding sites for EGFR has been hypothesized as an effective anti-cancer therapy. Chalcone derivative compounds have been shown to be highly effective anti-cancer agents, however there are still so many novel derivatives possible, one of which might get us the best targeted EGFR inhibitor. In this effort directed towards the discovery of novel, potent anti-tumor agents for the treatment of cancer, in the present study a library of novel chalcone series of compounds has been designed and evaluated for their anti-cancer activity targeting EGFR kinase domain using various computational approaches. Among the twenty five novel designed chalcone series of compounds, all of them have found to be successfully docking inside the active binding domain of EGFR receptor target with a binding energy in a range of -6.10 to -9.25 Kcal/mol with predicted IC50 value range of 33.50 micor molar to 164.66 nano molar respectively. On the other hand, calculated 2DQSAR molecular descriptor properties of the compounds showed promising ADME parameters and found to be well in compliance with Lipinski׳s rule of five. Among all the twenty five compounds tested, compound 21 ((2E)-3-(anthracen-9-yl)-1-phenylprop-2-2n-1- one) was found to be the best lead like molecule with a binding energy of -9.25 kcal/mol with predicted IC50 value of 164.66 nano molar. Conclusively, novel designed compound 21 of the present study have shown promising anti-cancer potential worth considering for further evaluations. PMID:26339147

  3. Activation of EGFR promotes squamous carcinoma SCC10A cell migration and invasion via inducing EMT-like phenotype change and MMP-9-mediated degradation of E-cadherin.

    Science.gov (United States)

    Zuo, Jian-Hong; Zhu, Wei; Li, Mao-Yu; Li, Xin-Hui; Yi, Hong; Zeng, Gu-Qing; Wan, Xun-Xun; He, Qiu-Yan; Li, Jian-Huang; Qu, Jia-Quan; Chen, Yu; Xiao, Zhi-Qiang

    2011-09-01

    EGFR is a potent stimulator of invasion and metastasis in head and neck squamous cell carcinomas (HNSCC). However, the mechanism by which EGFR may stimulate tumor cell invasion and metastasis still need to be elucidated. In this study, we showed that activation of EGFR by EGF in HNSCC cell line SCC10A enhanced cell migration and invasion, and induced loss of epitheloid phenotype in parallel with downregulation of E-cadherin and upregulation of N-cadherin and vimentin, indicating that EGFR promoted SCC10A cell migration and invasion possibly by an epithelial to mesenchymal transition (EMT)-like phenotype change. Interestingly, activation of EGFR by EGF induced production of matrix metalloproteinase-9 (MMP-9) and soluble E-cadherin (sE-cad), and knockdown of MMP-9 by siRNA inhibited sE-cad production induced by EGF in SCC10A. Moreover, both MMP-9 knockdown and E-cadherin overexpression inhibited cell migration and invasion induced by EGF in SCC10A. The results indicate that EGFR activation promoted cell migration and invasion through inducing MMP-9-mediated degradation of E-cadherin into sE-cad. Pharmacologic inhibition of EGFR, MEK, and PI3K kinase activity in SCC10A reduced phosphorylated levels of ERK-1/2 and AKT, production of MMP-9 and sE-cad, cell migration and invasion, and expressional changes of EMT markers (E-cadherin and N-cadherin) induced by EGF, indicating that EGFR activation promotes cell migration and invasion via ERK-1/2 and PI3K-regulated MMP-9/E-cadherin signaling pathways. Taken together, the data suggest that EGFR activation promotes HNSCC SCC10A cell migration and invasion by inducing EMT-like phenotype change and MMP-9-mediated degradation of E-cadherin into sE-cad related to activation of ERK-1/2 and PI3K signaling pathways.

  4. Synergistic antitumor effect between gefitinib and fractionated irradiation in anaplastic oligodendrogliomas cannot be predicted by the Egfr signaling activity.

    OpenAIRE

    Sophie Pinel; Jihane Mriouah; Marc Vandamme; Alicia Chateau; François Plénat; Eric Guérin; Luc Taillandier; Valérie Bernier-Chastagner; Jean-Louis Merlin; Pascal Chastagner

    2013-01-01

    International audience In high-grade gliomas, the identification of patients that could benefit from EGFR inhibitors remains a challenge, hindering the use of these agents. Using xenografts models, we evaluated the antitumor effect of the combined treatment "gefitinib + radiotherapy" and aimed to identify the profile of responsive tumors. Expression of phosphorylated proteins involved in the EGFR-dependent signaling pathways was analyzed in 10 glioma models. We focused on three models of a...

  5. EGFR-targeted TRAIL and a Smac mimetic synergize to overcome apoptosis resistance in KRAS mutant colorectal cancer cells.

    Directory of Open Access Journals (Sweden)

    Yvonne Möller

    Full Text Available TRAIL is a death receptor ligand that induces cell death preferentially in tumor cells. Recombinant soluble TRAIL, however, performs poorly as an anti-cancer therapeutic because oligomerization is required for potent biological activity. We previously generated a diabody format of tumor-targeted TRAIL termed Db(αEGFR-scTRAIL, comprising single-stranded TRAIL molecules (scTRAIL and the variable domains of a humanized variant of the EGFR blocking antibody Cetuximab. Here we define the bioactivity of Db(αEGFR-scTRAIL with regard to both EGFR inhibition and TRAIL receptor activation in 3D cultures of Caco-2 colorectal cancer cells, which express wild-type K-Ras. Compared with conventional 2D cultures, Caco-2 cells displayed strongly enhanced sensitivity toward Db(αEGFR-scTRAIL in these 3D cultures. We show that the antibody moiety of Db(αEGFR-scTRAIL not only efficiently competed with ligand-induced EGFR function, but also determined the apoptotic response by specifically directing Db(αEGFR-scTRAIL to EGFR-positive cells. To address how aberrantly activated K-Ras, which leads to Cetuximab resistance, affects Db(αEGFR-scTRAIL sensitivity, we generated stable Caco-2tet cells inducibly expressing oncogenic K-Ras(G12V. In the presence of doxycycline, these cells showed increased resistance to Db(αEGFR-scTRAIL, associated with the elevated expression of the anti-apoptotic proteins cIAP2, Bcl-xL and FlipS. Co-treatment of cells with the Smac mimetic SM83 restored the Db(αEGFR-scTRAIL-induced apoptotic response. Importantly, this synergy between Db(αEGFR-scTRAIL and SM83 also translated to 3D cultures of oncogenic K-Ras expressing HCT-116 and LoVo colorectal cancer cells. Our findings thus support the notion that Db(αEGFR-scTRAIL therapy in combination with apoptosis-sensitizing agents may be promising for the treatment of EGFR-positive colorectal cancers, independently of their KRAS status.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  7. Discovery of a series of novel phenylpiperazine derivatives as EGFR TK inhibitors

    Science.gov (United States)

    Sun, Juan; Wang, Xin-Yi; Lv, Peng-Cheng; Zhu, Hai-Liang

    2015-09-01

    Human epidermal growth factor receptor (EGFR) is an important drug target that plays a fundamental role in signal transduction pathways in oncology. We report herein the discovery of a novel class of phenylpiperazine derivatives with improved potency toward EGFR. The biological activity of compound 3p as inhibitor of EGFR was further investigated both in vitro and in vivo. Notably, compound 3p exhibited an IC50 in the nanomolar range in A549 cell cultures and induced a cessation of tumor growth with no toxicity, as determined by loss of body weight and death of treated mice. Compoutational docking studies also showed that compound 3p has interaction with EGFR key residues in the active site.

  8. EGFR and Bcl-2 in gastric mucosa of children infected with Helicobacter pylori

    Directory of Open Access Journals (Sweden)

    Ewa Ryszczuk

    2016-03-01

    Full Text Available Aim: The aim of the study was to evaluate the expression of EGFR and Bcl-2 proteins as inhibitory markers of apoptosis in surface epithelial cells and gland cells of antral gastric mucosa in children infected with Helicobacter pylori according to the severity and activity of antral gastritis and to assess the correlation between the number of cells expressing EGFR and the number of cells expressing Bcl-2 in H. pylori infected children. Materials and methods: The study included 44 children: 68.2% with chronic gastritis and positive IgG against H. pylori, and 31.8% with functional disorders of the gastrointestinal tract and with normal IgG against H. pylori. The evaluation of EGFR expression in gastric mucosa was performed immunohistochemically using monoclonal mouse anti-EGFR antibody. The polyclonal antibody was used to determine the expression of anti-Bcl-2. Results: A significant increase in the number of cells expressing EGFR and Bcl-2 protein was found in the epithelial cells in severe as well as mild and moderate gastritis in the group of children infected with H. pylori. An increase in the number of cells expressing EGFR and Bcl-2 protein was also found in the epithelial cells in group I according to the activity of gastritis. There was a statistically significant positive correlation between the numbers of cells expressing EGFR and Bcl-2 in H. pylori infected children. Conclusion: Increased expression of EGFR and Bcl-2 proteins in the epithelial cells and a statistically significant positive correlation between the numbers of cells expressing EGFR and Bcl-2 in H. pylori infected children could suggest increased regeneration abilities of gastric mucosa.

  9. Mutation analysis of circulating plasma DNA to determine response to EGFR tyrosine kinase inhibitor therapy of lung adenocarcinoma patients

    Science.gov (United States)

    Riediger, Anja Lisa; Dietz, Steffen; Schirmer, Uwe; Meister, Michael; Heinzmann-Groth, Ingrid; Schneider, Marc; Muley, Thomas; Thomas, Michael; Sültmann, Holger

    2016-01-01

    Long-lasting success in lung cancer therapy using tyrosine kinase inhibitors (TKIs) is rare since the tumors develop resistance due to the occurrence of molecularly altered subclones. The aim of this study was to monitor tumors over time based on the quantity of mutant plasma DNA and to identify early indications for therapy response and tumor progression. Serial plasma samples from lung adenocarcinoma patients treated with TKIs were used to quantify EGFR and KRAS mutations in circulating DNA by digital PCR. Mutant DNA levels were compared with the courses of responses to treatment with TKIs, conventional chemotherapy, radiotherapy, or combinations thereof. Variations in plasma DNA mutation levels over time were found in 15 patients. We categorize three major courses: First, signs of therapy response are associated with a fast clearing of plasma DNA mutations within a few days. Second, periods of stable disease are accompanied by either absence of mutations or fluctuation at low levels. Finally, dramatic increase of mutational load is followed by rapid tumor progression and poor patient survival. In summary, the serial assessment of EGFR mutations in the plasma of NSCLC patients allows conclusions about controlled disease and tumor progression earlier than currently available methods. PMID:27640882

  10. Immunohistochemical detections of EGFR status in NSCLC

    Directory of Open Access Journals (Sweden)

    Jie WANG

    2015-04-01

    Full Text Available Background and objective Patients with non-small cell lung cancer (NSCLC harboring mutations of the epidermal growth factor receptor (EGFR respond well to EGFR-tyrosine kinase inhibitor therapy. Immunohistochemistry (IHC is a simple and widely used technique in clinical pathology laboratories. IHC also features cost effectiveness and rapid detection of EGFR mutations compared with molecular methods. This study aims to determine the accuracy of IHC for EGFR mutation detection in NSCLC. Methods Specimens (obtained from surgery or biopsy from 97 NSCLC cases were stained through IHC with mutation-specific antibodies. The clinicopathological features of patients with positive immunostaining results were analyzed. Positive specimens were subjected to liquid chip technology to detect the actual EGFR status. Forty NSCLC specimens obtained from surgery and confirmed to have EGFR mutations through liquid chip technology were collected. These specimens were then subjected to IHC analyses with mutation-specific antibodies. The sensitivity of IHC in detecting EGFR mutations was calculated. Results Seventeen of the 97 NSCLC specimens were stained positive, and positive results were mostly observed in females, patients with adenocarcinoma, and non-smokers. About 76.9% of specimens with positive IHC results harbored mutations. The sensitivity of IHC was 40% among the 40 cases identified as containing EGFR mutations through liquid chip technology. Conclusion The strong positive immunostaining result is accurate, but the sensitivity of the method may not be optimal and significantly varies in different studies. The widespread application of IHC in clinics must be further investigated.

  11. EGFR as a therapeutic target in glioblastoma

    Directory of Open Access Journals (Sweden)

    David M Siebert

    2012-01-01

    Full Text Available The tyrosine kinase receptor epidermal growth factor receptor (EGFR can be activated by several ligands, thus triggering downstream pathways regulating cell growth and survival. Its dysregula­tion is particularly important for the development and progression of astrocytomas. After the description of its role in glioblastomas (WHO grade IV astrocytomas, an overview on the therapeutic strategies target­ing EGFR is provided. It analyzes the past and ongoing trials concerning the small molecule tyro­sine kinase inhibitors, i.e. gefitinib, erlotinib and the combination therapies, the EGFR vaccina­tion strategies, the antibodies directed against EGFR and finally the intracranially administered EGFR-targeted therapies. As our understanding of the underlying molecular aberrancies in glioblastoma grows, our ability to better target specific subtypes of glioblastoma should improve. Molecular biomarker enriched clinical trials may lead to improved patient outcomes.

  12. The relative expression of Mig6 and EGFR is associated with resistance to EGFR kinase inhibitors.

    Directory of Open Access Journals (Sweden)

    Xiaofei Chang

    Full Text Available The sensitivity of only a few tumors to anti-epidermal growth factor receptor EGFR tyrosine kinase inhibitors (TKIs can be explained by the presence of EGFR tyrosine kinase (TK domain mutations. In addition, such mutations were rarely found in tumor types other than lung, such as pancreatic and head and neck cancer. In this study we sought to elucidate mechanisms of resistance to EGFR-targeted therapies in tumors that do not harbor TK sensitizing mutations in order to identify markers capable of guiding the decision to incorporate these drugs into chemotherapeutic regimens. Here we show that EGFR activity was markedly decreased during the evolution of resistance to the EGFR tyrosine kinase inhibitor (TKI erlotinib, with a concomitant increase of mitogen-inducible gene 6 (Mig6, a negative regulator of EGFR through the upregulation of the PI3K-AKT pathway. EGFR activity, which was more accurately predicted by the ratio of Mig6/EGFR, highly correlated with erlotinib sensitivity in panels of cancer cell lines of different tissue origins. Blinded testing and analysis in a prospectively followed cohort of lung cancer patients treated with gefitinib alone demonstrated higher response rates and a marked increased in progression free survival for patients with a low Mig6/EGFR ratio (approximately 100 days, P = 0.01.

  13. Protein phosphorylation profiling using an in situ proximity ligation assay: phosphorylation of AURKA-elicited EGFR-Thr654 and EGFR-Ser1046 in lung cancer cells.

    Directory of Open Access Journals (Sweden)

    Tzu-Chi Chen

    Full Text Available The epidermal growth factor receptor (EGFR, which is up-regulated in lung cancer, involves the activation of mitogenic signals and triggers multiple signaling cascades. To dissect these EGFR cascades, we used 14 different phospho-EGFR antibodies to quantify protein phosphorylation using an in situ proximity ligation assay (in situ PLA. Phosphorylation at EGFR-Thr654 and -Ser1046 was EGF-dependent in the wild-type (WT receptor but EGF-independent in a cell line carrying the EGFR-L858R mutation. Using a ProtoAarray™ containing ∼5000 recombinant proteins on the protein chip, we found that AURKA interacted with the EGFR-L861Q mutant. Moreover, overexpression of EGFR could form a complex with AURKA, and the inhibitors of AURKA and EGFR decreased EGFR-Thr654 and -Ser1046 phosphorylation. Immunohistochemical staining of stage I lung adenocarcinoma tissues demonstrated a positive correlation between AURKA expression and phosphorylation of EGFR at Thr654 and Ser1046 in EGFR-mutant specimens, but not in EGFR-WT specimens. The interplay between EGFR and AURKA provides an explanation for the difference in EGF dependency between EGFR-WT and EGFR-mutant cells and may provide a new therapeutic strategy for lung cancer patients carrying EGFR mutations.

  14. Epitope mapping of epidermal growth factor receptor (EGFR) monoclonal antibody and induction of growth-inhibitory polyclonal antibodies by vaccination with EGFR mimotope.

    Science.gov (United States)

    Navari, Mohsen; Zare, Mehrak; Javanmardi, Masoud; Asadi-Ghalehni, Majid; Modjtahedi, Helmout; Rasaee, Mohammad Javed

    2014-10-01

    One of the proposed approaches in cancer therapy is to induce and direct the patient's own immune system against cancer cells. In this study, we determined the epitope mapping of the rat anti-human epidermal growth factor receptor (EGFR) monoclonal antibody ICR-62 using a phage display of random peptide library and identified a 12 amino acids peptide, which was recognized as a mimotope. The peptide was synthesized and conjugated to bovine serum albumin (BSA) as carrier protein (P-BSA). We have shown that ICR-62 can react specifically with P-BSA as well as native EGFR. Two rabbits were immunized either by BSA or P-BSA and the rabbits IgGs were purified and examined for binding to the antigens, mimotope and the EGFR protein purified from the EGFR overexpressing A431 cell line. We showed that the rabbit IgG generated against the mimotope is capable of inhibiting the growth of A431 cells by 15%, but does not have any effect on the growth of EGFR-negative MDA-MB-453 cell line in vitro. Our results support the need for further investigations on the potential of vaccination with either mimotope of the EGFR or epitope displayed on the surface of phage particles for use in active immunotherapy of cancer.

  15. EGFR-mediated carcinoma cell metastasis mediated by integrin αvβ5 depends on activation of c-Src and cleavage of MUC1.

    Directory of Open Access Journals (Sweden)

    Steven K M Lau

    Full Text Available Receptor tyrosine kinases and integrins play an essential role in tumor cell invasion and metastasis. We previously showed that EGF and other growth factors induce human carcinoma cell invasion and metastasis mediated by integrin αvβ5 that is prevented by Src blockade. MUC1, a transmembrane glycoprotein, is expressed in most epithelial tumors as a heterodimer consisting of an extracellular and a transmembrane subunit. The MUC1 cytoplasmic domain of the transmembrane subunit (MUC1.CD translocates to the nucleus where it promotes the transcription of a metastatic gene signature associated with epithelial to mesenchymal transition. Here, we demonstrate a requirement for MUC1 in carcinoma cell metastasis dependent on EGFR and Src without affecting primary tumor growth. EGF stimulates Src-dependent MUC1 cleavage and nuclear localization leading to the expression of genes linked to metastasis. Moreover, expression of MUC1.CD results in its nuclear localization and is sufficient for transcription of the metastatic gene signature and tumor cell metastasis. These results demonstrate that EGFR and Src activity contribute to carcinoma cell invasion and metastasis mediated by integrin αvβ5 in part by promoting proteolytic cleavage of MUC1 and highlight the ability of MUC1.CD to promote metastasis in a context-dependent manner. Our findings may have implications for the use and future design of targeted therapies in cancers known to express EGFR, Src, or MUC1.

  16. Plant polyphenols differentially modulate inflammatory responses of human keratinocytes by interfering with activation of transcription factors NFκB and AhR and EGFR-ERK pathway

    International Nuclear Information System (INIS)

    Molecular mechanisms underlying modulation of inflammatory responses in primary human keratinocytes by plant polyphenols (PPs), namely the glycosylated phenylpropanoid verbascoside, the stilbenoid resveratrol and its glycoside polydatin, and the flavonoid quercetin and its glycoside rutin were evaluated. As non-lethal stimuli, the prototypic ligand for epidermal growth factor receptor (EGFR) transforming growth factor alpha (TGFalpha), the combination of tumor necrosis factor (TNFalpha) and interferon (IFNgamma) (T/I), UVA + UVB irradiation, and bacterial lipopolysaccharide (LPS) were used. We demonstrated differential modulation of inflammatory responses in keratinocytes at signal transduction, gene transcription, and protein synthesis levels as a function of PP chemical structure, the pro-inflammatory trigger used, and PP interaction with intracellular detoxifying systems. The PPs remarkably inhibited constitutive, LPS- and T/I-induced but not TGFalpha-induced ERK phosphorylation. They also suppressed NFkappaB activation by LPS and T/I. Verbascoside and quercetin invariably impaired EGFR phosphorylation and UV-associated aryl hydrocarbon receptor (AhR)-mediated signaling, while rutin, polydatin and resveratrol did not affect EGFR phosphorylation and further activated AhR machinery in UV-exposed keratinocytes. In general, PPs down-regulated gene expression of pro-inflammatory cytokines/enzymes, except significant up-regulation of IL-8 observed under stimulation with TGFalpha. Both spontaneous and T/I-induced release of IL-8 and IP-10 was suppressed, although 50 μM resveratrol and polydatin up-regulated IL-8. At this concentration, resveratrol activated both gene expression and de novo synthesis of IL-8 and AhR-mediated mechanisms were involved. We conclude that PPs differentially modulate the inflammatory response of human keratinocytes through distinct signal transduction pathways, including AhR and EGFR. - Graphical abstract: Display Omitted Highlights:

  17. 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 intracellular region of the receptor. This tyrosine kinase phosphorylates a number of intracellular substrates that activates pathways leading to cell growth, DNA synthesis and the expression of oncogenes such as fos and jun. EGFR is thought to be involved the development of cancer, as the EGFR gene is often...... amplified, and/or mutated in cancer cells. In this review we will focus on: (I) the structure and function of EGFR, (II) implications of receptor/ligand coexpression and EGFR mutations or overexpression, (III) its effect on cancer cells, (IV) the development of the malignant phenotype and (V) the clinical...

  18. Quantifying the effects of co-expressing EGFR and HER2 on HER activation and trafficking

    Energy Technology Data Exchange (ETDEWEB)

    Shankaran, Harish; Zhang, Yi; Opresko, Lee; Resat, Haluk

    2008-06-27

    The integration of experimental measurements and computational predictions is a powerful means to understand the molecular mechanisms of complex biological systems. The human epidermal growth factor receptor (HER) system is an intricately regulated system that plays critical roles in development and tumorigenesis. Here, we apply integrated experimentation and modeling to analyze HER receptor activation in a panel of cell lines expressing different levels of HER1 and HER2. A mathematical model that includes the fundamental biochemical/biophysical processes involved in receptor activation was used to fit the experimental data, and 19 independent parameters including receptor dimerization affinities, trafficking rates and relative phosphorylation levels were estimated. The parameter values quantitatively support existing ideas on the effect of HER2 on HER1 phosphorylation dynamics, and enable us to predict receptor dimerization patterns in the cell lines. The integrated approach described here can be applied to obtain a predictive understanding of other biomolecular systems.

  19. Activation of EGFR/ERBB2 via Pathways Involving ERK1/2, P38 MAPK, AKT and FOXO Enhances Recovery of Diabetic Hearts from Ischemia-Reperfusion Injury

    Science.gov (United States)

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

    2012-01-01

    This study characterized the effects of diabetes and/or ischemia on epidermal growth factor receptor, EGFR, and/or erbB2 signaling pathways on cardiac function. Isolated heart perfusion model of global ischemia was used to study the effect of chronic inhibition or acute activation of EGFR/erbB2 signaling on cardiac function in a rat model of type-1 diabetes. Induction of diabetes with streptozotocin impaired recovery of cardiac function (cardiac contractility and hemodynamics) following 40 minutes of global ischemia in isolated hearts. Chronic treatment with AG825 or AG1478, selective inhibitors of erbB2 and EGFR respectively, did not affect hyperglycemia but led to an exacerbation whereas acute administration of the EGFR ligand, epidermal growth factor (EGF), led to an improvement in cardiac recovery in diabetic hearts. Diabetes led to attenuated dimerization and phosphorylation of cardiac erbB2 and EGFR receptors that was associated with reduced signaling via extracellular-signal-regulated kinase 1/2 (ERK1/2), p38 mitogen activated protein (MAP) kinase and AKT (protein kinase B). Ischemia was also associated with reduced cardiac signaling via these molecules whereas EGF-treatment opposed diabetes and/or ischemia induced changes in ERK1/2, p38 MAP kinase, and AKT-FOXO signaling. Losartan treatment improved cardiac function in diabetes but also impaired EGFR phosphorylation in diabetic heart. Co-administration of EGF rescued Losartan-mediated reduction in EGFR phosphorylation and significantly improved cardiac recovery more than with either agent alone. EGFR/erbB2 signaling is an important cardiac survival pathway whose activation, particularly in diabetes, ischemia or following treatment with drugs that inhibit this cascade, significantly improves cardiac function. These findings may have clinical relevance particularly in the treatment of diabetes-induced cardiac dysfunction. PMID:22720029

  20. Activation of EGFR/ERBB2 via pathways involving ERK1/2, P38 MAPK, AKT and FOXO enhances recovery of diabetic hearts from ischemia-reperfusion injury.

    Directory of Open Access Journals (Sweden)

    Saghir Akhtar

    Full Text Available This study characterized the effects of diabetes and/or ischemia on epidermal growth factor receptor, EGFR, and/or erbB2 signaling pathways on cardiac function. Isolated heart perfusion model of global ischemia was used to study the effect of chronic inhibition or acute activation of EGFR/erbB2 signaling on cardiac function in a rat model of type-1 diabetes. Induction of diabetes with streptozotocin impaired recovery of cardiac function (cardiac contractility and hemodynamics following 40 minutes of global ischemia in isolated hearts. Chronic treatment with AG825 or AG1478, selective inhibitors of erbB2 and EGFR respectively, did not affect hyperglycemia but led to an exacerbation whereas acute administration of the EGFR ligand, epidermal growth factor (EGF, led to an improvement in cardiac recovery in diabetic hearts. Diabetes led to attenuated dimerization and phosphorylation of cardiac erbB2 and EGFR receptors that was associated with reduced signaling via extracellular-signal-regulated kinase 1/2 (ERK1/2, p38 mitogen activated protein (MAP kinase and AKT (protein kinase B. Ischemia was also associated with reduced cardiac signaling via these molecules whereas EGF-treatment opposed diabetes and/or ischemia induced changes in ERK1/2, p38 MAP kinase, and AKT-FOXO signaling. Losartan treatment improved cardiac function in diabetes but also impaired EGFR phosphorylation in diabetic heart. Co-administration of EGF rescued Losartan-mediated reduction in EGFR phosphorylation and significantly improved cardiac recovery more than with either agent alone. EGFR/erbB2 signaling is an important cardiac survival pathway whose activation, particularly in diabetes, ischemia or following treatment with drugs that inhibit this cascade, significantly improves cardiac function. These findings may have clinical relevance particularly in the treatment of diabetes-induced cardiac dysfunction.

  1. Activation of EGFR/ERBB2 via pathways involving ERK1/2, P38 MAPK, AKT and FOXO enhances recovery of diabetic hearts from ischemia-reperfusion injury.

    Science.gov (United States)

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

    2012-01-01

    This study characterized the effects of diabetes and/or ischemia on epidermal growth factor receptor, EGFR, and/or erbB2 signaling pathways on cardiac function. Isolated heart perfusion model of global ischemia was used to study the effect of chronic inhibition or acute activation of EGFR/erbB2 signaling on cardiac function in a rat model of type-1 diabetes. Induction of diabetes with streptozotocin impaired recovery of cardiac function (cardiac contractility and hemodynamics) following 40 minutes of global ischemia in isolated hearts. Chronic treatment with AG825 or AG1478, selective inhibitors of erbB2 and EGFR respectively, did not affect hyperglycemia but led to an exacerbation whereas acute administration of the EGFR ligand, epidermal growth factor (EGF), led to an improvement in cardiac recovery in diabetic hearts. Diabetes led to attenuated dimerization and phosphorylation of cardiac erbB2 and EGFR receptors that was associated with reduced signaling via extracellular-signal-regulated kinase 1/2 (ERK1/2), p38 mitogen activated protein (MAP) kinase and AKT (protein kinase B). Ischemia was also associated with reduced cardiac signaling via these molecules whereas EGF-treatment opposed diabetes and/or ischemia induced changes in ERK1/2, p38 MAP kinase, and AKT-FOXO signaling. Losartan treatment improved cardiac function in diabetes but also impaired EGFR phosphorylation in diabetic heart. Co-administration of EGF rescued Losartan-mediated reduction in EGFR phosphorylation and significantly improved cardiac recovery more than with either agent alone. EGFR/erbB2 signaling is an important cardiac survival pathway whose activation, particularly in diabetes, ischemia or following treatment with drugs that inhibit this cascade, significantly improves cardiac function. These findings may have clinical relevance particularly in the treatment of diabetes-induced cardiac dysfunction.

  2. JNK Pathway Activation Modulates Acquired Resistance to EGFR/HER2-Targeted Therapies.

    Science.gov (United States)

    Manole, Simin; Richards, Edward J; Meyer, Aaron S

    2016-09-15

    Resistance limits the effectiveness of receptor tyrosine kinase (RTK)-targeted therapies. Combination therapies targeting resistance mechanisms can considerably improve response, but will require an improved understanding of when particular combinations will be effective. One common form of resistance is bypass signaling, wherein RTKs not targeted by an inhibitor can direct reactivation of pathways essential for survival. Although this mechanism of resistance is well appreciated, it is unclear which downstream signaling events are responsible. Here, we apply a combined experimental- and statistical modeling-based approach to identify a set of pathway reactivation essential for RTK-mediated bypass resistance. Differences in the downstream pathway activation provided by particular RTKs lead to qualitative differences in the capacity of each receptor to drive therapeutic resistance. We identify and validate that the JNK pathway is activated during and strongly modulates bypass resistance. These results identify effective therapeutic combinations that block bypass-mediated resistance and provide a basic understanding of this network-level change in kinase dependence that will inform the design of prognostic assays for identifying effective therapeutic combinations in individual patients. Cancer Res; 76(18); 5219-28. ©2016 AACR. PMID:27450453

  3. Sym004, a novel EGFR antibody mixture, can overcome acquired resistance to cetuximab.

    Science.gov (United States)

    Iida, Mari; Brand, Toni M; Starr, Megan M; Li, Chunrong; Huppert, Evan J; Luthar, Neha; Pedersen, Mikkel W; Horak, Ivan D; Kragh, Michael; Wheeler, Deric L

    2013-10-01

    The epidermal growth factor receptor (EGFR) is a central regulator of tumor progression in a variety of human cancers. Cetuximab is an anti-EGFR monoclonal antibody that has been approved for head and neck and colorectal cancer treatment, but many patients treated with cetuximab don't respond or eventually acquire resistance. To determine how tumor cells acquire resistance to cetuximab, we previously developed a model of acquired resistance using the non-small cell lung cancer line NCI-H226. These cetuximab-resistant (Ctx(R)) cells exhibit increased steady-state EGFR expression secondary to alterations in EGFR trafficking and degradation and, further, retained dependence on EGFR signaling for enhanced growth potential. Here, we examined Sym004, a novel mixture of antibodies directed against distinct epitopes on the extracellular domain of EGFR, as an alternative therapy for Ctx(R) tumor cells. Sym004 treatment of Ctx(R) clones resulted in rapid EGFR degradation, followed by robust inhibition of cell proliferation and down-regulation of several mitogen-activated protein kinase pathways. To determine whether Sym004 could have therapeutic benefit in vivo, we established de novo Ctx(R) NCI-H226 mouse xenografts and subsequently treated Ctx(R) tumors with Sym004. Sym004 treatment of mice harboring Ctx(R) tumors resulted in growth delay compared to mice continued on cetuximab. Levels of total and phospho-EGFR were robustly decreased in Ctx(R) tumors treated with Sym004. Immunohistochemical analysis of these Sym004-treated xenograft tumors further demonstrated decreased expression of Ki67, and phospho-rpS6, as well as a modest increase in cleaved caspase-3. These results indicate that Sym004 may be an effective targeted therapy for Ctx(R) tumors. PMID:24204198

  4. Sym004, a Novel EGFR Antibody Mixture, Can Overcome Acquired Resistance to Cetuximab

    Directory of Open Access Journals (Sweden)

    Mari Iida

    2013-10-01

    Full Text Available The epidermal growth factor receptor (EGFR is a central regulator of tumor progression in a variety of human cancers. Cetuximab is an anti-EGFR monoclonal antibody that has been approved for head and neck and colorectal cancer treatment, but many patients treated with cetuximab don't respond or eventually acquire resistance. To determine how tumor cells acquire resistance to cetuximab, we previously developed a model of acquired resistance using the non-small cell lung cancer line NCI-H226. These cetuximab-resistant (CtxR cells exhibit increased steady-state EGFR expression secondary to alterations in EGFR trafficking and degradation and, further, retained dependence on EGFR signaling for enhanced growth potential. Here, we examined Sym004, a novel mixture of antibodies directed against distinct epitopes on the extracellular domain of EGFR, as an alternative therapy for CtxR tumor cells. Sym004 treatment of CtxR clones resulted in rapid EGFR degradation, followed by robust inhibition of cell proliferation and down-regulation of several mitogen-activated protein kinase pathways. To determine whether Sym004 could have therapeutic benefit in vivo, we established de novo CtxR NCI-H226 mouse xenografts and subsequently treated CtxR tumors with Sym004. Sym004 treatment of mice harboring CtxR tumors resulted in growth delay compared to mice continued on cetuximab. Levels of total and phospho-EGFR were robustly decreased in CtxR tumors treated with Sym004. Immunohistochemical analysis of these Sym004-treated xenograft tumors further demonstrated decreased expression of Ki67, and phospho-rpS6, as well as a modest increase in cleaved caspase-3. These results indicate that Sym004 may be an effective targeted therapy for CtxR tumors.

  5. Activation of ErbB3, EGFR and Erk is essential for growth of human breast cancer cell lines with acquired resistance to fulvestrant

    DEFF Research Database (Denmark)

    Frogne, Thomas; Benjaminsen, Rikke V; Sonne-Hansen, Katrine;

    2008-01-01

    Seven fulvestrant resistant cell lines derived from the estrogen receptor alpha positive MCF-7 human breast cancer cell line were used to investigate the importance of epidermal growth factor receptor (ErbB1-4) signaling. We found an increase in mRNA expression of EGFR and the ErbB3/ErbB4 ligand...... cell lines concomitant with inhibition of Erk and unaltered Akt activation. In concert, inhibition of Erk with U0126 preferentially reduced growth of resistant cell lines. Treatment with ErbB3 neutralizing antibodies inhibited ErbB3 activation and resulted in a modest but statistically significant...

  6. Intratumoral distribution of EGFR-amplified and EGFR-mutated cells in pulmonary adenocarcinoma.

    Science.gov (United States)

    Soma, Shingo; Tsuta, Koji; Takano, Toshimi; Hatanaka, Yutaka; Yoshida, Akihiko; Suzuki, Kenji; Asamura, Hisao; Tsuda, Hitoshi

    2014-03-01

    Alterations in the epidermal growth factor receptor (EGFR) gene are associated with carcinogenesis in non-small cell lung cancer. However, the intratumoral distribution of these abnormalities has not been elucidated. This study included patients with surgically resected lung adenocarcinoma. The predominant histological growth pattern was determined. Chromogenic in situ hybridization (CISH) and EGFR-mutation specific-antibodies were used for analysis of changes in gene copy number and EGFR mutations, respectively. EGFR mutation detected immunohistochemistry (IHC) and amplification were identified in 31 (53%) and 30 (52%) cases, respectively. The predominant growth patterns in the 58 tumors evaluated were papillary (28, 48%), lepidic (8, 14%), acinar (15, 26%), and solid (7, 12%). EGFR mutations were the least common in cases with a solid predominant pattern. The incidence of EGFR amplification did not differ among predominant patterns. Analyzing each histological subtype, no differences were noted between the prevalence of EGFR-IHC positive and CISH-positive rates. In the analysis of EGFR amplification, CISH-positive status was more prevalent in IHC-positive cases than in IHC-negative cases. All 19 cases that were both IHC and CISH positive were analyzed. In 17 cases (90%), the IHC-positive area was equal to or larger than the CISH-positive area. Among the histological subtypes of lung adenocarcinoma, the solid predominant subtype was distinguishable by its infrequent EGFR mutations. EGFR gene mutations preceded changes in oncogenic drive, more so than did EGFR gene number alterations during the developmental process of lung adenocarcinoma. PMID:24355440

  7. Low prevalence of K-RAS, EGF-R and BRAF mutations in sinonasal adenocarcinomas. Implications for anti-EGFR treatments.

    Science.gov (United States)

    Franchi, Alessandro; Innocenti, Duccio Rossi Degli; Palomba, Annarita; Miligi, Lucia; Paiar, Fabiola; Franzese, Ciro; Santucci, Marco

    2014-07-01

    We have previously shown that a subset of sinonasal intestinal-type adenocarcinomas (ITAC) shows activation of the epidermal growth factor-receptor (EGFR) pathway. In this study we examine the status of the EGFR, KRAS and BRAF genes in a series of sinonasal intestinal (ITAC) and non-intestinal type adenocarcinomas (non-ITAC). Eighteen ITACs and 12 non-ITACs were studied immunohistochemically for EGFR expression. Point mutations were analyzed for EGFR exons 19 and 21, KRAS exon 2 and BRAF exon 15 by direct sequencing. Non-ITACs showed significantly higher expression of EGFR (p = 0.015). Mutation analysis revealed one ITAC with EGFR and one ITAC with KRAS mutation, while two non-ITACs presented mutation of BRAF. We conclude that a subset of sinonasal adenocarcinomas shows overexpression of EGFR, while activating mutations of the signaling cascade downstream of EGFR are rare, suggesting that these tumors could be good candidates for anti-EGFR therapies.

  8. Epidermal growth factor receptor (EGFR mutation status and Rad51 determine the response of glioblastoma (GBM to multimodality therapy with cetuximab, temozolomide and radiation

    Directory of Open Access Journals (Sweden)

    Phyllis Rachelle Wachsberger

    2013-02-01

    Full Text Available Purpose: EGFR amplification and mutation (i.e., EGFRvIII are found in 40% of primary GBM tumors and are believed to contribute to tumor development and therapeutic resistance. This study was designed to investigate how EGFR mutational status modulates response to multimodality treatment with cetuximab, an anti-EGFR inhibitor, the chemotherapeutic agent, temozolamide (TMZ and radiation therapy (RT Methods and Materials: In vitro and in vivo experiments were performed on two isogenic U87 GBM cell lines: one overexpressing wildtype EGFR (U87wtEGFR and the other overexpressing EGFRvIII (U87EGFRvIII. Results: Xenografts harboring EGFRvIII were more sensitive to TMZ alone and TMZ in combination with RT and/or cetuximab than xenografts expressing wtEGFR. In vitro experiments demonstrated that U87EGFRvIII-expressing tumors appear to harbor defective DNA homologous recombination repair in the form of Rad51 processing, Conclusions: The difference in sensitivity between EGFR-expressing and EGFRvIII-expressing tumors to combined modality treatment may help in the future tailoring of GBM therapy to subsets of patients expressing more or less of the EGFR mutant.

  9. Active G protein-coupled receptors (GPCR), matrix metalloproteinases 2/9 (MMP2/9), heparin-binding epidermal growth factor (hbEGF), epidermal growth factor receptor (EGFR), erbB2, and insulin-like growth factor 1 receptor (IGF-1R) are necessary for trenbolone acetate-induced alterations in protein turnover rate of fused bovine satellite cell cultures.

    Science.gov (United States)

    Thornton, K J; Kamanga-Sollo, E; White, M E; Dayton, W R

    2016-06-01

    Trenbolone acetate (TBA), a testosterone analog, increases protein synthesis and decreases protein degradation in fused bovine satellite cell (BSC) cultures. However, the mechanism through which TBA alters these processes remains unknown. Recent studies indicate that androgens improve rate and extent of muscle growth through a nongenomic mechanism involving G protein-coupled receptors (GPCR), matrix metalloproteinases (MMP), heparin-binding epidermal growth factor (hbEGF), the epidermal growth factor receptor (EGFR), erbB2, and the insulin-like growth factor-1 receptor (IGF-1R). We hypothesized that TBA activates GPCR, resulting in activation of MMP2/9 that releases hbEGF, which activates the EGFR and/or erbB2. To determine whether the proposed nongenomic pathway is involved in TBA-mediated alterations in protein turnover, fused BSC cultures were treated with TBA in the presence or absence of inhibitors for GPCR, MMP2/9, hbEGF, EGFR, erbB2, or IGF-1R, and resultant protein synthesis and degradation rates were analyzed. Assays were replicated at least 9 times for each inhibitor experiment utilizing BSC cultures obtained from at least 3 different steers that had no previous exposure to steroid compounds. As expected, fused BSC cultures treated with 10 n TBA exhibited increased ( BSC cultures with 10 n TBA in the presence of inhibitors for GPCR, MMP2/9, hbEGF, EGFR, erbB2, or IGF-1R suppressed ( 0.05) effect on TBA-mediated decreases in protein degradation. However, inhibition of both EGFR and erbB2 in the presence of 10 n TBA resulted in decreased ( BSC cultures treated with 10 n TBA exhibit increased ( BSC cultures.

  10. Safety and Activity of the First-in-Class Sym004 Anti-EGFR Antibody Mixture in Patients with Refractory Colorectal Cancer

    NARCIS (Netherlands)

    Dienstmann, R.; Patnaik, A.; Garcia-Carbonero, R.; Cervantes, A.; Benavent, M.; Rosello, S.; Tops, B.B.J.; Post, R.S. van der; Argiles, G.; Skartved, N.J.; Hansen, U.H.; Hald, R.; Pedersen, M.W.; Kragh, M.; Horak, I.D.; Braun, S.; Cutsem, E. Van; Tolcher, A.W.; Tabernero, J.

    2015-01-01

    Tumor growth in the context of EGFR inhibitor resistance may remain EGFR-dependent and is mediated by mechanisms including compensatory ligand upregulation and de novo gene alterations. Sym004 is a two-antibody mixture targeting nonoverlapping EGFR epitopes. In preclinical models, Sym004 causes sign

  11. Monitoring of Circulating Tumor Cells and Their Expression of EGFR/Phospho-EGFR During Combined Radiotherapy Regimens in Locally Advanced Squamous Cell Carcinoma of the Head and Neck

    Energy Technology Data Exchange (ETDEWEB)

    Tinhofer, Ingeborg, E-mail: ingeborg.tinhofer@charite.de [Translational Radiooncology Laboratory, Department of Radiooncology and Radiotherapy, Charite Campus Mitte, Charite Universitaetsmedizin Berlin, Berlin (Germany); Hristozova, Tsvetana; Stromberger, Carmen [Translational Radiooncology Laboratory, Department of Radiooncology and Radiotherapy, Charite Campus Mitte, Charite Universitaetsmedizin Berlin, Berlin (Germany); KeilhoIz, Ulrich [Department of Hematology and Oncology, Campus Benjamin Franklin, Charite Universitaetsmedizin Berlin, Berlin (Germany); Budach, Volker [Translational Radiooncology Laboratory, Department of Radiooncology and Radiotherapy, Charite Campus Mitte, Charite Universitaetsmedizin Berlin, Berlin (Germany)

    2012-08-01

    Purpose: The numbers of circulating tumor cells (CTCs) and their expression/activation of epidermal growth factor receptor (EGFR) during the course of combined chemo- or bioradiotherapy regimens as potential biomarkers of treatment efficacy in squamous cell carcinoma of the head and neck (SCCHN) were determined. Methods and Materials: Peripheral blood samples from SCCHN patients with locally advanced stage IVA/B disease who were treated with concurrent radiochemotherapy or induction chemotherapy followed by bioradiation with cetuximab were included in this study. Using flow cytometry, the absolute number of CTCs per defined blood volume as well as their expression of EGFR and its phosphorylated form (pEGFR) during the course of treatment were assessed. Results: Before treatment, we detected {>=}1 CTC per 3.75 mL blood in 9 of 31 patients (29%). Basal expression of EGFR was detected in 100% and pEGFR in 55% of the CTC+ cases. The frequency of CTC detection was not influenced by induction chemotherapy. However, the number of CTC+ samples significantly increased after radiotherapy. This radiation-induced increase in CTC numbers was less pronounced when radiotherapy was combined with cetuximab compared to its combination with cisplatin/5-fluorouracil. The former treatment regimen was also more effective in reducing pEGFR expression in CTCs. Conclusions: Definitive radiotherapy regimens of locally advanced SCCHN can increase the number of CTCs and might thus contribute to a systemic spread of tumor cells. Further studies are needed to evaluate the predictive value of the radiation-induced increase in CTC numbers and the persistent activation of the EGFR signalling pathway in individual CTC+ cases.

  12. Bufalin Reverses HGF-Induced Resistance to EGFR-TKIs in EGFR Mutant Lung Cancer Cells via Blockage of Met/PI3k/Akt Pathway and Induction of Apoptosis

    Directory of Open Access Journals (Sweden)

    Xiao-Hong Kang

    2013-01-01

    Full Text Available The epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs, such as gefitinib and erlotinib, have shown promising therapeutic efficacy in nonsmall cell lung cancer (NSCLC patients harboring epidermal growth factor receptor- (EGFR- activating mutation. However, the inevitable recurrence resulting from acquired resistance has limited the clinical improvement in therapy outcomes. Many studies demonstrate that hepatocyte growth factor- (HGF- Met axis plays an important role in tumor progression and drug sensitivity. HGF may induce resistance to EGFR-TKIs in EGFR mutant lung cancer cells by Met/PI3K/Akt signaling. The purpose of this study was to determine whether bufalin, a major bioactive component of Venenum Bufonis, could reverse HGF-induced resistance to reversible and irreversible EGFR-TKIs in mutant lung cancer cells PC-9, HCC827, and H1975. Our studies showed that bufalin could reverse resistance to reversible and irreversible EGFR-TKIs induced by exogenous HGF in EGFR mutant lung cancer cells by inhibiting the Met/PI3K/Akt pathway and inducing death signaling. These results suggested that bufalin might have a potential to overcome HGF-induced resistance to molecular-targeted drugs for lung cancer.

  13. Nimotuzumab enhances temozolomide-induced growth suppression of glioma cells expressing mutant EGFR in vivo.

    Science.gov (United States)

    Nitta, Yusuke; Shimizu, Saki; Shishido-Hara, Yukiko; Suzuki, Kaori; Shiokawa, Yoshiaki; Nagane, Motoo

    2016-03-01

    A mutant form of epidermal growth factor receptor (EGFR), EGFRvIII, is common in glioblastoma (GBM) and confers enhanced tumorigenic activity and drug resistance. Nimotuzumab, an anti-EGFR antibody, has shown preclinical and clinical activity to GBM, but its specific activity against EGFRvIII has not been fully investigated. Human glioma U87MG or LNZ308 cells overexpressing either wild-type (wt) EGFR or EGFRvIII were treated with nimotuzumab, temozolomide, or both. Expression and phosphorylation status of molecules were determined by Western blot analysis. Methylation status of promoter region of O(6) -methylguanine-DNA methyltransferase (MGMT) was detected by methylation-specific PCR. Antitumor activity was tested using nude mice bearing either subcutaneous or intracerebral xenografts along with analyses of EGFR phosphorylation status, proliferation, apoptosis, and vessel density. Nimotuzumab treatment resulted in reduction of EGFRvIII tyrosine phosphorylation with a decrease in Akt phosphorylation that was greater than that of wtEGFR. Correspondingly, antitumor effects, growth suppression and survival elongation, were more significant in mice bearing either subcutaneous or intracerebral tumor expressing EGFRvIII than in those expressing wtEGFR. These effects were markedly increased when temozolomide was combined with nimotuzumab. The post-treatment recurrent brain tumors exhibited a decrease in expression of the mismatch repair (MMR) proteins, MSH6 and MLH1, but their methylated MGMT status did not changed. Nimotuzumab has in vivo antitumor activity against GBM, especially those expressing EGFRvIII, when combined with temozolomide. This could provide a basis for preselection of patients with GBM by EGFR status who might benefit from the nimotuzumab and temozolomide combination therapy. PMID:26778701

  14. Current Approaches for Predicting a Lack of Response to Anti-EGFR Therapy in KRAS Wild-Type Patients

    Directory of Open Access Journals (Sweden)

    Tze-Kiong Er

    2014-01-01

    Full Text Available Targeting epidermal growth factor receptor (EGFR has been one of the most effective colorectal cancer strategies. Anti-EGFR antibodies function by binding to the extracellular domain of EGFR, preventing its activation, and ultimately providing clinical benefit. KRAS mutations in codons 12 and 13 are recognized prognostic and predictive biomarkers that should be analyzed at the clinic prior to the administration of anti-EGFR therapy. However, still an important fraction of KRAS wild-type patients do not respond to the treatment. The identification of additional genetic determinants of primary or secondary resistance to EGFR targeted therapy for further improving the selection of patients is urgent. Herein, we review the latest published literature highlighting the most important genes that may predict resistance to anti-EGFR monoclonal antibodies in colorectal cancer patients. According to the available findings, the evaluation of BRAF, NRAS, PIK3CA, and PTEN status could be the right strategy to select patients who are likely to respond to anti-EGFR therapies. In the future, the combination of those biomarkers will help establish consensus that can be introduced into clinical practice.

  15. A CLONALLY DERIVED CELL LINE,9L-EGFR IS USEFUL FOR THE STUDIES OF CANCER CELLS BEARING EGF RECEPTOR

    Institute of Scientific and Technical Information of China (English)

    Lin Qi; Rajesh Agarwal; Rana Singh; Gail S. Harrisona; L.Michael Glodea

    2003-01-01

    Since the epidermal growth factor receptor (EGFR) is a key regulator in cell signaling pathways of cancer cell. To investigate the mechanism between cancer cells survival and its EGFR expression, drug selection of cancer cells target therapy, we generated a cell line, 9L-EGFR, which stably expressed human EGFR; the parental rat glioma cell line, 9L, does not contain endogenous EGFR message or protein. Our results show that 9L-EGFR cells had high levels of EGFR on their cell surface by using RT-PCR, Western analysis and Flow cytometry analysis. The EGFR transfected into 9L cells was capable of being activated by EGF, in which either phosphorylated (p-EGFR) or total (EGFR) was showed by Western blot. This investigation may contribute to the further studies of cancer cells bearing EGFR.

  16. Molecular spectrum of somatic EGFR and KRAS gene mutations in non small cell lung carcinoma: determination of frequency, distribution pattern and identification of novel variations in Indian patients.

    Science.gov (United States)

    Das, Bibhu Ranjan; Bhaumik, Sangeet; Ahmad, Firoz; Mandsaurwala, Aziz; Satam, Heena

    2015-07-01

    Somatic mutations of EGFR and KRAS gene represent the most common alterations currently known in NSCLC patients. This study explored the frequency, distribution pattern of EGFR and KRAS mutations in Indian patients. The frequencies of EGFR and KRAS mutations were 29 % (116/400) and 4.5 % (6/132) respectively. Both EGFR and KRAS mutations were prevalent in females, and a trend towards higher mutation frequency was seen in patients under ≥ 60 years age. The presence of EGFR and KRAS mutations were higher in adenocarcinomas in comparison to other histological subtype. Sequencing analysis of EGFR exon 18 revealed Inframe deletion (G709_T710 > A) and missense mutation (K713R). Among exon 19 positive cases, 49.3 % (37/75) were in-frame deletions, of which E746_A750del was frequent. Similarly, ~47 % (35/75) cases showed complex mutation involving indel. Among mutations in exon 20 (N = 9), 8 were substitutions, one showed duplication, while all exon 21 mutations were of the missense types with L858R as the most recurrent type. Sequencing analysis of KRAS exon 1 revealed three different types codon 12 substitutions resulting in c34G > T (G12C) (n = 4), c.35G > A (G12D) (n = 1), and c.35G > T (G12V) (n = 1). In conclusion, the present study is an example of molecular diversity of EGFR and KRAS gene in Indian patients and further confirms that the frequency of EGFR and KRAS mutations varies considerably globally. To the best of our knowledge, this is the first Indian study to evaluate KRAS mutation. The current study also served to identify novel variations that added new insights into the genetic heterogeneity of NSCLC.

  17. Analyzing the Sensitivity of EGFR-L861Q Mutation to TKIs and A Case Report

    Directory of Open Access Journals (Sweden)

    Xingxing WANG

    2015-09-01

    Full Text Available Background and objective The significant efficacy of tyrosine kinase inhibitors (TKIs has been approved for advanced non-small cell lung cancer (NSCLC patients with activating epidermal growth factor receptor (EGFR mutations. No clear evidence exists that EGFR-L861Q is sensitive to TKIs, and the best treatment for NSCLC patients with EGFR-L861Q mutation is undetermined. This study aims to discuss the best treatment for advanced NSCLC patients with EGFR-L861Q mutation by analyzing the differences among the structures of wild-type EGFR, activating mutant EGFR-L858R, and EGFR-L861Q mutation. Method The protein structures of wild-type EGFR were reconstructed. EGFR-L858R and EGFR-L861Q mutation were activated. The differences among the three kinds of protein conformation were analyzed using homologous modeling technique. Results The structure of EGFR-L858R and wild-type EGFR exhibited notable distinctions. The structure of EGFR-L861Q mutation was different compared with wild-type EGFR and activating mutant EGFR-L858R protein conformations. NSCLC patients with EGFR-L861Q mutation were given chemotherapy as the first-line of therapy, and TKIs were applied to maintain treatment when the tumor is unchanged. Effect evaluation result was improved when the lung computed tomography lesions were reviewed. Conclusion The analysis of the protein conformation of EGFR-L861Q mutation and the curative effect of chemotherapy with TKIs could help predict the sensitivity of EGFR-L861Q to TKIs. Combining the analysis with a clinical case, maintenance treatment with TKIs may achieve satisfactory curative effect in advanced NSCLC patients who have achieved disease control after first-line chemotherapy.

  18. Radioresistance of human glioma spheroids and expression of HSP70, p53 and EGFr

    Directory of Open Access Journals (Sweden)

    Fedrigo Carlos A

    2011-11-01

    Full Text Available Abstract Background Radiation therapy is routinely prescribed for high-grade malignant gliomas. However, the efficacy of this therapeutic modality is often limited by the occurrence of radioresistance, reflected as a diminished susceptibility of the irradiated cells to undergo cell death. Thus, cells have evolved an elegant system in response to ionizing radiation induced DNA damage, where p53, Hsp70 and/or EGFr may play an important role in the process. In the present study, we investigated whether the content of p53, Hsp70 and EGFr are associated to glioblastoma (GBM cell radioresistance. Methods Spheroids from U-87MG and MO59J cell lines as well as spheroids derived from primary culture of tumor tissue of one GBM patient (UGBM1 were irradiated (5, 10 and 20 Gy, their relative radioresistance were established and the p53, Hsp70 and EGFr contents were immunohistochemically determined. Moreover, we investigated whether EGFr-phospho-Akt and EGFr-MEK-ERK pathways can induce GBM radioresistance using inhibitors of activation of ERK (PD098059 and Akt (wortmannin. Results At 5 Gy irradiation UGBM1 and U-87MG spheroids showed growth inhibition whereas the MO59J spheroid was relatively radioresistant. Overall, no significant changes in p53 and Hsp70 expression were found following 5 Gy irradiation treatment in all spheroids studied. The only difference observed in Hsp70 content was the periphery distribution in MO59J spheroids. However, 5 Gy treatment induced a significant increase on the EGFr levels in MO59J spheroids. Furthermore, treatment with inhibitors of activation of ERK (PD098059 and Akt (wortmannin leads to radiosensitization of MO59J spheroids. Conclusions These results indicate that the PI3K-Akt and MEK-ERK pathways triggered by EGFr confer GBM radioresistance.

  19. EGFR- and AKT-mediated reduction in PTEN expression contributes to tyrphostin resistance and is reversed by mTOR inhibition in endometrial cancer cells.

    Science.gov (United States)

    Li, Tian; Yang, Yuebo; Li, Xiaomao; Xu, Chengfang; Meng, Lirong

    2012-02-01

    Loss or mutation of the PTEN (phosphatase and tensin homologue deleted on chromosome 10) gene is associated with resistance to epidermal growth factor receptor (EGFR) inhibitors. However, the mechanism underlying remains elusive. In this study, we aimed to explore whether sensitivity to the EGFR tyrosine kinase inhibitor (TKI) is affected by PTEN status in endometrial cancer cells. PTEN siRNA and the PTEN gene were transfected into HEC-1A and Ishikawa endometrial cancer cells using lentiviral vectors. Cells were treated under various concentrations of RG14620 and rapamycin, which are EGFR and mammalian target of rapamycin (mTOR) inhibitors, respectively. The IC(50) of RG16420 was determined by using the MTT method. Cell apoptosis and the cell cycle were studied, and activation of EGFR, AKT, and p70S6 were detected by Western blot analysis. Loss of PTEN promoted cell proliferation and led to significant increases in the levels of EGFR, phospho-EGFR, AKT, phospho-AKT, and phospho-mTOR proteins. Ishikawa and HEC-1A(PTENkd) cells that displayed loss and inactivation of PTEN function were resistant to RG14620. HEC-1A and Ishikawa(PTEN) cells with intact PTEN were sensitive to RG14620. The combination of two inhibitors was more effective than both monotherapies, particularly in carcinoma cells with PTEN dysfunction. Decreased phospho-EGFR protein expression was observed in all cell lines that were sensitive to RG14620. Decreased phospho-AKT and phospho-p70S6 protein expression was observed in PTEN-intact cells that were sensitive to RG14620. PTEN loss results in resistance to EGFR TKI, which was reversed by PTEN reintroduction or mTOR inhibitor treatment. The combined treatment of EGFR TKI and the mTOR inhibitor provided a synergistic effect by promoting cell death in PTEN-deficient and PTEN-intact endometrial cancer cells, particularly in PTEN-deficient carcinoma cells with up-regulated EGFR activation.

  20. Activation of platelet-activating factor receptor and pleiotropic effects on tyrosine phospho-EGFR/Src/FAK/paxillin in ovarian cancer.

    Science.gov (United States)

    Aponte, Margarita; Jiang, Wei; Lakkis, Montaha; Li, Ming-Jiang; Edwards, Dale; Albitar, Lina; Vitonis, Allison; Mok, Samuel C; Cramer, Daniel W; Ye, Bin

    2008-07-15

    Among the proinflammatory mediators, platelet-activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphorylcholine) is a major primary and secondary messenger involved in intracellular and extracellular communication. Evidence suggests that PAF plays a significant role in oncogenic transformation, tumor growth, angiogenesis, and metastasis. However, PAF, with its receptor (PAFR) and their downstream signaling targets, has not been thoroughly studied in cancer. Here, we characterized the PAFR expression pattern in 4 normal human ovarian surface epithelial (HOSE) cell lines, 13 ovarian cancer cell lines, paraffin blocks (n = 84), and tissue microarrays (n = 230) from patients with ovarian cancer. Overexpression of PAFR was found in most nonmucinous types of ovarian cancer but not in HOSE and mucinous cancer cells. Correspondingly, PAF significantly induced cell proliferation and invasion only in PAFR-positive cells (i.e., OVCA429 and OVCA432), but not in PAFR-negative ovarian cells (HOSE and mucinous RMUG-L). The dependency of cell proliferation and invasion on PAFR was further confirmed using PAFR-specific small interfering RNA gene silencing probes, antibodies against PAFR and PAFR antagonist, ginkgolide B. Using quantitative multiplex phospho-antibody array technology, we found that tyrosine phosphorylation of EGFR/Src/FAK/paxillin was coordinately activated by PAF treatment, which was correlated with the activation of phosphatidylinositol 3-kinase and cyclin D1 as markers for cell proliferation, as well as matrix metalloproteinase 2 and 9 for invasion. Specific tyrosine Src inhibitor (PP2) reversibly blocked PAF-activated cancer cell proliferation and invasion. We suggest that PAFR is an essential upstream target of Src and other signal pathways to control the PAF-mediated cancer progression.

  1. 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...... tyrosine kinase inhibitors (EGFR-TKIs). The EGFR-TKI gefitinib has been approved in Europe for the treatment of adult patients with locally advanced or metastatic NSCLC with activating mutations of the EGFR TK. Because EGFR mutation testing is not yet well established across Europe, biomarker......-directed therapy only slowly emerges for the subset of NSCLC patients most likely to benefit: those with EGFR mutations....

  2. LL-37 via EGFR transactivation to promote high glucose-attenuated epithelial wound healing in organ-cultured corneas.

    Science.gov (United States)

    Yin, Jia; Yu, Fu-Shin X

    2010-04-01

    Purpose. Patients with diabetes are at higher risk for delayed corneal reepithelialization and infection. Previous studies indicated that high glucose (HG) impairs epidermal growth factor receptor (EGFR) signaling and attenuates ex vivo corneal epithelial wound healing. The authors investigated the effects of antimicrobial peptide LL-37 on HG-attenuated corneal epithelial EGFR signaling and wound closure. Methods. Human corneal epithelial cells (HCECs) were stimulated with LL-37. Heparin-binding EGF-like growth factor (HB-EGF) shedding was assessed by measuring the release of alkaline phosphatase (AP) in a stable HCEC line expressing HB-EGF-AP. Activation of EGFR, phosphoinositide 3-kinase (PI3K), and extracellular signal-regulated kinases 1/2 (ERK1/2) was determined by Western blot analysis. Corneal epithelial wound closure was assessed in cultured HCECs and porcine corneas. LL-37 expression was determined by immune dot blot. Results. LL-37 induced HB-EGF-AP release and EGFR activation in a dose-dependent manner. LL-37 prolonged EGFR signaling in response to wounding. LL-37 enhanced the closure of a scratch wound in cultured HCECs and partially rescued HG-attenuated wound healing in an EGFR- and a PI3K-dependent manner and restored HG-impaired EGFR signaling in cultured porcine corneas. HG attenuated wounding-induced LL-37 expression in cultured HCECs. Conclusions. LL-37 is a tonic factor promoting EGFR signaling and enhancing epithelial wound healing in normal and high glucose conditions. With both antimicrobial and regenerative capabilities, LL-37 may be a potential therapeutic for diabetic keratopathy.

  3. IGFBP2 potentiates nuclear EGFR-STAT3 signaling.

    Science.gov (United States)

    Chua, C Y; Liu, Y; Granberg, K J; Hu, L; Haapasalo, H; Annala, M J; Cogdell, D E; Verploegen, M; Moore, L M; Fuller, G N; Nykter, M; Cavenee, W K; Zhang, W

    2016-02-11

    Insulin-like growth factor binding protein 2 (IGFBP2) is a pleiotropic oncogenic protein that has both extracellular and intracellular functions. Despite a clear causal role in cancer development, the tumor-promoting mechanisms of IGFBP2 are poorly understood. The contributions of intracellular IGFBP2 to tumor development and progression are also unclear. Here we present evidence that both exogenous IGFBP2 treatment and cellular IGFBP2 overexpression lead to aberrant activation of epidermal growth factor receptor (EGFR), which subsequently activates signal transducer and activator of transcription factor 3 (STAT3) signaling. Furthermore, we demonstrate that IGFBP2 augments the nuclear accumulation of EGFR to potentiate STAT3 transactivation activities, via activation of the nuclear EGFR signaling pathway. Nuclear IGFBP2 directly influences the invasive and migratory capacities of human glioblastoma cells, providing a direct link between intracellular (and particularly nuclear) IGFBP2 and cancer hallmarks. These activities are also consistent with the strong association between IGFBP2 and STAT3-activated genes derived from The Cancer Genome Atlas database for human glioma. A high level of all three proteins (IGFBP2, EGFR and STAT3) was strongly correlated with poorer survival in an independent patient data set. These results identify a novel tumor-promoting function for IGFBP2 of activating EGFR/STAT3 signaling and facilitating EGFR accumulation in the nucleus, thereby deregulating EGFR signaling by two distinct mechanisms. As targeting EGFR in glioma has been relatively unsuccessful, this study suggests that IGFBP2 may be a novel therapeutic target.

  4. EGFR/cell membrane chromatography-online-high performance liquid chromatography/mass spectrometry method for screening EGFR antagonists from Radix Angelicae Pubescentis

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    The intracellular kinase domains of the epidermal growth factor receptor(EGFR) in some tumor cells are significant targets for drug discovery.We have developed a new EGFR cell membrane chromatography(EGFR/CMC)-online-high performance liquid chromatography/mass spectrometry(HPLC/MS) method for screening anti-EGFR antagonists from medicinal herbs such as Radix Angelicae Pubescentis.In this study,the HEK293 EGFR cells with high expression of EGFR were used to prepare cell membrane stationary phase(CMSP) in the EGFR/CMC model.The retention fractions on the EGFR/CMC model were directly analyzed by combining a 10 port columns switcher with a HPLC/MS system online.As a result,osthole from Radix Angelicae Pubescentis was found to be the active component acting on EGFR like dasatinib as the control drug.There was a good relationship between their inhibiting effects on EGFR secretion and HEK293 EGFR cell growth in vitro.This new EGFR/CMC-online-HPLC/MS method can be applied for screening anti-EGFR antagonists from TCMs,for instance,Radix Angelicae Pubescentis.It will be a useful method for drug discovery with natural medicinal herbs as a leading compound resource.

  5. Rational combination of targeted therapies as a strategy to overcome the mechanisms of resistance to inhibitors of EGFR signaling.

    Science.gov (United States)

    Bianco, Roberto; Damiano, Vincenzo; Gelardi, Teresa; Daniele, Gennaro; Ciardiello, Fortunato; Tortora, Giampaolo

    2007-01-01

    The epidermal growth factor receptor (EGFR) has been widely used as a target for novel anticancer agents, such as blocking antibodies and small molecular weight tyrosine kinase compounds. In spite of recent advances in cancer cell biology, leading to the introduction of clinically active new drugs, such as cetuximab, panitumumab and erlotinib, unfortunately disease control remains unsuccessful due to the presence of constitutive resistance to EGFR inhibitors in most patients and the development of acquired resistance in the responders. A large number of molecular abnormalities in tumor cells seem to partly contribute to their resistance to anti-EGFR therapy: increased angiogenesis, constitutive activation of downstream mediators, overexpression of other tyrosine kinase receptors. Moreover, some mutations in the EGFR receptor kinase domain seem to play a crucial role in determining the sensitivity of cancer cells to specific inhibitors by altering the conformation of the receptor and its activity. The development of rational combinations of anticancer agents and EGFR inhibitors, able to exert synergistic cytotoxic interactions, has been widely accepted and used in both preclinical and clinical studies. Although the failure of large clinical trial based on empirical combination of anti-EGFR and classic chemotherapeutic agents, several preclinical data seems to support the hypothesis that combining EGFR inhibitors and other novel agents could efficiently inhibit tumor growth and overcome intrinsic resistance to a single-agent based therapy. This review focuses on the role of complementary signalling pathways in the development of resistance to EGFR targeting agents and the rationale to combine novel inhibitors as anticancer therapy.

  6. No Incidence of BRAF Mutations in Salivary Gland Carcinomas—Implications for Anti-EGFR Therapies

    Directory of Open Access Journals (Sweden)

    Regine Dahse

    2009-01-01

    These findings imply that SGC rarely acquires mutations that result in a constitutive activation of the signaling cascade downstream of EGFR and this pleads in favor of further therapeutic trials with EGFR-targeting monoclonal antibodies.

  7. IGFBP2 potentiates nuclear EGFR-STAT3 signaling

    Science.gov (United States)

    Chua, Corrine Yingxuan; Liu, Yuexin; Granberg, Kirsi J.; Hu, Limei; Haapasalo, Hannu; Annala, Matti J.; Cogdell, David E.; Verploegen, Maartje; Moore, Lynette M.; Fuller, Gregory N.; Nykter, Matti; Cavenee, Webster K.; Zhang, Wei

    2015-01-01

    Insulin-like growth factor binding protein 2 (IGFBP2) is a pleiotropic oncogenic protein that has both extracellular and intracellular functions. Despite a clear causal role in cancer development, the tumor-promoting mechanisms of IGFBP2 are poorly understood. The contributions of intracellular IGFBP2 to tumor development and progression are also unclear. Here we present evidence that both exogenous IGFBP2 treatment and cellular IGFBP2 overexpression lead to aberrant activation of EGFR, which subsequently activates STAT3 signaling. Furthermore, we demonstrate that IGFBP2 augments the nuclear accumulation of EGFR to potentiate STAT3 transactivation activities, via activation of the nuclear EGFR signaling pathway. Nuclear IGFBP2 directly influences the invasive and migratory capacities of human glioblastoma cells, providing a direct link between intracellular (and particularly nuclear) IGFBP2 and cancer hallmarks. These activities are also consistent with the strong association between IGFBP2 and STAT3-activated genes derived from the TCGA database for human glioma. A high level of all 3 proteins (IGFBP2, EGFR and STAT3) was strongly correlated with poorer survival in an independent patient dataset. These results identify a novel tumor-promoting function for IGFBP2 of activating EGFR/STAT3 signaling and facilitating EGFR accumulation in the nucleus, thereby deregulating EGFR signaling by 2 distinct mechanisms. As targeting EGFR in glioma has been relatively unsuccessful, this study suggests that IGFBP2 may be a novel therapeutic target. PMID:25893308

  8. SKLB-287, a novel oral multikinase inhibitor of EGFR and VEGFR2, exhibits potent antitumor activity in LoVo colorectal tumor model.

    Science.gov (United States)

    Chen, X; Liu, Y; Yang, H-W; Zhou, S; Cheng, C; Zheng, M-W; Zhong, L; Fu, X-Y; Pan, Y-L; Ma, S; Tang, Y; Chen, Y-Z; Li, L-L; Yang, S-Y

    2014-01-01

    Colorectal cancer (CRC) is the third common cancer and most of the chemotherapies of CRC currently used often suffer limited efficacy and large side effects. Targeted small-molecule by anti-tumor drugs are thought a promising strategy for improving the efficacy and reducing the side effects. In this investigation, we report a novel multikinase inhibitor, termed SKLB-287, which was discovered by us recently. SKLB-287 could efficiently inhibit the activation of endothelial growth factor receptor (EGFR) and vascular endothelial growth factor receptor 2 (VEGFR2). It displayed very good anti-proliferative activity against LoVo CRC cells and considerable antiangiogenic potency in transgenic zebrafish embryos. Oral administration of SKLB-287 resulted in dose-dependent suppression of tumor growth in LoVo xenograft mouse model. Immunohistochemistry was adopted to examine the in vivo anti-tumor mechanism of action of SKLB-287.

  9. Ibrutinib selectively and irreversibly targets EGFR (L858R, Del19) mutant but is moderately resistant to EGFR (T790M) mutant NSCLC Cells

    OpenAIRE

    Wu, Hong; Wang, Aoli; Zhang, Wei; Wang, Beilei; Chen, Cheng; Wang, Wenchao; Hu, Chen; Ye, Zi; Zhao, Zheng; Wang, Li; Li, Xixiang; Yu, Kailin; Liu, Juan; Wu, Jiaxin; Yan, Xiao-E

    2015-01-01

    Through comprehensive comparison study, we found that ibrutinib, a clinically approved covalent BTK kinase inhibitor, was highly active against EGFR (L858R, del19) mutant driven NSCLC cells, but moderately active to the T790M ‘gatekeeper’ mutant cells and not active to wild-type EGFR NSCLC cells. Ibrutinib strongly affected EGFR mediated signaling pathways and induced apoptosis and cell cycle arrest (G0/G1) in mutant EGFR but not wt EGFR cells. However, ibrutinib only slowed down tumor progre...

  10. 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. PMID:25789707

  11. Silica nanoparticles induce cytokine responses in lung epithelial cells through activation of a p38/TACE/TGF-α/EGFR-pathway and NF-κΒ signalling

    Energy Technology Data Exchange (ETDEWEB)

    Skuland, Tonje, E-mail: tonje.skuland@fhi.no; Øvrevik, Johan; Låg, Marit; Schwarze, Per; Refsnes, Magne

    2014-08-15

    Amorphous silica nanoparticles (SiNPs) have previously been shown to induce marked cytokine (interleukin-6; IL-6 and interleukin-8; CXCL8/IL-8) responses independently of particle uptake in human bronchial epithelial BEAS-2B cells. In this study the involvement of the mitogen-activated protein kinases (MAP-kinases), nuclear factor-kappa Β (NF-κΒ) and in particular tumour necrosis factor-α converting enzyme (TACE) and—epidermal growth factor receptor (EGFR) signalling pathways were examined in triggering of IL-6 and CXCL8 release after exposure to a 50 nm silica nanoparticle (Si50). Exposure to Si50 increased phosphorylation of NF-κΒ p65 and MAP-kinases p38 and JUN-N-terminal protein kinase pathways (JNK), but not extracellular signal regulated kinases (ERK). Inhibition of NF-κΒ and p38 reduced the cytokine responses to Si50, whereas neither JNK- nor ERK-inhibition exerted any significant effect on the responses to Si50. Increases in membrane-bound transforming growth factor-α (TGF-α) release and EGFR phosphorylation were also observed after Si50 exposure, and pre-treatment with inhibitors of these pathways reduced the release of IL-6 and CXCL8, but did not affect the Si50-induced phosphorylation of p38 and p65. In contrast, p38-inhibition partially reduced Si50-induced TGF-α release, while the p65-inhibition was without effect. Overall, our results indicate that Si50-induced IL-6 and CXCL8 responses in BEAS-2B cells were regulated through combined activation of several pathways, including NF-κΒ and p38/TACE/TGF-α/EGFR signalling. The study identifies critical, initial events in the triggering of pro-inflammatory responses by nanoparticles. - Highlights: • Silica nanoparticles induce IL-6 and CXCL8 via NFκB and MAPKinase p38 in BEAS-2B • Silica nanoparticles induce release of the EGF-receptor ligand TGF-α • TGF-α release contributes to the IL-6 and CXCL8 release • Phosphorylation of p38 is involved in release of TGF-α.

  12. Nuclear EGFR as a molecular target in cancer

    International Nuclear Information System (INIS)

    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

  13. Nuclear trafficking of EGFR by Vps34 represses Arf expression to promote lung tumor cell survival.

    Science.gov (United States)

    Dayde, D; Guerard, M; Perron, P; Hatat, A-S; Barrial, C; Eymin, B; Gazzeri, S

    2016-07-28

    Epidermal growth factor receptor (EGFR) is a cell surface receptor that has an essential role in cell proliferation and survival, and overexpression of EGFR is a common feature of human cancers. In Non-small-cell lung cancer (NSCLC), activating mutations of EGFR have also been described. We recently showed that mutant EGFR-L858R inhibits the expression of the p14ARF tumor-suppressor protein to promote cell survival. In this study, we defined the molecular bases by which EGFR controls Arf expression. Using various lung tumor models, we showed that EGF stimulation inhibits Arf transcription by a mechanism involving the nuclear transport and recruitment of EGFR to the Arf promoter. We unraveled the vesicular trafficking protein Vps34 as a mediator of EGFR nuclear trafficking and showed that its neutralization prevents the accumulation of EGFR to the Arf promoter in response to ligand activation. Finally, in lung tumor cells that carry mutant EGFR-L858R, we demonstrated that inhibition of Vps34 using small interfering RNA restrains nuclear EGFR location and restores Arf expression leading to apoptosis. These findings identify the Arf tumor suppressor as a new transcriptional target of nuclear EGFR and highlight Vps34 as an important regulator of the nuclear EGFR/Arf survival pathway. As a whole, they provide a mechanistic explanation to the inverse correlation between nuclear expression of EGFR and overall survival in NSCLC patients. PMID:26686095

  14. DNA methylation down-regulates EGFR expression in chicken

    Science.gov (United States)

    The epidermal growth factor receptor (EGFR), a growth-factor-receptor tyrosine kinase, was found up-regulated in numerous tumors, which provides a good target for cancer therapy. Although it was documented that oncoviruses are responsible for the activation of EGFR in tumors, the impact of Marek’s d...

  15. Agonist-induced activation of histamine H3 receptor signals to extracellular signal-regulated kinases 1 and 2 through PKC-, PLD-, and EGFR-dependent mechanisms.

    Science.gov (United States)

    Lai, Xiangru; Ye, Lingyan; Liao, Yuan; Jin, Lili; Ma, Qiang; Lu, Bing; Sun, Yi; Shi, Ying; Zhou, Naiming

    2016-04-01

    The histamine H3 receptor (H3R), abundantly expressed in the central and the peripheral nervous system, has been recognized as a promising target for the treatment of various important CNS diseases including narcolepsy, Alzheimer's disease, and attention deficit hyperactivity disorder. The H3R acts via Gi/o -proteins to inhibit adenylate cyclase activity and modulate MAPK activity. However, the underlying molecular mechanisms for H3R mediation of the activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) remain to be elucidated. In this study, using HEK293 cells stably expressing human H3R and mouse primary cortical neurons endogenously expressing mouse H3R, we found that the H3R-mediated activation of ERK1/2 was significantly blocked by both the pertussis toxin and the MEK1/2 inhibitor U0126. Upon stimulation by H3R agonist histamine or imetit, H3R was shown to rapidly induce ERK1/2 phosphorylation via PLC/PKC-, PLDs-, and epidermal growth factor receptor (EGFR) transactivation-dependent pathways. Furthermore, it was also indicated that while the βγ-subunits play a key role in H3R-activated ERK1/2 phosphorylation, β-arrestins were not required for ERK1/2 activation. In addition, when the cultured mouse cortical neurons were exposed to oxygen and glucose deprivation conditions (OGD), imetit exhibited neuroprotective properties through the H3R. Treatment of cells with the inhibitor UO126 abolished these protective effects. This suggests a possible neuroprotective role of the H3R-mediated ERK1/2 pathway under hypoxia conditions. These observations may provide new insights into the pharmacological effects and the physiological functions modulated by the H3R-mediated activation of ERK1/2. Histamine H3 receptors are abundantly expressed in the brain and play important roles in various CNS physiological functions. However, the underlying mechanisms for H3R-induced activation of extracellular signal-regulated kinase (ERK)1/2 remain largely unknown. Here

  16. Neurotensin (NTS) and its receptor (NTSR1) causes EGFR, HER2 and HER3 over-expression and their autocrine/paracrine activation in lung tumors, confirming responsiveness to erlotinib.

    Science.gov (United States)

    Younes, Mohamad; Wu, Zherui; Dupouy, Sandra; Lupo, Audrey Mansuet; Mourra, Najat; Takahashi, Takashi; Fléjou, Jean François; Trédaniel, Jean; Régnard, Jean François; Damotte, Diane; Alifano, Marco; Forgez, Patricia

    2014-09-30

    Alterations in the signaling pathways of epidermal growth factor receptors (HERs) are associated with tumor aggressiveness. Neurotensin (NTS) and its high affinity receptor (NTSR1) are up regulated in 60% of lung cancers. In a previous clinical study, NTSR1 overexpression was shown to predict a poor prognosis for 5 year overall survival in a selected population of stage I lung adenocarcinomas treated by surgery alone. In a second study, shown here, the frequent and high expression of NTSR1 was correlated with a pejorative prognosis in 389 patients with stage I to III lung adenocarcinoma, and was an independent prognosis marker. Interactions between NTS and NTSR1 induce pro-oncogenic biological effects associated with neoplastic processes and tumor progression. Here we highlight the cellular mechanisms activated by Neurotensin (NTS) and its high affinity receptor (NTSR1) contributing to lung cancer cell aggressiveness. We show that the NTS autocrine and/or paracrine regulation causes EGFR, HER2, and HER3 over-expression and activation in lung tumor cells. The EGFR and HER3 autocrine activation is mediated by MMP1 activation and EGF "like" ligands (HB-EGF, Neuregulin 1) release. By establishing autocrine and/or paracrine NTS regulation, we show that tumor growth is modulated according to NTS expression, with a low growth rate in those tumors that do not express NTS. Accordingly, xenografted tumors expressing NTS and NTSR1 showed a positive response to erlotinib, whereas tumors void of NTSR1 expression had no detectable response. This is consistent with the presence of a NTS autocrine loop, leading to the sustained activation of EGFR and responsible for cancer aggressiveness. We propose the use of NTS/NTSR1 tumor expression, as a biomarker for the use of EGFR tyrosine kinase inhibitors in patients lacking EGFR mutation.

  17. Dermatologic Toxicities from Monoclonal Antibodies and Tyrosine Kinase Inhibitors against EGFR: Pathophysiology and Management

    OpenAIRE

    Shaad E. Abdullah; Missak Haigentz; Bilal Piperdi

    2012-01-01

    Epidermal growth factor receptor (EGFR) inhibition has now been well established as an effective treatment for various cancers. The EGFR belongs to the ErbB family of tyrosine kinase receptors which regulate tumor cell differentiation, survival and proliferation. Activation of EGFR drives tumorigenesis in lung, head and neck, colorectal and pancreatic cancers. Irrespective of the type of cancer being treated and the mechanism by which tumor EGFR drives tumorigenesis, the major side effect of ...

  18. Development of Cu-64 labeled EGF for In Vivo PET Imaging of EGFR Expression

    International Nuclear Information System (INIS)

    In this project we proposed to establish feasibility of the development of targeted tracers for radionuclide imaging of epidermal growth factor receptors (EGFR) in cancer patients. The significance and impact of the proposed radiotracers are determined by the crucial role that EGFR plays in many cancers and by the rapid entrance of EGFR-inhibiting drugs into clinic. Clinical experience, however, revealed that only 10-25% of patients that are defined as EGFR-positive by immunohistochemical analysis respond to EGFR-directed therapeutics and there is poor correlation between EGFR immunohistochemistry and treatment. Therefore, for more efficacious use of EGFR-targeting therapeutics, there is a need for information about EGFR activity in patients. We hypothesized that radionuclide imaging of functionally active EGFR will provide such information and would allow for (1) rational patient stratification, (2) rapid monitoring of responses to therapy, and (3) development of personalized treatment regimens. We hypothesized that tracers based epidermal growth factor (EGF), a natural EGFR ligand, as a targeting vector would be particularly advantageous. First, only functionally active and therefore critical for disease progression EGFRs will bind and internalize an EGF-based tracer. Second, continuous internalization of EGF-based tracers by recyclable EGFR would lead to intracellular accumulation of radionuclide and improved signal-to-background ratio. Third, small size of EGF relative to antibodies would facilitate tumor penetration with vastly better non-specific soft tissue and blood clearance rates. Fourth, as a human protein, EGF is not expected to be immunogenic. Finally, at the beginning of this project, we have already engineered and expressed functionally active EGF with an N-terminal Cys-tag for site-specific conjugation of various payloads, including radionuclide chelators. In the Phase I of this project, in collaboration with Dr. Blankenberg's group at Stanford

  19. Development of Cu-64 labeled EGF for In Vivo PET Imaging of EGFR Expression

    Energy Technology Data Exchange (ETDEWEB)

    Backer, Joseph M.

    2009-07-12

    In this project we proposed to establish feasibility of the development of targeted tracers for radionuclide imaging of epidermal growth factor receptors (EGFR) in cancer patients. The significance and impact of the proposed radiotracers are determined by the crucial role that EGFR plays in many cancers and by the rapid entrance of EGFR-inhibiting drugs into clinic. Clinical experience, however, revealed that only 10-25% of patients that are defined as EGFR-positive by immunohistochemical analysis respond to EGFR-directed therapeutics and there is poor correlation between EGFR immunohistochemistry and treatment. Therefore, for more efficacious use of EGFR-targeting therapeutics, there is a need for information about EGFR activity in patients. We hypothesized that radionuclide imaging of functionally active EGFR will provide such information and would allow for 1) rational patient stratification, 2) rapid monitoring of responses to therapy, and 3) development of personalized treatment regimens. We hypothesized that tracers based epidermal growth factor (EGF), a natural EGFR ligand, as a targeting vector would be particularly advantageous. First, only functionally active and therefore critical for disease progression EGFRs will bind and internalize an EGF-based tracer. Second, continuous internalization of EGF-based tracers by recyclable EGFR would lead to intracellular accumulation of radionuclide and improved signal-to-background ratio. Third, small size of EGF relative to antibodies would facilitate tumor penetration with vastly better non-specific soft tissue and blood clearance rates. Fourth, as a human protein, EGF is not expected to be immunogenic. Finally, at the beginning of this project, we have already engineered and expressed functionally active EGF with an N-terminal Cys-tag for site-specific conjugation of various payloads, including radionuclide chelators. In the Phase I of this project, in collaboration with Dr. Blankenberg’s group at Stanford

  20. LDA-SVM-Based EGFR Mutation Model for NSCLC Brain Metastases

    Science.gov (United States)

    Hu, Nan; Wang, Ge; Wu, Yu-Hao; Chen, Shi-Feng; Liu, Guo-Dong; Chen, Chuan; Wang, Dong; He, Zhong-Shi; Yang, Xue-Qin; He, Yong; Xiao, Hua-Liang; Huang, Ding-De; Xiong, Kun-Lin; Wu, Yan; Huang, Ming; Yang, Zhen-Zhou

    2015-01-01

    Abstract Epidermal growth factor receptor (EGFR) activating mutations are a predictor of tyrosine kinase inhibitor effectiveness in the treatment of non–small-cell lung cancer (NSCLC). The objective of this study is to build a model for predicting the EGFR mutation status of brain metastasis in patients with NSCLC. Observation and model set-up. This study was conducted between January 2003 and December 2011 in 6 medical centers in Southwest China. The study included 31 NSCLC patients with brain metastases. Eligibility requirements were histological proof of NSCLC, as well as sufficient quantity of paraffin-embedded lung and brain metastases specimens for EGFR mutation detection. The linear discriminant analysis (LDA) method was used for analyzing the dimensional reduction of clinical features, and a support vector machine (SVM) algorithm was employed to generate an EGFR mutation model for NSCLC brain metastases. Training-testing-validation (3 : 1 : 1) processes were applied to find the best fit in 12 patients (validation test set) with NSCLC and brain metastases treated with a tyrosine kinase inhibitor and whole-brain radiotherapy. Primary and secondary outcome measures: EGFR mutation analysis in patients with NSCLC and brain metastases and the development of a LDA-SVM-based EGFR mutation model for NSCLC brain metastases patients. EGFR mutation discordance between the primary lung tumor and brain metastases was found in 5 patients. Using LDA, 13 clinical features were transformed into 9 characteristics, and 3 were selected as primary vectors. The EGFR mutation model constructed with SVM algorithms had an accuracy, sensitivity, and specificity for determining the mutation status of brain metastases of 0.879, 0.886, and 0.875, respectively. Furthermore, the replicability of our model was confirmed by testing 100 random combinations of input values. The LDA-SVM-based model developed in this study could predict the EGFR status of brain metastases in this

  1. LDA-SVM-based EGFR mutation model for NSCLC brain metastases: an observational study.

    Science.gov (United States)

    Hu, Nan; Wang, Ge; Wu, Yu-Hao; Chen, Shi-Feng; Liu, Guo-Dong; Chen, Chuan; Wang, Dong; He, Zhong-Shi; Yang, Xue-Qin; He, Yong; Xiao, Hua-Liang; Huang, Ding-De; Xiong, Kun-Lin; Wu, Yan; Huang, Ming; Yang, Zhen-Zhou

    2015-02-01

    Epidermal growth factor receptor (EGFR) activating mutations are a predictor of tyrosine kinase inhibitor effectiveness in the treatment of non-small-cell lung cancer (NSCLC). The objective of this study is to build a model for predicting the EGFR mutation status of brain metastasis in patients with NSCLC. Observation and model set-up. This study was conducted between January 2003 and December 2011 in 6 medical centers in Southwest China. The study included 31 NSCLC patients with brain metastases. Eligibility requirements were histological proof of NSCLC, as well as sufficient quantity of paraffin-embedded lung and brain metastases specimens for EGFR mutation detection. The linear discriminant analysis (LDA) method was used for analyzing the dimensional reduction of clinical features, and a support vector machine (SVM) algorithm was employed to generate an EGFR mutation model for NSCLC brain metastases. Training-testing-validation (3 : 1 : 1) processes were applied to find the best fit in 12 patients (validation test set) with NSCLC and brain metastases treated with a tyrosine kinase inhibitor and whole-brain radiotherapy. Primary and secondary outcome measures: EGFR mutation analysis in patients with NSCLC and brain metastases and the development of a LDA-SVM-based EGFR mutation model for NSCLC brain metastases patients. EGFR mutation discordance between the primary lung tumor and brain metastases was found in 5 patients. Using LDA, 13 clinical features were transformed into 9 characteristics, and 3 were selected as primary vectors. The EGFR mutation model constructed with SVM algorithms had an accuracy, sensitivity, and specificity for determining the mutation status of brain metastases of 0.879, 0.886, and 0.875, respectively. Furthermore, the replicability of our model was confirmed by testing 100 random combinations of input values. The LDA-SVM-based model developed in this study could predict the EGFR status of brain metastases in this small cohort of

  2. Molecular Dynamics Analysis of Binding of Kinase Inhibitors to WT EGFR and the T790M Mutant.

    Science.gov (United States)

    Park, Jiyong; McDonald, Joseph J; Petter, Russell C; Houk, K N

    2016-04-12

    Epidermal growth factor receptor (EGFR) inhibitors interrupt EGFR-dependent cellular signaling pathways that lead to accelerated tumor growth and proliferation. Mutation of a threonine in the inhibitor binding pocket, known as the "gatekeeper", to methionine (T790M) confers acquired resistance to several EGFR-selective inhibitors. We studied interactions between EGFR inhibitors and the gatekeeper residues of the target protein. Thermodynamic integration (TI) with Amber14 indicates that the binding energies of gefitinib and AEE788 to the active state of the T790M mutant EGFR is 3 kcal/mol higher than to the wild type (WT), whereas ATP binding energy to the mutant is similar to the WT. Using metadynamics MD simulations with NAMD v2.9, the conformational equilibrium between the inactive resting state and the catalytically competent activate state was determined for the WT EGFR. When combined with the results obtained by Sutto and Gervasio, our simulations showed that the T790M point mutation lowers the free energy of the active state by 5 kcal/mol relative to the inactive state of the enzyme. Relative to the WT, the T790M mutant binds gefitinib more strongly. The T790M mutation is nevertheless resistant due to its increased binding of ATP. By contrast, the binding of AEE788 to the active state causes a conformational change in the αC-helix adjacent to the inhibitor binding pocket, that results in a 2 kcal/mol energy penalty. The energy penalty explains why the binding of AEE788 to the T790M mutant is unfavorable relative to binding to WT EGFR. These results establish the role of the gatekeeper mutation on inhibitor selectivity. Additional molecular dynamics (MD) simulations, TI, and metadynamics MD simulations reveal the origins of the changes in binding energy of WT and mutants. PMID:27010480

  3. XGFR*, a novel affinity-matured bispecific antibody targeting IGF-1R and EGFR with combined signaling inhibition and enhanced immune activation for the treatment of pancreatic cancer.

    Science.gov (United States)

    Schanzer, Juergen M; Wartha, Katharina; Moessner, Ekkehard; Hosse, Ralf J; Moser, Samuel; Croasdale, Rebecca; Trochanowska, Halina; Shao, Cuiying; Wang, Peng; Shi, Lei; Weinzierl, Tina; Rieder, Natascha; Bacac, Marina; Ries, Carola H; Kettenberger, Hubert; Schlothauer, Tilman; Friess, Thomas; Umana, Pablo; Klein, Christian

    2016-01-01

    The epidermal growth factor receptor (EGFR) and the insulin-like growth factor-1 receptor (IGF-1R) play critical roles in tumor growth, providing a strong rationale for the combined inhibition of IGF-1R and EGFR signaling in cancer therapy. We describe the design, affinity maturation, in vitro and in vivo characterization of the bispecific anti-IGF-1R/EGFR antibody XGFR*. XGFR* is based on the bispecific IgG antibody XGFR, which enabled heterodimerization of an IGF-1R binding scFab heavy chain with an EGFR-binding light and heavy chain by the "knobs-into-holes" technology. XGFR* is optimized for monovalent binding of human EGFR and IGF-1R with increased binding affinity for IGF-1R due to affinity maturation and highly improved protein stability to oxidative and thermal stress. It bears an afucosylated Fc-portion for optimal induction of antibody-dependent cell-mediated cytotoxicity (ADCC). Stable Chinese hamster ovary cell clones with production yields of 2-3 g/L were generated, allowing for large scale production of the bispecific antibody. XGFR* potently inhibits EGFR- and IGF-1R-dependent receptor phosphorylation, reduces tumor cell proliferation in cells with heterogeneous levels of IGF-1R and EGFR receptor expression and induces strong ADCC in vitro. A comparison of pancreatic and colorectal cancer lines demonstrated superior responsiveness to XGFR*-mediated signaling and tumor growth inhibition in pancreatic cancers that frequently show a high degree of IGF-1R/EGFR co-expression. XGFR* showed potent anti-tumoral efficacy in the orthotopic MiaPaCa-2 pancreatic xenograft model, resulting in nearly complete tumor growth inhibition with significant number of tumor remissions. In summary, the bispecific anti-IGF-1R/EGFR antibody XGFR* combines potent signaling and tumor growth inhibition with enhanced ADCC induction and represents a clinical development candidate for the treatment of pancreatic cancer. PMID:26984378

  4. Novel irreversible EGFR tyrosine kinase inhibitor 324674 sensitizes human colon carcinoma HT29 and SW480 cells to apoptosis by blocking the EGFR pathway

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Zhiwei; Cui, Binbin; Jin, Yinghu; Chen, Haipeng [Division of Colorectal Surgery, Third Affiliated Hospital of Harbin Medical University, Harbin (China); Wang, Xishan, E-mail: wxshan_oncologist@yahoo.com.cn [Division of Colorectal Surgery, Third Affiliated Hospital of Harbin Medical University, Harbin (China)

    2011-08-12

    Highlights: {yields} This article described the effects of the EGFR tyrosine kinase inhibitor on the cell proliferation and the apoptosis induction of the colon carcinoma cell lines. {yields} Demonstrated that 326474 is a more potent EGFR inhibitor on colon cancer cells than other three TKIs. {yields} It can be important when considering chemotherapy for colonic cancer patients. -- Abstract: Background: Epidermal growth factor receptor (EGFR) is widely expressed in multiple solid tumors including colorectal cancer by promoting cancer cell growth and proliferation. Therefore, the inhibition of EGFR activity may establish a clinical strategy of cancer therapy. Methods: In this study, using human colon adenocarcinoma HT29 and SW480 cells as research models, we compared the efficacy of four EGFR inhibitors in of EGFR-mediated pathways, including the novel irreversible inhibitor 324674, conventional reversible inhibitor AG1478, dual EGFR/HER2 inhibitor GW583340 and the pan-EGFR/ErbB2/ErbB4 inhibitor. Cell proliferation was assessed by MTT analysis, and apoptosis was evaluated by the Annexin-V binding assay. EGFR and its downstream signaling effectors were examined by western blotting analysis. Results: Among the four inhibitors, the irreversible EGFR inhibitor 324674 was more potent at inhibiting HT29 and SW480 cell proliferation and was able to efficiently induce apoptosis at lower concentrations. Western blotting analysis revealed that AG1478, GW583340 and pan-EGFR/ErbB2/ErbB4 inhibitors failed to suppress EGFR activation as well as the downstream mitogen-activated protein kinase (MAPK) and PI3K/AKT/mTOR (AKT) pathways. In contrast, 324674 inhibited EGFR activation and the downstream AKT signaling pathway in a dose-dependent manner. Conclusion: Our studies indicated that the novel irreversible EGFR inhibitor 324674 may have a therapeutic application in colon cancer therapy.

  5. A Novel Technique to Detect EGFR Mutations in Lung Cancer

    Directory of Open Access Journals (Sweden)

    Yuanbin Liu

    2016-05-01

    Full Text Available Epidermal growth factor receptor (EGFR gene mutations occur in multiple human cancers; therefore, the detection of EGFR mutations could lead to early cancer diagnosis. This study describes a novel EGFR mutation detection technique. Compared to direct DNA sequencing detection methods, this method is based on allele-specific amplification (ASA, recombinase polymerase amplification (RPA, peptide nucleic acid (PNA, and SYBR Green I (SYBR, referred to as the AS-RPA-PNA-SYBR (ARPS system. The principle of this technique is based on three continuous steps: ASA or ASA combined with PNA to prevent non-target sequence amplification (even single nucleotide polymorphisms, SNPs, the rapid amplification advantage of RPA, and appropriate SYBR Green I detection (the samples harboring EGFR mutations show a green signal. Using this method, the EGFR 19Del(2 mutation was detected in 5 min, while the EGFR L858R mutation was detected in 10 min. In this study, the detection of EGFR mutations in clinical samples using the ARPS system was compatible with that determined by polymerase chain reaction (PCR and DNA sequencing methods. Thus, this newly developed methodology that uses the ARPS system with appropriate primer sets is a rapid, reliable, and practical way to assess EGFR mutations in clinical samples.

  6. A Novel Technique to Detect EGFR Mutations in Lung Cancer

    Science.gov (United States)

    Liu, Yuanbin; Lei, Ting; Liu, Zhiyu; Kuang, Yanbin; Lyu, Jianxin; Wang, Qi

    2016-01-01

    Epidermal growth factor receptor (EGFR) gene mutations occur in multiple human cancers; therefore, the detection of EGFR mutations could lead to early cancer diagnosis. This study describes a novel EGFR mutation detection technique. Compared to direct DNA sequencing detection methods, this method is based on allele-specific amplification (ASA), recombinase polymerase amplification (RPA), peptide nucleic acid (PNA), and SYBR Green I (SYBR), referred to as the AS-RPA-PNA-SYBR (ARPS) system. The principle of this technique is based on three continuous steps: ASA or ASA combined with PNA to prevent non-target sequence amplification (even single nucleotide polymorphisms, SNPs), the rapid amplification advantage of RPA, and appropriate SYBR Green I detection (the samples harboring EGFR mutations show a green signal). Using this method, the EGFR 19Del(2) mutation was detected in 5 min, while the EGFR L858R mutation was detected in 10 min. In this study, the detection of EGFR mutations in clinical samples using the ARPS system was compatible with that determined by polymerase chain reaction (PCR) and DNA sequencing methods. Thus, this newly developed methodology that uses the ARPS system with appropriate primer sets is a rapid, reliable, and practical way to assess EGFR mutations in clinical samples. PMID:27223277

  7. JAK2 inhibition sensitizes resistant EGFR-mutant lung adenocarcinoma to tyrosine kinase inhibitors.

    Science.gov (United States)

    Gao, Sizhi P; Chang, Qing; Mao, Ninghui; Daly, Laura A; Vogel, Robert; Chan, Tyler; Liu, Shu Hui; Bournazou, Eirini; Schori, Erez; Zhang, Haiying; Red Brewer, Monica; Pao, William; Morris, Luc; Ladanyi, Marc; Arcila, Maria; Manova-Todorova, Katia; de Stanchina, Elisa; Norton, Larry; Levine, Ross L; Altan-Bonnet, Gregoire; Solit, David; Zinda, Michael; Huszar, Dennis; Lyden, David; Bromberg, Jacqueline F

    2016-01-01

    Lung adenocarcinomas with mutant epidermal growth factor receptor (EGFR) respond to EGFR-targeted tyrosine kinase inhibitors (TKIs), but resistance invariably occurs. We found that the Janus kinase (JAK)/signal transduction and activator of transcription 3 (STAT3) signaling pathway was aberrantly increased in TKI-resistant EGFR-mutant non-small cell lung cancer (NSCLC) cells. JAK2 inhibition restored sensitivity to the EGFR inhibitor erlotinib in TKI-resistant cell lines and xenograft models of EGFR-mutant TKI-resistant lung cancer. JAK2 inhibition uncoupled EGFR from its negative regulator, suppressor of cytokine signaling 5 (SOCS5), consequently increasing EGFR abundance and restoring the tumor cells' dependence on EGFR signaling. Furthermore, JAK2 inhibition led to heterodimerization of mutant and wild-type EGFR subunits, the activity of which was then blocked by TKIs. Our results reveal a mechanism whereby JAK2 inhibition overcomes acquired resistance to EGFR inhibitors and support the use of combination therapy with JAK and EGFR inhibitors for the treatment of EGFR-dependent NSCLC. PMID:27025877

  8. A fully automated two-step synthesis of an {sup 18}F-labelled tyrosine kinase inhibitor for EGFR kinase activity imaging in tumors

    Energy Technology Data Exchange (ETDEWEB)

    Kobus, D.; Giesen, Y.; Ullrich, R.; Backes, H. [Max Planck Institute for Neurological Research with Klaus-Joachim-Zuelch Laboratories of the Max Planck Society and the Faculty of Medicine of the University of Cologne, Cologne (Germany); Neumaier, B. [Max Planck Institute for Neurological Research with Klaus-Joachim-Zuelch Laboratories of the Max Planck Society and the Faculty of Medicine of the University of Cologne, Cologne (Germany)], E-mail: bernd.neumaier@nf.mpg.de

    2009-11-15

    Radiolabelled epidermal growth factor receptor (EGFR) tyrosine kinase (TK) inhibitors potentially facilitate the assessment of EGFR overexpression in tumors. Since elaborate multi-step radiosyntheses are required for {sup 18}F-labelling of EGFR-specific anilinoquinazolines we report on the development of a two-step click labelling approach that was adapted to a fully automated synthesis module. 6-(4-N,N-Dimethylaminocrotonyl)amido-4-(3-chloro-4-fluoro)phenylamino-7-{l_brace}3- [4-(2-[{sup 18}F]fluoroethyl)-2,3,4-triazol-1-yl]propoxy{r_brace}quinazoline ([{sup 18}F]6) was synthesized via Huisgen 1,3-dipolar cycloaddition between 2-[{sup 18}F]fluoroethylazide ([{sup 18}F]4) and the alkyne modified anilinoquinazoline precursor 5. PET images of PC9 tumor xenograft using the novel biomarker showed promising results to visualize EGFR overexpression.

  9. Cellular Functions Regulated by Phosphorylation of EGFR on Tyr845

    Directory of Open Access Journals (Sweden)

    Ken-ichi Sato

    2013-05-01

    Full Text Available The Src gene product (Src and the epidermal growth factor receptor (EGFR are prototypes of oncogene products and function primarily as a cytoplasmic non-receptor tyrosine kinase and a transmembrane receptor tyrosine kinase, respectively. The identification of Src and EGFR, and the subsequent extensive investigations of these proteins have long provided cutting edge research in cancer and other molecular and cellular biological studies. In 1995, we reported that the human epidermoid carcinoma cells, A431, contain a small fraction of Src and EGFR in which these two kinase were in physical association with each other, and that Src phosphorylates EGFR on tyrosine 845 (Y845 in the Src-EGFR complex. Y845 of EGFR is located in the activation segment of the kinase domain, where many protein kinases contain kinase-activating autophosphorylation sites (e.g., cAMP-dependent protein kinase, Src family kinases, transmembrane receptor type tyrosine kinases or trans-phosphorylation sites (e.g., cyclin-dependent protein kinase, mitogen-activated protein kinase, Akt protein kinase. A number of studies have demonstrated that Y845 phosphorylation serves an important role in cancer as well as normal cells. Here we compile the experimental facts involving Src phosphorylation of EGFR on Y845, by which cell proliferation, cell cycle control, mitochondrial regulation of cell metabolism, gamete activation and other cellular functions are regulated. We also discuss the physiological relevance, as well as structural insights of the Y845 phosphorylation.

  10. Therapeutic resistance in cancer:microRNA regulation of EGFR signaling networks

    Institute of Scientific and Technical Information of China (English)

    German G. Gomez; Jill Wykosky; Ciro Zanca; Frank B. Furnari; Webster K. Cavenee

    2013-01-01

    Receptor tyrosine kinases (RTKs) such as the epidermal growth factor receptor (EGFR) regulate cellular homeostatic processes. EGFR activates downstream signaling cascades that promote tumor cell survival, proliferation and migration. Dysregulation of EGFR signaling as a consequence of overexpression, amplification and mutation of the EGFR gene occurs frequently in several types of cancers and many become dependent on EGFR signaling to maintain their malignant phenotypes. Consequently, concerted efforts have been mounted to develop therapeutic agents and strategies to effectively inhibit EGFR. However, limited therapeutic beneifts to cancer patients have been derived from EGFR-targeted therapies. A well-documented obstacle to improved patient survival is the presence of EGFR-inhibitor resistant tumor cell variants within heterogeneous tumor cell masses. Here, we summarize the mechanisms by which tumors resist EGFR-targeted therapies and highlight the emerging role of microRNAs (miRs) as downstream effector molecules utilized by EGFR to promote tumor initiation, progression and that play a role in resistance to EGFR inhibitors. We also examine evidence supporting the utility of miRs as predictors of response to targeted therapies and novel therapeutic agents to circumvent EGFR-inhibitor resistance mechanisms.

  11. Spheroid Culture of Head and Neck Cancer Cells Reveals an Important Role of EGFR Signalling in Anchorage Independent Survival.

    Science.gov (United States)

    Braunholz, Diana; Saki, Mohammad; Niehr, Franziska; Öztürk, Merve; Borràs Puértolas, Berta; Konschak, Robert; Budach, Volker; Tinhofer, Ingeborg

    2016-01-01

    In solid tumours millions of cells are shed into the blood circulation each day. Only a subset of these circulating tumour cells (CTCs) survive, many of them presumable because of their potential to form multi-cellular clusters also named spheroids. Tumour cells within these spheroids are protected from anoikis, which allows them to metastasize to distant organs or re-seed at the primary site. We used spheroid cultures of head and neck squamous cell carcinoma (HNSCC) cell lines as a model for such CTC clusters for determining the role of the epidermal growth factor receptor (EGFR) in cluster formation ability and cell survival after detachment from the extra-cellular matrix. The HNSCC cell lines FaDu, SCC-9 and UT-SCC-9 (UT-SCC-9P) as well as its cetuximab (CTX)-resistant sub-clone (UT-SCC-9R) were forced to grow in an anchorage-independent manner by coating culture dishes with the anti-adhesive polymer poly-2-hydroxyethylmethacrylate (poly-HEMA). The extent of apoptosis, clonogenic survival and EGFR signalling under such culture conditions was evaluated. The potential of spheroid formation in suspension culture was found to be positively correlated with the proliferation rate of HNSCC cell lines as well as their basal EGFR expression levels. CTX and gefitinib blocked, whereas the addition of EGFR ligands promoted anchorage-independent cell survival and spheroid formation. Increased spheroid formation and growth were associated with persistent activation of EGFR and its downstream signalling component (MAPK/ERK). Importantly, HNSCC cells derived from spheroid cultures retained their clonogenic potential in the absence of cell-matrix contact. Addition of CTX under these conditions strongly inhibited colony formation in CTX-sensitive cell lines but not their resistant subclones. Altogether, EGFR activation was identified as crucial factor for anchorage-independent survival of HNSCC cells. Targeting EGFR in CTC cluster formation might represent an attractive anti

  12. Blockade of EGFR and MEK intercepts heterogeneous mechanisms of acquired resistance to anti-EGFR therapies in colorectal cancer.

    Science.gov (United States)

    Misale, Sandra; Arena, Sabrina; Lamba, Simona; Siravegna, Giulia; Lallo, Alice; Hobor, Sebastijan; Russo, Mariangela; Buscarino, Michela; Lazzari, Luca; Sartore-Bianchi, Andrea; Bencardino, Katia; Amatu, Alessio; Lauricella, Calogero; Valtorta, Emanuele; Siena, Salvatore; Di Nicolantonio, Federica; Bardelli, Alberto

    2014-02-19

    Colorectal cancers (CRCs) that are sensitive to the anti-epidermal growth factor receptor (EGFR) antibodies cetuximab or panitumumab almost always develop resistance within several months of initiating therapy. We report the emergence of polyclonal KRAS, NRAS, and BRAF mutations in CRC cells with acquired resistance to EGFR blockade. Regardless of the genetic alterations, resistant cells consistently displayed mitogen-activated protein kinase kinase (MEK) and extracellular signal-regulated kinase (ERK) activation, which persisted after EGFR blockade. Inhibition of MEK1/2 alone failed to impair the growth of resistant cells in vitro and in vivo. An RNA interference screen demonstrated that suppression of EGFR, together with silencing of MEK1/2, was required to hamper the proliferation of resistant cells. Indeed, concomitant pharmacological blockade of MEK and EGFR induced prolonged ERK inhibition and severely impaired the growth of resistant tumor cells. Heterogeneous and concomitant mutations in KRAS and NRAS were also detected in plasma samples from patients who developed resistance to anti-EGFR antibodies. A mouse xenotransplant from a CRC patient who responded and subsequently relapsed upon EGFR therapy showed exquisite sensitivity to combinatorial treatment with MEK and EGFR inhibitors. Collectively, these results identify genetically distinct mechanisms that mediate secondary resistance to anti-EGFR therapies, all of which reactivate ERK signaling. These observations provide a rational strategy to overcome the multifaceted clonal heterogeneity that emerges when tumors are treated with targeted agents. We propose that MEK inhibitors, in combination with cetuximab or panitumumab, should be tested in CRC patients who become refractory to anti-EGFR therapies.

  13. Pseudomonas aeruginosa pyocyanin activates NRF2-ARE-mediated transcriptional response via the ROS-EGFR-PI3K-AKT/MEK-ERK MAP kinase signaling in pulmonary epithelial cells.

    Science.gov (United States)

    Xu, Ying; Duan, Chaohui; Kuang, Zhizhou; Hao, Yonghua; Jeffries, Jayme L; Lau, Gee W

    2013-01-01

    The redox-active pyocyanin (PCN) secreted by the respiratory pathogen Pseudomonas aeruginosa generates reactive oxygen species (ROS) and causes oxidative stress to pulmonary epithelial cells. Nuclear factor (erythroid-derived 2)-like 2 (NRF2) confers protection against ROS-mediated cell death by inducing the expression of detoxifying enzymes and proteins via its binding to the cis-acting antioxidant response element (ARE). However, a clear relationship between NRF2 and PCN-mediated oxidative stress has not been established experimentally. In this study, we investigated the induction of NRF2-ARE response by PCN in the pulmonary epithelial cells. We analyzed the effect of PCN on NRF2 expression and nuclear translocation in cultured human airway epithelial cells, and in a mouse model of chronic PCN exposure. NRF2-dependent transcription of antioxidative enzymes was also assessed. Furthermore, we used inhibitors to examine the involvement of EGFR and its downstream signaling components that mediate NRF2-ARE-activation in response to PCN. PCN enhances the nuclear NRF2 accumulation and activates the transcription of ARE-mediated antioxidant genes. Furthermore, PCN activates NRF2 by inducing the EGFR-phosphoinositide-3-kinase (PI3K) signaling pathway and its main downstream effectors, AKT and MEK1/2-ERK1/2 MAP kinases. Inhibition of the EGFR-PI3K signaling markedly attenuates PCN-stimulated NRF2 accumulation in the nucleus. We demonstrate for the first time that PCN-mediated oxidative stress activates the EGFR-PI3K-AKT/MEK1/2-ERK1/2 MAP kinase signaling pathway, leading to nuclear NRF2 translocation and ARE responsiveness in pulmonary epithelial cells.

  14. A high expression EGFR/cell membrane chromatography and online high performance liquid chromatography/mass spectrometry method for screening EGFR antagonists from Rhizoma Polygoni Cuspidati

    OpenAIRE

    Meng Sun; Yan-min Zhang; Jie Zhang; Si-cen Wang; Lang-chong He

    2011-01-01

    The epidermal growth factor receptors (EGFRs) in some tumor cells are significant targets for drug discovery. In this work, we have developed an EGFR cell membrane chromatography and online high performance liquid chromatography/mass spectrometry system for screening active component from Rhizoma Polygoni Cuspidati. As a result, resveratrol from Rhizoma Polygoni Cuspidati was found to be the active component acting on EGFR like gefitinib. There was a good relationship between their inhibiting...

  15. Epidermal Growth Factor Receptor (EGFR) mutation analysis, gene expression profiling and EGFR protein expression in primary prostate cancer

    International Nuclear Information System (INIS)

    Activating mutations of the epidermal growth factor receptor (EGFR) confer sensitivity to the tyrosine kinase inhibitors (TKi), gefitinib and erlotinib. We analysed EGFR expression, EGFR mutation status and gene expression profiles of prostate cancer (PC) to supply a rationale for EGFR targeted therapies in this disease. Mutational analysis of EGFR TK domain (exons from 18 to 21) and immunohistochemistry for EGFR were performed on tumour tissues derived from radical prostatectomy from 100 PC patients. Gene expression profiling using oligo-microarrays was also carried out in 51 of the PC samples. EGFR protein overexpression (EGFRhigh) was found in 36% of the tumour samples, and mutations were found in 13% of samples. Patients with EGFRhigh tumours experienced a significantly increased risk of biochemical relapse (hazard ratio-HR 2.52, p=0.02) compared with patients with tumours expressing low levels of EGFR (EGFRlow). Microarray analysis did not reveal any differences in gene expression between EGFRhigh and EGFRlow tumours. Conversely, in EGFRhigh tumours, we were able to identify a 79 gene signature distinguishing mutated from non-mutated tumours. Additionally, 29 genes were found to be differentially expressed between mutated/EGFRhigh (n=3) and mutated/EGFRlow tumours (n=5). Four of the down-regulated genes, U19/EAF2, ABCC4, KLK3 and ANXA3 and one of the up-regulated genes, FOXC1, are involved in PC progression. Based on our findings, we hypothesize that accurate definition of the EGFR status could improve prognostic stratification and we suggest a possible role for EGFR-directed therapies in PC patients. Having been generated in a relatively small sample of patients, our results warrant confirmation in larger series

  16. Ghrelin promotes intestinal epithelial cell proliferation through PI3K/Akt pathway and EGFR trans-activation both converging to ERK 1/2 phosphorylation.

    Science.gov (United States)

    Waseem, Talat; Duxbury, Mark; Ashley, Stanley W; Robinson, Malcolm K

    2014-02-01

    Little is known about ghrelin's effects on intestinal epithelial cells even though it is known to be a mitogen for a variety of other cell types. Because ghrelin is released in close proximity to the proliferative compartment of the intestinal tract, we hypothesized that ghrelin may have potent pro-proliferative effect on intestinal epithelial cells as well. To test this hypothesis, we characterized the effects of ghrelin on FHs74Int and Caco-2 intestinal epithelial cell lines in vitro. We found that ghrelin has potent dose dependent proliferative effects in both cell lines through a yet to be characterized G protein coupled growth hormone secretagogue receptor (GHS-R) subtype. Consistent with above findings, cell cycle flowcytometric analyses demonstrated that ghrelin shifts cells from the G1 to S phase and thereby promotes cell cycle progression. Further characterization of subcellular events, suggested that ghrelin mediates its pro-proliferative effect through Adenylate cyclase (AC)-independent epidermal growth factor receptor (EGFR) trans-activation and PI3K-Akt phosphorylation. Both these pathways converge to stimulate MAPK, ERK 1/2 downstream. The role of ghrelin in states where intestinal mucosal injury and rapid mucosal repair occur warrants further investigation.

  17. Unexpected responses to EGFR inhibition in NSCLC

    Directory of Open Access Journals (Sweden)

    Giulia M. Stella

    2015-01-01

    Full Text Available The presence of activating mutations of the epidermal growth factor receptor (EGFR-gene identifies a distinct and clinically relevant molecular subset of non-small-cell lung cancer. It is now well demonstrated that EGFR tyrosine kinase inhibitors (TKIs gefitinib and erlotinib are superior to standard chemotherapy in this subset of tumors. Nevertheless, in many cases, responses are not durable and last for 6–12 months due to the occurrence of secondary or acquired resistance. Here we present three cases of EGFR-mutant lung adenocarcinomas (ADC, that showed an unexpected response to anti-EGFR small molecules. The first patient presented a continued 89 month-long response to erlotinib in a tumor recurred after surgery and conventional chemotherapy. In the other cases, subclinically persistent tumor in the lung tissue was documented histologically in lung resections performed after partial response to TKI treatment. The persistence of interstitial and endolymphatic tumor cells after TKI treatment might explain the common observation of tumor relapse after TKI discontinuation, and sustain the decision to continue treatment in responsive patients as in our first case.

  18. Unexpected responses to EGFR inhibition in NSCLC

    Science.gov (United States)

    Stella, Giulia M.; Valizia, Claudio; Zorzetto, Michele; Inghilleri, Simona; Valentini, Adele; Dore, Roberto; Colombo, Sara; Valentino, Francesco; Orlandoni, Giulio; Morbini, Patrizia

    2015-01-01

    The presence of activating mutations of the epidermal growth factor receptor (EGFR)-gene identifies a distinct and clinically relevant molecular subset of non-small-cell lung cancer. It is now well demonstrated that EGFR tyrosine kinase inhibitors (TKIs) gefitinib and erlotinib are superior to standard chemotherapy in this subset of tumors. Nevertheless, in many cases, responses are not durable and last for 6–12 months due to the occurrence of secondary or acquired resistance. Here we present three cases of EGFR-mutant lung adenocarcinomas (ADC), that showed an unexpected response to anti-EGFR small molecules. The first patient presented a continued 89 month-long response to erlotinib in a tumor recurred after surgery and conventional chemotherapy. In the other cases, subclinically persistent tumor in the lung tissue was documented histologically in lung resections performed after partial response to TKI treatment. The persistence of interstitial and endolymphatic tumor cells after TKI treatment might explain the common observation of tumor relapse after TKI discontinuation, and sustain the decision to continue treatment in responsive patients as in our first case. PMID:26744648

  19. EGFR soluble isoforms and their transcripts are expressed in meningiomas.

    Science.gov (United States)

    Guillaudeau, Angélique; Durand, Karine; Bessette, Barbara; Chaunavel, Alain; Pommepuy, Isabelle; Projetti, Fabrice; Robert, Sandrine; Caire, François; Rabinovitch-Chable, Hélène; Labrousse, François

    2012-01-01

    The EGFR (epidermal growth factor receptor) is involved in the oncogenesis of many tumors. In addition to the full-length EGFR (isoform a), normal and tumor cells produce soluble EGFR isoforms (sEGFR) that lack the intracellular domain. sEGFR isoforms b, c and d are encoded by EGFR variants 2 (v2), 3 (v3) and 4 (v4) mRNA resulting from gene alternative splicing. Accordingly, the results of EGFR protein expression analysis depend on the domain targeted by the antibodies. In meningiomas, EGFR expression investigations mainly focused on EGFR isoform a. sEGFR and EGFRvIII mutant, that encodes a constitutively active truncated receptor, have not been studied. In a 69 meningiomas series, protein expression was analyzed by immunohistochemistry using extracellular domain targeted antibody (ECD-Ab) and intracellular domain targeted antibody (ICD-Ab). EGFRv1 to v4 and EGFRvIII mRNAs were quantified by RT-PCR and EGFR amplification revealed by MLPA. Results were analyzed with respect to clinical data, tumor resection (Simpson grade), histological type, tumor grade, and patient outcome.Immunochemical staining was stronger with ECD-Ab than with ICD-Ab. Meningiomas expressed EGFRv1 to -v4 mRNAs but not EGFRvIII mutant. Intermediate or high ECD-Ab staining and high EGFRv1 to v4 mRNA levels were associated to a better progression free survival (PFS). PFS was also improved in women, when tumor resection was evaluated as Simpson 1 or 2, in grade I vs. grade II and III meningiomas and when Ki67 labeling index was lower than 10%. Our results suggest that, EGFR protein isoforms without ICD and their corresponding mRNA variants are expressed in meningiomas in addition to the whole isoform a. EGFRvIII was not expressed. High expression levels seem to be related to a better prognosis. These results indicate that the oncogenetic mechanisms involving the EGFR pathway in meningiomas could be different from other tumor types. PMID:22623992

  20. EGFR soluble isoforms and their transcripts are expressed in meningiomas.

    Directory of Open Access Journals (Sweden)

    Angélique Guillaudeau

    Full Text Available The EGFR (epidermal growth factor receptor is involved in the oncogenesis of many tumors. In addition to the full-length EGFR (isoform a, normal and tumor cells produce soluble EGFR isoforms (sEGFR that lack the intracellular domain. sEGFR isoforms b, c and d are encoded by EGFR variants 2 (v2, 3 (v3 and 4 (v4 mRNA resulting from gene alternative splicing. Accordingly, the results of EGFR protein expression analysis depend on the domain targeted by the antibodies. In meningiomas, EGFR expression investigations mainly focused on EGFR isoform a. sEGFR and EGFRvIII mutant, that encodes a constitutively active truncated receptor, have not been studied. In a 69 meningiomas series, protein expression was analyzed by immunohistochemistry using extracellular domain targeted antibody (ECD-Ab and intracellular domain targeted antibody (ICD-Ab. EGFRv1 to v4 and EGFRvIII mRNAs were quantified by RT-PCR and EGFR amplification revealed by MLPA. Results were analyzed with respect to clinical data, tumor resection (Simpson grade, histological type, tumor grade, and patient outcome.Immunochemical staining was stronger with ECD-Ab than with ICD-Ab. Meningiomas expressed EGFRv1 to -v4 mRNAs but not EGFRvIII mutant. Intermediate or high ECD-Ab staining and high EGFRv1 to v4 mRNA levels were associated to a better progression free survival (PFS. PFS was also improved in women, when tumor resection was evaluated as Simpson 1 or 2, in grade I vs. grade II and III meningiomas and when Ki67 labeling index was lower than 10%. Our results suggest that, EGFR protein isoforms without ICD and their corresponding mRNA variants are expressed in meningiomas in addition to the whole isoform a. EGFRvIII was not expressed. High expression levels seem to be related to a better prognosis. These results indicate that the oncogenetic mechanisms involving the EGFR pathway in meningiomas could be different from other tumor types.

  1. 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

  2. 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

  3. Effects of different ligands on epidermal growth factor receptor (EGFR) nuclear translocation.

    Science.gov (United States)

    Faria, Jerusa A Q A; de Andrade, Carolina; Goes, Alfredo M; Rodrigues, Michele A; Gomes, Dawidson A

    2016-09-01

    The epidermal growth factor receptor (EGFR) is activated through binding to specific ligands and generates signals for proliferation, differentiation, migration, and cell survival. Recent data show the role of nuclear EGFR in tumors. Although many EGFR ligands are upregulated in cancers, little is known about their effects on EGFR nuclear translocation. We have compared the effects of six EGFR ligands (EGF, HB-EGF, TGF-α, β-Cellulin, amphiregulin, and epiregulin) on nuclear translocation of EGFR, receptor phosphorylation, migration, and proliferation. Cell fractionation and confocal immunofluorescence detected EGFR in the nucleus after EGF, HB-EGF, TGF-α and β-Cellulin stimulation in a dose-dependent manner. In contrast, amphiregulin and epiregulin did not generate nuclear translocation of EGFR. EGF, HB-EGF, TGF-α and β-Cellulin showed correlations between a higher rate of wound closure and increased phosphorylation of residues in the carboxy-terminus of EGFR, compared to amphiregulin and epiregulin. The data indicate that EGFR is translocated to the nucleus after stimulation with EGF, HB-EGF, TGF-α and β-Cellulin, and that these ligands are related to increased phosphorylation of EGFR tyrosine residues, inducing migration of SkHep-1 cells. PMID:27462018

  4. Nuclear EGFR contributes to acquired resistance to cetuximab.

    Science.gov (United States)

    Li, C; Iida, M; Dunn, E F; Ghia, A J; Wheeler, D L

    2009-10-29

    Epidermal growth factor receptor (EGFR) is a ubiquitously expressed receptor tyrosine kinase involved in the etiology of several human cancers. Cetuximab is an EGFR-blocking antibody that has been approved for the treatment of patients with head and neck squamous cell carcinoma and metastatic colorectal cancer. Previous reports have shown that EGFR translocation to the nucleus is associated with cell proliferation. Here we investigated mechanisms of acquired resistance to cetuximab using a model derived from the non-small cell lung cancer line H226. We demonstrated that cetuximab-resistant cells overexpress HER family ligands including epidermal growth factor (EGF), amphiregulin, heparin-binding EGF and beta-cellulin. Overexpression of these ligands is associated with the nuclear translocation of the EGFR and this process was mediated by the Src family kinases (SFK). Treatment of cetuximab-resistant cells with the SFK inhibitor, dasatinib, resulted in loss of nuclear EGFR, increased membrane expression of the EGFR and resensitization to cetuximab. In addition, expression of a nuclear localization sequence-tagged EGFR in cetuximab-sensitive cells increased resistance to cetuximab both in vitro and in mouse xenografts. Collectively, these data suggest that nuclear expression of EGFR may be an important molecular determinant of resistance to cetuximab therapy and provides a rationale for investigating nuclear EGFR as a biomarker for cetuximab response. Further, these data suggest a rationale for the design of clinical trials that examine the value of treating patients with cetuximab-resistant tumors with inhibitors of SFKs in combination with cetuximab. PMID:19684613

  5. Epidermal Growth Factor Receptor (EGFR) Crosstalks in Liver Cancer

    International Nuclear Information System (INIS)

    Hepatocarcinogenesis is a complex multistep process in which many different molecular pathways have been implicated. Hepatocellular carcinoma (HCC) is refractory to conventional chemotherapeutic agents, and the new targeted therapies are meeting with limited success. Interreceptor crosstalk and the positive feedback between different signaling systems are emerging as mechanisms of targeted therapy resistance. The identification of such interactions is therefore of particular relevance to improve therapeutic efficacy. Among the different signaling pathways activated in hepatocarcinogenesis the epidermal growth factor receptor (EGFR) system plays a prominent role, being recognized as a “signaling hub” where different extracellular growth and survival signals converge. EGFR can be transactivated in response to multiple heterologous ligands through the physical interaction with multiple receptors, the activity of intracellular kinases or the shedding of EGFR-ligands. In this article we review the crosstalk between the EGFR and other signaling pathways that could be relevant to liver cancer development and treatment

  6. Epidermal Growth Factor Receptor (EGFR) Crosstalks in Liver Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Berasain, Carmen, E-mail: cberasain@unav.es; Latasa, María Ujue; Urtasun, Raquel; Goñi, Saioa; Elizalde, María; Garcia-Irigoyen, Oihane; Azcona, María [Division of Hepatology and Gene Therapy, CIMA, University of Navarra, Pamplona 31008 (Spain); Prieto, Jesús [Division of Hepatology and Gene Therapy, CIMA, University of Navarra, Pamplona 31008 (Spain); CIBERehd, University Clinic, University of Navarra, Pamplona 31080 (Spain); Ávila, Matías A. [Division of Hepatology and Gene Therapy, CIMA, University of Navarra, Pamplona 31008 (Spain)

    2011-05-18

    Hepatocarcinogenesis is a complex multistep process in which many different molecular pathways have been implicated. Hepatocellular carcinoma (HCC) is refractory to conventional chemotherapeutic agents, and the new targeted therapies are meeting with limited success. Interreceptor crosstalk and the positive feedback between different signaling systems are emerging as mechanisms of targeted therapy resistance. The identification of such interactions is therefore of particular relevance to improve therapeutic efficacy. Among the different signaling pathways activated in hepatocarcinogenesis the epidermal growth factor receptor (EGFR) system plays a prominent role, being recognized as a “signaling hub” where different extracellular growth and survival signals converge. EGFR can be transactivated in response to multiple heterologous ligands through the physical interaction with multiple receptors, the activity of intracellular kinases or the shedding of EGFR-ligands. In this article we review the crosstalk between the EGFR and other signaling pathways that could be relevant to liver cancer development and treatment.

  7. Ezrin Enhances EGFR Signaling and Modulates Erlotinib Sensitivity in Non–Small Cell Lung Cancer Cells

    Directory of Open Access Journals (Sweden)

    Yasemin Saygideğer-Kont

    2016-02-01

    Full Text Available Ezrin is a scaffolding protein that is involved in oncogenesis by linking cytoskeletal and membrane proteins. Ezrin interacts with epidermal growth factor receptor (EGFR in the cell membrane, but little is known about the effects of this interaction on EGFR signaling pathway. In this study, we established the biological and functional significance of ezrin-EGFR interaction in non–small cell lung cancer (NSCLC cells. Endogenous ezrin and EGRF interaction was confirmed by co-immunoprecipitation and immunofluorescent staining. When expression of ezrin was inhibited, EGFR activity and phosphorylation levels of downstream signaling pathway proteins ERK and STAT3 were decreased. Cell fractionation experiments revealed that nuclear EGFR was significantly diminished in ezrin-knockdown cells. Consequently, mRNA levels of EGFR target genes AURKA, COX-2, cyclin D1, and iNOS were decreased in ezrin-depleted cells. A small molecule inhibitor of ezrin, NSC305787, reduced EGF-induced phosphorylation of EGFR and downstream target proteins, EGFR nuclear translocation, and mRNA levels of nuclear EGFR target genes similar to ezrin suppression. NSC305787 showed synergism with erlotinib in wild-type EGFR-expressing NSCLC cells, whereas no synergy was observed in EGFR-null cells. Phosphorylation of ezrin on Y146 was found as an enhancer of ezrin-EGFR interaction and required for increased proliferation, colony formation, and drug resistance to erlotinib. These findings suggest that ezrin-EGFR interaction augments oncogenic functions of EGFR and that targeting ezrin may provide a potential novel approach to overcome erlotinib resistance in NSCLC cells.

  8. Analysis list: EGFR [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available EGFR Uterus + hg19 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target/EGFR.1.ts...v http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target/EGFR.5.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target/EGFR....10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/EGFR.Uterus.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/Uterus.gml ...

  9. 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.

  10. Targeting EGFR for Treatment of Glioblastoma: Molecular Basis to Overcome Resistance

    OpenAIRE

    Taylor, Tiffany E.; Furnari, Frank B.; Cavenee, Webster K.

    2012-01-01

    Glioblastoma (glioblastoma multiforme; GBM; WHO Grade IV) accounts for the majority of primary malignant brain tumors in adults. Amplification and mutation of the epidermal growth factor receptor (EGFR) gene represent signature genetic abnormalities encountered in GBM. A range of potential therapies that target EGFR or its mutant constitutively active form, ΔEGFR, including tyrosine kinase inhibitors (TKIs), monoclonal antibodies, vaccines, and RNA-based agents, are currently in development o...

  11. HER2+ Cancer Cell Dependence on PI3K vs. MAPK Signaling Axes Is Determined by Expression of EGFR, ERBB3 and CDKN1B.

    Directory of Open Access Journals (Sweden)

    Daniel C Kirouac

    2016-04-01

    Full Text Available Understanding the molecular pathways by which oncogenes drive cancerous cell growth, and how dependence on such pathways varies between tumors could be highly valuable for the design of anti-cancer treatment strategies. In this work we study how dependence upon the canonical PI3K and MAPK cascades varies across HER2+ cancers, and define biomarkers predictive of pathway dependencies. A panel of 18 HER2+ (ERBB2-amplified cell lines representing a variety of indications was used to characterize the functional and molecular diversity within this oncogene-defined cancer. PI3K and MAPK-pathway dependencies were quantified by measuring in vitro cell growth responses to combinations of AKT (MK2206 and MEK (GSK1120212; trametinib inhibitors, in the presence and absence of the ERBB3 ligand heregulin (NRG1. A combination of three protein measurements comprising the receptors EGFR, ERBB3 (HER3, and the cyclin-dependent kinase inhibitor p27 (CDKN1B was found to accurately predict dependence on PI3K/AKT vs. MAPK/ERK signaling axes. Notably, this multivariate classifier outperformed the more intuitive and clinically employed metrics, such as expression of phospho-AKT and phospho-ERK, and PI3K pathway mutations (PIK3CA, PTEN, and PIK3R1. In both cell lines and primary patient samples, we observed consistent expression patterns of these biomarkers varies by cancer indication, such that ERBB3 and CDKN1B expression are relatively high in breast tumors while EGFR expression is relatively high in other indications. The predictability of the three protein biomarkers for differentiating PI3K/AKT vs. MAPK dependence in HER2+ cancers was confirmed using external datasets (Project Achilles and GDSC, again out-performing clinically used genetic markers. Measurement of this minimal set of three protein biomarkers could thus inform treatment, and predict mechanisms of drug resistance in HER2+ cancers. More generally, our results show a single oncogenic transformation can

  12. The Potential Value of EGFR and P53 Immunostaining in Tumors of the Urinary Bladder

    Directory of Open Access Journals (Sweden)

    Ibrahim N

    2009-01-01

    Full Text Available The expression of EGFR and p53 has not been adequately studied as a prognostic tool in urinary bladder tumors. We analyzed 74 bladder cancer samples from Egypt for EGFR and p53 expression using immunohistochemistry. The tumors were of different histological types, grades and clinical stages, and with established lymph node status. Almost 61% of the tumors showed positive membranous EGFR expression and 74.3% had positive nuclear staining of p53. Analysis of correlation of the IHC staining with clinical variables showed a significant correlation only between EGFR expression and histological type (p=0.002, ANOVA, in that the expression was higher in squamous cell carcinomas than in other histological types. There were no significant correlations between p53 or EGFR with the other clinicopathological variables, including age, sex, staging, grading, and lymph node status. Further studies are needed to determine if EGFR and p53 might be used as prognostic tools in bladder cancer.

  13. NF-κB drives acquired resistance to a novel mutant-selective EGFR inhibitor.

    Science.gov (United States)

    Galvani, Elena; Sun, Jing; Leon, Leticia G; Sciarrillo, Rocco; Narayan, Ravi S; Sjin, Robert Tjin Tham; Lee, Kwangho; Ohashi, Kadoaki; Heideman, Daniëlle A M; Alfieri, Roberta R; Heynen, Guus J; Bernards, René; Smit, Egbert F; Pao, William; Peters, Godefridus J; Giovannetti, Elisa

    2015-12-15

    The clinical efficacy of EGFR tyrosine kinase inhibitors (TKIs) in non-small cell lung cancer (NSCLC) harbouring activating EGFR mutations is limited by the emergence of acquired resistance, mostly ascribed to the secondary EGFR-T790M mutation. Selective EGFR-T790M inhibitors have been proposed as a new, extremely relevant therapeutic approach. Here, we demonstrate that the novel irreversible EGFR-TKI CNX-2006, a structural analog of CO-1686, currently tested in a phase-1/2 trial, is active against in vitro and in vivo NSCLC models expressing mutant EGFR, with minimal effect on the wild-type receptor. By integration of genetic and functional analyses in isogenic cell pairs we provide evidence of the crucial role played by NF-κB1 in driving CNX-2006 acquired resistance and show that NF-κB activation may replace the oncogenic EGFR signaling in NSCLC when effective and persistent inhibition of the target is achieved in the presence of the T790M mutation. In this context, we demonstrate that the sole, either genetic or pharmacologic, inhibition of NF-κB is sufficient to reduce the viability of cells that adapted to EGFR-TKIs. Overall, our findings support the rational inhibition of members of the NF-κB pathway as a promising therapeutic option for patients who progress after treatment with novel mutant-selective EGFR-TKIs.

  14. Dacomitinib in lung cancer: a "lost generation" EGFR tyrosine-kinase inhibitor from a bygone era?

    Science.gov (United States)

    Ou, Sai-Hong Ignatius; Soo, Ross A

    2015-01-01

    EGFR tyrosine-kinase inhibitors (TKIs) have now been firmly established as the first-line treatment for non-small-cell lung cancer (NSCLC) patients harboring activating EGFR mutations, based on seven prospective randomized Phase III trials. However, despite significantly improved overall response rate and improved median progression-free survival when compared to platinum-doublet chemotherapy, EGFR-mutant NSCLC patients treated with EGFR TKIs invariably progress due to the emergence of acquired resistances, with the gatekeeper T790M mutation accounting for up to 60% of the resistance mechanisms. Second-generation irreversible EGFR TKIs were developed in part to inhibit the T790M mutation, in addition to the common activating EGFR mutations. Dacomitinib is one such second-generation EGFR TKI designed to inhibit both the wild-type (WT) EGFR and EGFR T790M. Afatinib is another second-generation EGR TKI that has been now been approved for the first-line treatment of EGFR-mutant NSCLC patients, while dacomitinib continues to undergo clinical evaluation. We will review the clinical development of dacomitinib from Phase I to Phase III trials, including the two recently published negative large-scale randomized Phase III trials (ARCHER 1009, NCIC-BR-26). Results from another large-scale randomized trial (ARCHER 1050) comparing dacomitinib to gefitinib as first-line treatment of advanced treatment-naïve EGFR-mutant NSCLC patients will soon be available and will serve as the lynchpin trial for the potential approval of dacomitinib in NSCLC. Meanwhile, third-generation EGFR TKIs (eg, CO-1686 [rociletinib], AZ9291, HM61713, EGF816, and ASP8273) that preferentially and potently inhibit EGFR T790M but not WT EGFR are in full-scale clinical development, and some of these EGFR TKIs have received "breakthrough" designation by the US Food and Drug Administration and will likely be approved in late 2015. Given the rapid development of third-generation EGFR TKIs and the approval of

  15. Simultaneous inhibition of epidermal growth factor receptor (EGFR) signaling and enhanced activation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor-mediated apoptosis induction by an scFv : sTRAIL fusion protein with specificity for human EGFR

    NARCIS (Netherlands)

    Bremer, E; Samplonius, DF; van Genne, L; Dijkstra, MH; Kroesen, BJ; de Leij, LFMH; Helfrich, W

    2005-01-01

    Epidermal growth factor receptor (EGFR) signaling inhibition by monoclonal antibodies and EGFR-specific tyrosine kinase inhibitors has shown clinical efficacy in cancer by restoring susceptibility of tumor cells to therapeutic apoptosis induction. Tumor necrosis factor-related apoptosis-inducing lig

  16. Advances in the management of acquired resistance to EGFR-TKI in non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    Fei Zhou; Caicun Zhou

    2015-01-01

    Drugs that specifical y target the tyrosine kinase domain of epidermal growth factor receptor (EGFR), such as erlotinib or gefitinib, have exhibited striking ef icacy in non-smal cel lung cancer (NSCLC) patients har-boring activating EGFR mutations. However, acquired resistance inevitably develops and remains a serious barrier for the successful management of patients with this disease. Multiple mechanisms are reportedly involved in the process of acquired resistance, which provide new insights into the management of EGFR-tyrosine kinase inhibitor (EGFR-TKI) resistance. Here, we provide an overview of the emerging treatment approaches for patients with EGFR-TKI resistance.

  17. Maintenance of EGFR and EGFRvIII expressions in an in vivo and in vitro model of human glioblastoma multiforme

    DEFF Research Database (Denmark)

    Stockhausen, Marie-Thérése; Broholm, Helle; Villingshøj, Mette;

    2011-01-01

    with mutation of EGFR, and the constitutive activated deletion variant EGFRvIII is the most common EGFR mutation found in GBM. Activated EGFR signaling, through overexpression and/or mutation, is involved in increased tumorigenic potential. As such, EGFR is an attractive target for GBM therapy. However...... the expressions of EGFR and EGFRvIII are maintained both in xenograft tumors growing subcutaneously on mice and in cell cultures established in stem cell conditions. With this model it will be possible to further study the role of EGFR and EGFRvIII, and response to targeted therapy, in GBM.......Glioblastoma multiforme (GBM) is the most common, and most aggressive primary brain tumor among adults. A vast majority of the tumors express high levels of the epidermal growth factor receptor (EGFR) as a consequence of gene amplification. Furthermore, gene amplification is often associated...

  18. Changes in sebum levels and the development of acneiform rash in patients with non–small cell lung cancer after treatment with EGFR inhibitors

    Directory of Open Access Journals (Sweden)

    Nakahara T

    2015-01-01

    Full Text Available Takeshi Nakahara,1,2 Yoichi Moroi,2 Koichi Takayama,3 Eriko Itoh,1,2 Makiko Kido-Nakahara,2 Yoichi Nakanishi,3 Masutaka Furue2 1Division of Skin Surface Sensing, 2Department of Dermatology, 3Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan Background: It has recently been shown that patients treated with epidermal growth factor receptor (EGFR inhibitors often develop various cutaneous adverse events. While the pathogenesis underlying these events remains unclear, the relationship between skin toxicity induced by EGFR inhibitors and the sebaceous glands that express EGFR has been previously reported. Objectives: The primary aim of this study was to determine the relationship between cutaneous sebum levels and acneiform rash, a typical skin toxicity of EGFR inhibitors, by measuring the sebum levels before and after EGFR inhibitor treatment. Methods: Eight patients diagnosed with non–small cell lung cancer (NSCLC (three men and five women with an average age of 69.3 years who were initiated on treatment with EGFR inhibitors (either gefitinib [Iressa®] or erlotinib [Tarceva®] were enrolled. Using a Sebumeter®, sebum levels in the face, chest, and back of each patient were measured before and after EGFR inhibitor treatment. The development of acneiform rash in each skin region was also assessed. Results: Changes in sebum level along with the development of an acneiform rash were observed after patients were started on EGFR inhibitor treatment. Patients who developed an EGFR inhibitor–induced acneiform rash tended to have higher pretreatment sebum levels (baseline than did patients who did not experience an acneiform rash. At each time point measurement, sebum levels were found to be significantly higher in patients who had developed an acneiform rash at that time. Patients who developed rash during treatment showed greater differences in sebum level compared with

  19. Role of TI-VAMP and CD82 in EGFR cell-surface dynamics and signaling.

    Science.gov (United States)

    Danglot, Lydia; Chaineau, Mathilde; Dahan, Maxime; Gendron, Marie-Claude; Boggetto, Nicole; Perez, Franck; Galli, Thierry

    2010-03-01

    The v-SNARE TI-VAMP (VAMP7) mediates exocytosis during neuritogenesis, phagocytosis and lysosomal secretion. It localizes to endosomes and lysosomes but also to the trans-Golgi network. Here we show that depletion of TI-VAMP enhances the endocytosis of activated EGF receptor (EGFR) without affecting constitutive endocytosis of EGFR, or transferrin uptake. This increased EGFR internalization is mainly clathrin dependent. Searching for defects in EGFR regulators, we found that TI-VAMP depletion reduces the cell surface amount of CD82, a tetraspanin known to control EGFR localization in microdomains. We further show that TI-VAMP is required for secretion from the Golgi apparatus to the cell surface, and that TI-VAMP-positive vesicles transport CD82. Quantum dots video-microscopy indicates that depletion of TI-VAMP, or its cargo CD82, restrains EGFR diffusion and the area explored by EGFR at the cell surface. Both depletions also impair MAPK signaling and enhance endocytosis of activated EGFR by increased recruitment of AP-2. These results highlight the role of TI-VAMP in the secretory pathway of a tetraspanin, and support a model in which CD82 allows EGFR entry in microdomains that control its clathrin-dependent endocytosis and signaling.

  20. A view on EGFR-targeted therapies from the oncogene-addiction perspective

    Directory of Open Access Journals (Sweden)

    Rolando ePerez

    2013-04-01

    Full Text Available Tumor cell growth and survival can often be impaired by inactivating a single oncogen – a phenomenon that has been called as 'oncogene addiction'. It is in such scenarios that molecular targeted therapies may succeed. Among known oncogenes, the epidermal growth factor receptor (EGFR has become the target of different cancer therapies. So far, however, the clinical benefit from EGFR-targeted therapies has been rather limited. A critical review of the large amount of clinical data obtained with anti-EGFR agents, carried out from the perspective of the oncogene addiction concept, may help to understand the causes of the unsatisfactory results. In this article we intend to do such an exercise taking as basis for the analysis a few case studies of anti-EGFR agents that are currently in the clinic. There, the 'EGFR addiction' phenomenon becomes apparent in high-responder patients. We further discuss how the concept of oncogene addiction needs to be interpreted on the light of emerging experimental evidences and ideas; in particular, that EGFR addiction may reflect the interconnection of several cellular pathways. In this regard we set forth several hypotheses; namely, that requirement of higher glucose uptake by hypoxic tumor cells may reinforce EGFR addiction; and that chronic use of EGFR-targeted antibodies in EGFR-addicted tumors would induce stable disease by reversing the malignant phenotype of cancer stem cells and also by sustaining an anti-tumor T cell response. Finally, we discuss possible reasons for the failure of certain combinatorial therapies involving anti-EGFR agents, arguing that some of these agents might produce either a negative or a positive trans-modulation effect on other oncogenes. It becomes evident that we need operational definitions of EGFR addiction in order to determine which patient populations may benefit from treatment with anti-EGFR drugs, and to improve the design of these therapies.

  1. Dacomitinib in lung cancer: a “lost generation” EGFR tyrosine-kinase inhibitor from a bygone era?

    Directory of Open Access Journals (Sweden)

    Ou SH

    2015-10-01

    Full Text Available Sai-Hong Ignatius Ou,1 Ross A Soo21Chao Family Comprehensive Cancer Center, Division of Hematology/Oncology, Department of Medicine, University of California, Irvine School of Medicine, Orange, CA, USA; 2National University Health System and Cancer Science Institute of Singapore, SingaporeAbstract: EGFR tyrosine-kinase inhibitors (TKIs have now been firmly established as the first-line treatment for non-small-cell lung cancer (NSCLC patients harboring activating EGFR mutations, based on seven prospective randomized Phase III trials. However, despite significantly improved overall response rate and improved median progression-free survival when compared to platinum-doublet chemotherapy, EGFR-mutant NSCLC patients treated with EGFR TKIs invariably progress due to the emergence of acquired resistances, with the gatekeeper T790M mutation accounting for up to 60% of the resistance mechanisms. Second-generation irreversible EGFR TKIs were developed in part to inhibit the T790M mutation, in addition to the common activating EGFR mutations. Dacomitinib is one such second-generation EGFR TKI designed to inhibit both the wild-type (WT EGFR and EGFR T790M. Afatinib is another second-generation EGR TKI that has been now been approved for the first-line treatment of EGFR-mutant NSCLC patients, while dacomitinib continues to undergo clinical evaluation. We will review the clinical development of dacomitinib from Phase I to Phase III trials, including the two recently published negative large-scale randomized Phase III trials (ARCHER 1009, NCIC-BR-26. Results from another large-scale randomized trial (ARCHER 1050 comparing dacomitinib to gefitinib as first-line treatment of advanced treatment-naïve EGFR-mutant NSCLC patients will soon be available and will serve as the lynchpin trial for the potential approval of dacomitinib in NSCLC. Meanwhile, third-generation EGFR TKIs (eg, CO-1686 [rociletinib], AZ9291, HM61713, EGF816, and ASP8273 that preferentially

  2. 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.

  3. Rational bases for the development of EGFR inhibitors for cancer treatment.

    Science.gov (United States)

    Bianco, Roberto; Gelardi, Teresa; Damiano, Vincenzo; Ciardiello, Fortunato; Tortora, Giampaolo

    2007-01-01

    Growth factor receptors and their ligands not only regulate normal cell processes but have been also identified as key regulators of human cancer formation. The epidermal growth factor receptor (EGFR/ErbB1/HER1) belongs to the ErbB/HER-family of tyrosine kinase receptors (RTKs). These trans-membrane proteins are activated following binding with peptide growth factors of the EGF-family of proteins. Several evidences suggest that cooperation of multiple ErbB receptors and ligands is required for the induction of cell transformation. In this respect, EGFR, upon activation, sustains a complex and redundant network of signal transduction pathways with the contribution of other trans-membrane receptors. EGFR has been found to be expressed and altered in a variety of malignancies and clearly it plays a significant role in tumor development and progression, including cell proliferation, regulation of apoptotic cell death, angiogenesis and metastatic spread. Moreover, amplification of the EGFR gene and mutations in the EGFR tyrosine kinase domain have been recently reported in human carcinomas. As a result, investigators have developed approaches to inhibit the effects of EGFR activation, with the aim of blocking tumor growth and invasion. A number of agents targeting EGFR, including specific antibodies directed against its ligand-binding domain and small molecules inhibiting its tyrosine kinase activity are either in clinical trials or are already approved for clinical treatment. This article reviews the EGFR role in carcinogenesis and tumor progression as rational bases for the development of specific therapeutic inhibitors.

  4. Radiation Response Modulation of GW572016 (EGFR/HER2 Dual Tyrosine Kinase Inhibitor) in Human Breast Cancer Xenografts

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Yeon Sil; Roh, Kwang Won; Chae, Soo Min; Yoon, Sei Chul; Jang, Hong Seok; Chung, Su Mi [The Catholic University of Korea, College of Medicine, Seoul (Korea, Republic of); Mun, Seong Kwon [Eulji University Hospital, Daejeon (Korea, Republic of)

    2007-12-15

    Purpose: We examined the effect of the dual EGFR/HER2 tyrosine kinase inhibitor, GW572016, on EGFR/HER2 receptor phosphorylation, inhibition of downstream signaling and radiosensitization in either an EGFR or HER2 overexpressing human breast cancer xenograft. Materials and Methods: We established SCID mice xenografts from 4 human breast cancer cell line that overexpressed EGFR or HER 2 (SUM 102, SUM 149, SUM 185, SUM 225). Two series of xenografts were established. One series was established for determining inhibition of the EGFR/HER2 receptor and downstream signaling activities by GW572016. The other series was established for determining the radiosensitization effect of GW572016. Inhibition of the receptor and downstream signaling proteins were measured by the use of immunoprecipitation and Western blotting. For determining the in vivo radiosensitization effect of GW572016, we compared tumor growth delay curves in the following four treatment arms: a) control; b) GW572016 alone; c) radiotherapy (RT) alone; d) GW572016 and RT. Results: GW572016 inhibited EGFR, HER2 receptor phosphorylation in SUM 149 and SUM 185 xenografts. In addition, the p44/42 MAPK (ERK 1/2) downstream signaling pathway was inactivated by GW572016 in the SUM 185 xenograft. In the SUM 225 xenograft, we could not observe inhibition of HER2 receptor phosphorylation by GW572016; both p44/42 MAPK (Erk1/2) and Akt downstream signal protein phosphorylation were inhibited by GW572016. GW572016 inhibited growth of the tumor xenograft of SUM 149 and SUM 185. The combination of GW572016 and RT enhanced growth inhibition greater than that with GW572016 alone or with RT alone in the SUM 149 xenograft. GW572016 appears to act as an in vivo radiosensitizer. Conclusion: GW572016 inhibited EGFR/HER2 receptor phosphorylation and downstream signaling pathway proteins. GW572016 modestly inhibited the growth of tumor in the SUM 185 xenograft and showed radiosensitization in the SUM 149 xenograft. Our results

  5. Co-targeting ALK and EGFR parallel signaling in oral squamous cell carcinoma.

    Science.gov (United States)

    Gonzales, Cara B; De La Chapa, Jorge J; Saikumar, Pothana; Singha, Prajjal K; Dybdal-Hargreaves, Nicholas F; Chavez, Jeffery; Horning, Aaron M; Parra, Jamie; Kirma, Nameer B

    2016-08-01

    Squamous cell carcinoma (SCC) comprises 90% of all head and neck cancers and has a poor survival rate due to late-stage disease that is refractive to traditional therapies. Epidermal growth factor receptor (EGFR) is over-expressed in greater than 80% of head and neck SCC (HNSCC). However, EGFR targeted therapies yielded little to no efficacy in clinical trials. This study investigated the efficacy of co-targeting EGFR and the anaplastic lymphoma kinase (ALK) whose promoter is hypomethylated in late-stage oral SCC (OSCC). We observed increased ALK activity in late-stage human OSCC tumors and invasive OSCC cell lines. We also found that while ALK inhibition alone had little effect on proliferation, co-targeting ALK and EGFR significantly reduced OSCC cell proliferation in vitro. Further analysis showed significant efficacy of combined treatment in HSC3-derived xenografts resulting in a 30% decrease in tumor volumes by 14days (panalysis showed that co-targeting ALK and EGFR significantly reduced EGFR phosphorylation (Y1148) in HSC3 cells but not Cal27 cells. ALK and EGFR downstream signaling interactions are also demonstrated by Western blot analysis in which lone EGFR and ALK inhibitors attenuated AKT activity whereas co-targeting ALK and EGFR completely abolished AKT activation. No effects were observed on ERK1/2 activation. STAT3 activity was significantly induced by lone ALK inhibition in HSC3 cells and to a lower extent in Cal27 cells. Together, these data illustrate that ALK inhibitors enhance anti-tumor activity of EGFR inhibitors in susceptible tumors that display increased ALK expression, most likely through abolition of AKT activation. PMID:27424178

  6. Effect of EGFR-TKI retreatment following chemotherapy for advanced non-small cell lung cancer patients who underwent EGFR-TKI

    International Nuclear Information System (INIS)

    Non-small cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR)-activating mutations have higher response rate and more prolonged survival following treatment with single-agent EGFR tyrosine kinase inhibitor (EGFR-TKI) compared with patients with wild-type EGFR. However, all patients treated with reversible inhibitors develop acquired resistance over time. The mechanisms of resistance are complicated. The lack of established therapeutic options for patients after a failed EGFR-TKI treatment poses a great challenge to physicians in managing this group of lung cancer patients. This study evaluates the influence of EGFR-TKI retreatment following chemotherapy after failure of initial EGFR-TKI within at least 6 months on NSCLC patients. The data of 27 patients who experienced treatment failure from their initial use of EGFR-TKI within at least 6 months were analyzed. After chemotherapy, the patients were retreated with EGFR-TKI (gefitinib 250 mg qd or erlotinib 150 mg qd), and the tumor progression was observed. The patients were assessed for adverse events and response to therapy. Targeted tumor lesions were assessed with CT scan. Of the 27 patients who received EGFR-TKI retreatment, 1 (3.7%) patient was observed in complete response (CR), 8 (29.6%) patients in partial response (PR), 14 (51.9%) patients in stable disease (SD), and 4 (14.8%) patients in progressive disease (PD). The disease control rate (DCR) was 85.2% (95% CI: 62%-94%). The median progression-free survival (mPFS) was 6 months (95% CI: 1-29). Of the 13 patients who received the same EGFR-TKI, 1 patient in CR, 3 patients in PR, 8 patients in SD, and 2 patients in PD were observed. The DCR was 84.6%, and the mPFS was 5 months. Of the 14 patients who received another EGFR-TKI, no patient in CR, 6 patients in PR, 6 patients in SD, and 2 patients in PD were observed. The DCR was 85.7%, and the mPFS was 9.5 months. Significant difference was found between the two groups in PFS but not

  7. Effect of EGFR-TKI retreatment following chemotherapy for advanced non-small cell lung cancer patients who under went EGFR-TKI

    Institute of Scientific and Technical Information of China (English)

    Guo-Hao Xia; Ji-Feng Feng; Yun Zeng; Ying Fang; Shao-Rong Yu; Li Wang; Mei-Qi Shi; Wei-Li Sun; Xin-En Huang; Jia Chen

    2014-01-01

    Objective:Non-small cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR)-activating mutations have higher response rate and more prolonged survival following treatment with single-agent EGFR tyrosine kinase inhibitor (EGFR-TKI) compared with patients with wild-type EGFR. However, all patients treated with reversible inhibitors develop acquired resistance over time. hTe mechanisms of resistance are complicated. hTe lack of established therapeutic options for patients atfer a failed EGFR-TKI treatment poses a great challenge to physicians in managing this group of lung cancer patients. hTis study evaluates the inlfuence of EGFR-TKI retreatment following chemotherapy atfer failure of initial EGFR-TKI within at least 6 months on NSCLC patients. Methods:hTe data of 27 patients who experienced treatment failure from their initial use of EGFR-TKI within at least 6 months were analyzed. Atfer chemotherapy, the patients were retreated with EGFR-TKI (geiftinib 250 mg qd or erlotinib 150 mg qd), and the tumor progression was observed. The patients were assessed for adverse events and response to therapy. Targeted tumor lesions were assessed with CT scan. Results:Of the 27 patients who received EGFR-TKI retreatment, 1 (3.7%) patient was observed in complete response (CR), 8 (29.6%) patients in partial response (PR), 14 (51.9%) patients in stable disease (SD), and 4 (14.8%) patients in progressive disease (PD). hTe disease control rate (DCR) was 85.2%(95%CI:62%-94%). hTe median progression-free survival (mPFS) was 6 months (95%CI:1-29). Of the 13 patients who received the same EGFR-TKI, 1 patient in CR, 3 patients in PR, 8 patients in SD, and 2 patients in PD were observed. hTe DCR was 84.6%, and the mPFS was 5 months. Of the 14 patients who received another EGFR-TKI, no patient in CR, 6 patients in PR, 6 patients in SD, and 2 patients in PD were observed. hTe DCR was 85.7%, and the mPFS was 9.5 months. Signiifcant difference was found between the two

  8. Detection and clinical significance of intratumoral EGFR mutational heterogeneity in Chinese patients with advanced non-small cell lung cancer.

    Directory of Open Access Journals (Sweden)

    Hua Bai

    Full Text Available PURPOSE: This study evaluated occurrence and potential clinical significance of intratumoral EGFR mutational heterogeneity in Chinese patients with non-small cell lung cancer (NSCLC. MATERIALS AND METHODS: Eighty-five stage IIIa-IV NSCLC patients who had undergone palliative surgical resection were included in this study. Of these, 45 patients carried EGFR mutations (group-M and 40 patients were wild-type (group-W. Each tumor sample was microdissected to yield 28-34 tumor foci and Intratumoral EGFR mutation were determined using Denaturing High Performance Liquid Chromatography (DHPLC and Amplification Refractory Mutation System (ARMS. EGFR copy numbers were measured using fluorescence in situ hybridization (FISH. RESULTS: Microdissection yielded 1,431 tumor foci from EGFR mutant patients (group-M and 1,238 foci from wild-type patients (group-W. The EGFR mutant frequencies in group-M were 80.6% (1,154/1,431 and 87.1% (1,247/1,431 using DHPLC and ARMS, respectively. A combination of EGFR-mutated and wild-type cells was detected in 32.9% (28/85 of samples by DHPLC and 28.2% (24/85 by ARMS, supporting the occurrence of intratumoral heterogeneity. Thirty-one patients (36.5% were identified as EGFR FISH-positive. Patients harboring intratumoral mutational heterogeneity possessed lower EGFR copy numbers than those tumors contained mutant cells alone (16.7% vs. 71.0%, P<0.05. Among 26 patients who had received EGFR-TKIs, the mean EGFR mutation content was higher in patients showing partial response (86.1% or stable disease (48.7% compared with patients experiencing progressive disease (6.0% (P = 0.001. There also showed relationship between progression-free survival (PFS and different content of EGFR mutation groups (pure wild type EGFR, EGFR mutation with heterogeneity and pure mutated EGFR (P = 0.001. CONCLUSION: Approximately 30% of patients presented intratumoral EGFR mutational heterogeneity, accompanying with relatively low EGFR copy

  9. Bile acids regulate intestinal cell proliferation by modulating EGFR and FXR signaling.

    Science.gov (United States)

    Dossa, Avafia Y; Escobar, Oswaldo; Golden, Jamie; Frey, Mark R; Ford, Henri R; Gayer, Christopher P

    2016-01-15

    Bile acids (BAs) are synthesized in the liver and secreted into the intestine. In the lumen, enteric bacteria metabolize BAs from conjugated, primary forms into more toxic unconjugated, secondary metabolites. Secondary BAs can be injurious to the intestine and may contribute to disease. The epidermal growth factor receptor (EGFR) and the nuclear farnesoid X receptor (FXR) are known to interact with BAs. In this study we examined the effects of BAs on intestinal epithelial cell proliferation and investigated the possible roles for EGFR and FXR in these effects. We report that taurine-conjugated cholic acid (TCA) induced proliferation, while its unconjugated secondary counterpart deoxycholic acid (DCA) inhibited proliferation. TCA stimulated phosphorylation of Src, EGFR, and ERK 1/2. Pharmacological blockade of any of these pathways or genetic ablation of EGFR abrogated TCA-stimulated proliferation. Interestingly, Src or EGFR inhibitors eliminated TCA-induced phosphorylation of both molecules, suggesting that their activation is interdependent. In contrast to TCA, DCA exposure diminished EGFR phosphorylation, and pharmacological or siRNA blockade of FXR abolished DCA-induced inhibition of proliferation. Taken together, these results suggest that TCA induces intestinal cell proliferation via Src, EGFR, and ERK activation. In contrast, DCA inhibits proliferation via an FXR-dependent mechanism that may include downstream inactivation of the EGFR/Src/ERK pathway. Since elevated secondary BA levels are the result of specific bacterial modification, this may provide a mechanism through which an altered microbiota contributes to normal or abnormal intestinal epithelial cell proliferation.

  10. Detection of EGFR mutations in plasma and biopsies from non-small cell lung cancer patients by allele-specific PCR assays

    DEFF Research Database (Denmark)

    Weber, Britta; Meldgaard, Peter; Hager, Henrik;

    2014-01-01

    BACKGROUND: Lung cancer patients with mutations in the epidermal growth factor receptor (EGFR) are primary candidates for EGFR-targeted therapy. Reliable analyses of such mutations have previously been possible only in tumour tissue. Here, we demonstrate that mutations can be detected in plasma...... of mutations in exons 18-21 of the EGFR gene, employing the cobas(®) EGFR Tissue Test and cobas(®) EGFR Blood Test (in development, Roche Molecular Systems, Inc., CA, USA). RESULTS: Test results were obtained in all 199 (100%) plasma samples and 196/199 (98%) of the biopsies. EGFR-activating mutations were...... identified in 24/199 (12%) plasma samples and 28/196 (14%) biopsy samples, and 17/196 (9%) matched pairs contained the same mutation. Six EGFR mutations were present only in plasma samples but not in the biopsy samples. The overall concordance of the EGFR gene mutations detected in plasma and biopsy tissue...

  11. Utilization of Structure-Based Design to Identify Novel, Irreversible Inhibitors of EGFR Harboring the T790M Mutation.

    Science.gov (United States)

    Hennessy, Edward J; Chuaqui, Claudio; Ashton, Susan; Colclough, Nicola; Cross, Darren A E; Debreczeni, Judit É; Eberlein, Cath; Gingipalli, Lakshmaiah; Klinowska, Teresa C M; Orme, Jonathan P; Sha, Li; Wu, Xiaoyun

    2016-05-12

    A novel series of covalent inhibitors of EGFR (epidermal growth factor receptor) kinase was discovered through a combination of subset screening and structure-based design. These compounds preferentially inhibit mutant forms of EGFR (activating mutant and T790M mutant) over wild-type EGFR in cellular assays measuring EGFR autophosphorylation and proliferation, suggesting an improved therapeutic index in non-small cell lung cancer patients would be achievable relative to established EGFR inhibitors. We describe our design approaches, resulting in the identification of the lead compound 5, and our efforts to develop an understanding of the structure-activity relationships within this series. In addition, strategies to overcome challenges around metabolic stability and aqueous solubility are discussed. Despite limitations in its physical properties, 5 is orally bioavailable in mice and demonstrates pronounced antitumor activity in in vivo models of mutant EGFR-driven cancers.

  12. EGFR Regulates the Development and Microarchitecture of Intratumoral Angiogenic Vasculature Capable of Sustaining Cancer Cell Intravasation

    Directory of Open Access Journals (Sweden)

    Petra Minder

    2015-08-01

    Full Text Available Many malignant characteristics of cancer cells are regulated through pathways induced by the tyrosine kinase activity of the epidermal growth factor receptor (EGFR. Herein, we show that besides directly affecting the biology of cancer cells per se, EGFR also regulates the primary tumor microenvironment. Specifically, our findings demonstrate that both the expression and signaling activity of EGFR are required for the induction of a distinct intratumoral vasculature capable of sustaining tumor cell intravasation, a critical rate-limiting step in the metastatic cascade. An intravasation-sustaining mode of intratumoral angiogenic vessels depends on high levels of tumor cell EGFR and the interplay between EGFR-regulated production of interleukin 8 by tumor cells, interleukin-8–induced influx of tumor-infiltrating neutrophils delivering their unique matrix metalloproteinase-9, and neutrophil matrix metalloproteinase-9–dependent release of the vascular permeability and endothelial growth factor, VEGF. Our data indicate that through VEGF-mediated disruption of endothelial layer integrity and increase of intratumoral vasculature permeability, EGFR activity significantly facilitates active intravasation of cancer cells. Therefore, this study unraveled an important but overlooked function of EGFR in cancer, namely, its ability to create an intravasation-sustaining microenvironment within the developing primary tumor by orchestrating several interrelated processes required for the initial steps of cancer metastasis through vascular routes. Our findings also suggest that EGFR-targeted therapies might be more effective when implemented in cancer patients with early-staged primary tumors containing a VEGF-dependent angiogenic vasculature. Accordingly, early EGFR inhibition combined with various anti-VEGF approaches could synergistically suppress tumor cell intravasation through inhibiting the highly permeable angiogenic vasculature induced by EGFR

  13. Periodic mechanical stress activates EGFR-dependent Rac1 mitogenic signals in rat nucleus pulpous cells via ERK1/2.

    Science.gov (United States)

    Gao, Gongming; Shen, Nan; Jiang, Xuefeng; Sun, Huiqing; Xu, Nanwei; Zhou, Dong; Nong, Luming; Ren, Kewei

    2016-01-15

    The mitogenic effects of periodic mechanical stress on nucleus pulpous cells have been studied extensively but the mechanisms whereby nucleus pulpous cells sense and respond to mechanical stimulation remain a matter of debate. We explored this question by performing cell culture experiments in our self-developed periodic stress field and perfusion culture system. Under periodic mechanical stress, rat nucleus pulpous cell proliferation was significantly increased (p mechanical stress-induced nucleus pulpous cell proliferation (p mechanical stress (p mechanical stress (p mechanical stress promotes nucleus pulpous cell proliferation in part through the EGFR-Rac1-ERK1/2 signaling pathway, which links these three important signaling molecules into a mitogenic cascade.

  14. Quantification and kinetic analysis of Grb2-EGFR interaction on micro-patterned surfaces for the characterization of EGFR-modulating substances.

    Directory of Open Access Journals (Sweden)

    Peter Lanzerstorfer

    Full Text Available The identification of the epidermal growth factor receptor (EGFR as an oncogene has led to the development of several anticancer therapeutics directed against this receptor tyrosine kinase. However, drug resistance and low efficacy remain a severe challenge, and have led to a demand for novel systems for an efficient identification and characterization of new substances. Here we report on a technique which combines micro-patterned surfaces and total internal reflection fluorescence (TIRF microscopy (μ-patterning assay for the quantitative analysis of EGFR activity. It does not simply measure the phosphorylation of the receptor, but instead quantifies the interaction of the key signal transmitting protein Grb2 (growth factor receptor-bound protein 2 with the EGFR in a live cell context. It was possible to demonstrate an EGF dependent recruitment of Grb2 to the EGFR, which was significantly inhibited in the presence of clinically tested EGFR inhibitors, including small tyrosine kinase inhibitors and monoclonal antibodies targeting the EGF binding site. Importantly, in addition to its potential use as a screening tool, our experimental setup offers the possibility to provide insight into the molecular mechanisms of bait-prey interaction. Recruitment of the EGFR together with Grb2 to clathrin coated pits (CCPs was found to be a key feature in our assay. Application of bleaching experiments enabled calculation of the Grb2 exchange rate, which significantly changed upon stimulation or the presence of EGFR activity inhibiting drugs.

  15. Plant stanols induce intestinal tumor formation by up-regulating Wnt and EGFR signaling in Apc Min mice.

    Science.gov (United States)

    Marttinen, Maija; Päivärinta, Essi; Storvik, Markus; Huikko, Laura; Luoma-Halkola, Heli; Piironen, Vieno; Pajari, Anne-Maria; Mutanen, Marja

    2013-01-01

    The rate of APC mutations in the intestine increases in middle-age. At the same period of life, plant sterol and stanol enriched functional foods are introduced to diet to lower blood cholesterol. This study examined the effect of plant stanol enriched diet on intestinal adenoma formation in the Apc(Min) mouse. Apc(Min) mice were fed 0.8% plant stanol diet or control diet for nine weeks. Cholesterol, plant sterols and plant stanols were analyzed from the caecum content and the intestinal mucosa. Levels of β-catenin, cyclin D1, epidermal growth factor receptor (EGFR) and extracellular signal-regulated kinase 1/2 (ERK1/2) were measured from the intestinal mucosa by Western blotting. Gene expression was determined from the intestinal mucosa using Affymetrix and the data were analyzed for enriched categories and pathways. Plant stanols induced adenoma formation in the small intestine, however, the adenoma size was not affected. We saw increased levels of nuclear β-catenin, phosphorylated β-catenin (Ser675 and Ser552), nuclear cyclin D1, total and phosphorylated EGFR and phosphorylated ERK1/2 in the intestinal mucosa after plant stanol feeding. The Affymetrix data demonstrate that several enzymes of cholesterol synthesis pathway were up-regulated, although the cholesterol level in the intestinal mucosa was not altered. We show that plant stanols induce adenoma formation by activating Wnt and EGFR signaling. EGFR signaling seems to have promoted β-catenin phosphorylation and its translocation into the nucleus, where the expression of cyclin D1 was increased. Up-regulated cholesterol synthesis may partly explain the increased EGFR signaling in the plant stanol-fed mice.

  16. Lung Adenocarcinoma with Pulmonary Miliary Metastases and Complex Somatic Heterozygous EGFR Mutation

    Directory of Open Access Journals (Sweden)

    Alexandre Schaller

    2014-11-01

    Full Text Available The pretreatment detection of an activating mutation of EGFR is now routinely performed in metastatic nonsquamous non-small cell lung cancer (NSCLC. The therapeutic impact of such a detection is major, as patients with advanced NSCLC exhibiting a mutation of exon 19 or 21 will benefit from EGFR-tyrosine kinase inhibitors (TKI. The presence of an EGFR resistance mutation, such as T790M in EGFR-TKI-naïve patients, is seldom looked for and is related either to a germinal mutation or to somatically mutated subclones. It has a negative predictive impact. We present the case of a patient with a lung papillary adenocarcinoma and miliary intrapulmonary metastases whose tumor displays a somatic complex heterozygous EGFR mutation, combining L858R (exon 21 and a primary resistance mutation T790M (exon 20, both detected by direct sequencing.

  17. Amplification of the MET receptor drives resistance to anti-EGFR therapies in colorectal cancer

    Science.gov (United States)

    Bardelli, Alberto; Corso, Simona; Bertotti, Andrea; Hobor, Sebastijan; Valtorta, Emanuele; Siravegna, Giulia; Sartore-Bianchi, Andrea; Scala, Elisa; Cassingena, Andrea; Zecchin, Davide; Apicella, Maria; Migliardi, Giorgia; Galimi, Francesco; Lauricella, Calogero; Zanon, Carlo; Perera, Timothy; Veronese, Silvio; Corti, Giorgio; Amatu, Alessio; Gambacorta, Marcello; Diaz, Luis A.; Sausen, Mark; Velculescu, Victor E.; Comoglio, Paolo; Trusolino, Livio; Di Nicolantonio, Federica; Giordano, Silvia; Siena, Salvatore

    2014-01-01

    EGFR targeted monoclonal antibodies are effective in a subset of metastatic colorectal tumors (mCRC). Inevitably, all patients develop resistance, which occurs through emergence of KRAS mutations in approximately 50% of the cases. We show that amplification of the MET proto-oncogene is associated with acquired resistance in patients who do not develop KRAS mutations during anti-EGFR therapy. Amplification of the MET locus was present in circulating tumor DNA before relapse was clinically evident. Functional studies demonstrate that MET activation confers resistance to anti-EGFR therapy both in vitro and in vivo. Notably, in patient-derived CRC xenografts, MET amplification correlated with resistance to EGFR blockade which could be overcome by MET kinase inhibitors. These results highlight the role of MET in mediating primary and secondary resistance to anti-EGFR therapies in CRC and encourage the use of MET inhibitors in patients displaying resistance as a result of MET amplification. PMID:23729478

  18. Routine implementation of EGFR mutation testing in clinical practice in Flanders: 'HERMES' project.

    Science.gov (United States)

    Janssens, A; De Droogh, E; Lefebure, A; Kockx, M; Pauwels, P; Germonpre, P; van Meerbeeck, J P

    2014-04-01

    Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) is the recommended first-line treatment in metastatic EGFR-mutation-positive non-small cell lung cancer (NSCLC) patients. Such a personalized treatment requires fast EGFR mutation testing. This study was performed to determine the turn around time (TAT) for EGFR mutation testing on tumour samples of NSCLC in the clinical care in the region of Antwerp (Belgium). The secondary aim was to determine the frequency of EGFR mutations in this Flemish population. Tumour tissue was prospectively obtained from lung cancer patients in participating hospitals and sent from the local pathology laboratory (lab) to two central laboratories (labs) where EGFR-mutation analysis was performed. Results were returned from the central labs to the clinicians and the local pathology lab. TAT was defined as the interval between the request from the oncologist and the result obtained by the oncologist. One hundred and seven specimens were analysed. The clinician got the result from the local lab in a median time of 10 days (3-37 days) and from the central lab in 9 days (3-29 days). We detected seven mutations (7%) in this study population, all occurring in tumours with an adenocarcinoma histology, four (57%) in men and five (71%) in (ex-)smokers. There were six exon 19 deletions and one L858R mutation. It is possible to implement EGFR-mutation testing with timely reporting of the EGFR-mutation status. EGFR-mutation occurs in 7% of Flemish patients with NSCLC. Patients with advanced non-squamous NSCLC should be tested for EGFR mutation regardless of their gender and smoking history. PMID:24724747

  19. Epidermal Growth Factor Receptor (EGFR gene copy number (GCN correlates with clinical activity of irinotecan-cetuximab in K-RAS wild-type colorectal cancer: a fluorescence in situ (FISH and chromogenic in situ hybridization (CISH analysis

    Directory of Open Access Journals (Sweden)

    Scartozzi Mario

    2009-08-01

    Full Text Available Abstract Background K-RAS wild type colorectal tumors show an improved response rate to anti-EGFR monoclonal antibodies. Nevertheless 70% to 40% of these patients still does not seem to benefit from this therapeutic approach. FISH EGFR GCN has been previously demonstrated to correlate with clinical outcome of colorectal cancer treated with anti-EGFR monoclonal antibodies. CISH also seemed able to provide accurate EGFR GCN information with the advantage of a simpler and reproducible technique involving immunohistochemistry and light microscopy. Based on these findings we investigated the correlation between both FISH and CISH EGFR GCN and clinical outcome in K-RAS wild-type colorectal cancer treated with irinotecan-cetuximab. Methods Patients with advanced K-RAS wild-type, colorectal cancer receiving irinotecan-cetuximab after failure of irinotecan-based chemotherapy were eligible. A cut-off value for EGFR GCN of 2.6 and 2.12 for FISH and CISH respectively was derived from ROC curve analysis. Results Forty-four patients were available for analysis. We observed a partial remission in 9 (60% and 2 (9% cases with a FISH EGFR GCN ≥ 2.6 and Conclusion FISH and CISH EGFR GCN may both represent effective tools for a further patients selection in K-RAS wild-type colorectal cancer treated with cetuximab.

  20. RAB-7 antagonizes LET-23 EGFR signaling during vulva development in Caenorhabditis elegans.

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    Olga Skorobogata

    Full Text Available The Rab7 GTPase regulates late endosome trafficking of the Epidermal Growth Factor Receptor (EGFR to the lysosome for degradation. However, less is known about how Rab7 activity, functioning late in the endocytic pathway, affects EGFR signaling. Here we used Caenorhabditis elegans vulva cell fate induction, a paradigm for genetic analysis of EGFR/Receptor Tyrosine Kinase (RTK signaling, to assess the genetic requirements for rab-7. Using a rab-7 deletion mutant, we demonstrate that rab-7 antagonizes LET-23 EGFR signaling to a similar extent, but in a distinct manner, as previously described negative regulators such as sli-1 c-Cbl. Epistasis analysis places rab-7 upstream of or in parallel to lin-3 EGF and let-23 EGFR. However, expression of gfp::rab-7 in the Vulva Presursor Cells (VPCs is sufficient to rescue the rab-7(- VPC induction phenotypes indicating that RAB-7 functions in the signal receiving cell. We show that components of the Endosomal Sorting Complex Required for Transport (ESCRT-0, and -I, complexes, hgrs-1 Hrs, and vps-28, also antagonize signaling, suggesting that LET-23 EGFR likely transits through Multivesicular Bodies (MVBs en route to the lysosome. Consistent with RAB-7 regulating LET-23 EGFR trafficking, rab-7 mutants have increased number of LET-23::GFP-positive endosomes. Our data imply that Rab7, by mediating EGFR trafficking and degradation, plays an important role in downregulation of EGFR signaling. Failure to downregulate EGFR signaling contributes to oncogenesis, and thus Rab7 could possess tumor suppressor activity in humans.

  1. Missense Mutations in Exons 18–24 of EGFR in Hepatocellular Carcinoma Tissues

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    Ravat Panvichian

    2015-01-01

    Full Text Available Epidermal growth factor receptor (EGFR, a transmembrane tyrosine kinase receptor, plays important roles in various cancers. In nonsmall cell lung cancer (NSCLC, EGFR mutations cluster around the ATP-binding pocket (exons 18–21 and some of these mutations activate the kinase and induce an increased sensitivity to EGFR-tyrosine kinase inhibitors. Nevertheless, data of EGFR mutations in HCC are limited. In this study, we investigated EGFR expression by immunohistochemistry and EGFR mutations (exons 18–24 by PCR cloning and sequencing. EGFR overexpression in HCC and matched nontumor tissues were detected in 13/40 (32.5% and 10/35 (28.6%, respectively. Moreover, missense and silent mutations were detected in 13/33 (39.4% and 11/33 (33.3% of HCC tissues, respectively. The thirteen different missense mutations were p.L730P, p.V742I, p.K757E, p.I780T, p.N808S, p.R831C, p.V851A, p.V897A, p.S912P, p.P937L, p.T940A, p.M947V, and p.M947T. We also found already known SNP, p.Q787Q (CAG>CAA, in 13/33 (39.4% of HCC tissues. However, no significant association was detected between EGFR mutations and EGFR overexpression, tissue, age, sex, tumor size, AFP, HBsAg, TP53, and Ki-67. Further investigation is warranted to validate the frequency and activity of these missense mutations, as well as their roles in HCC tumorigenesis and in EGFR-targeted therapy.

  2. Molecular determinants of epidermal growth factor binding: a molecular dynamics study.

    Science.gov (United States)

    Sanders, Jeffrey M; Wampole, Matthew E; Thakur, Mathew L; Wickstrom, Eric

    2013-01-01

    The epidermal growth factor receptor (EGFR) is a member of the receptor tyrosine kinase family that plays a role in multiple cellular processes. Activation of EGFR requires binding of a ligand on the extracellular domain to promote conformational changes leading to dimerization and transphosphorylation of intracellular kinase domains. Seven ligands are known to bind EGFR with affinities ranging from sub-nanomolar to near micromolar dissociation constants. In the case of EGFR, distinct conformational states assumed upon binding a ligand is thought to be a determining factor in activation of a downstream signaling network. Previous biochemical studies suggest the existence of both low affinity and high affinity EGFR ligands. While these studies have identified functional effects of ligand binding, high-resolution structural data are lacking. To gain a better understanding of the molecular basis of EGFR binding affinities, we docked each EGFR ligand to the putative active state extracellular domain dimer and 25.0 ns molecular dynamics simulations were performed. MM-PBSA/GBSA are efficient computational approaches to approximate free energies of protein-protein interactions and decompose the free energy at the amino acid level. We applied these methods to the last 6.0 ns of each ligand-receptor simulation. MM-PBSA calculations were able to successfully rank all seven of the EGFR ligands based on the two affinity classes: EGF>HB-EGF>TGF-α>BTC>EPR>EPG>AR. Results from energy decomposition identified several interactions that are common among binding ligands. These findings reveal that while several residues are conserved among the EGFR ligand family, no single set of residues determines the affinity class. Instead we found heterogeneous sets of interactions that were driven primarily by electrostatic and Van der Waals forces. These results not only illustrate the complexity of EGFR dynamics but also pave the way for structure-based design of therapeutics targeting EGF

  3. Molecular determinants of epidermal growth factor binding: a molecular dynamics study.

    Directory of Open Access Journals (Sweden)

    Jeffrey M Sanders

    Full Text Available The epidermal growth factor receptor (EGFR is a member of the receptor tyrosine kinase family that plays a role in multiple cellular processes. Activation of EGFR requires binding of a ligand on the extracellular domain to promote conformational changes leading to dimerization and transphosphorylation of intracellular kinase domains. Seven ligands are known to bind EGFR with affinities ranging from sub-nanomolar to near micromolar dissociation constants. In the case of EGFR, distinct conformational states assumed upon binding a ligand is thought to be a determining factor in activation of a downstream signaling network. Previous biochemical studies suggest the existence of both low affinity and high affinity EGFR ligands. While these studies have identified functional effects of ligand binding, high-resolution structural data are lacking. To gain a better understanding of the molecular basis of EGFR binding affinities, we docked each EGFR ligand to the putative active state extracellular domain dimer and 25.0 ns molecular dynamics simulations were performed. MM-PBSA/GBSA are efficient computational approaches to approximate free energies of protein-protein interactions and decompose the free energy at the amino acid level. We applied these methods to the last 6.0 ns of each ligand-receptor simulation. MM-PBSA calculations were able to successfully rank all seven of the EGFR ligands based on the two affinity classes: EGF>HB-EGF>TGF-α>BTC>EPR>EPG>AR. Results from energy decomposition identified several interactions that are common among binding ligands. These findings reveal that while several residues are conserved among the EGFR ligand family, no single set of residues determines the affinity class. Instead we found heterogeneous sets of interactions that were driven primarily by electrostatic and Van der Waals forces. These results not only illustrate the complexity of EGFR dynamics but also pave the way for structure-based design of

  4. A case of EGFR mutant lung adenocarcinoma that acquired resistance to EGFR-tyrosine kinase inhibitors with MET amplification and epithelial-to-mesenchymal transition

    Directory of Open Access Journals (Sweden)

    Miyoshi S

    2015-04-01

    Full Text Available Seigo Miyoshi,1 Takahide Kato,1 Hitoshi Katayama,1 Ryoji Ito,1 Yosuke Mizuno,2 Takafumi Okura,1 Jitsuo Higaki1 1Department of Cardiology, Pulmonology, Hypertension and Nephrology, Ehime University Graduate School of Medicine, Ehime, Japan; 2Department of Molecular Pathology, Ehime University Graduate School of Medicine, Ehime, Japan Abstract: EGFR mutant lung cancer responds to EGFR tyrosine kinase inhibitors (TKIs, but all patients eventually develop resistance to EGFR-TKIs. Herein we report a case of EGFR mutant lung adenocarcinoma that acquired resistance to EGFR-TKI with MET amplification and epithelial-to-mesenchymal transition (EMT. A 73-year-old woman was diagnosed with adenocarcinoma harboring an EGFR exon 19 deletion. She received gefitinib as second-line therapy. Tumors were reduced 1 month after gefitinib therapy. However, only a few months later, chest computed tomography results indicated cancer progression. Gefitinib therapy was stopped and docetaxel therapy started. However, she died 13 days after admission. Microscopic examination of postmortem specimens revealed a diffuse proliferation of atypical giant cells in primary and metastatic lesions, but no adenocarcinomatous components as in the antemortem specimens. Immunohistochemical analyses showed that antemortem tumor specimens were positive for CDH1 but negative for VIM. In contrast, postmortem tumor specimens were positive for VIM but negative for CDH1. Genetic analyses revealed MET amplification. We concluded that resistance to EGFR-TKI might be caused by MET amplification and EMT. To our knowledge, no clinical studies have reported that MET amplification and EMT together may be associated with acquired resistance to EGFR-TKI. Second biopsy after the development of EGFR-TKI resistance may be recommended to determine the best therapeutic strategy. Keywords: epidermal growth factor receptor tyrosine kinase inhibitor, MET amplification, epithelial-to-mesenchymal transition

  5. Quantitative PET of EGFR expression in xenograft-bearing mice using {sup 64}Cu-labeled cetuximab, a chimeric anti-EGFR monoclonal antibody

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    Cai, Weibo; Chen, Kai; He, Lina; Cao, Qizhen; Chen, Xiaoyuan [Stanford University School of Medicine, The Molecular Imaging Program at Stanford (MIPS), Department of Radiology and Bio-X Program, Stanford, CA (United States); Koong, Albert [Stanford University School of Medicine, Department of Radiation Oncology, Stanford, CA (United States)

    2007-06-15

    Cetuximab, a chimeric monoclonal antibody targeting epidermal growth factor receptor (EGFR) on the surface of cancer cells, was approved by the FDA to treat patients with metastatic colorectal cancer. It is currently also in advanced-stage development for the treatment of several other solid tumors. Here we report for the first time the quantitative positron emission tomography (PET) imaging of EGFR expression in xenograft-bearing mice using {sup 64}Cu-labeled cetuximab. We conjugated cetuximab with macrocyclic chelating agent 1,4,7,10-tetraazadodecane-N,N',N'',N'''-tetraacetic acid (DOTA), labeled with {sup 64}Cu, and tested the resulting {sup 64}Cu-DOTA-cetuximab in seven xenograft tumor models. The tracer uptake measured by PET was correlated with the EGFR expression quantified by western blotting. The estimated human dosimetry based on the PET data in Sprague-Dawley rats was also calculated. MicroPET imaging showed that {sup 64}Cu-DOTA-cetuximab had increasing tumor activity accumulation over time in EGFR-positive tumors but relatively low uptake in EGFR-negative tumors at all times examined (<5%ID/g). There was a good correlation (R {sup 2} = 0.80) between the tracer uptake (measured by PET) and the EGFR expression level (measured by western blotting). Human dosimetry estimation indicated that the tracer may be safely administered to human patients for tumor diagnosis, with the dose-limiting organ being the liver. The success of EGFR-positive tumor imaging using {sup 64}Cu-DOTA-cetuximab can be translated into the clinic to characterize the pharmacokinetics, to select the right population of patients for EGFR-targeted therapy, to monitor the therapeutic efficacy of anti-EGFR treatment, and to optimize the dosage of either cetuximab alone or cetuximab in combination with other therapeutic agents. (orig.)

  6. Dacomitinib in lung cancer: a “lost generation” EGFR tyrosine-kinase inhibitor from a bygone era?

    Science.gov (United States)

    Ou, Sai-Hong Ignatius; Soo, Ross A

    2015-01-01

    EGFR tyrosine-kinase inhibitors (TKIs) have now been firmly established as the first-line treatment for non-small-cell lung cancer (NSCLC) patients harboring activating EGFR mutations, based on seven prospective randomized Phase III trials. However, despite significantly improved overall response rate and improved median progression-free survival when compared to platinum-doublet chemotherapy, EGFR-mutant NSCLC patients treated with EGFR TKIs invariably progress due to the emergence of acquired resistances, with the gatekeeper T790M mutation accounting for up to 60% of the resistance mechanisms. Second-generation irreversible EGFR TKIs were developed in part to inhibit the T790M mutation, in addition to the common activating EGFR mutations. Dacomitinib is one such second-generation EGFR TKI designed to inhibit both the wild-type (WT) EGFR and EGFR T790M. Afatinib is another second-generation EGR TKI that has been now been approved for the first-line treatment of EGFR-mutant NSCLC patients, while dacomitinib continues to undergo clinical evaluation. We will review the clinical development of dacomitinib from Phase I to Phase III trials, including the two recently published negative large-scale randomized Phase III trials (ARCHER 1009, NCIC-BR-26). Results from another large-scale randomized trial (ARCHER 1050) comparing dacomitinib to gefitinib as first-line treatment of advanced treatment-naïve EGFR-mutant NSCLC patients will soon be available and will serve as the lynchpin trial for the potential approval of dacomitinib in NSCLC. Meanwhile, third-generation EGFR TKIs (eg, CO-1686 [rociletinib], AZ9291, HM61713, EGF816, and ASP8273) that preferentially and potently inhibit EGFR T790M but not WT EGFR are in full-scale clinical development, and some of these EGFR TKIs have received “breakthrough” designation by the US Food and Drug Administration and will likely be approved in late 2015. Given the rapid development of third-generation EGFR TKIs and the approval

  7. NOX4 mediates cytoprotective autophagy induced by the EGFR inhibitor erlotinib in head and neck cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Sobhakumari, Arya [Interdisciplinary Graduate Program in Human Toxicology, The University of Iowa, Iowa City, IA (United States); Department of Pathology, The University of Iowa, Iowa City, IA (United States); Schickling, Brandon M. [Department of Internal Medicine, The University of Iowa, Iowa City, IA (United States); Love-Homan, Laurie; Raeburn, Ayanna [Department of Pathology, The University of Iowa, Iowa City, IA (United States); Fletcher, Elise V.M. [Interdisciplinary Graduate Program in Human Toxicology, The University of Iowa, Iowa City, IA (United States); Department of Pathology, The University of Iowa, Iowa City, IA (United States); Case, Adam J. [Free Radical and Radiation Biology Program, Department of Radiation Oncology, The University of Iowa, Iowa City, IA (United States); Domann, Frederick E. [Interdisciplinary Graduate Program in Human Toxicology, The University of Iowa, Iowa City, IA (United States); Department of Pathology, The University of Iowa, Iowa City, IA (United States); Free Radical and Radiation Biology Program, Department of Radiation Oncology, The University of Iowa, Iowa City, IA (United States); Holden Comprehensive Cancer Center, University of Iowa Hospitals and Clinics (UIHC), Iowa City, IA (United States); Miller, Francis J. [Department of Internal Medicine, The University of Iowa, Iowa City, IA (United States); Free Radical and Radiation Biology Program, Department of Radiation Oncology, The University of Iowa, Iowa City, IA (United States); Holden Comprehensive Cancer Center, University of Iowa Hospitals and Clinics (UIHC), Iowa City, IA (United States); and others

    2013-11-01

    Most head and neck squamous cell carcinomas (HNSCCs) overexpress epidermal growth factor receptor (EGFR) and EGFR inhibitors are routinely used in the treatment of HNSCC. However, many HNSCC tumors do not respond or become refractory to EGFR inhibitors. Autophagy, which is a stress-induced cellular self-degradation process, has been reported to reduce the efficacy of chemotherapy in various disease models. The purpose of this study is to determine if the efficacy of the EGFR inhibitor erlotinib is reduced by activation of autophagy via NOX4-mediated oxidative stress in HNSCC cells. Erlotinib induced the expression of the autophagy marker LC3B-II and autophagosome formation in FaDu and Cal-27 cells. Inhibition of autophagy by chloroquine and knockdown of autophagy pathway genes Beclin-1 and Atg5 sensitized both cell lines to erlotinib-induced cytotoxicity, suggesting that autophagy may serve as a protective mechanism. Treatment with catalase (CAT) and diphenylene iodonium (DPI) in the presence of erlotinib suppressed the increase in LC3B-II expression in FaDu and Cal-27 cells. Erlotinib increased NOX4 mRNA and protein expression by increasing its promoter activity and mRNA stability in FaDu cells. Knockdown of NOX4 using adenoviral siNOX4 partially suppressed erlotinib-induced LC3B-II expression, while overexpression of NOX4 increased expression of LC3B-II. These studies suggest that erlotinib may activate autophagy in HNSCC cells as a pro-survival mechanism, and NOX4 may play a role in mediating this effect. - Highlights: • Erlotinib increased LC3B-II and autophagosome formation in HNSCC cells. • Inhibition of autophagy sensitized HNSCC cells to erlotinib. • Erlotinib increased NOX4 promoter and 3′UTR luciferase activity. • Manipulating NOX4 decreases or increases autophagy.

  8. Concurrent hypermethylation of DNMT1, MGMT and EGFR genes in progression of gliomas

    Directory of Open Access Journals (Sweden)

    Gömöri Éva

    2012-01-01

    Full Text Available Abstract Background Gliomas are the most common neoplasm of the brain. High-grade gliomas often resist treatment even with aggressive surgical resection and adjuvant radiation and chemotherapy. Despite the combined treatment, they frequently recur with the same or higher-grade histology. Genetic instability is commonly associated with inactivation of the normal DNA repair function and tumour suppressor genes as well as activation of oncogenes resulting from alterations of promoter hypermethylation, but the molecular mechanisms of the histological and clinical progression of gliomas are still poorly understood. Methods This study involved longitudinal analysis samples of primary and recurrent gliomas to determine whether the progression of low- and high-grade gliomas is associated with the promoter methylation of the DNMT1, MGMT and EGFR genes by PCR-based restriction enzyme assay. Epigenetic inactivation of these three important glioma-associated genes was analyzed in paired biopsy samples from 18 patients with tumour recurrence. Results The methylation analysis of the CpG sites in the DNA methyltransferase (DNMT1 promoter revealed a total of 6 hypermethylations (6/18, the methylguanine-DNA methyltransferase (MGMT promoter revealed a total of 10 hypermethylations (10/18 and the epithelial grow factor receptor (EGFR promoter revealed a total of 12 (12/18 hypermethylations respectively in recurrent gliomas. The results demonstrated that DNMT1 promoter hypermethylation does not occur in low-grade gliomas, it was mainly observed in secondary glioblastomas. Additionally, the MGMT and EGFR promoter was hypermethylated in both low-and high-grade GLs and their corresponding histological transformed GLs. Conclusion This study has provided further evidence that the histological transformation and progression of gliomas may be associated with the inactivation of the EGFR and MGMT genes. It seems that EGFR and MGMT promoter hypermethylations are early

  9. Thrombin stimulates VSMC proliferation through an EGFR-dependent pathway: involvement of MMP-2.

    Science.gov (United States)

    Smiljanic, Katarina; Obradovic, Milan; Jovanovic, Aleksandra; Djordjevic, Jelena; Dobutovic, Branislava; Jevremovic, Danimir; Marche, Pierre; Isenovic, Esma R

    2014-11-01

    In this study, the role of epidermal growth factor receptor (EGFR), extracellular signal-regulated kinase (ERK1/2), heparin-binding EGF-like growth factor (HB-EGF), general metalloproteinases, matrix metalloproteinases-2 (MMP-2) in mediating the mitogenic action of thrombin in rat vascular smooth muscle cells (VSMC) was investigated. The incubation of rat VSMC with thrombin (1 U/ml) for 5 min resulted in significant (p EGFR phosphorylation by 8.5 ± 1.3-fold (p EGFR tyrosine kinase irreversible inhibitor, 10 µM PD169540 (PD), and 20 µM anti-HB-EGF antibody significantly reduced thrombin-stimulated EGFR and ERK1/2 phosphorylation by 81, 72 % and by 48 and 61 %, respectively. Furthermore, the same pretreatments with PD or anti-HB-EGF antibody reduced thrombin-induced VSMC proliferation by 44 and 45 %, respectively. In addition, 30-min pretreatments with 10 µM specific MMP-2 inhibitor significantly reduced thrombin-stimulated phosphorylation of both EGFR and ERK1/2 by 25 %. Moreover, the same pretreatment with MMP-2 inhibitor reduced thrombin-induced VSMC proliferation by 45 %. These results show that the thrombin-induced DNA synthesis correlates with the level of ERK1/2 activation rather than EGFR activation. These results further suggest that thrombin acts through EGFR and ERK 1/2 signaling pathways involving MMP-2 to upregulate proliferation of VSMC.

  10. 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

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

    Science.gov (United States)

    Akhtar, Saghir; Al-Zaid, Bashayer; El-Hashim, Ahmed Z; Chandrasekhar, Bindu; Attur, Sreeja; Yousif, Mariam H M; Benter, Ibrahim F

    2015-01-01

    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 dendrimers

  12. Ultraviolet-B induced expression of hypoxia-inducible factor 1α,transferrin receptor through EGFR/PI3K/AKT/DEC1 pathway

    Institute of Scientific and Technical Information of China (English)

    LI Yanhua; BI Zhigang

    2007-01-01

    The aim of this research was to explore the effects and signaling pathway of ultraviolet-B (UVB) irradiation on the expression of hypoxia-inducible factor 1α (HIF-1α) and transferrin receptor (TfR).HIF-1α protein was measured by Western blot method.Expressions of epidermal growth factor receptor (EGFR),phosphor-EGF-R and TfR after UVB irradiation were determined with flow cytometry.After UVB irradiation,mRNA levels of HIF-1α and TfR were detected by real time-PCR.Results showed that compared with control groups,UVB was able to induce HIF1α and TfR protein expression in a dose- and time-dependent manner in HaCat cells (P < 0.05).TfR mRNA was expressed in a dose-dependent manner and reached a peak at the 8th hour in HaCat cells (P<0.05) whereas HIF-1α mRNA expression was not affected by UVB treatment (P>0.05).The EGFR/PI3K/AKT signaling pathway was required for the induction of HIF-lcx and TfR expression induced by UVB.UVB induced activation of EGFR in HaCat cells and EGFR regulated expression of TfR and HIF-1α.EGFR (-/-) MEF did not increase the HIF 1 expression following UVB irradiation (P>0.05).In contrast,EGFR (+/+) MEF strongly enhanced HIF 1α expression after UVB irradiation (P < 0.05).PD153035,a selective inhibitor of EGFR tyrosine kinase,inhibited the TfR protein expression in UVB-treated cells in a dose-dependent manner (P<0.05).PI3K inhibitors,LY294002 and wortmannin,inhibited HIF-1α and TfR expressions induced by UVB (P < 0.05).The DEC1 (-/-) Ha-Cat cells did not increase their TfR and HIF-1α expressions following UVB irradiation (P>0.05).In contrast,DEC1 (+/+) HaCat cells strongly enhanced TfR and HIF-1α protein expression after UVB irradiation (P<0.05).We conclude that UVB induces TfR and HIF-1α expressions via EGFR/PI3K/AKT/DEC1 signaling pathway.

  13. Using the MCF10A/MCF10CA1a Breast Cancer Progression Cell Line Model to Investigate the Effect of Active, Mutant Forms of EGFR in Breast Cancer Development and Treatment Using Gefitinib.

    Directory of Open Access Journals (Sweden)

    Darrell C Bessette

    Full Text Available Basal-like and triple negative breast cancer (TNBC share common molecular features, poor prognosis and a propensity for metastasis to the brain. Amplification of epidermal growth factor receptor (EGFR occurs in ~50% of basal-like breast cancer, and mutations in the epidermal growth factor receptor (EGFR have been reported in up to ~ 10% of Asian TNBC patients. In non-small cell lung cancer several different mutations in the EGFR tyrosine kinase domain confer sensitivity to receptor tyrosine kinase inhibitors, but the tumourigenic potential of EGFR mutations in breast cells and their potential for targeted therapy is unknown.Constructs containing wild type, G719S or E746-A750 deletion mutant forms of EGFR were transfected into the MCF10A breast cells and their tumorigenic derivative, MCF10CA1a. The effects of EGFR over-expression and mutation on proliferation, migration, invasion, response to gefitinib, and tumour formation in vivo was investigated. Copy number analysis and whole exome sequencing of the MCF10A and MCF10CA1a cell lines were also performed.Mutant EGFR increased MCF10A and MCF10CA1a proliferation and MCF10A gefitinib sensitivity. The EGFR-E746-A750 deletion increased MCF10CA1a cell migration and invasion, and greatly increased MCF10CA1a xenograft tumour formation and growth. Compared to MCF10A cells, MCF10CA1a cells exhibited large regions of gain on chromosomes 3 and 9, deletion on chromosome 7, and mutations in many genes implicated in cancer.Mutant EGFR enhances the oncogenic properties of MCF10A cell line, and increases sensitivity to gefitinib. Although the addition of EGFR E746-A750 renders the MCF10CA1a cells more tumourigenic in vivo it is not accompanied by increased gefitinib sensitivity, perhaps due to additional mutations, including the PIK3CA H1047R mutation, that the MCF10CA1a cell line has acquired. Screening TNBC/basal-like breast cancer for EGFR mutations may prove useful for directing therapy but, as in non

  14. Targeting the EGFR/PCNA signaling suppresses tumor growth of triple-negative breast cancer cells with cell-penetrating PCNA peptides.

    Directory of Open Access Journals (Sweden)

    Yung-Luen Yu

    Full Text Available Tyrosine 211 (Y211 phosphorylation of proliferation cell nuclear antigen (PCNA coincides with pronounced cancer cell proliferation and correlates with poor survival of breast cancer patients. In epidermal growth factor receptor (EGFR tyrosine kinase inhibitor (TKI-resistant cells, both nuclear EGFR (nEGFR expression and PCNA Y211 phosphorylation are increased. Moreover, the resistance to EGFR TKI is a major clinical problem in treating EGFR-overexpressing triple-negative breast cancer (TNBC. Thus, effective treatment to combat resistance is urgently needed. Here, we show that treatment of cell-penetrating PCNA peptide (CPPP inhibits growth and induces apoptosis of human TNBC cells. The Y211F CPPP specifically targets EGFR and competes directly for PCNA tyrosine Y211 phosphorylation and prevents nEGFR from binding PCNA in vivo; it also suppresses tumor growth by sensitizing EGFR TKI resistant cells, which have enhanced nEGFR function and abrogated classical EGFR membrane signaling. Furthermore, we identify an active motif of CPPP, RFLNFF (RF6 CPPP, which is necessary and sufficient to inhibit TKI-resistant TNBC cell growth of orthotopic implanted tumor in mice. Finally, the activity of its synthetic retro-inverted derivative, D-RF6 CPPP, on an equimolar basis, is more potent than RF6 CPPP. Our study reveals a drug candidate with translational potential for the future development of safe and effective therapeutic for EGFR TKI resistance in TNBC.

  15. Rationale and Study Design of the IRENE-Trial (NVALT-16) : A Phase II Trial to Evaluate Iressa Rechallenge in Advanced NSCLC Patients With an Activating EGFR Mutation Who Responded to an EGFR-TKI Used As First-Line or Previous Treatment

    NARCIS (Netherlands)

    Kuiper, Justine L.; Heideman, Danielle A. M.; Wurdinger, Tom; Grunberg, Katrien; Groen, Harry J. M.; Smit, Egbert F.

    2015-01-01

    Background: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have shown improved progression-free survival (PFS) and overall survival (OS) over chemotherapy in a molecularly defined subgroup of advanced non-small-cell lung cancer (NSCLC) patients (ie, patients with an activa

  16. 5-Fluorocytosine combined with Fcy-hEGF fusion protein targets EGFR-expressing cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Lan, Keng-Hsueh [Division of Radiation Oncology, Department of Oncology, National Taiwan University Hospital, Taipei 100, Taiwan (China); Shih, Yi-Sheng [Cancer Center, Taipei Veterans General Hospital, Taipei 112, Taiwan (China); Chang, Cheng Allen [Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, Taipei 112, Taiwan (China); School of Biomedical Science and Engineering, National Yang-Ming University, Taipei 112, Taiwan (China); Yen, Sang-Hue [Cancer Center, Taipei Veterans General Hospital, Taipei 112, Taiwan (China); Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, Taipei 112, Taiwan (China); Lan, Keng-Li, E-mail: kllan@vghtpe.gov.tw [Cancer Center, Taipei Veterans General Hospital, Taipei 112, Taiwan (China); Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, Taipei 112, Taiwan (China)

    2012-11-16

    Highlights: Black-Right-Pointing-Pointer EGFR-expressing epithelial cancers account for significant portion of cancer deaths. Black-Right-Pointing-Pointer EGF-EGFR signaling pathway is validated as an important anticancer drug target. Black-Right-Pointing-Pointer EGF and Fcy fusion protein (Fcy-hEGF) can bind to EGFR and convert 5-FC to 5-FU. Black-Right-Pointing-Pointer Fcy-hEGF combined with 5-FC preferentially inhibits EGFR-expressing cells viability. -- Abstract: Human epithelial cancers account for approximately 50% of all cancer deaths. This type of cancer is characterized by excessive activation and expression of the epidermal growth factor receptor (EGFR). The EGFR pathway is critical for cancer cell proliferation, survival, metastasis and angiogenesis. The EGF-EGFR signaling pathway has been validated as an important anticancer drug target. Increasing numbers of targeted therapies against this pathway have been either approved or are currently under development. Here, we adopted a prodrug system that uses 5-fluorocytosine (5-FC) and human EGF (hEGF) fused with yeast cytosine deaminase (Fcy) to target EGFR-overexpressing cancer cells and to convert 5-FC to a significantly more toxic chemotherapeutic, 5-fluorouracil (5-FU). We cloned and purified the Fcy-hEGF fusion protein from Pichia pastoris yeast. This fusion protein specifically binds to EGFR with a similar affinity as hEGF, approximately 10 nM. Fcy-hEGF binds tightly to A431 and MDA-MB-468 cells, which overexpress EGFR, but it binds with a lower affinity to MDA-MB-231 and MCF-7, which express lower levels of EGFR. Similarly, the viability of EGFR-expressing cells was suppressed by Fcy-hEGF in the presence of increasing concentrations of 5-FC, and the IC{sub 50} values for A431 and MDA-MB-468 were approximately 10-fold lower than those of MDA-MB-231 and MCF-7. This novel prodrug system, Fcy-hEGF/5-FC, might represent a promising addition to the available class of inhibitors that specifically target EGFR

  17. 5-Fluorocytosine combined with Fcy–hEGF fusion protein targets EGFR-expressing cancer cells

    International Nuclear Information System (INIS)

    Highlights: ► EGFR-expressing epithelial cancers account for significant portion of cancer deaths. ► EGF–EGFR signaling pathway is validated as an important anticancer drug target. ► EGF and Fcy fusion protein (Fcy–hEGF) can bind to EGFR and convert 5-FC to 5-FU. ► Fcy–hEGF combined with 5-FC preferentially inhibits EGFR-expressing cells viability. -- Abstract: Human epithelial cancers account for approximately 50% of all cancer deaths. This type of cancer is characterized by excessive activation and expression of the epidermal growth factor receptor (EGFR). The EGFR pathway is critical for cancer cell proliferation, survival, metastasis and angiogenesis. The EGF–EGFR signaling pathway has been validated as an important anticancer drug target. Increasing numbers of targeted therapies against this pathway have been either approved or are currently under development. Here, we adopted a prodrug system that uses 5-fluorocytosine (5-FC) and human EGF (hEGF) fused with yeast cytosine deaminase (Fcy) to target EGFR-overexpressing cancer cells and to convert 5-FC to a significantly more toxic chemotherapeutic, 5-fluorouracil (5-FU). We cloned and purified the Fcy–hEGF fusion protein from Pichia pastoris yeast. This fusion protein specifically binds to EGFR with a similar affinity as hEGF, approximately 10 nM. Fcy–hEGF binds tightly to A431 and MDA-MB-468 cells, which overexpress EGFR, but it binds with a lower affinity to MDA-MB-231 and MCF-7, which express lower levels of EGFR. Similarly, the viability of EGFR-expressing cells was suppressed by Fcy–hEGF in the presence of increasing concentrations of 5-FC, and the IC50 values for A431 and MDA-MB-468 were approximately 10-fold lower than those of MDA-MB-231 and MCF-7. This novel prodrug system, Fcy–hEGF/5-FC, might represent a promising addition to the available class of inhibitors that specifically target EGFR-expressing cancers.

  18. Psychological determinants of information searching activity.

    Science.gov (United States)

    Gorunova, L

    2012-01-01

    The paper deals with the application of the activity theory in describing psychological determinants of the information searching activity. The notions of information behavior, information retrieval, information competence, information retrieval activity given in Russian and English scientific literature are compared. The research approach to the information retrieval activity based on the principles developed in the Russian theory of activity is described; and the fundamentals of G. Sukhodolsky's generalized conception of activity are presented for the first time. Analysis of empirical researches showed that specific features of information search depend on how the user evaluates information resources, information, conditions and results of search. Psychological determiners of information search may be detected as the system of evaluative alternatives, which is generated by the user during the process of his experience growth. We discovered that user's evaluation system is also related to his individual typological and personal regulative features and determines the choice of the search strategy.

  19. Reggies/flotillins regulate E-cadherin-mediated cell contact formation by affecting EGFR trafficking.

    Science.gov (United States)

    Solis, Gonzalo P; Schrock, Yvonne; Hülsbusch, Nikola; Wiechers, Marianne; Plattner, Helmut; Stuermer, Claudia A O

    2012-05-01

    The reggie/flotillin proteins are implicated in membrane trafficking and, together with the cellular prion protein (PrP), in the recruitment of E-cadherin to cell contact sites. Here, we demonstrate that reggies, as well as PrP down-regulation, in epithelial A431 cells cause overlapping processes and abnormal formation of adherens junctions (AJs). This defect in cell adhesion results from reggie effects on Src tyrosine kinases and epidermal growth factor receptor (EGFR): loss of reggies reduces Src activation and EGFR phosphorylation at residues targeted by Src and c-cbl and leads to increased surface exposure of EGFR by blocking its internalization. The prolonged EGFR signaling at the plasma membrane enhances cell motility and macropinocytosis, by which junction-associated E-cadherin is internalized and recycled back to AJs. Accordingly, blockage of EGFR signaling or macropinocytosis in reggie-deficient cells restores normal AJ formation. Thus, by promoting EGFR internalization, reggies restrict the EGFR signaling involved in E-cadherin macropinocytosis and recycling and regulate AJ formation and dynamics and thereby cell adhesion.

  20. Dermatologic Toxicities from Monoclonal Antibodies and Tyrosine Kinase Inhibitors against EGFR: Pathophysiology and Management

    Directory of Open Access Journals (Sweden)

    Shaad E. Abdullah

    2012-01-01

    Full Text Available Epidermal growth factor receptor (EGFR inhibition has now been well established as an effective treatment for various cancers. The EGFR belongs to the ErbB family of tyrosine kinase receptors which regulate tumor cell differentiation, survival and proliferation. Activation of EGFR drives tumorigenesis in lung, head and neck, colorectal and pancreatic cancers. Irrespective of the type of cancer being treated and the mechanism by which tumor EGFR drives tumorigenesis, the major side effect of EGFR inhibition is a papulopustular (also described as maculopapular or acneiform rash which occurs in about two thirds of treated patients. Interestingly, this rash has been commonly correlated with better clinical outcomes (objective tumor response and patient survival. The pathophysiology of dermatological toxicity from EGFR inhibitors is an important area of clinical research, and the proper management of the rash is essential to increase the therapeutic index from this class of drugs. In this paper, we review the dermatologic toxicities associated with EGFR inhibitors with an emphasis on its pathophysiology and clinical management.

  1. Evaluation of radiolabeled ML04, a putative irreversible inhibitor of epidermal growth factor receptor, as a bioprobe for PET imaging of EGFR-overexpressing tumors

    Energy Technology Data Exchange (ETDEWEB)

    Abourbeh, Galith [Department of Medical Biophysics and Nuclear Medicine, Hadassah Hebrew University, Jerusalem 91120 (Israel); Unit of Cellular Signaling, Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Hebrew University, Jerusalem 91904 (Israel); Dissoki, Samar [Department of Medical Biophysics and Nuclear Medicine, Hadassah Hebrew University, Jerusalem 91120 (Israel); Jacobson, Orit [Department of Medical Biophysics and Nuclear Medicine, Hadassah Hebrew University, Jerusalem 91120 (Israel); Litchi, Amir [Department of Medical Biophysics and Nuclear Medicine, Hadassah Hebrew University, Jerusalem 91120 (Israel); Daniel, Revital Ben [Department of Medical Biophysics and Nuclear Medicine, Hadassah Hebrew University, Jerusalem 91120 (Israel); Laki, Desirediu [Department of Medical Biophysics and Nuclear Medicine, Hadassah Hebrew University, Jerusalem 91120 (Israel); Levitzki, Alexander [Unit of Cellular Signaling, Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Hebrew University, Jerusalem 91904 (Israel); Mishani, Eyal [Department of Medical Biophysics and Nuclear Medicine, Hadassah Hebrew University, Jerusalem 91120 (Israel)]. E-mail: mishani@md.huji.ac.il

    2007-01-15

    Overexpression of epidermal growth factor receptor (EGFR) has been implicated in tumor development and malignancy. Evaluating the degree of EGFR expression in tumors could aid in identifying patients for EGFR-targeted therapies and in monitoring treatment. Nevertheless, no currently available assay can reliably quantify receptor content in tumors. Radiolabeled inhibitors of EGFR-TK could be developed as bioprobes for positron emission tomography imaging. Such imaging agents would not only provide a noninvasive quantitative measurement of EGFR content in tumors but also serve as radionuclide carriers for targeted radiotherapy. The potency, reversibility, selectivity and specific binding characteristics of ML04, an alleged irreversible inhibitor of EGFR, were established in vitro. The distribution of the F-18-labeled compound and the extent of EGFR-specific tumor uptake were evaluated in tumor-bearing mice. ML04 demonstrated potent, irreversible and selective inhibition of EGFR, combined with specific binding to the receptor in intact cells. In vivo distribution of the radiolabeled compound revealed tumor/blood and tumor/muscle activity uptake ratios of about 7 and 5, respectively, 3 h following administration of a radiotracer. Nevertheless, only minor EGFR-specific uptake of the compound was detected in these studies, using either EGFR-negative tumors or blocking studies as controls. To improve the in vivo performance of ML04, administration via prolonged intravenous infusion is proposed. Detailed pharmacokinetic characterization of this bioprobe could assist in the development of a kinetic model that would afford accurate measurement of EGFR content in tumors.

  2. AZD8931, an equipotent, reversible inhibitor of signaling by epidermal growth factor receptor (EGFR), HER2, and HER3: preclinical activity in HER2 non-amplified inflammatory breast cancer models

    OpenAIRE

    Mu, Zhaomei; Klinowska, Teresa; Dong, Xiaoshen; Foster, Emily; Womack, Chris; Fernandez, Sandra V.; Cristofanilli, Massimo

    2014-01-01

    Introduction Epidermal growth factor receptor (EGFR) overexpression has been associated with prognostic and predictive value in inflammatory breast cancer (IBC). Epidermal growth factor receptor 2 (HER2) overexpression is observed at a higher rate in IBC compared with noninflammatory breast cancer. Current clinically available anti-HER2 therapies are effective only in patients with HER2 amplified breast cancer, including IBC. AZD8931 is a novel small-molecule equipotent inhibitor of EGFR, HER...

  3. Protein tyrosine phosphatase SHP-1 sensitizes EGFR/HER-2 positive breast cancer cells to trastuzumab through modulating phosphorylation of EGFR and HER-2

    Directory of Open Access Journals (Sweden)

    Wu YF

    2015-09-01

    Full Text Available Yifen Wu,1,2,* Rong Li,3,* Junyi Zhang,4 Gang Wang,5 Bin Liu,6 Xiaofang Huang,7 Tao Zhang,7 Rongcheng Luo8 1Traditional Chinese Medicine-Integrated Hospital, Southern Medical University, Guangzhou, 2Department of Oncology, Dongguan People’s Hospital, Dongguan, 3Department of Oncology, Nanfang Hospital, 4Department of Oncology, Traditional Chinese Medicine-Integrated Hospital, Southern Medical University, Guangzhou, 5Department of Radiology, Dongguan People’s Hospital, Dongguan, 6Second Affiliated Hospital of Guangzhou Medical College, 7College of Traditional Chinese medicine, Southern Medical University, 8Cancer Center, Traditional Chinese Medicine-Integrated Hospital, Southern Medical University, Guangzhou, Guangdong, People’s Republic of China *These authors contributed equally to this work Background: Trastuzumab resistance in HER-2 positive breast cancer cells is closely related to overexpression of both epidermal growth factor receptor (EGFR and human epidermal receptor (HER-2. SHP-1 has been demonstrated to downregulate tyrosine kinase activity including EGFR via its phosphatase function, but its effect on HER-2 activity is still unknown. Here, we examined the hypothesis that SHP-1 enhances the anticancer efficacy of trastuzumab in EGFR/HER-2 positive breast cancer cells through combining dual inhibition of EGFR and HER-2.Methods: Trastuzumab-resistant breast cancer SKBr-3 cells were generated by long-term in vitro culture of SKBr-3cells in the presence of trastuzumab. The SHP-1 was ectopically expressed by stable transfection. The activity and expression of EGFR, HER-2, and downstream signaling pathways were tested by Western blot. Cell viability was examined by the 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay, and apoptosis was examined by flow cytometry. The binding between SHP-1 and EGFR/HER-2 was evaluated by immunoprecipitation assay and bimolecular fluorescence complementation. The effects of SHP-1

  4. Suppression of PAI-1 expression through inhibition of the EGFR-mediated signaling cascade in rat kidney fibroblast by ascofuranone.

    Science.gov (United States)

    Cho, Hyun-Ji; Kang, Jeong-Han; Kim, Teoan; Park, Kwang-Kyun; Kim, Cheorl-Ho; Lee, In-Seon; Min, Kwan-Sik; Magae, Junji; Nakajima, Hiroo; Bae, Young-Seuk; Chang, Young-Chae

    2009-05-15

    Fibrosis in glomerulosclerosis causes progressive loss of renal function. Transforming growth factor (TGF)-beta, one of the major profibrotic cytokines, induces the synthesis of plasminogen activator inhibitor (PAI)-1, a factor that plays a crucial role in the development of fibrosis. Here, we found that an isoprenoid antibiotic, ascofuranone, suppresses expression of profibrotic factors including matrix proteins and PAI-1 induced by TGF-beta in renal fibroblasts. Ascofuranone selectively inhibits phosphorylation of epidermal growth factor receptor (EGFR), and downstream kinases such as Raf-1, MEK-1/2, and ERK-1/2. PAI-1 transcription also is suppressed by treatment with kinase inhibitors for MEK-1/2 or EGFR, and with small interfering RNA for EGFR. Ascofuranone inhibits cellular metalloproteinase activity, and an inhibitor of metalloproteinases suppresses EGFR phosphorylation and PAI-1 transcription. These results suggest that ascofuranone suppresses expression of profibrotic factors through the inhibition of an EGFR-dependent signal transduction pathway activated by metalloproteinases.

  5. 雌激素激活GPER-EGFR-ERK通路促进人乳腺癌SKBR-3细胞系增殖%Estrogen activates GPER-EGFR-ERK pathway to promote the proliferation of human breast cancer cell line SKBR-3

    Institute of Scientific and Technical Information of China (English)

    李维东; 罗浩军; 李振华; 余腾华; 杨光伦; 涂刚

    2012-01-01

    Objective To explore the effect of estrogen on the proliferation of human breast cancer cell line SKBR-3 mediated fay G protein-coupled estrogen receptor (GPER). Methods Calcium influx, cell growth ability and cell cycle were examined by confocal laser scanning microscopy, CCK-8 assay and flow cytometry in the presence of different concentrations of drugs, respectively. The relative expression level of phospho-extracellular signal-regulated kinase (p-ERK) was detected by Western blotting. Results After the cells were treated with 17-β estradiol (E2) or GFER specific agonist (Gl) , intracellular calcium ion concentration shifted quickly from 120 seconds, and cell proliferation increased significantly with the relative cell numbers (2.08 ±0.07) times and (2.0 ±0.04) times more than those of the control group. The proliferation index (PI) (43. 12 ±0. 38)% and (42.43 ±0.52)% at S phase and G2-M phase increased significantly as compared to the control group (29.19 ± 0. 29) % (P 〈 0. 05). The protein level of p-ERK was higher in the treatment group than in the control group (P 〈 0, 05). GFPE antagonist G15, EGFR antagonist AG1478 and ERK antagonist U0126 rather than PI3K antagonist wortmannin could inhibit the changes induced by E2 or G1. Conclusion Estrogen activates GPER-EGFR-ERK pathway to promote the proliferation of human breast cancer cell line SKBR-3.%目的 探讨G蛋白偶联雌激素受体(G protein-coupled estrogen receptor,GPER)介导雌激素对人乳癌细胞系SKBR-3增殖的影响.方法 激光共聚焦显微镜扫描检测钙离子探针标记的细胞内钙离子浓度随时间的变化,CCK-8法观测细胞的增殖生长,流式细胞术检测细胞周期,Western blot检测磷酸化细胞外信号调节激酶(pbospho-extracellular regulate kinase,p-ERK)的相对表达量.结果 17-β雌二醇(E2)与GPER激动剂(G1)(E2、G1处理组)刺激SKBR-3细胞后,细胞内钙离子浓度从120 s开始迅速升高,细胞增殖与

  6. Determining Enzyme Activity by Radial Diffusion

    Science.gov (United States)

    Davis, Bill D.

    1977-01-01

    Discusses advantages of radial diffusion assay in determining presence of enzyme and/or rough approximation of amount of enzyme activities. Procedures are included for the preparation of starch-agar plates, and the application and determination of enzyme. Techniques using plant materials (homogenates, tissues, ungerminated embryos, and seedlings)…

  7. Sym004, a Novel EGFR Antibody Mixture, Can Overcome Acquired Resistance to Cetuximab1

    OpenAIRE

    Iida, Mari; Brand, Toni M; Starr, Megan M.; Li, Chunrong; Huppert, Evan J; Luthar, Neha; Pedersen, Mikkel W; Horak, Ivan D.; Kragh, Michael; Wheeler, Deric L

    2013-01-01

    The epidermal growth factor receptor (EGFR) is a central regulator of tumor progression in a variety of human cancers. Cetuximab is an anti-EGFR monoclonal antibody that has been approved for head and neck and colorectal cancer treatment, but many patients treated with cetuximab don't respond or eventually acquire resistance. To determine how tumor cells acquire resistance to cetuximab, we previously developed a model of acquired resistance using the non-small cell lung cancer line NCI-H226. ...

  8. Sym004, a Novel EGFR Antibody Mixture, Can Overcome Acquired Resistance to Cetuximab

    OpenAIRE

    Mari Iida; Brand, Toni M; Starr, Megan M.; Chunrong Li; Huppert, Evan J; Neha Luthar; Pedersen, Mikkel W; Horak, Ivan D.; Michael Kragh; Wheeler, Deric L

    2013-01-01

    The epidermal growth factor receptor (EGFR) is a central regulator of tumor progression in a variety of human cancers. Cetuximab is an anti-EGFR monoclonal antibody that has been approved for head and neck and colorectal cancer treatment, but many patients treated with cetuximab don't respond or eventually acquire resistance. To determine how tumor cells acquire resistance to cetuximab, we previously developed a model of acquired resistance using the non-small cell lung cancer line NCI-H226. ...

  9. Determination of proteolytic activity in cereals

    Directory of Open Access Journals (Sweden)

    D. Wojciechowska

    2015-05-01

    Full Text Available Results have been presented indicating that native proteolytic enzymes and bromelain acted variously ion wheat proteins differentiated in quality, they also act nonspecifically ton haemoglobin. It was thought useful to elaborate a new method of proteolytic activity determination based on the application of a standard natural substrate. The latter consists of gluten extract in acetic acid from a definite flour sample. The method of substrate selection and the determination procedure are described and some examples of wheat proteolytic activity determination are given.

  10. AZD9291 overcomes T790 M-mediated resistance through degradation of EGFR(L858R/T790M) in non-small cell lung cancer cells.

    Science.gov (United States)

    Ku, Bo Mi; Bae, Yeon-Hee; Koh, Jiae; Sun, Jong-Mu; Lee, Se-Hoon; Ahn, Jin Seok; Park, Keunchil; Ahn, Myung-Ju

    2016-08-01

    The discovery of activating mutations of epidermal growth factor receptor (EGFR) has resulted in the development of more effective treatments for non-small cell lung cancer (NSCLC). Although first-generation EGFR tyrosine kinase inhibitors (EGFR TKIs) provide significant clinical benefit, acquired resistance often occurs, most commonly (>50 %) via a T790 M resistance mutation. Although AZD9291 is selective for both T790 M and activating EGFR mutations over wild-type EGFR, it is highly active when T790 M is present, especially EGFR(L858R/T790M), and modestly active when T790 M is absent. The aim of this study was to elucidate the underlying mechanism of the high sensitivity of NSCLC cells harboring EGFR(L858R/T790M) to AZD9291. In H1975 cells harboring EGFR(L858R/T790M), AZD9291 potently inhibited cellular growth and EGFR signaling pathways together with depletion of mutant EGFR protein. AZD9291-induced depletion of EGFR(L858R/T790M) protein was abrogated through inhibition of the proteasome with MG132. However, AZD9291 had no effect on protein levels of EGFR(WT) and EGFR(L858R). In addition, AZD9291 induced apoptosis and caused expression changes in cell cycle-related genes. Moreover, oral administration of AZD9291 as a single agent induced tumor regression in vivo in a H1975 tumor xenograft model and reduced EGFR(L858R/T790M) protein levels in xenograft tumors. Taken together, our results provide a potential mechanism for the sensitivity of EGFR(L858R/T790M) cells to AZD9291 and suggest that AZD9291 may be effective in cases of T790 M-positive EGFR resistance. PMID:27044261

  11. Mycoplasma pneumoniae modulates STAT3-STAT6/EGFR-FOXA2 signaling to induce overexpression of airway mucins.

    Science.gov (United States)

    Hao, Yonghua; Kuang, Zhizhou; Jing, Jia; Miao, Jinfeng; Mei, Li Yu; Lee, Ryan J; Kim, Susie; Choe, Shawn; Krause, Duncan C; Lau, Gee W

    2014-12-01

    Aberrant mucin secretion and accumulation in the airway lumen are clinical hallmarks associated with various lung diseases such as asthma, chronic obstructive pulmonary disease, and cystic fibrosis. Mycoplasma pneumoniae, long appreciated as one of the triggers of acute exacerbations of chronic pulmonary diseases, has recently been reported to promote excessive mucus secretion. However, the mechanism of mucin overproduction induced by M. pneumoniae remains unclear. This study aimed to determine the mechanism by which M. pneumoniae induces mucus hypersecretion by using M. pneumoniae infection of mouse lungs, human primary bronchial epithelial (NHBE) cells cultured at the air-liquid interface, and the conventionally cultured airway epithelial NCI-H292 cell line. We demonstrated that M. pneumoniae induced the expression of mucins MUC5AC and MUC5B by activating the STAT6-STAT3 and epidermal growth factor receptor (EGFR) signal pathways, which in turn downregulated FOXA2, a transcriptional repressor of mucin biosynthesis. The upstream stimuli of these pathways, including interleukin-4 (IL-4), IL-6, and IL-13, increased dramatically upon exposure to M. pneumoniae. Inhibition of the STAT6, STAT3, and EGFR signaling pathways significantly restored the expression of FOXA2 and attenuated the expression of airway mucins MUC5AC and MUC5B. Collectively, these studies demonstrated that M. pneumoniae induces airway mucus hypersecretion by modulating the STAT/EGFR-FOXA2 signaling pathways. PMID:25287927

  12. Pharmacological inhibition of EGFR signaling enhances G-CSF-induced hematopoietic stem cell mobilization.

    Science.gov (United States)

    Ryan, Marnie A; Nattamai, Kalpana J; Xing, Ellen; Schleimer, David; Daria, Deidre; Sengupta, Amitava; Köhler, Anja; Liu, Wei; Gunzer, Matthias; Jansen, Michael; Ratner, Nancy; Le Cras, Timothy D; Waterstrat, Amanda; Van Zant, Gary; Cancelas, Jose A; Zheng, Yi; Geiger, Hartmut

    2010-10-01

    Mobilization of hematopoietic stem and progenitor cells (HSPCs) from bone marrow into peripheral blood by the cytokine granulocyte colony-stimulating factor (G-CSF) has become the preferred source of HSPCs for stem cell transplants. However, G-CSF fails to mobilize sufficient numbers of stem cells in up to 10% of donors, precluding autologous transplantation in those donors or substantially delaying transplant recovery time. Consequently, new regimens are needed to increase the number of stem cells in peripheral blood upon mobilization. Using a forward genetic approach in mice, we mapped the gene encoding the epidermal growth factor receptor (Egfr) to a genetic region modifying G-CSF-mediated HSPC mobilization. Amounts of EGFR in HSPCs inversely correlated with the cells' ability to be mobilized by G-CSF, implying a negative role for EGFR signaling in mobilization. In combination with G-CSF treatment, genetic reduction of EGFR activity in HSPCs (in waved-2 mutant mice) or treatment with the EGFR inhibitor erlotinib increased mobilization. Increased mobilization due to suppression of EGFR activity correlated with reduced activity of cell division control protein-42 (Cdc42), and genetic Cdc42 deficiency in vivo also enhanced G-CSF-induced mobilization. Our findings reveal a previously unknown signaling pathway regulating stem cell mobilization and provide a new pharmacological approach for improving HSPC mobilization and thereby transplantation outcomes. PMID:20871610

  13. IGFBP2 potentiates nuclear EGFR-STAT3 signaling

    OpenAIRE

    Chua, Corrine Yingxuan; Liu, Yuexin; Granberg, Kirsi J.; Hu, Limei; Haapasalo, Hannu; Annala, Matti J.; Cogdell, David E.; Verploegen, Maartje; Moore, Lynette M.; Fuller, Gregory N.; Nykter, Matti; Cavenee, Webster K.; Zhang, Wei

    2015-01-01

    Insulin-like growth factor binding protein 2 (IGFBP2) is a pleiotropic oncogenic protein that has both extracellular and intracellular functions. Despite a clear causal role in cancer development, the tumor-promoting mechanisms of IGFBP2 are poorly understood. The contributions of intracellular IGFBP2 to tumor development and progression are also unclear. Here we present evidence that both exogenous IGFBP2 treatment and cellular IGFBP2 overexpression lead to aberrant activation of EGFR, which...

  14. Identification of potent EGFR inhibitors from TCM Database@Taiwan.

    Directory of Open Access Journals (Sweden)

    Shun-Chieh Yang

    2011-10-01

    Full Text Available Overexpression of epidermal growth factor receptor (EGFR has been associated with cancer. Targeted inhibition of the EGFR pathway has been shown to limit proliferation of cancerous cells. Hence, we employed Traditional Chinese Medicine Database (TCM Database@Taiwan (http://tcm.cmu.edu.tw to identify potential EGFR inhibitor. Multiple Linear Regression (MLR, Support Vector Machine (SVM, Comparative Molecular Field Analysis (CoMFA, and Comparative Molecular Similarities Indices Analysis (CoMSIA models were generated using a training set of EGFR ligands of known inhibitory activities. The top four TCM candidates based on DockScore were 2-O-caffeoyl tartaric acid, Emitine, Rosmaricine, and 2-O-feruloyl tartaric acid, and all had higher binding affinities than the control Iressa®. The TCM candidates had interactions with Asp855, Lys716, and Lys728, all which are residues of the protein kinase binding site. Validated MLR (r² = 0.7858 and SVM (r² = 0.8754 models predicted good bioactivity for the TCM candidates. In addition, the TCM candidates contoured well to the 3D-Quantitative Structure-Activity Relationship (3D-QSAR map derived from the CoMFA (q² = 0.721, r² = 0.986 and CoMSIA (q² = 0.662, r² = 0.988 models. The steric field, hydrophobic field, and H-bond of the 3D-QSAR map were well matched by each TCM candidate. Molecular docking indicated that all TCM candidates formed H-bonds within the EGFR protein kinase domain. Based on the different structures, H-bonds were formed at either Asp855 or Lys716/Lys728. The compounds remained stable throughout molecular dynamics (MD simulation. Based on the results of this study, 2-O-caffeoyl tartaric acid, Emitine, Rosmaricine, and 2-O-feruloyl tartaric acid are suggested to be potential EGFR inhibitors.

  15. EGFR CA repeat polymorphism predict clinical outcome in EGFR mutation positive NSCLC patients treated with erlotinib

    DEFF Research Database (Denmark)

    Winther Larsen, Anne; Nissen, Peter Henrik; Meldgaard, Peter;

    2014-01-01

    OBJECTIVES: Somatic mutations in the epidermal growth factor receptor (EGFR) are predictors of efficacy for treatment with the EGFR tyrosine kinase inhibitor erlotinib in non-small cell lung cancer (NSCLC). A CA repeat polymorphism in intron 1 of the EGFR gene influences the transcription...... of the EGFR gene. This study evaluates the association between the CA repeat polymorphism and outcome in NSCLC patients treated with erlotinib.MATERIALS AND METHODS: Number of CA repeats in the EGFR gene was evaluated with PCR-fragment length analysis by capillary electrophoresis in 432 advanced NSCLC...... patients treated with erlotinib irrespective of EGFR mutation status. Patients were dichotomized into harboring short allele (CA≤16 in any allele) or long alleles (CA>16 in both alleles). Number of repeats was correlated with clinical characteristic and outcome. A subgroup analysis was performed based...

  16. Determination of proteolytic activity in cereals

    OpenAIRE

    D. Wojciechowska; J. Kączkowski

    2015-01-01

    Results have been presented indicating that native proteolytic enzymes and bromelain acted variously ion wheat proteins differentiated in quality, they also act nonspecifically ton haemoglobin. It was thought useful to elaborate a new method of proteolytic activity determination based on the application of a standard natural substrate. The latter consists of gluten extract in acetic acid from a definite flour sample. The method of substrate selection and the determination procedure are descri...

  17. Nuclear EGFR characterize still controlled proliferation retained in better differentiated clear cell RCC.

    Science.gov (United States)

    Ahel, J; Dordevic, G; Markic, D; Mozetic, V; Spanjol, J; Grahovac, B; Stifter, S

    2015-08-01

    Renal cell carcinoma (RCC) is the most common solid kidney tumor representing 2-3% of all cancers, with the highest frequency occurring in Western countries. There was a worldwide and European annual increase in incidence of approximately 2% although incidence has been stabilized in last few years. One third of the patients already have metastases in the time of the diagnosis with poor prognosis because RCC are radio and chemoresistant. The prognostic value of EGFR over-expression in RCC is a controversial issue that could be explained by different histological types of study tumors and non-standardized criteria for evaluation of expression. Recent evidences points to a new mode of EGFR signaling pathway in which activated EGFR undergoes nuclear translocalization and then, as transcription factor, mediates gene expression and other cellular events required for highly proliferating activities. According to our observations, the membranous expression of EGFR associates with high nuclear grade and poor differentiated tumors. On the other hand, nuclear EGFR expression was high in low nuclear graded and well differentiated tumors with good prognosis. We hypothesize that this mode of EGFR signaling characterizes still controlled proliferation retained in well differentiated RCC with Furhman nuclear grade I or II.

  18. High specificity but low sensitivity of mutation-specific antibodies against EGFR mutations in non-small-cell lung cancer

    DEFF Research Database (Denmark)

    Bondgaard, Anna-Louise; Høgdall, Estrid; Mellemgaard, Anders;

    2014-01-01

    Determination of epidermal growth factor receptor (EGFR) mutations has a pivotal impact on treatment of non-small-cell lung cancer (NSCLC). A standardized test has not yet been approved. So far, Sanger DNA sequencing has been widely used. Its rather low sensitivity has led to the development...... of more sensitive methods including real-time PCR (RT-PCR). Immunohistochemistry with mutation-specific antibodies might be a promising detection method. We evaluated 210 samples with NSCLC from an unselected Caucasian population. Extracted DNA was analyzed for EGFR mutations by RT-PCR (Therascreen EGFR...... was demonstrated. However, sensitivity was low, especially for exon19 deletions, and thus these antibodies cannot yet be used as screening method for EGFR mutations in NSCLC. Refinement of sensitivity for the mutation-specific antibodies is warranted to improve molecular diagnosis using EGFR immunohistochemistry....

  19. 18F-FDG uptake for prediction EGFR mutation status in non-small cell lung cancer.

    Science.gov (United States)

    Guan, Jian; Xiao, Nan J; Chen, Min; Zhou, Wen L; Zhang, Yao W; Wang, Shuang; Dai, Yong M; Li, Lu; Zhang, Yue; Li, Qin Y; Li, Xiang Z; Yang, Mi; Wu, Hu B; Chen, Long H; Liu, Lai Y

    2016-07-01

    Epidermal growth factor receptor (EGFR) mutations in non-small cell lung cancer (NSCLC) are a response to EGFR-tyrosine kinase inhibitor. However, a lack of sufficient tumor tissue has been a limitation for determining EGFR mutation status in clinical practice. The objective of this study was to predict EGFR mutation status in NSCLC patients based on a model including maximum standardized uptake value (SUVmax) and clinical features.We retrospectively reviewed NSCLC patients undergoing EGFR mutation testing and pretreatment positron emission tomography/computed tomography between March 2009 and December 2013. The relationships of EGFR mutations with both SUVmax and patient characteristics were evaluated, and a multivariate logistic regression analysis was performed. The model was assessed by area under the receiver-operating characteristic curve (AUC) and was prospectively validated during January to June 2014.Three hundred and sixteen patients meeting the criteria were enrolled for model construction. The SUVmax values were significantly lower for EGFR mutations (mean, 9.5 ± 5.74) than for EGFR wild-type (mean, 12.7 ± 6.43; P smoking history and primary tumor size when genetic tests are not available. PMID:27472739

  20. egr-4, a target of EGFR signaling, is required for the formation of the brain primordia and head regeneration in planarians.

    Science.gov (United States)

    Fraguas, Susanna; Barberán, Sara; Iglesias, Marta; Rodríguez-Esteban, Gustavo; Cebrià, Francesc

    2014-05-01

    During the regeneration of freshwater planarians, polarity and patterning programs play essential roles in determining whether a head or a tail regenerates at anterior or posterior-facing wounds. This decision is made very soon after amputation. The pivotal role of the Wnt/β-catenin and Hh signaling pathways in re-establishing anterior-posterior (AP) polarity has been well documented. However, the mechanisms that control the growth and differentiation of the blastema in accordance with its AP identity are less well understood. Previous studies have described a role of Smed-egfr-3, a planarian epidermal growth factor receptor, in blastema growth and differentiation. Here, we identify Smed-egr-4, a zinc-finger transcription factor belonging to the early growth response gene family, as a putative downstream target of Smed-egfr-3. Smed-egr-4 is mainly expressed in the central nervous system and its silencing inhibits anterior regeneration without affecting the regeneration of posterior regions. Single and combinatorial RNA interference to target different elements of the Wnt/β-catenin pathway, together with expression analysis of brain- and anterior-specific markers, revealed that Smed-egr-4: (1) is expressed in two phases - an early Smed-egfr-3-independent phase and a late Smed-egfr-3-dependent phase; (2) is necessary for the differentiation of the brain primordia in the early stages of regeneration; and (3) that it appears to antagonize the activity of the Wnt/β-catenin pathway to allow head regeneration. These results suggest that a conserved EGFR/egr pathway plays an important role in cell differentiation during planarian regeneration and indicate an association between early brain differentiation and the proper progression of head regeneration. PMID:24700819

  1. Comparative analysis of clinicoradiologic characteristics of lung adenocarcinomas with ALK rearrangements or EGFR mutations

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, J.Y.; Zheng, J.; Chen, X.; Zhou, J.Y. [Zhejiang University, Department of Respiratory Disease, Thoracic Disease Center, First Affiliated Hospital, College of Medicine, Hangzhou (China); Yu, Z.F.; Xiao, W.B.; Jiang, L.N. [Zhejiang University, Department of Radiology, First Affiliated Hospital, College of Medicine, Hangzhou (China); Zhao, J.; Sun, K.; Wang, B.; Ding, W. [Zhejiang University, Department of Pathology, First Affiliated Hospital, College of Medicine, Hangzhou (China)

    2015-05-01

    To compare the clinicoradiologic features of tumours with echinoderm anaplastic lymphoma kinase (ALK) rearrangements, epidermal growth factor receptor (EGFR) mutations, or wild type (WT) for both genes in a cohort of patients with lung adenocarcinoma to identify useful characteristics of different gene statuses. In 346 lung adenocarcinoma patients, ALK rearrangements were confirmed with fluorescence in situ hybridisation, and EGFR mutations were determined by pyrosequencing assay. Patients were divided into three groups: ALK rearrangement (ALK+ group, n = 48), EGFR mutation (EGFR+ group, n = 166), and WT for both genes (WT group, n = 132). Chest computed tomography (CT) examinations were performed in all patients. The percentages of ground-glass opacity volume (pGGO) and tumour shadow disappearance rate (TDR) were measured using semi-automated nodule assessment software. The pGGO was significantly lower in the ALK+ group (25.1 % ± 24.3) than in the EGFR+ group (37.2 % ± 25.7, p < 0.001) and the WT group (36.1 % ± 24.6, p = 0.001). The TDR in the ALK+ group (17.3 % ± 25.1) was significantly lower than in the EGFR+ group (26.8 % ± 24.9, p = 0.002) and the WT group (25.7 % ± 24.6, p = 0.003). Solid pattern with lower incidence of lobulated border, finely spiculated margins, pleural retraction, and bubble-like lucency on CT imaging are the main characteristics of ALK rearrangement tumours. (orig.)

  2. Fluctuations in eGFR in relation to unenhanced and enhanced MRI and CT outpatients

    DEFF Research Database (Denmark)

    Azzouz, Manal; Rømsing, Janne; Thomsen, Henrik S

    2014-01-01

    OBJECTIVE: To study fluctuations in estimated glomerular filtration rate (eGFR) in relation to contrast medium (CM) enhanced magnetic resonance imaging (MRI) and computed tomography (CT) compared to control groups in outpatients. MATERIALS AND METHODS: eGFR was determined right before the imaging...... relevant rise in eGFR after three days in all four groups. The average eGFR variation was 4.8ml/min/1.73m(2). There were large variations in eGFR between the two measurements in 45.8% of the patients as they had a change greater than ±10ml/min/1.73m(2). Only three patients fulfilled the contrast......-induced nephropathy (CIN) requirement when the definition s-creatinine ≥44μmol/l (0.5mg/dl) was used. CONCLUSIONS: eGFR in outpatients undergoing MRI or CT did vary independently of whether the patient received contrast or not. The findings probably reflect the natural variations in s-creatinine levels. This should...

  3. Fluctuations in eGFR in relation to unenhanced and enhanced MRI and CT outpatients

    Energy Technology Data Exchange (ETDEWEB)

    Azzouz, Manal, E-mail: manalazzouz@gmail.com [Department of Diagnostic Radiology, Copenhagen University Hospital Herlev, Herlev Ringvej 75, DK 2730 Herlev (Denmark); Rømsing, Janne [Department of Drug Design and Pharmacology, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen Ø (Denmark); Thomsen, Henrik S. [Department of Diagnostic Radiology, Copenhagen University Hospital Herlev, Herlev Ringvej 75, DK 2730 Herlev (Denmark)

    2014-06-15

    Objective: To study fluctuations in estimated glomerular filtration rate (eGFR) in relation to contrast medium (CM) enhanced magnetic resonance imaging (MRI) and computed tomography (CT) compared to control groups in outpatients. Materials and methods: eGFR was determined right before the imaging procedure and three days later at the department or at the patient's home. The iodine-based and gadolinium-based contrast media were the same as used for all other examinations at the department. Results: A total of 716 patients completed the study. There was a statistically significant, but not clinically relevant rise in eGFR after three days in all four groups. The average eGFR variation was 4.8 ml/min/1.73 m{sup 2}. There were large variations in eGFR between the two measurements in 45.8% of the patients as they had a change greater than ±10 ml/min/1.73 m{sup 2}. Only three patients fulfilled the contrast-induced nephropathy (CIN) requirement when the definition s-creatinine ≥44 μmol/l (0.5 mg/dl) was used. Conclusions: eGFR in outpatients undergoing MRI or CT did vary independently of whether the patient received contrast or not. The findings probably reflect the natural variations in s-creatinine levels. This should be taken into consideration when CIN is studied.

  4. Genetic variations of the A13/A14 repeat located within the EGFR 3′ untranslated region have no oncogenic effect in patients with colorectal cancer

    International Nuclear Information System (INIS)

    The EGFR 3′ untranslated region (UTR) harbors a polyadenine repeat which is polymorphic (A13/A14) and undergoes somatic deletions in microsatellite instability (MSI) colorectal cancer (CRC). These mutations could be oncogenic in colorectal tissue since they were shown to result into increased EGFR mRNA stability in CRC cell lines. First, we determined in a case control study including 429 CRC patients corresponding to different groups selected or not on age of tumor onset and/or familial history and/or MSI, whether or not, the germline EGFR A13/A14 polymorphism constitutes a genetic risk factor for CRC; second, we investigated the frequency of somatic mutations of this repeat in 179 CRC and their impact on EGFR expression. No statistically significant difference in allelic frequencies of the EGFR polyA repeat polymorphism was observed between CRC patients and controls. Somatic mutations affecting the EGFR 3′UTR polyA tract were detected in 47/80 (58.8%) MSI CRC versus 0/99 microsatellite stable (MSS) tumors. Comparative analysis in 21 CRC samples of EGFR expression, between tumor and non malignant tissues, using two independent methods showed that somatic mutations of the EGFR polyA repeat did not result into an EGFR mRNA increase. Germline and somatic genetic variations occurring within the EGFR 3′ UTR polyA tract have no impact on CRC genetic risk and EGFR expression, respectively. Genotyping of the EGFR polyA tract has no clinical utility to identify patients with a high risk for CRC or patients who could benefit from anti-EGFR antibodies

  5. 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.

  6. Phosphoproteomics-based modeling defines the regulatory mechanism underlying aberrant EGFR signaling.

    Directory of Open Access Journals (Sweden)

    Shinya Tasaki

    Full Text Available BACKGROUND: Mutation of the epidermal growth factor receptor (EGFR results in a discordant cell signaling, leading to the development of various diseases. However, the mechanism underlying the alteration of downstream signaling due to such mutation has not yet been completely understood at the system level. Here, we report a phosphoproteomics-based methodology for characterizing the regulatory mechanism underlying aberrant EGFR signaling using computational network modeling. METHODOLOGY/PRINCIPAL FINDINGS: Our phosphoproteomic analysis of the mutation at tyrosine 992 (Y992, one of the multifunctional docking sites of EGFR, revealed network-wide effects of the mutation on EGF signaling in a time-resolved manner. Computational modeling based on the temporal activation profiles enabled us to not only rediscover already-known protein interactions with Y992 and internalization property of mutated EGFR but also further gain model-driven insights into the effect of cellular content and the regulation of EGFR degradation. Our kinetic model also suggested critical reactions facilitating the reconstruction of the diverse effects of the mutation on phosphoproteome dynamics. CONCLUSIONS/SIGNIFICANCE: Our integrative approach provided a mechanistic description of the disorders of mutated EGFR signaling networks, which could facilitate the development of a systematic strategy toward controlling disease-related cell signaling.

  7. 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.

  8. Properties of resistant cells generated from lung cancer cell lines treated with EGFR inhibitors

    International Nuclear Information System (INIS)

    Epidermal growth factor receptor (EGFR) signaling plays an important role in non-small cell lung cancer (NSCLC) and therapeutics targeted against EGFR have been effective in treating a subset of patients bearing somatic EFGR mutations. However, the cancer eventually progresses during treatment with EGFR inhibitors, even in the patients who respond to these drugs initially. Recent studies have identified that the acquisition of resistance in approximately 50% of cases is due to generation of a secondary mutation (T790M) in the EGFR kinase domain. In about 20% of the cases, resistance is associated with the amplification of MET kinase. In the remaining 30-40% of the cases, the mechanism underpinning the therapeutic resistance is unknown. An erlotinib resistant subline (H1650-ER1) was generated upon continuous exposure of NSCLC cell line NCI-H1650 to erlotinib. Cancer stem cell like traits including expression of stem cell markers, enhanced ability to self-renew and differentiate, and increased tumorigenicity in vitro were assessed in erlotinib resistant H1650-ER1 cells. The erlotinib resistant subline contained a population of cells with properties similar to cancer stem cells. These cells were found to be less sensitive towards erlotinib treatment as measured by cell proliferation and generation of tumor spheres in the presence of erlotinib. Our findings suggest that in cases of NSCLC accompanied by mutant EGFR, treatment targeting inhibition of EGFR kinase activity in differentiated cancer cells may generate a population of cancer cells with stem cell properties

  9. 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

  10. 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.

  11. A proposed EGFR inhibitor dermatologic adverse event-specific grading scale from the MASCC skin toxicity study group

    NARCIS (Netherlands)

    Lacouture, Mario E.; Maitland, Michael L.; Segaert, Siegfried; Setser, Ann; Baran, Robert; Fox, Lindy P.; Epstein, Joel B.; Barasch, Andrei; Einhorn, Lawrence; Wagner, Lynne; West, Dennis P.; Rapoport, Bernardo L.; Kris, Mark G.; Basch, Ethan; Eaby, Beth; Kurtin, Sandra; Olsen, Elise A.; Chen, Alice; Dancey, Janet E.; Trotti, Andy

    2010-01-01

    Accurate grading of dermatologic adverse events (AE) due to epidermal growth factor receptor (EGFR) inhibitors (EGFRIs) is necessary for drug toxicity determinations, interagent comparisons, and supportive care trials. The most widely used severity grading scale, the National Cancer Institute's Comm

  12. Meta-Analysis of First-Line Therapies in Advanced Non–Small-Cell Lung Cancer Harboring EGFR-Activating Mutations

    OpenAIRE

    Haaland, Benjamin; Tan, Pui San; Castro, Gilberto de; Lopes, Gilberto

    2014-01-01

    Introduction: Tyrosine kinase inhibitors gefitinib, erlotinib, and afatinib have been compared with chemotherapy as first-line therapies for patients with advanced non–small-cell lung cancer harboring epidermal growth factor receptor–activating mutations. This meta-analysis compares gefitinib, erlotinib, afatinib, and chemotherapy. Methods: Literature search was performed using relevant keywords. Direct and indirect meta-estimates were generated using log-linear mixed-effects models, with ran...

  13. Simultaneous Inhibition of EGFR and PI3K Enhances Radiosensitivity in Human Breast Cancer

    International Nuclear Information System (INIS)

    Purpose: Mutations in the epidermal growth factor receptor (EGFR)/phosphoinositide 3-kinase (PI3K)/Akt signaling transduction pathway are common in cancer. This pathway is imperative to the radiosensitivity of cancer cells. We aimed to investigate the radiosensitizing effects of the simultaneous inhibition of EGFR and PI3K in breast cancer cells. Methods and Materials: MCF-7 cell lines with low expression of EGFR and wild-type PTEN and MDA-MB-468 cell lines with high expression of EGFR and mutant PTEN were used. The radiosensitizing effects by the inhibition of EGFR with AG1478 and/or PI3K with Ly294002 were determined by colony formation assay, Western blot was used to investigate the effects on downstream signaling. Flow cytometry was used for apoptosis and cell cycle analysis. Mice-bearing xenografts of MDA-MB-468 breast cancer cells were also used to observe the radiosensitizing effect. Results: Simultaneous inhibition of EGFR and PI3K greatly enhanced radiosensitizing effect in MDA-MB-468 in terms of apoptosis and mitotic death, either inhibition of EGFR or PI3K alone could enhance radiosensitivity with a dose-modifying factor (DMFSF2) of 1.311 and 1.437, radiosensitizing effect was further enhanced by simultaneous inhibition of EGFR and PI3K with a DMFSF2 at 2.698. DNA flow cytometric analysis indicated that dual inhibition combined with irradiation significantly induced G0/G1 phase arrest in MDA-MB-468 cells. The expression of phosphor-Akt and phosphor-Erk1/2 (induced by irradiation and PI3K inhibitor) were fully attenuated by simultaneous treatment with both inhibitors in combination with irradiation. In addition, dual inhibition combined with irradiation induced dramatic tumor growth delay in MDA-MB-468 xenografts. Conclusions: Our study indicated that simultaneous inhibition of EGFR and PI3K could further sensitize the cancer cells to irradiation compared to the single inhibitor with irradiation in vitro and in vivo. The approach may have important

  14. EGFR inhibitors identified as a potential treatment for chordoma in a focused compound screen.

    Science.gov (United States)

    Scheipl, Susanne; Barnard, Michelle; Cottone, Lucia; Jorgensen, Mette; Drewry, David H; Zuercher, William J; Turlais, Fabrice; Ye, Hongtao; Leite, Ana P; Smith, James A; Leithner, Andreas; Möller, Peter; Brüderlein, Silke; Guppy, Naomi; Amary, Fernanda; Tirabosco, Roberto; Strauss, Sandra J; Pillay, Nischalan; Flanagan, Adrienne M

    2016-07-01

    Chordoma is a rare malignant bone tumour with a poor prognosis and limited therapeutic options. We undertook a focused compound screen (FCS) against 1097 compounds on three well-characterized chordoma cell lines; 154 compounds were selected from the single concentration screen (1 µm), based on their growth-inhibitory effect. Their half-maximal effective concentration (EC50 ) values were determined in chordoma cells and normal fibroblasts. Twenty-seven of these compounds displayed chordoma selective cell kill and 21/27 (78%) were found to be EGFR/ERBB family inhibitors. EGFR inhibitors in clinical development were then studied on an extended cell line panel of seven chordoma cell lines, four of which were sensitive to EGFR inhibition. Sapitinib (AstraZeneca) emerged as the lead compound, followed by gefitinib (AstraZeneca) and erlotinib (Roche/Genentech). The compounds were shown to induce apoptosis in the sensitive cell lines and suppressed phospho-EGFR and its downstream pathways in a dose-dependent manner. Analysis of substituent patterns suggested that EGFR-inhibitors with small aniline substituents in the 4-position of the quinazoline ring were more effective than inhibitors with large substituents in that position. Sapitinib showed significantly reduced tumour growth in two xenograft mouse models (U-CH1 xenograft and a patient-derived xenograft, SF8894). One of the resistant cell lines (U-CH2) was shown to express high levels of phospho-MET, a known bypass signalling pathway to EGFR. Neither amplifications (EGFR, ERBB2, MET) nor mutations in EGFR, ERBB2, ERBB4, PIK3CA, BRAF, NRAS, KRAS, PTEN, MET or other cancer gene hotspots were detected in the cell lines. Our findings are consistent with the reported (p-)EGFR expression in the majority of clinical samples, and provide evidence for exploring the efficacy of EGFR inhibitors in the treatment of patients with chordoma and studying possible resistance mechanisms to these compounds in vitro and in vivo. © 2016

  15. Simultaneous Inhibition of EGFR and PI3K Enhances Radiosensitivity in Human Breast Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Li Ping; Zhang Qing [Department of Radiation Oncology, 6th People' s Hospital of Jiao Tong University, Shanghai 200233 (China); Torossian, Artour [Vanderbilt University, School of Medicine, Nashville, TN (United States); Li Zhaobin; Xu Wencai [Department of Radiation Oncology, 6th People' s Hospital of Jiao Tong University, Shanghai 200233 (China); Lu Bo [Department of Radiation Oncology, Thomas Jefferson University and Hospitals, Inc. Philadelphia, PA (United States); Fu Shen, E-mail: fushen1117@gmail.com [Department of Radiation Oncology, 6th People' s Hospital of Jiao Tong University, Shanghai 200233 (China)

    2012-07-01

    Purpose: Mutations in the epidermal growth factor receptor (EGFR)/phosphoinositide 3-kinase (PI3K)/Akt signaling transduction pathway are common in cancer. This pathway is imperative to the radiosensitivity of cancer cells. We aimed to investigate the radiosensitizing effects of the simultaneous inhibition of EGFR and PI3K in breast cancer cells. Methods and Materials: MCF-7 cell lines with low expression of EGFR and wild-type PTEN and MDA-MB-468 cell lines with high expression of EGFR and mutant PTEN were used. The radiosensitizing effects by the inhibition of EGFR with AG1478 and/or PI3K with Ly294002 were determined by colony formation assay, Western blot was used to investigate the effects on downstream signaling. Flow cytometry was used for apoptosis and cell cycle analysis. Mice-bearing xenografts of MDA-MB-468 breast cancer cells were also used to observe the radiosensitizing effect. Results: Simultaneous inhibition of EGFR and PI3K greatly enhanced radiosensitizing effect in MDA-MB-468 in terms of apoptosis and mitotic death, either inhibition of EGFR or PI3K alone could enhance radiosensitivity with a dose-modifying factor (DMF{sub SF2}) of 1.311 and 1.437, radiosensitizing effect was further enhanced by simultaneous inhibition of EGFR and PI3K with a DMF{sub SF2} at 2.698. DNA flow cytometric analysis indicated that dual inhibition combined with irradiation significantly induced G0/G1 phase arrest in MDA-MB-468 cells. The expression of phosphor-Akt and phosphor-Erk1/2 (induced by irradiation and PI3K inhibitor) were fully attenuated by simultaneous treatment with both inhibitors in combination with irradiation. In addition, dual inhibition combined with irradiation induced dramatic tumor growth delay in MDA-MB-468 xenografts. Conclusions: Our study indicated that simultaneous inhibition of EGFR and PI3K could further sensitize the cancer cells to irradiation compared to the single inhibitor with irradiation in vitro and in vivo. The approach may have

  16. 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.

  17. Clinical Characteristics and Outcomes of Lung Cancer Patients 
with EGFR Mutations in Exons 19 and 21

    Directory of Open Access Journals (Sweden)

    Renwang LIU

    2014-11-01

    Full Text Available Background and objective Studies on the epidermal growth factor receptor (EGFR signaling pathways and the therapeutic effects of EGFR-tyrosine kinase inhibitors (EGFR-TKIs have recently proven that targeted therapy has a major role in the treatment of lung cancer. However, the therapeutic effects of EGFR-TKIs on lung cancers with different EGFR mutation subtypes remain unclear. And if there is a significant difference in the effects of EGFR-TKIs, the mechanisms for the difference remain unclear. The aim of this study was to investigate the clinical importance of EGFR mutations in exons 19 and 21 of lung cancer patients and to compare the outcomes of these patients. Methods The study recruited 113 patients who had non-small cell lung cancer (NSCLC with EGFR mutations. EGFR mutations were detected for 47 patients using Real-time PCR or DNA sequencinag. The mutations of the remaining patients were determined using xTag-EGFR liquid chip technology. All stages I-III patients underwent radical resection followed by 4 cycles of postoperative chemotherapy. Patients with pleural metastases underwent pleural biopsy, pleurodesis, and chemotherapy only. Patients with distant metastases underwent biopsy and chemotherapy only. Collected clinical data were analyzed using SPSS 19.0 software. Results EGFR exon mutations 19 and 21 were found in 56 and 57 patients, respectively. The mean age of patients with exon 19 mutations was lower than the age of the patients with exon 21 mutations (57.02±11.31 years vs 62.25±7.76 years, respectively; P0.05 between the patients with exon 19 and 21 mutations; and survival analysis of 91 (80.5% patients with complete clinical data found no differences in overall survival. Stratification analysis found out that patients with exon 19 mutations had longer overall survival associated with age>61 years, male gender, ever smoking, and stage IV disease; although the differences were not significant. Conclusion Compared to the lung

  18. The epidermal growth factor receptor (EGFR / HER-1 gatekeeper mutation T790M is present in European patients with early breast cancer.

    Directory of Open Access Journals (Sweden)

    Vahid Bemanian

    Full Text Available The epidermal growth factor receptor (EGFR is one of the major oncogenes identified in a variety of human malignancies including breast cancer (BC. EGFR-mutations have been studied in lung cancer for some years and are established as important markers in guiding therapy with tyrosine kinase inhibitors (TKIs. In contrast, EGFR-mutations have been reported to be rare if not absent in human BC, although recent evidence has suggested a significant worldwide variation in somatic EGFR-mutations. Therefore, we investigated the presence of EGFR-mutations in 131 norwegian patients diagnosed with early breast cancer using real-time PCR methods. In the present study we identified three patients with an EGFR-T790M-mutation. The PCR-findings were confirmed by direct Sanger sequencing. Two patients had triple-negative BC (TNBC while the third was classified as luminal-A subtype. The difference in incidence of T790M mutations comparing the TNBC subgroup with the other BC subgroups was statistical significant (P = 0.023. No other EGFR mutations were identified in the entire cohort. Interestingly, none of the patients had received any previous cancer treatment. To our best knowledge, the EGFR-T790M-TKI-resistance mutation has not been previously detected in breast cancer patients. Our findings contrast with the observations made in lung cancer patients where the EGFR-T790M-mutation is classified as a typical "second mutation"causing resistance to TKI-therapy during ongoing anticancer therapy. In conclusion, we have demonstrated for the first time that the EGFR-T790M-mutation occurs in primary human breast cancer patients. In the present study the EGFR-T790M mutation was not accompanied by any simultaneous EGFR-activating mutation.

  19. Phase III study of afatinib or cisplatin plus pemetrexed in patients with metastatic lung adenocarcinoma with EGFR mutations.

    LENUS (Irish Health Repository)

    Sequist, Lecia V

    2013-09-20

    The LUX-Lung 3 study investigated the efficacy of chemotherapy compared with afatinib, a selective, orally bioavailable ErbB family blocker that irreversibly blocks signaling from epidermal growth factor receptor (EGFR\\/ErbB1), human epidermal growth factor receptor 2 (HER2\\/ErbB2), and ErbB4 and has wide-spectrum preclinical activity against EGFR mutations. A phase II study of afatinib in EGFR mutation-positive lung adenocarcinoma demonstrated high response rates and progression-free survival (PFS).

  20. Expressions of c-Cbl, Cbl-b and EGFR and Its Role of Prognosis in NSCLC

    OpenAIRE

    Jiao, Xin; JIN, BO; Qu, Xiujuan; Shunchao YAN; Hou, Kezuo; Liu, Yunpeng; Hu, Xuejun

    2011-01-01

    Background and objective Epidermal growth factor receptor (EGFR) is closely correlated with the progression of lung cancer. Its activity is modulated by Casitas B-lineage lymphoma (Cbl) family. The aim of this study is to investigate the expression and clinical relevance of c-Cbl, Cbl-b and EGFR in non-small cell lung cancer (NSCLC). Methods Expressions of c-Cbl, Cbl-b and EGFR protein were detected with tissue microarrays and immunohistochemistry technique in 94 cases of NSCLC. The correlati...

  1. [Determination of riboflavin kinase activity in yeast].

    Science.gov (United States)

    Shavlovsky, G M; Kashchenko, V E

    1975-01-01

    It is established that the main reason of the riboflavin kinase (RFK, EC 2.7.1.26) low specific activity in the cell-free extracts of the yeast Pichia guillermondii Wickerham ATCC 9058 is the presence of alkaline phosphatase (EC 3.1.3.1), effectively destructing flaven mononucleotide. By chromatography of the cell-free extracts of P. guillermondii on DEAE-Sephadex A-50, CM-Sphadex C-50, CM-cellulose, Sephadexes G-75 and G-100 RFK and alkaline phosphatase may be separated completely. Any of these procedures results in a several times increase of the RFK activity as compared with the initial preparation. One failed to obtain a similar effect by fractionation of the extracts with amminium sulphate and by hydroxylapatite chromatography. A simple method is developed for determining the activity of RFK in the cell-free extracts of yeast on the basis of negative adsorption of this enzyme on DEAE-Sephadex A-50. A selective inhibition of alkaline phosphatase by ions Be2+ and F- yields a less satisfactory result. The data are presented on the PFK activity of certain species of flavinogenic (Pichia guillermondii, Torulopsis camdida) and non-flavinogenic (Pichia ohmeri, Candida utilis, Saccharomyces cervisiae) yeast. PMID:174262

  2. 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

  3. Role of Pgrmc1 in estrogen maintenance of meiotic arrest in zebrafish oocytes through Gper/Egfr.

    Science.gov (United States)

    Aizen, Joseph; Thomas, Peter

    2015-04-01

    The regulation of receptor trafficking to the cell surface and its effect on responses of target cells to growth factors and hormones remain poorly understood. Initial evidence has been recently obtained using cancer cells that surface expression of the epidermal growth factor receptor (EGFR) is dependent on its association with progesterone receptor membrane component 1 (PGRMC1). Estrogen inhibition of oocyte maturation (OM) in zebrafish is mediated through G-protein-coupled estrogen membrane receptor 1 (Gper1) and involves activation of Egfr. Therefore, in this study, the potential roles of Pgrmc1 in the cell surface expression and functions of Egfr in normal cells were investigated in this in vitro OM model of Egfr action using an inhibitor of PGMRC1 signaling, AG205. A single ∼60 kDa protein band, which corresponds to the size of the Pgrmc1 dimer, was detected on plasma membranes of fully grown oocytes by western blotting. Co-treatment with the PGRMC1 inhibitor AG205 (20 μM) blocked the inhibitory effects of 100 nM estradiol-17β and the GPER agonist, G-1, on spontaneous maturation of denuded zebrafish oocytes. Moreover, reversal of these estrogen effects on OM by the EGFR inhibitors AG1478 and AG825 (50 μM) was prevented by co-incubation with the PGRMC1 inhibitor. Inhibition of Pgrmc1 signaling with AG205 also caused a decrease in Egfr-dependent signaling and Egfr expression on oocyte cell membranes. These results indicate that maintenance of Pgrmc1 signaling is required for Egfr expression on zebrafish oocyte cell membranes and for conserving the functions of Egfr in maintaining meiotic arrest through estrogen activation of Gper. PMID:25720537

  4. Tumor-penetrating peptide fused EGFR single-domain antibody enhances cancer drug penetration into 3D multicellular spheroids and facilitates effective gastric cancer therapy

    Science.gov (United States)

    Sha, Huizi; Zou, Zhengyun; Xin, Kai; Bian, Xinyu; Cai, Xueting; Lu, Wuguang; Chen, Jiao; Chen, Gang; Huang, Leaf; Blair, Andrew M.; Cao, Peng; Liu, Baorui

    2016-01-01

    Human tumors, including gastric cancer, frequently express high levels of epidermal growth factor receptors (EGFRs), which are associated with a poor prognosis. Targeted delivery of anticancer drugs to cancerous tissues shows potential in sparing unaffected tissues. However, it has been a major challenge for drug penetration in solid tumor tissues due to the complicated tumor microenvironment. We have constructed a recombinant protein named anti-EGFR-iRGD consisting of an anti-EGFR VHH (the variable domain from the heavy chain of the antibody) fused to iRGD, a tumor-specific binding peptide with high permeability. Anti-EGFR-iRGD, which targets EGFR and αvβ3, spreads extensively throughout both the multicellular spheroids and the tumor mass. The recombinant protein anti-EGFR-iRGD also exhibited antitumor activity in tumor cell lines, multicellular spheroids, and mice. Moreover, anti-EGFR-iRGD could improve anticancer drugs, such as doxorubicin (DOX), bevacizumab, nanoparticle permeability and efficacy in multicellular spheroids. This study draws attention to the importance of iRGD peptide in the therapeutic approach of anti-EGFR-iRGD. As a consequence, anti-EGFR-iRGD could be a drug candidate for cancer treatment and a useful adjunct of other anticancer drugs. PMID:25553823

  5. Pharmacogenomics of EGFR-targeted therapies in non-small cell lung cancer:EGFR and beyond

    Institute of Scientific and Technical Information of China (English)

    Christopher Delaney; Samuel Frank; R Stephanie Huang

    2015-01-01

    Commonly observed aberrations in epidermal growth factor receptor (EGFR) signaling have led to the development of EGFR-targeted therapies for various cancers, including non–small cell lung cancer (NSCLC). EGFR mutations and overexpression have further been shown to modulate sensitivity to these EGFR-targeted therapies in NSCLC and several other types of cancers. However, it is clear that mutations and/or genetic variations in EGFR alone cannot explain all of the variability in the responses of patients with NSCLC to EGFR-targeted therapies. For instance, in addition to EGFR genotype, genetic variations in other members of the signaling pathway downstream of EGFR or variations in paral el receptor tyrosine kinase (RTK) pathways are now recognized to have a significant impact on the efficacy of certain EGFR-targeted therapies. In this review, we highlight the mutations and genetic variations in such genes downstream of EGFR and in parallel RTK pathways. Specifically, the directional effects of these pharmacogenetic factors are discussed with a focus on two commonly prescribed EGFR inhibitors:cetuximab and erlotinib. The results of this comprehensive review can be used to optimize the treatment of NSCLC with EGFR inhibitors. Furthermore, they may provide the rationale for the design of subsequent combination therapies that involve the inhibition of EGFR.

  6. Past Decline Versus Current eGFR and Subsequent Mortality Risk.

    Science.gov (United States)

    Naimark, David M J; Grams, Morgan E; Matsushita, Kunihiro; Black, Corri; Drion, Iefke; Fox, Caroline S; Inker, Lesley A; Ishani, Areef; Jee, Sun Ha; Kitamura, Akihiko; Lea, Janice P; Nally, Joseph; Peralta, Carmen Alicia; Rothenbacher, Dietrich; Ryu, Seungho; Tonelli, Marcello; Yatsuya, Hiroshi; Coresh, Josef; Gansevoort, Ron T; Warnock, David G; Woodward, Mark; de Jong, Paul E

    2016-08-01

    A single determination of eGFR associates with subsequent mortality risk. Prior decline in eGFR indicates loss of kidney function, but the relationship to mortality risk is uncertain. We conducted an individual-level meta-analysis of the risk of mortality associated with antecedent eGFR slope, adjusting for established risk factors, including last eGFR, among 1.2 million subjects from 12 CKD and 22 other cohorts within the CKD Prognosis Consortium. Over a 3-year antecedent period, 12% of participants in the CKD cohorts and 11% in the other cohorts had an eGFR slope 5 ml/min per 1.73 m(2) per year, respectively. Compared with a slope of 0 ml/min per 1.73 m(2) per year, a slope of -6 ml/min per 1.73 m(2) per year associated with adjusted hazard ratios for all-cause mortality of 1.25 (95% confidence interval [95% CI], 1.09 to 1.44) among CKD cohorts and 1.15 (95% CI, 1.01 to 1.31) among other cohorts during a follow-up of 3.2 years. A slope of +6 ml/min per 1.73 m(2) per year also associated with higher all-cause mortality risk, with adjusted hazard ratios of 1.58 (95% CI, 1.29 to 1.95) among CKD cohorts and 1.43 (95% CI, 1.11 to 1.84) among other cohorts. Results were similar for cardiovascular and noncardiovascular causes of death and stronger for longer antecedent periods (3 versus rise in eGFR associates with an increased risk of mortality, independent of current eGFR. PMID:26657865

  7. Concordance of KRAS/BRAF Mutation Status in Metastatic Colorectal Cancer before and after Anti-EGFR Therapy

    Directory of Open Access Journals (Sweden)

    S. Gattenlöhner

    2009-01-01

    Full Text Available Anti-EGFR targeted therapy is a potent strategy in the treatment of metastatic colorectal cancer (mCRC but activating mutations in the KRAS gene are associated with poor response to this treatment. Therefore, KRAS mutation analysis is employed in the selection of patients for EGFR-targeted therapy and various studies have shown a high concordance between the mutation status in primary CRC and corresponding metastases. However, although development of therapy related resistance occurs also in the context of novel drugs such as tyrosine kinase-inhibitors the effect of the anti-EGFR treatment on the KRAS/BRAF mutation status itself in recurrent mCRC has not yet been clarified. Therefore, we analyzed 21 mCRCs before/after anti-EGFR therapy and found a pre-/posttherapeutic concordance of the KRAS/BRAF mutation status in 20 of the 21 cases examined. In the one discordant case, further analyses revealed that a tumor mosaicism or multiple primary tumors were present, indicating that anti-EGFR therapy has no influence on KRAS/BRAF mutation status in mCRC. Moreover, as the preselection of patients with a KRASwt genotype for anti-EGFR therapy has become a standard procedure, sample sets such ours might be the basis for future studies addressing the identification of potential anti-EGFR therapy induced genetic alterations apart from KRAS/BRAF mutations.

  8. Expressions of c-Cbl, Cbl-b and EGFR and Its Role of Prognosis in NSCLC

    Directory of Open Access Journals (Sweden)

    Xin JIAO

    2011-06-01

    Full Text Available Background and objective Epidermal growth factor receptor (EGFR is closely correlated with the progression of lung cancer. Its activity is modulated by Casitas B-lineage lymphoma (Cbl family. The aim of this study is to investigate the expression and clinical relevance of c-Cbl, Cbl-b and EGFR in non-small cell lung cancer (NSCLC. Methods Expressions of c-Cbl, Cbl-b and EGFR protein were detected with tissue microarrays and immunohistochemistry technique in 94 cases of NSCLC. The correlations between the expression of the three proteins and clinicopathological parameters were analyzed. Results The positive expression rates of EGFR, c-Cbl and Cbl-b were 60.6% (57/94, 30.9% (29/94 and 84.0% (79/94, respectively. The expression of EGFR, c-Cbl and Cbl-b was not associated with age, pathological type, TNM stage, lymph node metastasis, and smoking history. c-Cbl and Cbl-b status was not significantly correlated with overall survival. Subgroup analyses showed that c-Cbl-positive patients had longer survival than c-Cbl-negative patients in EGFR-positive group (P=0.014. Conclusion Detection of c-Cbl protein levels might contribute to the prognosis evaluation of EGFR-positive NSCLC.

  9. Nimotuzumab promotes radiosensitivity of EGFR-overexpression esophageal squamous cell carcinoma cells by upregulating IGFBP-3

    Directory of Open Access Journals (Sweden)

    Zhao Lei

    2012-12-01

    Full Text Available Abstract Background Epidermal growth factor receptor (EGFR is suggested to predict the radiosensitivity and/or prognosis of human esophageal squamous cell carcinoma (ESCC. The objective of this study was to investigate the efficacy of Nimotuzumab (an anti-EGFR monoclonal antibody on ESCC radiotherapy (RT and underlying mechanisms. Methods Nimotuzumab was administrated to 2 ESCC cell lines KYSE30 and TE-1 treated with RT. Cell growth, colony formation and apoptosis were used to measure anti-proliferation effects. The method of RNA interference was used to investigate the role of insulin-like growth factor binding protein-3 (IGFBP-3 in ESCC cells radiosensitivity treated with Nimotuzumab. In vivo effect of Nimotuzumab on ESCC radiotherapy was done using a mouse xenograft model. Results Nimotuzumab enhanced radiation response of KYSE30 cells (with high EGFR expression in vitro, as evidenced by increased radiation-inhibited cell growth and colony formation and radiation-mediated apoptosis. Mechanism study revealed that Nimotuzumab inhibited phosphorylated EGFR (p-EGFR induced by EGF in KYSE30 cells. In addition, knockdown of IGFBP-3 by short hairpin RNA significantly reduced KYSE30 cells radiosensitivity (PP>0.05. In KYSE30 cell xenografts, Nimotuzumab combined with radiation led to significant tumor growth delay, compared with that of radiation alone (P=0.029, and also with IGFBP-3 up-regulation in tumor tissue. Conclusions Nimotuzumab could enhance the RT effect of ESCC cells with a functional active EGFR pathway. In particular, the increased ESCC radiosensitivity by Nimotuzumab might be dependent on the up-regulation of IGFBP-3 through EGFR-dependent pathway.

  10. Simultaneous molecular imaging of EGFR and HER2 using hyperspectral darkfield microscopy and immunotargeted nanoparticles

    Science.gov (United States)

    Crow, Matthew J.; Marinakos, Stella; Chilkoti, Ashutosh; Wax, Adam P.

    2009-02-01

    Epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor (HER2) contribute to the regulation of cell proliferation, and when jointly over-expressed are associated with several types of cancer. The ability to monitor both receptors simultaneously results in a more accurate indicator of degree of cancerous activity than either receptor alone. Plasmonic nanoparticles (NPs) show promise as a potential EGFR and HER2 biomarker over alternatives such as fluorophores and quantum dots, which are limited by their cytotoxicity and photobleaching. To observe immunolabeled NPs bound to receptor-expressing cells, our past experiments were conducted using a novel optical darkfield microspectroscopy system. We implemented an epi-illumination darkfield broadband light train, which allows for darkfield analysis of live cells in culture with enhanced NP contrast. Under this setup, molecularly specific binding of NPs immunolabeled with anti-EGFR was confirmed. We have since adapted our darkfield setup, which previously only obtained spectral information from a line imaging spectrometer, to incorporate hyperspectral imaging capabilities, allowing widefield data acquisition within seconds. The new system has been validated through observation of shifts in the peak wavelength of scattering by gold NPs on silanated cover glasses using several immersion media. Peak resonant scattering wavelengths match well with that predicted by Mie theory. We will further demonstrate the potential of the system with simultaneous molecular imaging of multiple receptors in vitro using labeled EGFR+/HER2+ SK-BR-3 human breast cancer cells with anti-EGFR immunolabeled gold nanospheres and anti-HER2 immunolabeled gold nanorods, with each scattering in different spectral windows. Additional trials will be performed to demonstrate molecularly specific binding using EGFR+/HER2- MDA-MB-468 and HER2+/EGFR- MDA-MB-453 breast cancer cells.

  11. Epidermal to Mesenchymal Transition and Failure of EGFR-Targeted Therapy in Glioblastoma

    Energy Technology Data Exchange (ETDEWEB)

    Pala, Andrej; Karpel-Massler, Georg [Department of Neurosurgery, University of Ulm School of Medicine, Steinhövelstrasse 9, Ulm D-89077 (Germany); Kast, Richard Eric [Department of Psychiatry, University of Vermont, 22 Church Street, Burlington, VT 05401 (United States); Wirtz, Christian Rainer; Halatsch, Marc-Eric, E-mail: marc-eric.halatsch@uniklinik-ulm.de [Department of Neurosurgery, University of Ulm School of Medicine, Steinhövelstrasse 9, Ulm D-89077 (Germany)

    2012-05-08

    Glioblastoma multiforme (GBM), the most common primary brain tumor in adults, is almost never curable with the current standard treatment consisting of surgical resection, irradiation and temozolomide. The prognosis remains poor despite undisputable advances in the understanding of this tumor’s molecular biology and pathophysiology, which unfortunately has so far failed to translate into a meaningful clinical benefit. Dysregulation and a resulting prominent pathophysiological role of the epidermal growth factor receptor (EGFR) have been identified in several different malignant tumor entities, GBM among them. The EGFR is overexpressed in about 40% of GBM cases, and half of these coexpress a mutant, constitutively activated subtype, EGFRvIII. Unfortunately, recent trials studying with therapeutic approaches targeted against the EGFR and EGFRvIII have failed to meet expectations, with only a minority of patients responding despite evidence of good in vitro and rodent model activity. Having potentially high relevance within this context, epithelial to mesenchymal transition (EMT) is a phenomenon associated with early stages of carcinogenesis, cancer invasion and recurrence. During EMT, epithelial cells lose many of their epithelial characteristics, prominently E-cadherin expression, and acquire properties that are typical for mesenchymal cells such as the expression of vimentin. Epithelial to mesenchymal transition has been specifically demonstrated in GBM. In this review, we summarize the evidence that EMT may precipitate GBM resistance to EGFR-targeted therapy, and may thus be among the principal factors contributing to the clinical failure of targeted therapy against EGFR and EGFRvIII.

  12. Inhibition of tumor growth by targeted anti-EGFR/IGF-1R Nanobullets depends on efficient blocking of cell survival pathways

    NARCIS (Netherlands)

    Meel, van der Roy; Oliveira, Sabrina; Altintas, Isil; Heukers, Raimond; Pieters, Ebel H.E.; Bergen en Henegouwen, van Paul M.P.; Storm, Gert; Hennink, Wim E.; Kok, Robbert J.; Schiffelers, Raymond M.

    2013-01-01

    The clinical efficacy of epidermal growth factor receptor (EGFR)-targeted inhibitors is limited due to resistance mechanisms of the tumor such as activation of compensatory pathways. Crosstalk between EGFR and insulin-like growth factor 1 (IGF-1R) signaling has been frequently described to be involv

  13. Delphinidin reduces cell proliferation and induces apoptosis of non-small-cell lung cancer cells by targeting EGFR/VEGFR2 signaling pathways.

    Directory of Open Access Journals (Sweden)

    Harish Chandra Pal

    Full Text Available Epidermal growth factor receptor (EGFR and vascular endothelial growth factor receptor 2 (VEGFR2 have emerged as two effective clinical targets for non-small-cell lung cancer (NSCLC. In the present study, we found that delphinidin, an anthocyanidin, present in pigmented fruits and vegetables, is a potent inhibitor of both EGFR and VEGFR2 in NSCLC cells that overexpress EGFR/VEGFR2. Using these cells, we next determined the effects of delphinidin on cell growth and apoptosis in vitro and on tumor growth and angiogenesis in vivo. Delphinidin (5-60 µM treatment of NSCLC cells inhibited the activation of PI3K, and phosphorylation of AKT and MAPKs. Additionally, treatment of NSCLC cells with delphinidin resulted in inhibition of cell growth without having significant toxic effects on normal human bronchial epithelial cells. Specifically, treatment of NCI-H441 and SK-MES-1 cells with delphindin (5-60 µM resulted in (i cleavage of PARP protein, (ii activation of caspase-3 and -9, (iii downregulation of anti-apoptotic proteins (Bcl2, Bcl-xL and Mcl-1, (iv upregulation of pro-apoptotic proteins (Bax and Bak, and (v decreased expression of PCNA and cyclin D1. Furthermore, in athymic nude mice subcutaneously implanted with human NSCLC cells, delphinidin treatment caused a (i significant inhibition of tumor growth, (ii decrease in the expression of markers for cell proliferation (Ki67 and PCNA and angiogenesis (CD31 and VEGF, and (iii induction of apoptosis, when compared with control mice. Based on these observations, we suggest that delphinidin, alone or as an adjuvant to current therapies, could be used for the management of NSCLC, especially those that overexpress EGFR and VEGFR2.

  14. 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.

  15. mTOR inhibitors control the growth of EGFR mutant lung cancer even after acquiring resistance by HGF.

    Directory of Open Access Journals (Sweden)

    Daisuke Ishikawa

    Full Text Available Resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs, gefitinib and erlotinib, is a critical problem in the treatment of EGFR mutant lung cancer. Several mechanisms, including bypass signaling by hepatocyte growth factor (HGF-triggered Met activation, are implicated as mediators of resistance. The mammalian target of rapamycin (mTOR, is a downstream conduit of EGFR and MET signaling, and is thus considered a therapeutically attractive target in the treatment of various types of cancers. The purpose of this study was to examine whether 2 clinically approved mTOR inhibitors, temsirolimus and everolimus, overcome HGF-dependent resistance to EGFR-TKIs in EGFR mutant lung cancer cells. Both temsirolimus and everolimus inhibited the phosphorylation of p70S6K and 4E-BP1, which are downstream targets of the mTOR pathway, and reduced the viability of EGFR mutant lung cancer cells, PC-9, and HCC827, even in the presence of HGF in vitro. In a xenograft model, temsirolimus suppressed the growth of PC-9 cells overexpressing the HGF-gene; this was associated with suppression of the mTOR signaling pathway and tumor angiogenesis. In contrast, erlotinib did not suppress this signaling pathway or tumor growth. Multiple mechanisms, including the inhibition of vascular endothelial growth factor production by tumor cells and suppression of endothelial cell viability, contribute to the anti-angiogenic effect of temsirolimus. These findings indicate that mTOR inhibitors may be useful for controlling HGF-triggered EGFR-TKI resistance in EGFR mutant lung cancer, and they provide the rationale for clinical trials of mTOR inhibitors in patients stratified by EGFR mutation and HGF expression status.

  16. Galbanic acid inhibits HIF-1α expression via EGFR/HIF-1α pathway in cancer cells.

    Science.gov (United States)

    Eskandani, Morteza; Abdolalizadeh, Jalal; Hamishehkar, Hamed; Nazemiyeh, Hossein; Barar, Jaleh

    2015-03-01

    Hypoxia Inducible Factor-1 plays a key transcriptional role in the adaptation of hypoxic solid tumors to low oxygen environment. Here, we aimed to investigate galbanic acid (GBA) inhibitory effects on HIF-1 activation during hypoxia and normoxia. MTT survival and Annexin V assays were used to evaluate GBA cytotoxicity and apoptosis in treated cells. Quantitative real time PCR and western blotting were used to estimate mRNA expression and translated protein, respectively. Results showed that GBA dose- and time-dependently decreased the in vitro growth of OVCAR-3 human epithelial carcinoma cells with an IC50 of approximately 37, 12.1 and 10μM GBA at 24, 48 and 72h, respectively. Following phosphatidylserine of outer leaflet of the plasma membrane revealed occurrence of early/late apoptosis in GBA treated cells. In addition, we found that GBA down-regulates HIF-1α and HIF-1β mRNA expression in both hypoxia and normoxia. To determine the mechanism of action, we showed that GBA did not inhibit Akt and EGFR mRNA expression, yet protein degradation investigation showed that GBA shortened the half-life of EGFR through decreasing its stability with a decrease of nearly 2 and 3h in A549 and OVCAR-3 cell lines, respectively. We also found that downstream genes contributed in glycolysis, including Eno 1 and GluT-1, are underexpressed in GBA treated cells in hypoxia. Conclusively, GBA may inhibit HIF-1 activation through down-regulation of its subunit expression in hypoxia, and increasing of EGFR degradation in normoxia. PMID:25510323

  17. EGFR expression level predicts response and overall survival in gastric cancer PDTX model treated with cetuximab

    Institute of Scientific and Technical Information of China (English)

    2015-01-01

    Objective:hTe aim of this study was to determine whether the EGFR statuscould significantly predict some benefit in overall survival and response to cetuximab in advanced GC xenografts.Methods: Two hundred xenografts derived from 20 GC patients were established. Then they were divided into cetuximab treated group and control group randomly.Results:Among the cetuximab treated group, 4 GC cases were identified responded to cetuximab.hTose cetuximab treated PDX models had longer OS than non-treated. High EGFR mRNA expression and immunohistochemistry score are more prone to response to cetuximab. EGFR amplification, mRNA and protein overexpression were associated with the OS in cetuximab treated PDX models. Moreover, in the PDX models derived from EGFR ampliifcation, mRNA or protein overexpression cases, the OS is signiifcantly different between the cetuximab treated and control group, while the OS in not statistically different in other cases.Conclusion:EGFR status predicts sensitivity to therapy and survival in GC treated with cetuximab, especially the mRNA and protein expression level.

  18. Inhibition of EGFR/MAPK signaling reduces microglial inflammatory response and the associated secondary damage in rats after spinal cord injury

    Directory of Open Access Journals (Sweden)

    Qu Wen-sheng

    2012-07-01

    Full Text Available Abstract Background Emerging evidence indicates that reactive microglia-initiated inflammatory responses are responsible for secondary damage after primary traumatic spinal cord injury (SCI; epidermal growth factor receptor (EGFR signaling may be involved in cell activation. In this report, we investigate the influence of EGFR signaling inhibition on microglia activation, proinflammatory cytokine production, and the neuronal microenvironment after SCI. Methods Lipopolysaccharide-treated primary microglia/BV2 line cells and SCI rats were used as model systems. Both C225 and AG1478 were used to inhibit EGFR signaling activation. Cell activation and EGFR phosphorylation were observed after fluorescent staining and western blot. Production of interleukin-1beta (IL-1β and tumor necrosis factor alpha (TNFα was tested by reverse transcription PCR and ELISA. Western blot was performed to semi-quantify the expression of EGFR/phospho-EGFR, and phosphorylation of Erk, JNK and p38 mitogen-activated protein kinases (MAPK. Wet-dry weight was compared to show tissue edema. Finally, axonal tracing and functional scoring were performed to show recovery of rats. Results EGFR phosphorylation was found to parallel microglia activation, while EGFR blockade inhibited activation-associated cell morphological changes and production of IL-1β and TNFα. EGFR blockade significantly downregulated the elevated MAPK activation after cell activation; selective MAPK inhibitors depressed production of cytokines to a certain degree, suggesting that MAPK mediates the depression of microglia activation brought about by EGFR inhibitors. Subsequently, seven-day continual infusion of C225 or AG1478 in rats: reduced the expression of phospho-EGFR, phosphorylation of Erk and p38 MAPK, and production of IL-1β and TNFα; lessened neuroinflammation-associated secondary damage, like microglia/astrocyte activation, tissue edema and glial scar/cavity formation; and enhanced axonal

  19. Structural Determinants of Sleeping Beauty Transposase Activity.

    Science.gov (United States)

    Abrusán, György; Yant, Stephen R; Szilágyi, András; Marsh, Joseph A; Mátés, Lajos; Izsvák, Zsuzsanna; Barabás, Orsolya; Ivics, Zoltán

    2016-08-01

    Transposases are important tools in genome engineering, and there is considerable interest in engineering more efficient ones. Here, we seek to understand the factors determining their activity using the Sleeping Beauty transposase. Recent work suggests that protein coevolutionary information can be used to classify groups of physically connected, coevolving residues into elements called "sectors", which have proven useful for understanding the folding, allosteric interactions, and enzymatic activity of proteins. Using extensive mutagenesis data, protein modeling and analysis of folding energies, we show that (i) The Sleeping Beauty transposase contains two sectors, which span across conserved domains, and are enriched in DNA-binding residues, indicating that the DNA binding and endonuclease functions of the transposase coevolve; (ii) Sector residues are highly sensitive to mutations, and most mutations of these residues strongly reduce transposition rate; (iii) Mutations with a strong effect on free energy of folding in the DDE domain of the transposase significantly reduce transposition rate. (iv) Mutations that influence DNA and protein-protein interactions generally reduce transposition rate, although most hyperactive mutants are also located on the protein surface, including residues with protein-protein interactions. This suggests that hyperactivity results from the modification of protein interactions, rather than the stabilization of protein fold.

  20. Structural Determinants of Sleeping Beauty Transposase Activity.

    Science.gov (United States)

    Abrusán, György; Yant, Stephen R; Szilágyi, András; Marsh, Joseph A; Mátés, Lajos; Izsvák, Zsuzsanna; Barabás, Orsolya; Ivics, Zoltán

    2016-08-01

    Transposases are important tools in genome engineering, and there is considerable interest in engineering more efficient ones. Here, we seek to understand the factors determining their activity using the Sleeping Beauty transposase. Recent work suggests that protein coevolutionary information can be used to classify groups of physically connected, coevolving residues into elements called "sectors", which have proven useful for understanding the folding, allosteric interactions, and enzymatic activity of proteins. Using extensive mutagenesis data, protein modeling and analysis of folding energies, we show that (i) The Sleeping Beauty transposase contains two sectors, which span across conserved domains, and are enriched in DNA-binding residues, indicating that the DNA binding and endonuclease functions of the transposase coevolve; (ii) Sector residues are highly sensitive to mutations, and most mutations of these residues strongly reduce transposition rate; (iii) Mutations with a strong effect on free energy of folding in the DDE domain of the transposase significantly reduce transposition rate. (iv) Mutations that influence DNA and protein-protein interactions generally reduce transposition rate, although most hyperactive mutants are also located on the protein surface, including residues with protein-protein interactions. This suggests that hyperactivity results from the modification of protein interactions, rather than the stabilization of protein fold. PMID:27401040

  1. Novel targets in HPV-negative head and neck cancer: overcoming resistance to EGFR inhibition.

    Science.gov (United States)

    Burtness, Barbara; Bauman, Julie E; Galloway, Thomas

    2013-07-01

    Cancers of the head and neck that arise from habitual exposure to carcinogens have lower cure rates than those that arise from infection with human papillomavirus (HPV), and intensification of cytotoxic chemotherapy and radiation has not improved outcomes. HPV-negative head and neck cancers abundantly express EGFR, and the monoclonal antibody cetuximab, directed against EGFR, is the only targeted therapy that has improved disease survival so far. However, response rates to single-agent cetuximab are lower than 15%, and cetuximab given with chemotherapy or radiation leads to only a modest effect on survival. Thus, investigating the mechanisms of resistance to EGFR inhibition in HPV-negative head and neck cancer might help identify novel and active therapies. In this Review, we focus on therapies in development that target redundant receptor tyrosine kinases (eg, HER2 and MET), reduce or abrogate nuclear functions of EGFR, affect cellular trafficking by inhibition of histone deacetylase, or treatments that might address resistance that arises in the EGFR signalling stream (eg, aurora-kinase inhibitors and STAT decoys). PMID:23816296

  2. Individualized therapies in colorectal cancer: KRAS as a marker for response to EGFR-targeted therapy

    Directory of Open Access Journals (Sweden)

    Li Kuiyuan

    2009-04-01

    Full Text Available Abstract Individualized therapies that are tailored to a patient's genetic composition will be of tremendous value for treatment of cancer. Recently, Kirsten ras (KRAS status has emerged as a predictor of response to epidermal growth factor receptor (EGFR targeted therapies. In this article, we will discuss targeted therapies for colorectal cancers (CRC based on EGFR signaling pathway and review published data about the potential usefulness of KRAS as a biological marker for response to these therapies. Results from relevant studies published since 2005 and unpublished results presented at national meetings were retrieved and summarized. These studies reflected response (or lack of response to EGFR-targeted therapies in patients with metastatic CRC as a function of KRAS status. It has become clear that patients with colorectal cancer whose tumor has an activating mutation in KRAS do not respond to monoclonal antibody therapies targeting EGFR. It should now become a standard practice that any patients being considered for EGFR targeted therapies have their tumors tested for KRAS status and only those with wild-type KRAS being offered such therapies.

  3. EGFR mutation frequency and effectiveness of erlotinib

    DEFF Research Database (Denmark)

    Weber, Britta; Hager, Henrik; Sorensen, Boe S;

    2014-01-01

    OBJECTIVES: In 2008, we initiated a prospective study to explore the frequency and predictive value of epidermal growth factor receptor (EGFR) mutations in an unselected population of Danish patients with non-small cell lung cancer offered treatment with erlotinib, mainly in second-line. MATERIALS...

  4. EGFR, KRAS, BRAF, and HER-2 molecular status in brain metastases from 77 NSCLC patients

    OpenAIRE

    Villalva, Claire; Duranton-Tanneur, Valérie; Guilloteau, Karline; Burel-Vandenbos, Fanny; Wager, Michel; Doyen, Jérôme,; Levillain, Pierre Marie; Fontaine, Denys; Blons, Hélène; Pedeutour, Florence; Karayan-Tapon, Lucie

    2013-01-01

    The aim of this study was to determine the frequency of EGFR, KRAS, BRAF, and HER-2 mutations in brain metastases from non-small cell lung carcinomas (BM-NSCLC). A total of 77 samples of BM-NSCLC were included and 19 samples of BM from breast, kidney, and colorectal tumors were also studied as controls. These samples were collected from patients followed between 2008 and 2011 at Poitiers and Nice University Hospitals in France. The frequencies of EGFR, KRAS, BRAF, and HER-2 mutations in BM-NS...

  5. 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%.

  6. Expression and significance of EGFR protein in model of radiation injury in mouse skin

    International Nuclear Information System (INIS)

    Objective: The expression of EGFR protein was studied by SABC immunohistochemistry in 40 cases of model of radiation injury in mouse skin. Methods: Experiment animals were divided into four groups according to radiation dose. Results: The positive rates were 27.0%, 49.3%, 72.2%, 87.6% in 5 Gy group, 15 Gy group, 30 Gy group, 45 Gy group respectively, showing significant difference (P < 0.01). While the positive rate was 10.8% in normal control group, with significant difference (P < 0.01) compared with each radiation group. Conclusion: The enhancement of expression of EGFR in accordance with the increasing of radiation dose in certain dose range might be one important factor related to c-erbB-1 gene activated and enlarged by radiation, and the overexpression of EGFR protein might be related to poor healing in radiation skin injury

  7. Synthetic Genistein Glycosides Inhibiting EGFR Phosphorylation Enhance the Effect of Radiation in HCT 116 Colon Cancer Cells

    Directory of Open Access Journals (Sweden)

    Aleksandra Gruca

    2014-11-01

    Full Text Available The need to find new EGFR inhibitors for use in combination with radiotherapy in the treatment of solid tumors has drawn our attention to compounds derived from genistein, a natural isoflavonoid. The antiproliferative potential of synthetic genistein derivatives used alone or in combination with ionizing radiation was evaluated in cancer cell lines using clonogenic assay. EGFR phosphorylation was assessed with western blotting. Genistein derivatives inhibited clonogenic growth of HCT 116 cancer cells additively or synergistically when used in combination with ionizing radiation, and decreased EGFR activation. Our preclinical evaluation of genistein-derived EGFR inhibitors suggests that these compounds are much more potent sensitizers of cells to radiation than the parent isoflavonoid, genistein and indicate that these compounds may be useful in the treatment of colon cancer with radiation therapy.

  8. Quantification of EGFR autoantibodies in the amplification phenomenon of HER2 in breast cancer

    DEFF Research Database (Denmark)

    Olsen, Dorte Aa; Jakobsen, Erik H; Brandslund, Ivan

    2013-01-01

    Abstract Background: Gene amplification or overexpression of human epidermal growth factor receptor HER2/ErB2 is seen in 25-30% of patients with breast cancer and is related to an aggressive disease. The mechanism behind the HER2 gene amplification is unknown, but it may be caused by continuous...... stimulation and activation. We hypothesised that autoantibodies against EGFR might have a stimulatory effect. To investigate this we developed a quantitative method to measure autoantibodies against EGFR in serum (S-EGFRAb). Methods: Serum samples from primary breast cancer patients were selected based...

  9. EGFR-Mutated Breast Metastasis of Lung Adenocarcinoma: A Case Report.

    Science.gov (United States)

    Dansin, Eric; Carnot, Aurélien; Servent, Véronique; Daussay, Dorothée; Robin, Yves-Marie; Surmei-Pintilie, Ecaterina; Lauridant, Géraldine; Descarpentries, Clothilde; Révillion, Françoise; Delattre, Claire

    2015-01-01

    Breast metastasis from other primary carcinoma is very rare and could be difficult to identify despite immunohistochemistry analysis. Breast metastasis from lung adenocarcinoma can mimic triple-negative breast cancer. Given the prognosis and therapeutic challenges, a correct diagnosis appears essential, and molecular biomarkers could be useful. We report the case of a 52-year-old woman with a breast mass initially diagnosed as primary breast cancer and secondarily attached to breast metastasis from an EGFR-mutated lung adenocarcinoma. The same activating EGFR mutations were identified in both the primary lung carcinoma and the breast metastasis.

  10. EGFR-Mutated Breast Metastasis of Lung Adenocarcinoma: A Case Report

    Directory of Open Access Journals (Sweden)

    Eric Dansin

    2015-03-01

    Full Text Available Breast metastasis from other primary carcinoma is very rare and could be difficult to identify despite immunohistochemistry analysis. Breast metastasis from lung adenocarcinoma can mimic triple-negative breast cancer. Given the prognosis and therapeutic challenges, a correct diagnosis appears essential, and molecular biomarkers could be useful. We report the case of a 52-year-old woman with a breast mass initially diagnosed as primary breast cancer and secondarily attached to breast metastasis from an EGFR-mutated lung adenocarcinoma. The same activating EGFR mutations were identified in both the primary lung carcinoma and the breast metastasis.

  11. Clinical Developments for the EGFR-TKI Combined with Radiotherapy in Advanced Non-small Cell Lung Cancer%EGFR-TKI联合放疗治疗晚期非小细胞肺癌的研究进展

    Institute of Scientific and Technical Information of China (English)

    李夏南; 朱广迎

    2014-01-01

    肺癌是全球最常见的恶性肿瘤之一,严重威胁人类生命。近年来,以表皮生长因子受体酪氨酸激酶抑制剂(epidermal growth factor receptor tyrosine kinase inhibitors, EGFR-TKIs)为首的靶向药物在肺癌治疗中取得巨大进展。因其具有高选择性和低毒性的优势,目前已成为IV期非小细胞肺癌(non-small cell lung cancer, NSCLC)EGFR突变患者的一线治疗方案。然而随着临床的广泛应用,继发耐药成为临床亟待解决的问题。近年来,基础研究证实, EGFR-TKI具有放射增敏性,理论上二者联合不但可以解决放疗后期肿瘤的放射抵抗以及EGFR-TKI继发耐药,还可以增加对肿瘤杀伤能力,同时副反应较同步放化疗小。因此,EGFR-TKI与放疗联合成为晚期NSCLC(IIIb期/IV期)极具探索的治疗模式。本文就EGFR-TKI与放疗联合治疗晚期NSCLC的基础与临床研究进展进行综述。%Lung cancer is one of the most common malignant tumor in the world, severely threatening human life. Recently, targeted therapy such as the epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) made huge progress in treatment of lung cancer. EGFR-TKIs, with its high selectivity and low toxicity, is the ifrst choice for EGFR-mutated patients in stage IV non-small cell lung cancer (NSCLC). However, secondary drug resistance becomes a clinical problem to be urgently resolved. In recent years, a series of preclinical studies showed that EGFR-TKI can enhance the antitumor activ-ity of ionizing radiation. hTerefore, EGFR-TKI combined with radiation is extremely promising therapy pattern for advanced NSCLC. hTis review will discuss the research status in EGFR-TKI and radiotherapy for advanced NSCLC.

  12. Impact of the 3D microenvironment on phenotype, gene expression, and EGFR inhibition of colorectal cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Anna C Luca

    Full Text Available Three-dimensional (3D tumor cell cultures grown in laminin-rich-extracellular matrix (lrECM are considered to reflect human tumors more realistic as compared to cells grown as monolayer on plastic. Here, we systematically investigated the impact of ECM on phenotype, gene expression, EGFR signaling pathway, and on EGFR inhibition in commonly used colorectal cancer (CRC cell lines. LrECM on-top (3D culture assays were performed with the CRC cell lines SW-480, HT-29, DLD-1, LOVO, CACO-2, COLO-205 and COLO-206F. Morphology of lrECM cultivated CRC cell lines was determined by phase contrast and confocal laser scanning fluorescence microscopy. Proliferation of cells was examined by MTT assay, invasive capacity of the cell lines was assayed using Matrigel-coated Boyden chambers, and migratory activity was determined employing the Fence assay. Differential gene expression was analyzed at the transcriptional level by the Agilent array platform. EGFR was inhibited by using the specific small molecule inhibitor AG1478. A specific spheroid growth pattern was observed for all investigated CRC cell lines. DLD-1, HT-29 and SW-480 and CACO-2 exhibited a clear solid tumor cell formation, while LOVO, COLO-205 and COLO-206F were characterized by forming grape-like structures. Although the occurrence of a spheroid morphology did not correlate with an altered migratory, invasive, or proliferative capacity of CRC cell lines, gene expression was clearly altered in cells grown on lrECM as compared to 2D cultures. Interestingly, in KRAS wild-type cell lines, inhibition of EGFR was less effective in lrECM (3D cultures as compared to 2D cell cultures. Thus, comparing both 2D and 3D cell culture models, our data support the influence of the ECM on cancer growth. Compared to conventional 2D cell culture, the lrECM (3D cell culture model offers the opportunity to investigate permanent CRC cell lines under more physiological conditions, i.e. in the context of molecular

  13. Epidermal growth factor receptor (EGFR mutations and expression in squamous cell carcinoma of the esophagus in central Asia

    Directory of Open Access Journals (Sweden)

    Abedi-Ardekani Behnoush

    2012-12-01

    Full Text Available Abstract Background Esophageal squamous cell carcinoma (ESCC shows geographic variations in incidence, with high incidences (>50/105 person-years in central Asia, including North Eastern Iran (Golestan and Northern India (Kashmir. In contrast to Western countries, smoking does not appear to be a significant risk factor for ESCC in central Asia. In lung adenocarcinoma, activating mutations in the gene encoding epidermal growth factor receptor (EGFR are frequent in tumors of never smokers of Asian origin, predicting therapeutic sensitivity to Egfr-targeting drugs. Methods In this study 152 cases of histologically confirmed ESCC from Iran (Tehran and Golestan Province and North India (Kashmir Valley have been analyzed for EGFR mutation by direct sequencing of exons 18–21. Egfr protein expression was evaluated by immunohistochemistry in 34 samples from Tehran and HER2 mutations were analyzed in 54 cases from Kashmir. Results A total of 14 (9.2% EGFR variations were detected, including seven variations in exons. Among those, four (2.6% were already documented in lung cancers, two were reported as polymorphisms and one was a potentially new activating mutation. All but one variation in introns were previously identified as polymorphisms. Over-expression of Egfr was detected in 22/34 (65% of tested cases whereas no HER2 mutation was found in 54 cases from Kashmir. Conclusion Overall, EGFR mutations appear to be a rare event in ESCC in high incidence areas of central Asia, although a very small proportion of cases may harbor mutations predicting sensitivity to anti-Egfr drugs.

  14. Screening for EGFR Mutations in Patients with Head and Neck Cancer Treated with Gefitinib on a Compassionate-Use Program: A Hellenic Cooperative Oncology Group Study

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    Samuel Murray

    2010-01-01

    Full Text Available Background and Aim. EGFR is commonly expressed in cancers of the head and neck (H and N, and anti-EGFR agents have demonstrated improvements in outcomes (TTP and OS. The aim of this study was to determine EGFR gene status in H and N cancer patients treated with gefitinib and to correlate mutational status with clinico-pathological data and response. Patients and Methods. Patients with histologically confirmed H and N cancer having failed prior treatment for advanced disease entered this compassionate-use-program. Nineteen patients received gefitinib. EGFR expression was assessed by IHC, gene copy number by FISH, and mutation analysis was conducted for EGFR (18-21, KRAS, BRAF (V600E, and HER-2 exon 20. An additional TKI naive cohort of 73 patients was also screened. Results. Mutations were detected in 6/19 patients (3× EGFR, 1× KRAS, and 2× HER2-exon 20. There were no significant differences in TTP or OS for patients with somatic EGFR mutations. No BRAF mutations were detected. Conclusions. The incidence of EGFR mutations in H and N cancer in this study was 5.3%. No statistically relevant correlations between mutation or gene gain and response or survival were observed. Due to the limited number of patients and low incidence of genetic aberrations in the genes analyzed, additional studies are warranted.

  15. 树突状细胞活化TCR基因转染记忆性T细胞联合化疗治疗EGFR-TKI耐药性晚期非小细胞肺癌的疗效与安全性%Efficacy and safety of combining infusion of antigen-activated,TCR gene-transfected memory T cells combined with chemotherapy for treating advanced non-small cell lung cancer resistant to EGFR-TKIs

    Institute of Scientific and Technical Information of China (English)

    江冠铭; 袁领勤; 贾筠; 林顺欢; 蔡彦敏; 刘淳; 吴依芬; 王在国; 黄树林

    2014-01-01

    Objective To examine whether infusion of antigen-activated memory T cells transfected with TCR genes alters the clinical efficacy and toxicity of pemetrexed-cisplatin combination chemotherapy to treatin the treatment of advanced non-small cell lung cancer resistant to EGFR-TKIs. Methods A total of 41 patients with non-small cell lung cancer were randomized into a group (n=20)that received standard chemotherapy involving a 3-week regime of pemetrexed (500 mg/m2)and cisplatin (75 mg/m2),or into a group (n=21)that received the same chemotherapy after harvesting,culturing and intravenously infusing DC-activated T cells.Curative effects,immune function,quality of life and adverse reactions were compared between the two treatments. Results Combining infu-sion of DC-activated T cells with chemotherapy significantly increased serum populations of CD3+,CD4+,CD8+,CD95+,and CD122+cells,as well as the CD4+/CD8+ ratio.Combination therapy and standard chemotherapy alone gave similar overall response rates of 33.33%and 30.00%(P>0.05),respectively,but combination therapy led to a significantly higher disease control rate(76.19%vs 60.00%, P0.05),疾病控制率分别为76.19%和60.00%(P<0.05)。中位疾病无进展时间联合治疗组为7.1个月,明显高于化疗组的3.7个月(P<0.05)。联合治疗组患者KPS评分改善率为85.71%,高于化疗组45.00%(P<0.05)。联合治疗组患者粒细胞减少、恶心呕吐和疲乏的发生率分别为46.6%、52.4%和23.8%,明显低于化疗组的75.0%、90.0%和65.0%(P<0.05)。结论负载抗原DC活化TCR基因转染记忆性T细胞联合培美曲塞、顺铂治疗EGFR-TKI耐药性晚期NSCLC的疗效较好,能提高患者免疫功能和改善生活质量,减低化疗毒副反应,安全性较好。

  16. Multiplex RT-PCR-based detections of CEA, CK20 and EGFR in colorectal cancer patients

    Science.gov (United States)

    Tsouma, Aikaterini; Aggeli, Chrysanthi; Lembessis, Panagiotis; Zografos, George N; Korkolis, Dimitris P; Pectasides, Dimitrios; Skondra, Maria; Pissimissis, Nikolaos; Tzonou, Anastasia; Koutsilieris, Michael

    2010-01-01

    AIM: To develop a multiplex reverse transcription polymerase chain reaction (RT-PCR) method detecting circulating tumor cells in the peripheral blood of colorectal cancer (CRC) patients. METHODS: Peripheral blood samples were collected from 88 CRC patients and 40 healthy individuals from the blood donors’ clinic and subsequently analyzed by multiplex RT-RCR for the expression of carcinoembryonic antigen (CEA), cytokeratin 20 (CK20) and epidermal growth factor receptor (EGFR) mRNA. The analysis involved determining the detection rates of CEA, CK20 and EGFR transcripts vs disease stage and overall survival. Median follow-up period was 19 mo (range 8-28 mo). RESULTS: Rates of CEA, CK20 and EGFR detection in CRC patients were 95.5%, 78.4% and 19.3%, respectively. CEA transcripts were detected in 3 healthy volunteer samples (7.5%), whereas all control samples were tested negative for CK20 and EGFR transcripts. The increasing number of positive detections for CEA, CK20 and EGFR transcripts in each blood sample was positively correlated with Astler-Coller disease stage (P < 0.001) and preoperative serum levels of CEA (P = 0.029) in CRC patients. Data analysis using Kaplan-Meier estimator documented significant differences in the overall survival of the different CRC patient groups as formed according to the increasing number of positivity for CEA, CK20 and EGFR transcripts. CONCLUSION: These data suggest that multiplex RT-PCR assay can provide useful information concerning disease stage and overall survival of CRC patients. PMID:21157973

  17. Focal Adhesion Kinase Inhibitors in Combination with Erlotinib Demonstrate Enhanced Anti-Tumor Activity in Non-Small Cell Lung Cancer.

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    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

  18. EGFR and HER2 expression in primary cervical cancers and corresponding lymph node metastases: Implications for targeted radiotherapy

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    Yang Zhengyan

    2008-08-01

    Full Text Available Abstract Background Proteins overexpressed on the surface of tumor cells can be selectively targeted. Epidermal growth factor receptor (EGFR and human epidermal growth factor receptor 2 (HER2 are among the most often targeted proteins. The level and stability of expression in both primary tumors and corresponding metastases is crucial in the assessment of a receptor as target for imaging in nuclear medicine and for various forms of therapy. So far, the expression of EGFR and HER2 has only been determined in primary cervical cancers, and we have not found published data regarding the receptor status in corresponding metastatic lesions. The goal of this study was to evaluate whether any of these receptors are suitable as target for clinical diagnosis and therapy. Methods Expression of EGFR and HER2 was investigated immunohistochemically in both lymph node metastases and corresponding primary cervical cancers (n = 53. HER2 and EGFR expression was scored using HercepTest criteria (0, 1+, 2+ or 3+. Results EGFR overexpression (2+ or 3+ was found in 64% (35/53 of the primary cervical tumors and 60% (32/53 of the corresponding lymph node metastases. There was a good concordance between the primary tumors and the paired metastases regarding EGFR expression. Only four patients who had 2+ or 3+ in the primary tumors changed to 0 or 1+ in lymph node metastases, and another two cases changed the other way around. None of the primary tumors or the lymph node metastases expressed HER2 protein. Conclusion The EGFR expression seems to be common and stable during cervical cancer metastasis, which is encouraging for testing of EGFR targeted radiotherapy. HER2 appears to be of poor interest as a potential target in the treatment of cervical cancer.

  19. EGFR and HER2 expression in primary cervical cancers and corresponding lymph node metastases: Implications for targeted radiotherapy

    International Nuclear Information System (INIS)

    Proteins overexpressed on the surface of tumor cells can be selectively targeted. Epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) are among the most often targeted proteins. The level and stability of expression in both primary tumors and corresponding metastases is crucial in the assessment of a receptor as target for imaging in nuclear medicine and for various forms of therapy. So far, the expression of EGFR and HER2 has only been determined in primary cervical cancers, and we have not found published data regarding the receptor status in corresponding metastatic lesions. The goal of this study was to evaluate whether any of these receptors are suitable as target for clinical diagnosis and therapy. Expression of EGFR and HER2 was investigated immunohistochemically in both lymph node metastases and corresponding primary cervical cancers (n = 53). HER2 and EGFR expression was scored using HercepTest criteria (0, 1+, 2+ or 3+). EGFR overexpression (2+ or 3+) was found in 64% (35/53) of the primary cervical tumors and 60% (32/53) of the corresponding lymph node metastases. There was a good concordance between the primary tumors and the paired metastases regarding EGFR expression. Only four patients who had 2+ or 3+ in the primary tumors changed to 0 or 1+ in lymph node metastases, and another two cases changed the other way around. None of the primary tumors or the lymph node metastases expressed HER2 protein. The EGFR expression seems to be common and stable during cervical cancer metastasis, which is encouraging for testing of EGFR targeted radiotherapy. HER2 appears to be of poor interest as a potential target in the treatment of cervical cancer

  20. EGFR participates downstream of ERα in estradiol-17β-D-glucuronide-induced impairment of Abcc2 function in isolated rat hepatocyte couplets.

    Science.gov (United States)

    Barosso, Ismael R; Zucchetti, Andrés E; Miszczuk, Gisel S; Boaglio, Andrea C; Taborda, Diego R; Roma, Marcelo G; Crocenzi, Fernando A; Sánchez Pozzi, Enrique J

    2016-04-01

    Estradiol-17β-D-glucuronide (E17G) induces acute endocytic internalization of canalicular transporters, including multidrug resistance-associated protein 2 (Abcc2) in rat, generating cholestasis. Several proteins organized in at least two different signaling pathways are involved in E17G cholestasis: one pathway involves estrogen receptor alpha (ERα), Ca(2+)-dependent protein kinase C and p38-mitogen activated protein kinase, and the other pathway involves GPR30, PKA, phosphoinositide 3-kinase/AKT and extracellular signal-related kinase 1/2. EGF receptor (EGFR) can potentially participate in both pathways since it interacts with GPR30 and ERα. Hence, the aim of this study was to analyze the potential role of this receptor and its downstream effectors, members of the Src family kinases in E17G-induced cholestasis. In vitro, EGFR inhibition by Tyrphostin (Tyr), Cl-387785 or its knockdown with siRNA strongly prevented E17G-induced impairment of Abcc2 function and localization. Activation of EGFR was necessary but not sufficient to impair the canalicular transporter function, whereas the simultaneous activation of EGFR and GPR30 could impair Abcc2 transport. The protection of Tyr was not additive to that produced by the ERα inhibitor ICI neither with that produced by Src kinase inhibitors, suggesting that EGFR shared the signaling pathway of ERα and Src. Further analysis of ERα, EGFR and Src activations induced by E17G, demonstrated that ERα activation precedes that of EGFR and EGFR activation precedes that of Src. In conclusion, activation of EGFR is a key factor in the alteration of canalicular transporter function and localization induced by E17G and it occurs before that of Src and after that of ERα.

  1. EPIDERMAL GROWTH FACTOR RECEPTOR (EGFR AND HUMAN PAPILLOMAVIRUS (HPV L1 CAPSID PROTEIN IN CERVICAL SQUAMOUS INTRAEPITHELIAL LESIONS

    Directory of Open Access Journals (Sweden)

    Balan Raluca

    2010-09-01

    Full Text Available We analyzed the immunohistochemical pattern of epidermal growth factor receptor (EGFR in cervical squamous intraepithelial lesions (SILs in correlation with L1 HPV capsid protein, in order to determine the relationship between EGFR expression and the infection status of human papillomavirus (HPV. The study included 40 cases, 24 LSIL (low grade SIL (CIN1, cervical intraepithelial neoplasia and 16 HSIL (high grade SIL (6 cases of CIN2 and 10 cases of CIN3. The immunoexpression of L1 HPV protein was assessed on conventional cervico-vaginal smears and EGFR was immunohistochemically evaluated on the corresponding cervical biopsies. The HPV L1 capsid protein was expressed in 45.83% of LSIL and 25% of HSIL. EGFR was overexpressed in 62,4% of HSIL (58,4% CIN2 and 41,6% CIN3 and 37,6% LSIL. The immunoexpression of L1 HPV has clinical application in the progression assessment of the cervical precancerous lesions without a correlation to the grade of the cervical SIL. EGFR is expressed by all proliferating squamous epithelial cells, thus corresponding with the grade of SIL. The evaluation of EGFR status, correlated with L1 HPV protein expression, can provide useful data of progression risk of cervical squamous intraepithelial lesions

  2. Temporal changes of EGFR mutations and T790M levels in tumour and plasma DNA following AZD9291 treatment.

    Science.gov (United States)

    Chia, Puey Ling; Do, Hongdo; Morey, Adrienne; Mitchell, Paul; Dobrovic, Alexander; John, Thomas

    2016-08-01

    AZD9291, a T790M specific epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), has demonstrated impressive response rates in tumours harbouring the EGFR T790M resistance mutation. Emergence of resistance to AZD9291 has been shown to occur through several different mechanisms including the development of new mutations (e.g. C797S) in the EGFR tyrosine kinase domain. We studied two patients with paired tumour biopsies and blood samples pre- and post-progression on AZD9291 to explore possible resistance mechanisms. Pre- and Post-AZD9291 tumour biopsies as well as serial plasma samples were collected from two patients on the AURA clinical study (AZD9291 First Time in Patients Ascending Dose study). Droplet digital PCR (ddPCR) assays were used to quantify T790M, the driver EGFR mutation, and the C797S mutation in genomic DNA from paired tumour biopsies and plasma cell-free DNA. In the first patient, both EGFR T790M and L858R became undetectable in the plasma within 1 month after treatment with AZD9291. However, the T790M and the original L858R mutation re-emerged with radiologically confirmed resistance to AZD9291. In patient two, the levels of T790M were undetectable at the time of radiological resistance to AZD9291 but increasing levels of the original EGFR exon 19 deletion was detected. MET amplification was detected in a biopsy performed on progression. The EGFR C797S mutation was not detected in either patient at the time of relapse. ddPCR of cell free DNA enables real time monitoring of patients on 3rd generation TKIs. As resistance mechanisms are variable, monitoring levels of the initial activating EGFR mutation may facilitate more reliable detection of progression. PMID:27393503

  3. Targeting TORC1/2 enhances sensitivity to EGFR inhibitors in head and neck cancer preclinical models.

    Science.gov (United States)

    Cassell, Andre; Freilino, Maria L; Lee, Jessica; Barr, Sharon; Wang, Lin; Panahandeh, Mary C; Thomas, Sufi M; Grandis, Jennifer R

    2012-11-01

    Head and neck squamous cell carcinoma (HNSCC) is characterized by overexpression of the epidermal growth factor receptor (EGFR) where treatments targeting EGFR have met with limited clinical success. Elucidation of the key downstream-pathways that remain activated in the setting of EGFR blockade may reveal new therapeutic targets. The present study was undertaken to test the hypothesis that inhibition of the mammalian target of rapamycin (mTOR) complex would enhance the effects of EGFR blockade in HNSCC preclinical models. Treatment of HNSCC cell lines with the newly developed TORC1/TORC2 inhibitor OSI-027/ASP4876 resulted in dose-dependent inhibition of proliferation with abrogation of phosphorylation of known downstream targets including phospho-AKT (Ser473), phospho-4E-BP1, phospho-p70s6K, and phospho-PRAS40. Furthermore, combined treatment with OSI-027 and erlotinib resulted in enhanced biochemical effects and synergistic growth inhibition in vitro. Treatment of mice bearing HNSCC xenografts with a combination of the Food and Drug Administration (FDA)-approved EGFR inhibitor cetuximab and OSI-027 demonstrated a significant reduction of tumor volumes compared with either treatment alone. These findings suggest that TORC1/TORC2 inhibition in conjunction with EGFR blockade represents a plausible therapeutic strategy for HNSCC. PMID:23226094

  4. Targeting TORC1/2 Enhances Sensitivity to EGFR Inhibitors in Head and Neck Cancer Preclinical Models

    Directory of Open Access Journals (Sweden)

    Andre Cassell

    2012-11-01

    Full Text Available Head and neck squamous cell carcinoma (HNSCC is characterized by overexpression of the epidermal growth factor receptor (EGFR where treatments targeting EGFR have met with limited clinical success. Elucidation of the key downstream-pathways that remain activated in the setting of EGFR blockade may reveal new therapeutic targets. The present study was undertaken to test the hypothesis that inhibition of the mammalian target of rapamycin (mTOR complex would enhance the effects of EGFR blockade in HNSCC preclinical models. Treatment of HNSCC cell lines with the newly developed TORC1/TORC2 inhibitor OSI-027/ASP4876 resulted in dose-dependent inhibition of proliferation with abrogation of phosphorylation of known downstream targets including phospho-AKT (Ser473, phospho-4E-BP1, phospho-p70s6K, and phospho-PRAS40. Furthermore, combined treatment with OSI-027 and erlotinib resulted in enhanced biochemical effects and synergistic growth inhibition in vitro. Treatment of mice bearing HNSCC xenografts with a combination of the Food and Drug Administration (FDA-approved EGFR inhibitor cetuximab and OSI-027 demonstrated a significant reduction of tumor volumes compared with either treatment alone. These findings suggest that TORC1/TORC2 inhibition in conjunction with EGFR blockade represents a plausible therapeutic strategy for HNSCC.

  5. Clinical Observation of Icotinib Hydrochloride for Advanced Non-small Cell Lung Cancer Patients with EGFR Status Identified

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    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.

  6. Long-Term Treatment with Erlotinib for EGFR Wild-Type Non-Small Cell Lung Cancer: A Case Report.

    Science.gov (United States)

    Polychronidou, Genovefa; Papakotoulas, Pavlos

    2013-01-01

    The epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) gefitinib and erlotinib are known to have greater efficacy in EGFR mutation-positive non-small cell lung cancer (NSCLC), although erlotinib also has activity in wild-type disease. We report the successful long-term maintenance treatment of a patient with EGFR wild-type NSCLC with gefitinib and later erlotinib. The patient (male; 44 years old; smoker) was diagnosed with EGFR wild-type NSCLC after computer tomography had revealed a mediastinal mass, and histology and mutation testing had identified the tumor as an EGFR wild-type grade 3 adenocarcinoma. The patient received multiple rounds of chemotherapy, followed by gefitinib maintenance (3 years). Later on, he received erlotinib maintenance and developed a persistent rash (grade 1/2) that lasted throughout the treatment. The patient's condition has remained stable on erlotinib for more than 5 years, with no evidence of progression. We describe the patient's disease course and treatment in the context of EGFR TKI therapy and the prognostic factors for long-term clinical outcomes of NSCLC, including the development of erlotinib-induced rash.

  7. Targeting EGFR and COX-2 as a potential treatment improvement strategy in cancer radiotherapy

    International Nuclear Information System (INIS)

    Molecular targeting, i.e. use of agents that counteract molecular processes that are dysregulated in cancer cells, which may be responsible for tumor radio-or chemoresistance, is a recently extensively investigated approach to further improve radiotherapy, chemotherapy or radiochemotherapy. Many potential targets for augmentation of radio (or chemo) response have been identified including epidermal growth factor receptor (EGFR), cyclooxygenase-2 (COX-2) enzyme, mutated ras and angiogenic molecules. Agents that selectively inhibit these molecules are becoming available at a rapid rate, and many of them have been shown in preclinical testing to be highly effective in improving tumor radioresponse or chemoresponse, without significantly affecting normal tissues. The interaction of EGFR and COX-2 inhibitors with radiation has attracted a flare of investigational interest, and is over-viewed here. EGFR is frequently overexpressed or mutated in many types of cancer, which is associated with more aggressive tumor behavior and poorer tumor response to cytotoxic agents, including radiation. Blockade of EGFR or interference with its downstream signaling processes can improve tumor treatment. Our own studies, using human tumor xenografts, demonstrated that blocking EGFR with C225 anti-EGFR antibody produces a dramatic enhancement of tumor radioresponse, and even more dramatic response when radiation is combined with chemotherapeutic agents. C225 acts by a number of mechanisms including inhibition of DNA repair from radiation damage, enhancement of apoptosis and tumor necrosis, and by inhibition of tumor angiogenesis. Inhibition of tyrosine kinase activation by small molecule agents, such as Iressa or Tarceva, is another approach in interfering with EGFR-signaling, which also showed potent enhancing effect on tumor radioresponse. There are two COX enzymes: COX-1 and COX-2. While COX-1 is a ubiquitous constitutive enzyme having housekeeping physiological function, COX-2 is an

  8. Thrombin stimulation of inflammatory breast cancer cells leads to aggressiveness via the EGFR-PAR1-Pak1 pathway.

    Science.gov (United States)

    Ohshiro, Kazufumi; Bui-Nguyen, Tri M; Divijendra Natha, Reddy S; Schwartz, Arnold M; Levine, Paul; Kumar, Rakesh

    2012-12-27

    Inflammatory breast cancer (IBC) accounts for a small fraction but aggressive form of epithelial breast cancer. Although the role of thrombin in cancer is beginning to be unfolded, its impact on the biology of IBC remains unknown. The purpose of this study was to establish the role of thrombin on the invasiveness of IBC cells. The IBC SUM149 cell line was treated with thrombin in the absence or presence of the epidermal growth factor receptor (EGFR) inhibitor erlotinib and protease-activated receptor 1 (PAR1) inhibitor. The effects of pharmacological inhibitors on the ability of thrombin to stimulate the growth rate and invasiveness were examined. We found that the inhibition of putative cellular targets of thrombin action suppresses both the growth and invasiveness of SUM149 cells in a concentration-dependent manner. In addition, thrombin-mediated increased invasion of SUM149 cells was routed through EGFR phosphorylation, and in turn, stimulation of the p21-activated kinase (Pak1) activity in a EGFR-sensitive manner. Interestingly, thrombin-mediated activation of the Pak1 pathway stimulation was blocked by erlotinib and PAR1 inhibitor. For proof-of-principle studies, we found immunohistochemical evidence of Pak1 activation as well as expression of PAR1 in IBC. Thrombin utilizes EGFR to relay signals promoting SUM149 cell growth and invasion via the Pak1 pathway. The study provides the rationale for future therapeutic approaches in mitigating the invasive nature of IBC by targeting Pak1 and/or EGFR.

  9. 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

  10. Nuclear EGFR Contributes to Acquired Resistance to Cetuximab

    OpenAIRE

    Li, Chunrong; Iida, Mari; Dunn, Emily F.; Ghia, Amol J.; Wheeler, Deric L

    2009-01-01

    Epidermal growth factor receptor (EGFR) is a ubiquitously expressed receptor tyrosine kinase involved in the etiology of several human cancers. Cetuximab is an EGFR blocking-antibody that has been approved for the treatment of patients with cancers of the head and neck (HNSCC) and metastatic colorectal cancer (mCRC). Previous reports have shown that EGFR translocation to the nucleus is associated with cell proliferation. Here we investigated mechanisms of acquired resistance to cetuximab usin...

  11. FIH-1 disrupts an LRRK1/EGFR complex to positively regulate keratinocyte migration.

    Science.gov (United States)

    Peng, Han; Kaplan, Nihal; Yang, Wending; Getsios, Spiro; Lavker, Robert M

    2014-12-01

    Factor inhibiting hypoxia-inducible factor 1 (FIH-1; official symbol HIF1AN) is a hydroxylase that negatively regulates hypoxia-inducible factor 1α but also targets other ankyrin repeat domain-containing proteins such as Notch receptor to limit epithelial differentiation. We show that FIH-1 null mutant mice exhibit delayed wound healing. Importantly, in vitro scratch wound assays demonstrate that the positive role of FIH-1 in migration is independent of Notch signaling, suggesting that this hydroxylase targets another ankyrin repeat domain-containing protein to positively regulate motogenic signaling pathways. Accordingly, FIH-1 increases epidermal growth factor receptor (EGFR) signaling, which in turn enhances keratinocyte migration via mitogen-activated protein kinase pathway, leading to extracellular signal-regulated kinase 1/2 activation. Our studies identify leucine-rich repeat kinase 1 (LRRK1), a key regulator of the EGFR endosomal trafficking and signaling, as an FIH-1 binding partner. Such an interaction prevents the formation of an EGFR/LRRK1 complex, necessary for proper EGFR turnover. The identification of LRRK1 as a novel target for FIH-1 provides new insight into how FIH-1 functions as a positive regulator of epithelial migration.

  12. Role and regulation of EGFR in actin remodeling in sperm capacitation and the acrosome reaction

    Institute of Scientific and Technical Information of China (English)

    Haim Breitbart; Nir Etkovitz

    2011-01-01

    To bind and fertilize the egg,the spermatozoon should undergo few biochemical and motility changes in the female reproductive tract collectively called capacitation.The capacitated spermatozoon binds to the egg zona pellucida,and then undergoes the acrosome reaction(AR),which allows its penetration into the egg.The mechanisms regulating sperm capacitation and the AR are not completely understood.In the present review,we summarize some data regarding the role and regulation of the epidermal growth factor receptor(EGFR)in these processes.In the capacitation process,the EGFR is partially activated by protein kinase A(PKA),resulting in phospholipase D(PLD)activation and actin polymerization.Protein kinase C alpha(PKCα),which is already activated at the beginning of the capacitation,also participates in PLD activation.Further activation of the EGFR at the end of the capacitation enhances intracellular Ca2+concentration leading to F-actin breakdown and allows the AR to take place.Under in vivo conditions,the EGFR can be directly activated by its known ligand epidermal growth factor(EGF),and indirectly by activating PKA or by transactivation mediated by G protein-coupled receptors(GPCRs)activation or by ouabain.Under physiological conditions,sperm PKA is activated mainly by bicarbonate,which activates the soluble adenylyl cyclase to produce cyclic adenosine monophosphate(cAMP),the activator of PKA.The GPCR activators angiotensin ll or lysophosphatidic acid,as well as ouabain and EGF are physiological components present in the female reproductive tract.

  13. Discovery of AZD8931, an Equipotent, Reversible Inhibitor of Signaling by EGFR, HER2, and HER3 Receptors.

    Science.gov (United States)

    Barlaam, Bernard; Anderton, Judith; Ballard, Peter; Bradbury, Robert H; Hennequin, Laurent F A; Hickinson, D Mark; Kettle, Jason G; Kirk, George; Klinowska, Teresa; Lambert-van der Brempt, Christine; Trigwell, Cath; Vincent, John; Ogilvie, Donald

    2013-08-01

    Deregulation of HER family signaling promotes proliferation and tumor cell survival and has been described in many human cancers. Simultaneous, equipotent inhibition of EGFR-, HER2-, and HER3-mediated signaling may be of clinical utility in cancer settings where the selective EGFR or HER2 therapeutic agents are ineffective or only modestly active. We describe the discovery of AZD8931 (2), an equipotent, reversible inhibitor of EGFR-, HER2-, and HER3-mediated signaling and the structure-activity relationships within this series. Docking studies based on a model of the HER2 kinase domain helped rationalize the increased HER2 activity seen with the methyl acetamide side chain present in AZD8931. AZD8931 exhibited good pharmacokinetics in preclinical species and showed superior activity in the LoVo tumor growth efficacy model compared to close analogues. AZD8931 is currently being evaluated in human clinical trials for the treatment of cancer. PMID:24900741

  14. Nuclear FABP7 immunoreactivity is preferentially expressed in infiltrative glioma and is associated with poor prognosis in EGFR-overexpressing glioblastoma

    Directory of Open Access Journals (Sweden)

    Aldape Ken D

    2006-04-01

    Full Text Available Abstract Background We previously identified brain type fatty acid-binding protein (FABP7 as a prognostic marker for patients with glioblastoma (GBM. Increased expression of FABP7 is associated with reduced survival. To investigate possible molecular mechanisms underlying this association, we compared the expression and subcellular localization of FABP7 in non-tumor brain tissues with different types of glioma, and examined the expression of FABP7 and epidermal growth factor receptor (EGFR in GBM tumors. Methods Expression of FABP7 in non-tumor brain and glioma specimens was examined using immunohistochemistry, and its correlation to the clinical behavior of the tumors was analyzed. We also analyzed the association between FABP7 and EGFR expression in different sets of GBM specimens using published DNA microarray datasets and semi-quantitative immunohistochemistry. In vitro migration was examined using SF763 glioma cell line. Results FABP7 was present in a unique population of glia in normal human brain, and its expression was increased in a subset of reactive astrocytes. FABP7 immunoreactivity in grade I pilocytic astrocytoma was predominantly cytoplasmic, whereas nuclear FABP7 was detected in other types of infiltrative glioma. Nuclear, not cytoplasmic, FABP7 immunoreactivity was associated with EGFR overexpression in GBM (N = 61, p = 0.008. Expression of the FABP7 gene in GBM also correlated with the abundance of EGFR mRNA in our previous microarray analyses (N = 34, p = 0.016 and an independent public microarray dataset (N = 28, p = 0.03. Compared to those negative for both markers, nuclear FABP7-positive/EGFR-positive and nuclear FABP7-positive/EGFR-negative GBM tumors demonstrated shortest survival, whereas those only positive for EGFR had intermediate survival. EGFR activation increased nuclear FABP7 immunoreactivity in a glioma cell line in vitro, and inhibition of FABP7 expression suppressed EGF-induced glioma-cell migration. Our data

  15. EMT-induced stemness and tumorigenicity are fueled by the EGFR/Ras pathway.

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    Dominic Chih-Cheng Voon

    Full Text Available Recent studies have revealed that differentiated epithelial cells would acquire stem cell-like and tumorigenic properties following an Epithelial-Mesenchymal Transition (EMT. However, the signaling pathways that participate in this novel mechanism of tumorigenesis have not been fully characterized. In Runx3 (-/- p53 (-/- murine gastric epithelial (GIF-14 cells, EMT-induced plasticity is reflected in the expression of the embryonal proto-oncogene Hmga2 and Lgr5, an exclusive gastrointestinal stem cell marker. Here, we report the concurrent activation of an EGFR/Ras gene expression signature during TGF-β1-induced EMT in GIF-14 cells. Amongst the altered genes was the induction of Egfr, which corresponded with a delayed sensitization to EGF treatment in GIF-14. Co-treatment with TGF-β1 and EGF or the expression of exogenous KRas led to increased Hmga2 or Lgr5 expression, sphere initiation and colony formation in soft agar assay. Interestingly, the gain in cellular plasticity/tumorigenicity was not accompanied by increased EMT. This uncoupling of EMT and the induction of plasticity reveals an involvement of distinct signaling cues, whereby the EGFR/Ras pathway specifically promotes stemness and tumorigenicity in EMT-altered GIF-14 cells. These data show that the EGFR/Ras pathway requisite for the sustenance of gastric stem cells in vivo and in vitro is involved in the genesis and promotion of EMT-induced tumor-initiating cells.

  16. Cross-talk between AMPK and EGFR dependent Signaling in Non-Small Cell Lung Cancer

    Science.gov (United States)

    Praveen, Paurush; Hülsmann, Helen; Sültmann, Holger; Kuner, Ruprecht; Fröhlich, Holger

    2016-06-01

    Lung cancers globally account for 12% of new cancer cases, 85% of these being Non Small Cell Lung Cancer (NSCLC). Therapies like erlotinib target the key player EGFR, which is mutated in about 10% of lung adenocarcinoma. However, drug insensitivity and resistance caused by second mutations in the EGFR or aberrant bypass signaling have evolved as a major challenge in controlling these tumors. Recently, AMPK activation was proposed to sensitize NSCLC cells against erlotinib treatment. However, the underlying mechanism is largely unknown. In this work we aim to unravel the interplay between 20 proteins that were previously associated with EGFR signaling and erlotinib drug sensitivity. The inferred network shows a high level of agreement with protein-protein interactions reported in STRING and HIPPIE databases. It is further experimentally validated with protein measurements. Moreover, predictions derived from our network model fairly agree with somatic mutations and gene expression data from primary lung adenocarcinoma. Altogether our results support the role of AMPK in EGFR signaling and drug sensitivity.

  17. miR-27a suppresses EV71 replication by directly targeting EGFR.

    Science.gov (United States)

    Zhang, Lianglu; Chen, Xiong; Shi, Yingying; Zhou, Bingfei; Du, Chen; Liu, Yongjuan; Han, Song; Yin, Jun; Peng, Biwen; He, Xiaohua; Liu, Wanhong

    2014-12-01

    Enterovirus 71 (EV71), a major causative agent of hand, foot, and mouth disease, has broken out several times and was accompanied by neurological disease. microRNAs, a class of small non-coding RNAs that are approximately 20 nucleotides long, play important roles in the regulation of various biological processes, including antiviral defense. However, the roles of miRNAs in EV71 replication and pathogenesis are not well understood. In this study, we found that the expression of miR-27a was significantly decreased in EV71-infected cells. Interestingly, the over-expression of miR-27a could inhibit EV71 replication, as measured by virus titration, qPCR, and Western blotting. We identified EGFR mRNA is a bona fide target of miR-27a by computational analysis and luciferase reporter assays. Furthermore, miR-27a could decrease EGFR expression, as measured by qPCR and Western blotting. Moreover, the inhibition of EGFR expression by miR-27a decreased the phosphorylation of Akt and ERK, which facilitate EV71 replication. These results suggest that miR-27a may have antiviral activity against EV71 by inhibiting EGFR.

  18. The challenge of targeting EGFR: experience with gefitinib in nonsmall cell lung cancer

    Directory of Open Access Journals (Sweden)

    C.L. Watkins

    2010-09-01

    Full Text Available As the first approved epidermal growth factor receptor (EGFR-targeted therapy for nonsmall cell lung cancer (NSCLC, the clinical development of gefitinib was complex. Advances in scientific understanding of the target biology during its clinical development enabled the identification of a biomarker to define patients most likely to derive benefit from gefitinib. Initial phase II trials showed clinically meaningful anti-tumour activity in 12–18% of unselected pretreated patients with advanced NSCLC at the optimum biological dose (250 mg. Subgroup analyses of these and subsequent phase III trials in unselected patients suggested that EGFR mutation and some clinical characteristics associated with a higher incidence of EGFR mutation (Asian ethnicity, adenocarcinoma histology, never-smoking and female sex were linked with increased response to gefitinib. Consequently, the IRESSA Pan-Asia Study (IPASS was conducted in never-smokers or former light-smokers in East Asia who had adenocarcinoma of the lung. IPASS showed that EGFR mutation was the strongest predictor of improved progression-free survival (mutation-positive subgroup hazard ratio (HR 0.48, 95% CI 0.36–0.64 (p<0.001, n = 261; mutation-negative subgroup HR 2.85, 95% CI 2.05–3.98 (p<0.001, n = 176; interaction test p<0.001 with gefitinib versus carboplatin/paclitaxel as first-line therapy for advanced NSCLC. Important lessons for the development of future personalised medicines are discussed.

  19. Protein determination in seeds by proton activation

    Science.gov (United States)

    Morales, J. R.; Dinator, M. I.; Cerda, P.

    1989-04-01

    A proton beam of 6.6 MeV has been used to produce 11C and 13N in Araucaria Araucana seeds. Their positron decay allows determination of the N/C ratio. In seeds the nitrogen content is associated to proteins while carbon is spread in the organic material. Samples were irradiated for about 10 min with a beam intensity of 5 nA on areas of 1 mm 2. Slices of the seed were radially explored, showing a larger concentration of protein in the center.

  20. Protein determination in seeds by proton activation

    Energy Technology Data Exchange (ETDEWEB)

    Morales, J.R.; Dinator, M.I. (Chile Univ., Santiago (Chile). Lab. de Fisica); Cerda, P. (Bio-Bio Univ., Chillan (Chile))

    1989-04-01

    A proton beam of 6.6 MeV has been used to produce /sup 11/C and /sup 13/N in Araucaria Araucana seeds. Their positron decay allows determination of the N/C ratio. In seeds the nitrogen content is associated to proteins while carbon is spread in the organic material. Samples were irradiated for about 10 min with a beam intensity of 5 nA on areas of 1 mm/sup 2/. Slices of the seed were radially explored, showing a larger concentration of protein in the center. (orig.).

  1. 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

  2. Prognostic Value of Baseline 18F-FDG PET/CT Functional Parameters in Patients with Advanced Lung Adenocarcinoma Stratified by EGFR Mutation Status.

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    Dalong Wang

    Full Text Available The study objective was to retrospectively analyze the metabolic variables derived from 18 F-fluorodeoxyglucose (18F-FDG positron emission tomography/computed tomography (PET/CT as predictors of progression-free survival (PFS and overall survival (OS in advanced lung adenocarcinoma stratified by epidermal growth factor receptor (EGFR mutation status. A total of 176 patients (91, EGFR mutation; 85, wild-type EGFR who underwent 18F-FDG PET/CT before treatment were enrolled. The main 18F-FDG PET/CT-derived variables: primary tumor maximum standardized uptake value (SUVmaxT, primary tumor total lesion glycolysis (TLGT, the maximum SUVmax of all selected lesions in whole body determined using the Response Evaluation Criteria In Solid Tumors (RECIST 1.1 criteria (SUVmaxWBR, and whole-body total TLG determined using the RECIST 1.1 criteria (TLGWBR were measured. Survival analysis regarding TLGWBR, and other factors in advanced lung adenocarcinoma patients stratified using EGFR mutation status, were evaluated. The results indicated that high TLGWBR (≥259.85, EGFR wild-type, and high serum LDH were independent predictors of worse PFS and OS in all patients with advanced lung adenocarcinoma. Among patients with wild-type EGFR, only TLGWBR retained significance as an independent predictor of both PFS and OS. Among patients with the EGFR mutation, high serum LDH level was an independent predictor of worse PFS and OS, and high TLGWBR (≥259.85 was an independent predictor of worse PFS but not worse OS. In conclusion, TLGWBR is a promising parameter for prognostic stratification of patients with advanced lung adenocarcinoma and EGFR status; however, it cannot be used to further stratify the risk of worse OS for patients with the EGFR mutation. Further prospective studies are needed to validate our findings.

  3. [Research Progress of EGFR-TKI Therapy for Patients with Central Nervous System 
Metastases from Non-small Cell Lung Cancer].

    Science.gov (United States)

    Jin, Yinghua; Xin, Tao

    2016-08-20

    Approximately half of all patients with non-small cell lung cancer (NSCLC) develop central nervous system metastases during the course of their disease which indicate poor prognosis. A part of NSCLC patients demonstrates activating epidermal growth factor receptor gene (EGFR) mutations who represent effectiveness and well tolerance of EGFR-specific tyrosine kinase inhibitors (TKIs) therapy. Although the systemic efficacy of targeted agents is established, the efficacy of central nervous system (CNS) metastases is not as well characterized. In this article, we review recent data on the use of EGFR inhibitors for treatment of patients with NSCLC and CNS metastases. PMID:27561797

  4. Differentiation of glioblastoma multiforme stem-like cells leads to downregulation of EGFR and EGFRvIII and decreased tumorigenic and stem-like cell potential

    DEFF Research Database (Denmark)

    Stockhausen, Marie-Thérése; Kristoffersen, Karina; Stobbe, Louise;

    2014-01-01

    Glioblastoma multiforme (GBM) is the most common and devastating primary brain tumor among adults. Despite recent treatment progress, most patients succumb to their disease within 2 years of diagnosis. Current research has highlighted the importance of a subpopulation of cells, assigned brain...... and activation of the epidermal growth factor receptor (EGFR) and expression of a deletion variant EGFRvIII. In the normal brain, EGFR is expressed in neurogenic areas where also NSC are located and it has been shown that EGFR is involved in regulation of NSC proliferation, migration, and differentiation...

  5. 晚期非小细胞肺癌 EGFR-TKI 获得性耐药机制%Acquired resistance mechanisms of EGFR-TKI in advanced non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    郑悦; 魏素菊

    2015-01-01

    Since the development of molecular biology,the treatment of advanced non-small cell cancer is shifting from traditional chemotherapy into molecular targeted therapy with genotyping as a guide′s help.The most widely used is epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs).With the appli-cation of EGFR-TKIs,the resistances to EGFR inhibitors are paid more and more attention,in recent years. The main mechanisms of acquired resistances are as follows:secondary mutation of the EGFR gene,amplifica-tion of c-MET,Her2 and other target genes,histological transformation,activation of the bypass mechanisms, loss of p53,the relief of negative feedback loops,overlap of mechanisms,etc.%随着分子生物学的发展,晚期非小细胞肺癌的治疗由传统的化疗转向以基因分型为指导的分子靶向治疗。其中应用较多的是表皮生长因子受体酪氨酸激酶抑制剂(EGFR-TKI)类药物。然而EGFR-TKI 的耐药问题近年来受到越来越多的关注,其机制主要有 EGFR 基因二次突变,c-MET、人类表皮生长因子受体2(Her2)等靶基因的扩增,组织表型的改变,旁路途径的激活,p53基因的缺失,细胞信号通路负反馈的减弱以及多重机制叠加等。

  6. Controlled physical activity for functional operability determination

    Directory of Open Access Journals (Sweden)

    Luchenkov А.А.

    2014-06-01

    Full Text Available The aim: to use veloergometry test at patients of high anaeshtesiology-operative risk before traumatic operations for functional operability determination. Material and methods: Randomized prospective research with the double "blind" control has been performed at 95 patients of high risk (ASA>III, undergoing long and traumatic operations on thoracic and abdominal organs. Patients have divided into groups without complications and with complications (51 and 44 patients which one day prior to operation have spent veloergometry test (VEMT. Average dynamic pressure (ADP, the general peripheral resistance of vessels (GPRV, a core index (Cl; arterio-venous (a-v difference on oxygen, oxygen delivery to tissues, consumption of oxygen and coefficients extraction oxygen in tissues; energy consumption have been investigated. Statistics has been done by nonparametric methods. Results. In response to veloergometry test in both groups growth of Cl at the expense of a tachycardia and GPRV fall which in complication group remains above norm is noted. In group without complications coefficients extraction oxygen in tissues were normalised, a-v difference on 02 became above norm, in other group — coefficients extraction oxygen in tissues and a-v difference on 02 began to exceed norm, and oxygen consumption has grown almost in 2 times. After VEMT a-v a difference on О and oxygen consumption were essentially above in group with complications. Under the influence of VEMT markecT stabilization of function of vegetative nervous systems (VNS. The number of complications made 39: intraoperation cardiovascular — 23, postoperative respiratory — 16. Clinically important connection (p=0,069 of perioperative complications with growth a-v differences on oxygen and a power interchanging have been received. Conclusion. Thus, the oxygen-energy exchange and the vegetative status can be referred to clinical functional operability determination.

  7. EGFR kinase promotes acquisition of stem cell-like properties: a potential therapeutic target in head and neck squamous cell carcinoma stem cells.

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    Eric L Abhold

    Full Text Available Members of the EGFR/ErbB family of tyrosine kinases are found to be highly expressed and deregulated in many cancers, including head and neck squamous cell carcinoma (HNSCC. The ErbB family, including EGFR, has been demonstrated to play key roles in metastasis, tumorigenesis, cell proliferation, and drug resistance. Recently, these characteristics have been linked to a small subpopulation of cells classified as cancer stem cells (CSCs which are believed to be responsible for tumor initiation and maintenance. In this study, we investigated the possible role of EGFR as a regulator of "stemness" in HNSCC cells. Activation of EGFR by the addition of EGF ligand or ectopic expression of EGFR in two established HNSCC cell lines (UMSCC-22B and HN-1 resulted in the induction of CD44, BMI-1, Oct-4, NANOG, CXCR4, and SDF-1. Activation of EGFR also resulted in increased tumorsphere formation, a characteristic ability of cancer stem cells. Conversely, treatment with the EGFR kinase inhibitor, Gefinitib (Iressa, resulted in decreased expression of the aforementioned genes, and loss of tumorsphere-forming ability. Similar trends were observed in a 99.9% CD44 positive stem cell culture derived from a fresh HNSCC tumor, confirming our findings for the cell lines. Additionally, we found that these putative cancer stem cells, when treated with Gefitinib, possessed a lower capacity to invade and became more sensitive to cisplatin-induced death in vitro. These results suggest that EGFR plays critical roles in the survival, maintenance, and function of cancer stem cells. Drugs that target EGFR, perhaps administered in combination with conventional chemotherapy, might be an effective treatment for HNSCC.

  8. Radiosensitivity of tumor cell lines after pretreatment with the EGFR tyrosine kinase inhibitor ZD1839 (Iressa {sup registered})

    Energy Technology Data Exchange (ETDEWEB)

    Burdak-Rothkamm, S. [Dept. of Radiotherapy, Saarland Univ. Hospital, Homburg/Saar (Germany); Gray Cancer Inst., Northwood, Middlesex (United Kingdom); Ruebe, C.E.; Nguyen, T.P.; Ludwig, D.; Ruebe, C. [Dept. of Radiotherapy, Saarland Univ. Hospital, Homburg/Saar (Germany); Feldmann, K. [AstraZeneca GmbH, Wedel (Germany); Wiegel, T. [Dept. of Radiotherapy, Univ. Hospital Benjamin Franklin, Berlin (Germany)

    2005-03-01

    Background and purpose: the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor ZD1839 (Iressa registered) reduces survival and augments radiation response of certain tumor cells. The aim of this study was to identify cellular events that are associated with the modulation of radiosensitivity by ZD1839. Material and methods: three tumor cell lines (A549, H596, FaDu) were exposed to ionizing radiation, treatment with ZD1839, and combined treatment. Clonogenic cell survival was determined by colony assays, EGFR and transforming growth factor-(TGF-){alpha} expression by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR), cell cycle distribution and apoptosis by flow cytometry. Results: in A549 and H596 cells ZD1839 had little effect on clonogenic growth, but survival curves revealed a radiosensitizing effect of 5 {mu}M ZD1839 on A549 cells. Both cell lines expressed moderate amounts of EGFR mRNA and very low levels of TGF-{alpha} mRNA. FaDu cells expressed relatively high amounts of EGFR and TGF-{alpha} transcripts and showed marked inhibition of clonogenic growth, reduction of S-phase cells, and induction of apoptosis after treatment with 1 {mu}M ZD1839 and combined treatment. Surprisingly, the subpopulation of FaDu cells surviving ZD1839 pretreatment was more radioresistant. Exposure to ZD1839 caused a decrease in EGFR mRNA expression in A549 cells, no change in H596, and even an increase in FaDu cells. Conclusion: the sensitivity to ZD1839 correlated with the EGFR expression level, an inhibition of cell proliferation, and induction of apoptosis in the cell lines analyzed. A radiosensitizing effect of ZD1839 was associated with downregulation of EGFR mRNA expression. (orig.)

  9. The human epidermal growth factor receptor (EGFR gene in European patients with advanced colorectal cancer harbors infrequent mutations in its tyrosine kinase domain

    Directory of Open Access Journals (Sweden)

    Delvenne Philippe

    2011-10-01

    Full Text Available Abstract Background The epidermal growth factor receptor (EGFR, a member of the ErbB family of receptors, is a transmembrane tyrosine kinase (TK activated by the binding of extracellular ligands of the EGF-family and involved in triggering the MAPK signaling pathway, which leads to cell proliferation. Mutations in the EGFR tyrosine kinase domain are frequent in non-small-cell lung cancer (NSCLC. However, to date, only very few, mainly non-European, studies have reported rare EGFR mutations in colorectal cancer (CRC. Methods We screened 236 clinical tumor samples from European patients with advanced CRC by direct DNA sequencing to detect potential, as yet unknown mutations, in the EGFR gene exons 18 to 21, mainly covering the EGFR TK catalytic domain. Results EGFR sequences showed somatic missense mutations in exons 18 and 20 at a frequency of 2.1% and 0.4% respectively. Somatic SNPs were also found in exons 20 and 21 at a frequency of about 3.1% and 0.4% respectively. Of these mutations, four have not yet been described elsewhere. Conclusions These mutation frequencies are higher than in a similarly sized population characterized by Barber and colleagues, but still too low to account for a major role played by the EGFR gene in CRC.

  10. Detecting and treating breast cancer resistance to EGFR inhibitors

    Science.gov (United States)

    Moonlee, Sun-Young; Bissell, Mina J.; Furuta, Saori; Meier, Roland; Kenny, Paraic A.

    2016-04-05

    The application describes therapeutic compositions and methods for treating cancer. For example, therapeutic compositions and methods related to inhibition of FAM83A (family with sequence similarity 83) are provided. The application also describes methods for diagnosing cancer resistance to EGFR inhibitors. For example, a method of diagnosing cancer resistance to EGFR inhibitors by detecting increased FAM83A levels is described.

  11. Expression of growth factor receptors and targeting of EGFR in cholangiocarcinoma cell lines

    International Nuclear Information System (INIS)

    Cholangiocarcinoma (CC) is a malignant neoplasm of the bile ducts or the gallbladder. Targeting of growth factor receptors showed therapeutic potential in palliative settings for many solid tumors. The aim of this study was to determine the expression of seven growth factor receptors in CC cell lines and to assess the effect of blocking the EGFR receptor in vitro. Expression of EGFR (epithelial growth factor receptor), HGFR (hepatocyte growth factor receptor) IGF1R (insulin-like growth factor 1 receptor), IGF2R (insulin-like growth factor 2 receptor) and VEGFR1-3 (vascular endothelial growth factor receptor 1-3) were examined in four human CC cell lines (EGI-1, HuH28, OZ and TFK-1). The effect of the anti-EGFR-antibody cetuximab on cell growth and apoptosis was studied and cell lines were examined for KRAS mutations. EGFR, HGFR and IGFR1 were present in all four cell lines tested. IGFR2 expression was confirmed in EGI-1 and TFK-1. No growth-inhibitory effect was found in EGI-1 cells after incubation with cetuximab. Cetuximab dose-dependently inhibited growth in TFK-1. Increased apoptosis was only seen in TFK-1 cells at the highest cetuximab dose tested (1 mg/ml), with no dose-response-relationship at lower concentrations. In EGI-1 a heterozygous KRAS mutation was found in codon 12 (c.35G>A; p.G12D). HuH28, OZ and TFK-1 lacked KRAS mutation. CC cell lines express a pattern of different growth receptors in vitro. Growth factor inhibitor treatment could be affected from the KRAS genotype in CC. The expression of EGFR itself does not allow prognoses on growth inhibition by cetuximab

  12. EGFR, FLT1 and heparanase as markers identifying patients at risk of short survival in cholangiocarcinoma.

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    Andreas-Claudius Hoffmann

    Full Text Available BACKGROUND: Cholangiocarcinoma remains to be a tumor with very few treatment choices and limited prognosis. In this study, we sought to determine the prognostic role of fms-related tyrosine kinase 1/vascular endothelial growth factor receptor 1 (FLT1/VEGFR1, heparanase (HPSE and epidermal growth factor receptor (EGFR gene expression in patients with resected CCC. METHODS: 47 formalin-fixed paraffin embedded FFPE tumor samples from patients with resected CCC were analyzed. FFPE tissues were dissected using laser-captured microdissection and analyzed for FLT1, FLT4, HPSE, Hif1a, VEGFA/C, HB-EGF, PDGFA, PDGF-RA and EGFR mRNA expression using a quantitative real-time RT-PCR method. Gene expression values (relative mRNA levels are expressed as ratios between the target gene and internal reference genes (beta-actin, b2mg, rplp2, sdha. RESULTS: EGFR, FLT1 and HPSE expression levels were significantly associated with overall survival (OS. FLT1 showed the strongest significant independent association with overall survival in a multivariate cox regression analysis when compared to the other genes and clinicopathological factors with a nearly 5 times higher relative risk (4.74 of dying earlier when expressed in low levels (p = 0.04. ROC Curve Analysis revealed that measuring EGFR potentially identifies patients at risk of a worsened outcome with a sensitivity of 80% and a specificity of 75% (p = 0.01. CONCLUSIONS: EGFR and FLT1 seem to be potential markers to identify those patients at high risk of dying from cholangiocarcinoma. Therefore these markers may help to identify patient subgroups in need for a more aggressive approach in a disease that is in desperate need for new approaches.

  13. Systems biology modeling reveals a possible mechanism of the tumor cell death upon oncogene inactivation in EGFR addicted cancers.

    Directory of Open Access Journals (Sweden)

    Jian-Ping Zhou

    Full Text Available Despite many evidences supporting the concept of "oncogene addiction" and many hypotheses rationalizing it, there is still a lack of detailed understanding to the precise molecular mechanism underlying oncogene addiction. In this account, we developed a mathematic model of epidermal growth factor receptor (EGFR associated signaling network, which involves EGFR-driving proliferation/pro-survival signaling pathways Ras/extracellular-signal-regulated kinase (ERK and phosphoinositol-3 kinase (PI3K/AKT, and pro-apoptotic signaling pathway apoptosis signal-regulating kinase 1 (ASK1/p38. In the setting of sustained EGFR activation, the simulation results show a persistent high level of proliferation/pro-survival effectors phospho-ERK and phospho-AKT, and a basal level of pro-apoptotic effector phospho-p38. The potential of p38 activation (apoptotic potential due to the elevated level of reactive oxygen species (ROS is largely suppressed by the negative crosstalk between PI3K/AKT and ASK1/p38 pathways. Upon acute EGFR inactivation, the survival signals decay rapidly, followed by a fast increase of the apoptotic signal due to the release of apoptotic potential. Overall, our systems biology modeling together with experimental validations reveals that inhibition of survival signals and concomitant release of apoptotic potential jointly contribute to the tumor cell death following the inhibition of addicted oncogene in EGFR addicted cancers.

  14. Pedometer-determined physical activity and active transport in girls

    Directory of Open Access Journals (Sweden)

    Schofield Grant

    2008-01-01

    Full Text Available Abstract Background It is well established that the risk of insufficient physical activity is greater in girls than in boys, especially during the adolescent years. The promotion of active transport (AT to and from school has been posited as a practical and convenient solution for increasing girls' total daily activity. However, there is limited information describing the associations between AT choices and girls' physical activity across a range of age, ethnic, and socioeconomic groups. The objectives of this study were to (1 investigate physical activity patterns in a large multiethnic sample of female children and adolescents, and to (2 estimate the physical activity associated with AT to and from school. Methods A total of 1,513 girls aged 5–16 years wore sealed multiday memory (MDM pedometers for three weekdays and two weekend days. The ethnic composition of this sample was 637 European (42.1%, 272 Pacific Island (18.0%, 207 East Asian (13.7%, 179 Maori (11.8%, 142 South Asian (9.4%, and 76 from other ethnic groups (5%. Pedometer compliance and school-related AT were assessed by questionnaire. Results Mean weekday step counts (12,597 ± 3,630 were higher and less variable than mean weekend steps (9,528 ± 4,407. A consistent decline in daily step counts was observed with age: after adjustment for ethnicity and SES, girls in school years 9–10 achieved 2,469 (weekday and 4,011 (weekend fewer steps than girls in years 1–2. Daily step counts also varied by ethnicity, with Maori girls the most active and South Asian girls the least active. Overall, 44.9% of participants used AT for school-related travel. Girls who used AT to and from school averaged 1,052 more weekday steps than those who did not use AT. However, the increases in steps associated with AT were significant only in older girls (school years 5–10 and in those of Maori or European descent. Conclusion Our data suggest that adolescent-aged girls and girls of Asian descent are

  15. EGFR and microvessel density in canine malignant mammary tumours.

    Science.gov (United States)

    Carvalho, Maria Isabel; Guimarães, Maria João; Pires, Isabel; Prada, Justina; Silva-Carvalho, Ricardo; Lopes, Carlos; Queiroga, Felisbina L

    2013-12-01

    The epidermal growth factor receptor (EGFR) is a transmembrane tyrosine kinase receptor which has been shown to have an important role in human breast cancer. Its role appears to be associated with increased angiogenesis and metastasis. In order to clarify its role in canine mammary tumours (CMT), 61 malignant neoplasms were studied by using immunohistochemistry, comparing expression of EGFR, microvessel density (MVD) by CD31 immunolabelling and characteristics of tumour aggressiveness. High EGFR immunoexpression was statistically significantly associated with tumour size, tumour necrosis, mitotic grade, histological grade of malignancy and clinical stage. High CD31 immunoreactivity was statistically significantly associated with tubule formation, histological grade of malignancy and clinical stage. A positive correlation between EGFR and CD31 immunoexpression (r = 0.843; P aggression in malignant CMT, presenting the possibility of using EGFR inhibitors in the context of metastatic disease treatment. PMID:24091029

  16. TAE226, a Bis-Anilino Pyrimidine Compound, Inhibits the EGFR-Mutant Kinase Including T790M Mutant to Show Anti-Tumor Effect on EGFR-Mutant Non-Small Cell Lung Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Hiroki Otani

    Full Text Available TAE226, a bis-anilino pyrimidine compound, has been developed as an inhibitor of focal adhesion kinase (FAK and insulin-like growth factor-I receptor (IGF-IR. In this study, we investigated the effect of TAE226 on non-small-cell lung cancer (NSCLC, especially focusing on the EGFR mutational status. TAE226 was more effective against cells with mutant EGFR, including the T790M mutant, than against cells with wild-type one. TAE226 preferentially inhibited phospho-EGFR and its downstream signaling mediators in the cells with mutant EGFR than in those with wild-type one. Phosphorylation of FAK and IGF-IR was not inhibited at the concentration at which the proliferation of EGFR-mutant cells was inhibited. Results of the in vitro binding assay indicated significant differences in the affinity for TAE226 between the wild-type and L858R (or delE746_A750 mutant, and the reduced affinity of ATP to the L858R (or delE746_A750 mutant resulted in good responsiveness of the L858R (or delE746_A750 mutant cells to TAE226. Of interest, the L858R/T790M or delE746_A750/T790M mutant enhanced the binding affinity for TAE226 compared with the L858R or delE746_A750 mutant, resulting in the effectiveness of TAE226 against T790M mutant cells despite the T790M mutation restoring the ATP affinity for the mutant EGFR close to that for the wild-type. TAE226 also showed higher affinity of about 15-fold for the L858R/T790M mutant than for the wild-type one by kinetic interaction analysis. The anti-tumor effect against EGFR-mutant tumors including T790M mutation was confirmed in mouse models without any significant toxicity. In summary, we showed that TAE226 inhibited the activation of mutant EGFR and exhibited anti-proliferative activity against NSCLCs carrying EGFR mutations, including T790M mutation.

  17. Conservation of protein abundance patterns reveals the regulatory architecture of the EGFR-MAPK pathway.

    Science.gov (United States)

    Shi, Tujin; Niepel, Mario; McDermott, Jason E; Gao, Yuqian; Nicora, Carrie D; Chrisler, William B; Markillie, Lye M; Petyuk, Vladislav A; Smith, Richard D; Rodland, Karin D; Sorger, Peter K; Qian, Wei-Jun; Wiley, H Steven

    2016-01-01

    Various genetic mutations associated with cancer are known to alter cell signaling, but it is not clear whether they dysregulate signaling pathways by altering the abundance of pathway proteins. Using a combination of RNA sequencing and ultrasensitive targeted proteomics, we defined the primary components-16 core proteins and 10 feedback regulators-of the epidermal growth factor receptor (EGFR)-mitogen-activated protein kinase (MAPK) pathway in normal human mammary epithelial cells and then quantified their absolute abundance across a panel of normal and breast cancer cell lines as well as fibroblasts. We found that core pathway proteins were present at very similar concentrations across all cell types, with a variance similar to that of proteins previously shown to display conserved abundances across species. In contrast, EGFR and transcriptionally controlled feedback regulators were present at highly variable concentrations. The absolute abundance of most core proteins was between 50,000 and 70,000 copies per cell, but the adaptors SOS1, SOS2, and GAB1 were found at far lower amounts (2000 to 5000 copies per cell). MAPK signaling showed saturation in all cells between 3000 and 10,000 occupied EGFRs, consistent with the idea that adaptors limit signaling. Our results suggest that the relative stoichiometry of core MAPK pathway proteins is very similar across different cell types, with cell-specific differences mostly restricted to variable amounts of feedback regulators and receptors. The low abundance of adaptors relative to EGFR could be responsible for previous observations that only a fraction of total cell surface EGFR is capable of rapid endocytosis, high-affinity binding, and mitogenic signaling. PMID:27405981

  18. Conservation of protein abundance patterns reveals the regulatory architecture of the EGFR-MAPK pathway

    Science.gov (United States)

    Shi, Tujin; Niepel, Mario; McDermott, Jason E.; Gao, Yuqian; Nicora, Carrie D.; Chrisler, William B.; Markillie, Lye M.; Petyuk, Vladislav A.; Smith, Richard D.; Rodland, Karin D.; Sorger, Peter K.; Qian, Wei-Jun; Wiley, H. Steven

    2016-01-01

    Various genetic mutations associated with cancer are known to alter cell signaling, but it is not clear whether they dysregulate signaling pathways by altering the abundance of pathway proteins. Using a combination of RNA sequencing and ultrasensitive targeted proteomics, we defined the primary components—16 core proteins and 10 feedback regulators—of the epidermal growth factor receptor (EGFR)–mitogen-activated protein kinase (MAPK) pathway in normal human mammary epithelial cells and then quantified their absolute abundance across a panel of normal and breast cancer cell lines as well as fibroblasts. We found that core pathway proteins were present at very similar concentrations across all cell types, with a variance similar to that of proteins previously shown to display conserved abundances across species. In contrast, EGFR and transcriptionally controlled feedback regulators were present at highly variable concentrations. The absolute abundance of most core proteins was between 50,000 and 70,000 copies per cell, but the adaptors SOS1, SOS2, and GAB1 were found at far lower amounts (2000 to 5000 copies per cell). MAPK signaling showed saturation in all cells between 3000 and 10,000 occupied EGFRs, consistent with the idea that adaptors limit signaling. Our results suggest that the relative stoichiometry of core MAPK pathway proteins is very similar across different cell types, with cell-specific differences mostly restricted to variable amounts of feedback regulators and receptors. The low abundance of adaptors relative to EGFR could be responsible for previous observations that only a fraction of total cell surface EGFR is capable of rapid endocytosis, high-affinity binding, and mitogenic signaling. PMID:27405981

  19. Abrogating phosphorylation of eIF4B is required for EGFR and mTOR inhibitor synergy in triple-negative breast cancer.

    Science.gov (United States)

    Madden, Julie M; Mueller, Kelly L; Bollig-Fischer, Aliccia; Stemmer, Paul; Mattingly, Raymond R; Boerner, Julie L

    2014-09-01

    Triple-negative breast cancer (TNBC) patients suffer from a highly malignant and aggressive disease. They have a high rate of relapse and often develop resistance to standard chemotherapy. Many TNBCs have elevated epidermal growth factor receptor (EGFR) but are resistant to EGFR inhibitors as monotherapy. In this study, we sought to find a combination therapy that could sensitize TNBC to EGFR inhibitors. Phospho-mass spectrometry was performed on the TNBC cell line, BT20, treated with 0.5 μM gefitinib. Immunoblotting measured protein levels and phosphorylation. Colony formation and growth assays analyzed the treatment on cell proliferation, while MTT assays determined the synergistic effect of inhibitor combination. A Dual-Luciferase reporter gene plasmid measured translation. All statistical analysis was done on CalucuSyn and GraphPad Prism using ANOVAs. Phospho-proteomics identified the mTOR pathway to be of interest in EGFR inhibitor resistance. In our studies, combining gefitinib and temsirolimus decreased cell growth and survival in a synergistic manner. Our data identified eIF4B, as a potentially key fragile point in EGFR and mTOR inhibitor synergy. Decreased eIF4B phosphorylation correlated with drops in growth, viability, clonogenic survival, and cap-dependent translation. Taken together, these data suggest EGFR and mTOR inhibitors abrogate growth, viability, and survival via disruption of eIF4B phosphorylation leading to decreased translation in TNBC cell lines. Further, including an mTOR inhibitor along with an EGFR inhibitor in TNBC with increased EGFR expression should be further explored. Additionally, translational regulation may play an important role in regulating EGFR and mTOR inhibitor synergy and warrant further investigation.

  20. Structural and institutional determinants of investment activity in Africa

    OpenAIRE

    Chuku, Chuku; Onye, Kenneth; Ajah, Hycent

    2015-01-01

    This paper considers the structural and institutional determinants of investment activity in selected African countries within a neoclassical framework. Generalized method of moments and a family of panel data estimation techniques are utilized in addition to nonparametric kernel regression techniques to uncover the relationship. Three main findings emerge; (i) financial openness and institutional quality are reasonably robust structural and institutional determinants of investment activity...

  1. Determinants of Change in Physical Activity in Children and Adolescents

    OpenAIRE

    Craggs, Christopher; Corder, Kirsten; van Sluijs, Esther M. F.; Simon J Griffin

    2011-01-01

    Context Data are available on correlates of physical activity in children and adolescents, less is known about the determinants of change. This review aims to systematically review the published evidence regarding determinants of change in physical activity in children and adolescents. Evidence acquisition Prospective quantitative studies investigating change in physical activity in children and adolescents aged 4–18 years were identified from seven databases (to November 2010): PubMed, SCOPU...

  2. 表皮生长因子受体抑制剂耐药机制的研究进展%Research Progress of Mechanism of Resistance to EGFR Inhibitors

    Institute of Scientific and Technical Information of China (English)

    冯昌怡

    2012-01-01

    The epidermal growth factor receptor( EGFR )is an important molecular target in cancer treatment, and two classes of anti-EGFR agents, the monoclonal antibodies and the small molecular tyro-sine kinase inhibitors,have developed in clinical successfully. Howeverthe primary and secondary resistance to anti-EGFR agents have been paid increasing attention. The mechanisms of resistance to EGFR inhibitors in solid tumors may include:EGFR,K-ras,Braf mutations;autocrine/paracrine production of lig-and;MET amplification; constitutive activation of a downstream pathway; activation of alternative pathways; activation of EGFR-independent, tumour-induced angiogenesis and etc. .%表皮生长因子受体(EGFR)是抗肿瘤治疗中重要的分子靶点,两类抗表皮生长因子受体(anti-EGFR)药物:单克隆抗体及小分子酪氨酸激酶抑制剂已成功应用于临床,但其原发和继发耐药问题也日益受到关注.肿瘤对anti-EGFR药物耐药的可能机制包括:EGFR、K-ras、Braf突变;配体的自分泌或旁分泌;原癌基因MET扩增;下游通路持续活化;激活替代通路;肿瘤诱导不依赖于EGFR活化的血管生成等.

  3. The T790M mutation in EGFR kinase causes drug resistance by increasing the affinity for ATP

    Energy Technology Data Exchange (ETDEWEB)

    Yun, C.H.; Mengwasser, K.E.; Toms, A.V.; Woo, M.S.; Greulich, H.; Wong, K.K.; Meyerson, M.; Eck, M.J. (Harvard-Med); (DFCI)

    2008-07-15

    Lung cancers caused by activating mutations in the epidermal growth factor receptor (EGFR) are initially responsive to small molecule tyrosine kinase inhibitors (TKIs), but the efficacy of these agents is often limited because of the emergence of drug resistance conferred by a second mutation, T790M. Threonine 790 is the 'gatekeeper' residue, an important determinant of inhibitor specificity in the ATP binding pocket. The T790M mutation has been thought to cause resistance by sterically blocking binding of TKIs such as gefitinib and erlotinib, but this explanation is difficult to reconcile with the fact that it remains sensitive to structurally similar irreversible inhibitors. Here, we show by using a direct binding assay that T790M mutants retain low-nanomolar affinity for gefitinib. Furthermore, we show that the T790M mutation activates WT EGFR and that introduction of the T790M mutation increases the ATP affinity of the oncogenic L858R mutant by more than an order of magnitude. The increased ATP affinity is the primary mechanism by which the T790M mutation confers drug resistance. Crystallographic analysis of the T790M mutant shows how it can adapt to accommodate tight binding of diverse inhibitors, including the irreversible inhibitor HKI-272, and also suggests a structural mechanism for catalytic activation. We conclude that the T790M mutation is a 'generic' resistance mutation that will reduce the potency of any ATP-competitive kinase inhibitor and that irreversible inhibitors overcome this resistance simply through covalent binding, not as a result of an alternative binding mode.

  4. Proximate analysis for determination of micropores in granulated activated carbon

    Energy Technology Data Exchange (ETDEWEB)

    Berman, Ya. G.; Nikolaev, V.B.; Shepelev, A.N.

    1987-02-01

    A method is discussed for determining the specific micropore volume of granulated activated carbon used for water treatment in Soviet coking plants. Toluene molecules with a diameter of 0.67 nm are sorbed by activated carbon with micropore diameter ranging from 0.7 to 1.4 nm. Therefore, sorptive properties of activated carbon in relation to toluene supply information on micropore volume in carbon. A formula which describes this relation is derived. The method for determining micropore volume on the basis of toluene adsorption was tested using 8 types of activated carbon produced from coal and petroleum. Types of activated carbon characterized by the highest adsorption were selected. 1 ref.

  5. Oncogenic EGFR Represses the TET1 DNA Demethylase to Induce Silencing of Tumor Suppressors in Cancer Cells

    Directory of Open Access Journals (Sweden)

    Matteo Forloni

    2016-07-01

    Full Text Available Oncogene-induced DNA methylation-mediated transcriptional silencing of tumor suppressors frequently occurs in cancer, but the mechanism and functional role of this silencing in oncogenesis are not fully understood. Here, we show that oncogenic epidermal growth factor receptor (EGFR induces silencing of multiple unrelated tumor suppressors in lung adenocarcinomas and glioblastomas by inhibiting the DNA demethylase TET oncogene family member 1 (TET1 via the C/EBPα transcription factor. After oncogenic EGFR inhibition, TET1 binds to tumor suppressor promoters and induces their re-expression through active DNA demethylation. Ectopic expression of TET1 potently inhibits lung and glioblastoma tumor growth, and TET1 knockdown confers resistance to EGFR inhibitors in lung cancer cells. Lung cancer samples exhibited reduced TET1 expression or TET1 cytoplasmic localization in the majority of cases. Collectively, these results identify a conserved pathway of oncogenic EGFR-induced DNA methylation-mediated transcriptional silencing of tumor suppressors that may have therapeutic benefits for oncogenic EGFR-mediated lung cancers and glioblastomas.

  6. EGFR基因突变检测的临床应用%Clinical Application of Detecting EGFR Mutations

    Institute of Scientific and Technical Information of China (English)

    周瑞芳; 卢仁泉

    2013-01-01

    EGFR(epidermal growth factor receptor)is a transmembrane tyrosine kinase receptor. Mutations that lead to EGFR overexpression or overactivity have been associated with a number of cancers. Mutations of EGFR could lead to its constant activation,which plays an important role in the proliferation,apoptosis and angiogenesis of cancerous cells. Recently, the detection of EGFR mutations in lung cancer is paid enough attention. Mastering the test methods and common sample pretreatment could provide valuable insights into its clinical application.%表皮生长因子受体(epidermal growth factor receptor,EGFR)是一种跨膜受体酪氨酸激酶,在许多肿瘤中过表达或发生突变,通过信号转导对癌细胞增殖、凋亡和血管生成具有重要意义,已成为抗肿瘤药物的重要靶点之一.近年来,尤其是肺癌患者中开展该基因突变的检测已逐渐受到重视.掌握EGFR基因突变检测的临床意义,EGFR基因突变检测常用方法以及EGFR基因突变检测常用的样本及其处理,可为其今后在临床中的广泛应用提供更广的前景.

  7. Crosstalk between EGFR and integrin affects invasion and proliferation of gastric cancer cell line, SGC7901

    Directory of Open Access Journals (Sweden)

    Dan L

    2012-10-01

    Full Text Available Li Dan,1,* Ding Jian,2,* Lin Na,1 Wang Xiaozhong,1 1Digestive Department, the Union Hospital of Fujian Medical University, Fujian, People’s Republic of China; 2Digestive Department, the First Affiliated Hospital of Fujian Medical University, Fuzhou, Fujian, People’s Republic of China*These authors contributed equally to this workBackground/objective: To investigate the crosstalk between epidermal growth factor receptor (EGFR and integrin-mediated signal transduction pathways in human gastric adenocarcinoma cells.Methods: EGF was used as a ligand of EGFR to stimulate the gastric adenocarcinoma cell, SGC7901. Signal molecules downstream of the integrin, FAK(Y397 and p130cas(Y410 phosphorylation, were measured by immunoprecipitation and western blot. Fibronectin (Fn was used as a ligand of integrin to stimulate the same cell line. Signal molecules downstream of EGFR and extracellular signal-regulated kinase (ERK general phosphorylation were also measured. Focal adhesion kinase (FAK small-interfering RNA was designed and transfected into SGC7901 cells to decrease the expression of FAK. Modified Boyden chambers and MTT assay were used to examine the effect of FAK inhibition on the invasiveness and proliferation of SGC7901.Results: EGF activated FAK(Y397 and p130cas(Y410 phosphorylation, while Fn activated ERK general phosphorylation. Inhibition of FAK expression decreased p130cas(Y410 phosphorylation activated by EGF and ERK general phosphorylation activated by Fn, also decreased the invasiveness and proliferation of SGC7901 cells activated by EGF or Fn.Conclusion: There is crosstalk between EGFR and integrin signal transduction. FAK may be a key cross point of the two signal pathways and acts as a potential target for human gastric cancer therapy.Keywords: gastric adenocarcinoma, epidermal growth factor receptor, integrin, focal adhesion kinase, crosstalk

  8. PD-1 Axis Inhibitors in EGFR- and ALK-Driven Lung Cancer: Lost Cause?

    Science.gov (United States)

    Gettinger, Scott; Politi, Katerina

    2016-09-15

    Programmed death axis 1 (PD-1) inhibitors have ushered in a new error of cancer immunotherapeutics for advanced smoking-associated non-small cell lung cancer. Their role in treating EGFR-mutant and ALK-rearranged lung cancer has yet to be determined. Clin Cancer Res; 22(18); 4539-41. ©2016 AACRSee related article by Gainor et al., p. 4585. PMID:27470969

  9. EGFR Amplification and Glioblastoma Stem-Like Cells

    Directory of Open Access Journals (Sweden)

    Katrin Liffers

    2015-01-01

    Full Text Available Glioblastoma (GBM, the most common malignant brain tumor in adults, contains a subpopulation of cells with a stem-like phenotype (GS-cells. GS-cells can be maintained in vitro using serum-free medium supplemented with epidermal growth factor, basic fibroblast growth factor-2, and heparin. However, this method does not conserve amplification of the Epidermal Growth Factor Receptor (EGFR gene, which is present in over 50% of all newly diagnosed GBM cases. GS-cells with retained EGFR amplification could overcome the limitations of current in vitro model systems and contribute significantly to preclinical research on EGFR-targeted therapy. This review recapitulates recent methodological approaches to expand stem-like cells from GBM with different EGFR status in order to maintain EGFR-dependent intratumoral heterogeneity in vitro. Further, it will summarize the current knowledge about the impact of EGFR amplification and overexpression on the stem-like phenotype of GBM-derived GS-cells and different approaches to target the EGFR-dependent GS-cell compartment of GBM.

  10. Impact of bevacizumab in combination with erlotinib on EGFR-mutated non-small cell lung cancer xenograft models with T790M mutation or MET amplification.

    Science.gov (United States)

    Furugaki, Koh; Fukumura, Junko; Iwai, Toshiki; Yorozu, Keigo; Kurasawa, Mitsue; Yanagisawa, Mieko; Moriya, Yoichiro; Yamamoto, Kaname; Suda, Kenichi; Mizuuchi, Hiroshi; Mitsudomi, Tetsuya; Harada, Naoki

    2016-02-15

    Erlotinib (ERL), an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, shows notable efficacy against non-small cell lung cancer (NSCLC) harboring EGFR mutations. Bevacizumab (BEV), a humanized monoclonal antibody to vascular endothelial cell growth factor (VEGF), in combination with ERL (BEV+ERL) significantly extended progression-free survival in patients with EGFR-mutated NSCLC compared with ERL alone. However, the efficacy of BEV+ERL against EGFR-mutated NSCLC harboring T790M mutation or MET amplification, is unclear. Here, we examined the antitumor activity of BEV+ERL in four xenograft models of EGFR-mutated NSCLC (three harboring ERL resistance mutations). In the HCC827 models (exon 19 deletion: DEL), ERL significantly inhibited tumor growth by blocking EGFR signal transduction. Although there was no difference between ERL and BEV+ERL in maximum tumor growth inhibition, BEV+ERL significantly suppressed tumor regrowth during a drug-cessation period. In the HCC827-EPR model (DEL+T790M) and HCC827-vTR model (DEL+MET amplification), ERL reduced EGFR signal transduction and showed less pronounced but still significant tumor growth inhibition than in the HCC827 model. In these models, tumor growth inhibition was significantly stronger with BEV+ERL than with each single agent. In the NCI-H1975 model (L858R+T790M), ERL did not inhibit growth or EGFR signal transduction, and BEV+ERL did not inhibit growth more than BEV. BEV alone significantly decreased microvessel density in each tumor. In conclusion, addition of BEV to ERL did not enhance antitumor activity in primarily ERL-resistant tumors with T790M mutation; however, BEV+ERL enhanced antitumor activity in T790M mutation- or MET amplification-positive tumors as long as their growth remained significantly suppressed by ERL. PMID:26370161

  11. EGFR testing and clinical management of advanced NSCLC: a Galician Lung Cancer Group study (GGCP 048-10

    Directory of Open Access Journals (Sweden)

    Vázquez S

    2016-02-01

    %, respectively. All but two patients received gefitinib. Median progression-free survival and overall survival were 10 (95% confidence interval: 4.8–15.3 months and 17.8 (95% confidence interval: 13.9–21.6 months, respectively, in patients carrying sensitizing mutations. Conclusion: The incidence of EGFR mutations in Galicia is consistent with previous data in Spain. Our results also support the feasibility of EGFR testing to guide treatment decision making using tumor tissue or cytology samples, or serum samples if tumor specimens are unavailable. These findings also confirm that first-line gefitinib is an active treatment option in Caucasians with EGFR mutation-positive NSCLC. Keywords: epidermal growth factor receptor, EGFR tyrosine inhibitors, TKIs, EGFR gene mutation, EGFR mutation testing, non-small-cell lung cancer

  12. Progress of EGFR Mutation and EGFR-targeted Therapy%EGFR基因突变与肿瘤靶向治疗

    Institute of Scientific and Technical Information of China (English)

    薛丽; 白玉杰

    2012-01-01

    表皮生长因子受体(epidermal growth factor receptor,EGFR)属于受体酪氨酸激酶超家族,在多种恶性肿瘤中表达.配体与EGFR结合诱导形成二聚体和构象变化,活化酪氨酸激酶及信号转导途径,产生细胞增殖、侵润、转移及抗凋亡等效应.EGFR酪氨酸激酶抑制剂(tyrosine kinase inhibitors,TKIs)类靶向药物,如吉非替尼和厄洛替尼等已应用于临床.临床研究显示仅10%~30%患者对TKIs敏感,部分位于EGFR激酶结构域的活化突变与药物敏感性相关.检测EGFR基因突变有助于预测对药物敏感性和提高疗效.随着治疗绝大多数敏感的患者获得继发耐药性,其中约半数有继发突变T790M,降低药物对靶分子的亲和力,其他许多位于EGFR下游信号途径或旁激活途径的分子也参与耐药形成.因此,未来个体化用药和准确预测敏感性,不仅仅要分析EGFR基因,而且要综合考虑下游和其他信号途径的基因,如PI3K,K-RAS,BRAF,MET和PTEN等.%The epidermal growth factor receptor(EGFR) belongs to the super-family of receptor tyrosine kinase and is expressed in many malignancies. Upon their ligand-induced dimerization and conformational change, it initiates activation of intracellular tyrosine kinase and a vast array of cell signaling pathways. The cascade of intracellular activation results in cell proliferation, invasion, metastasis and decreased apoptosis. EGFR represents a critical player in many types of malignancies and becomes a natural goal of the targeted therapy. Both monoclonal antibodies and tyrosine kinase inhibitors (TKIs), such as erlotinib and gefitinib, have already entered clinical application. Clinical trials have shown that only 10%-30% of patients responded to the TKIs treatment. Further studies have found that some mutations within the kinase domain activated the kinase and conferred sensitivity to the treatments. The detection of mutations will likely predict the sensitive patients and

  13. Immunohistochemical expression of SP-NK-1R-EGFR pathway and VDR in colonic inflammation and neoplasia

    OpenAIRE

    Isidro, Raymond A; Cruz, Myrella L; Isidro, Angel A; Baez, Axel; Arroyo, Axel; González-Marqués, William A; González-Keelan, Carmen; Torres, Esther A.; Appleyard, Caroline B.

    2015-01-01

    AIM: To determine the expression of neurokinin-1 receptor (NK-1R), phosphorylated epidermal growth factor receptor (pEGFR), cyclooxygenase-2 (Cox-2), and vitamin D receptor (VDR) in normal, inflammatory bowel disease (IBD), and colorectal neoplasia tissues from Puerto Ricans.

  14. Phase I/II clinical and pharmacokinetic study evaluating a fully human monoclonal antibody against EGFr (HuMax-EGFr) in patients with advanced squamous cell carcinoma of the head and neck

    DEFF Research Database (Denmark)

    Bastholt, Lars; Specht, Lena; Jensen, Kenneth;

    2007-01-01

    PURPOSE: To assess safety, tolerability, pharmacokinetics and clinical activity of HuMax-EGFr in patients with SCCHN. PATIENTS AND METHODS: Twenty-eight patients with SCCHN were enrolled. The study comprised a single-dose escalation part for assessment of safety issues followed by a repeat dose...

  15. EGFR-independent autophagy induction with gefitinib and enhancement of its cytotoxic effect by targeting autophagy with clarithromycin in non-small cell lung cancer cells.

    Science.gov (United States)

    Sugita, Shohei; Ito, Kentaro; Yamashiro, Yutaro; Moriya, Shota; Che, Xiao-Fang; Yokoyama, Tomohisa; Hiramoto, Masaki; Miyazawa, Keisuke

    2015-05-22

    Gefitinib (GEF), an inhibitor for EGFR tyrosine kinase, potently induces autophagy in non-small cell lung cancer (NSCLC) cell lines such as PC-9 cells expressing constitutively activated EGFR kinase by EGFR gene mutation as well as A549 and H226 cells with wild-type EGFR. Unexpectedly, GEF-induced autophagy was also observed in non-NSCLC cells such as murine embryonic fibroblasts (MEF) and leukemia cell lines K562 and HL-60 without EGFR expression. Knockout of EGFR gene in A549 cells by CRISPR/Cas9 system still exhibited autophagy induction after treatment with GEF, indicating that the autophagy induction by GEF is not mediated through inhibiting EGFR kinase activity. Combined treatment with GEF and clarithromycin (CAM), a macrolide antibiotic having the effect of inhibiting autophagy flux, enhances the cytotoxic effect in NSCLC cell lines, although treatment with CAM alone exhibits no cytotoxicity. GEF treatment induced up-regulation of endoplasmic reticulum (ER)-stress related genes such as CHOP/GADD153 and GRP78. Knockdown of CHOP in PC-9 cells and Chop-knockout MEF both exhibited less sensitivity to GEF than controls. Addition of CAM in culture medium resulted in further pronounced GEF-induced ER stress loading, while CAM alone exhibited no effect. These data suggest that GEF-induced autophagy functions as cytoprotective and indicates the potential therapeutic possibility of using CAM for GEF therapy. Furthermore, it is suggested that the intracellular signaling for autophagy initiation in response to GEF can be completely dissociated from EGFR, but unknown target molecule(s) of GEF for autophagy induction might exist.

  16. EGFR-independent autophagy induction with gefitinib and enhancement of its cytotoxic effect by targeting autophagy with clarithromycin in non-small cell lung cancer cells.

    Science.gov (United States)

    Sugita, Shohei; Ito, Kentaro; Yamashiro, Yutaro; Moriya, Shota; Che, Xiao-Fang; Yokoyama, Tomohisa; Hiramoto, Masaki; Miyazawa, Keisuke

    2015-05-22

    Gefitinib (GEF), an inhibitor for EGFR tyrosine kinase, potently induces autophagy in non-small cell lung cancer (NSCLC) cell lines such as PC-9 cells expressing constitutively activated EGFR kinase by EGFR gene mutation as well as A549 and H226 cells with wild-type EGFR. Unexpectedly, GEF-induced autophagy was also observed in non-NSCLC cells such as murine embryonic fibroblasts (MEF) and leukemia cell lines K562 and HL-60 without EGFR expression. Knockout of EGFR gene in A549 cells by CRISPR/Cas9 system still exhibited autophagy induction after treatment with GEF, indicating that the autophagy induction by GEF is not mediated through inhibiting EGFR kinase activity. Combined treatment with GEF and clarithromycin (CAM), a macrolide antibiotic having the effect of inhibiting autophagy flux, enhances the cytotoxic effect in NSCLC cell lines, although treatment with CAM alone exhibits no cytotoxicity. GEF treatment induced up-regulation of endoplasmic reticulum (ER)-stress related genes such as CHOP/GADD153 and GRP78. Knockdown of CHOP in PC-9 cells and Chop-knockout MEF both exhibited less sensitivity to GEF than controls. Addition of CAM in culture medium resulted in further pronounced GEF-induced ER stress loading, while CAM alone exhibited no effect. These data suggest that GEF-induced autophagy functions as cytoprotective and indicates the potential therapeutic possibility of using CAM for GEF therapy. Furthermore, it is suggested that the intracellular signaling for autophagy initiation in response to GEF can be completely dissociated from EGFR, but unknown target molecule(s) of GEF for autophagy induction might exist. PMID:25858318

  17. Integrated genomic approaches identify upregulation of SCRN1 as a novel mechanism associated with acquired resistance to erlotinib in PC9 cells harboring oncogenic EGFR mutation

    Science.gov (United States)

    Kim, Nayoung; Cho, Ahye; Watanabe, Hideo; Choi, Yoon-La; Aziz, Meraj; Kassner, Michelle; Joung, Je-Gun; Park, Angela KJ; Francis, Joshua M.; Bae, Joon Seol; Ahn, Soo-min; Kim, Kyoung-Mee; Park, Joon Oh; Park, Woong-Yang; Ahn, Myung-Ju; Park, Keunchil; Koo, Jaehyung; Yin, Hongwei Holly; Cho, Jeonghee

    2016-01-01

    Therapies targeting the tyrosine kinase activity of Epidermal Growth Factor Receptor (EGFR) have been proven to be effective in treating a subset of non-small cell lung cancer (NSCLC) patients harboring activating EGFR mutations. Inevitably these patients develop resistance to the EGFR-targeted tyrosine kinase inhibitors (TKIs). Here, we performed integrated genomic analyses using an in vitro system to uncover alternative genomic mechanisms responsible for acquired resistance to EGFR-TKIs. Specifically, we identified 80 genes whose expression is significantly increased in the erlotinib-resistant clones. RNAi-based systematic synthetic lethal screening of these candidate genes revealed that suppression of one upregulated transcript, SCRN1, a secernin family member, restores sensitivity to erlotinib by enhancing inhibition of PI3K/AKT signaling pathway. Furthermore, immunohistochemical analysis revealed increased levels of SCRN1 in 5 of 11 lung tumor specimens from EGFR-TKIs resistant patients. Taken together, we propose that upregulation of SCRN1 is an additional mechanism associated with acquired resistance to EGFR-TKIs and that its suppression serves as a novel therapeutic strategy to overcome drug resistance in these patients. PMID:26883194

  18. Determination of rhenium in molybdenite by neutron-activation analysis.

    Science.gov (United States)

    Terada, K; Yoshimura, Y; Osaki, S; Kiba, T

    1967-01-01

    A neutron-activation method is described for the determination of rhenium in molybdenite. Radiochemical separation by a carrier technique was carried out very rapidly by means of successive liquid-liquid extraction processes. The recovery of rhenium, which was determined by a spectrophotometric method, was about 93%. About 10 samples could be analysed within 6 hr in parallel runs. PMID:18960067

  19. Gold determination in massive samples by neutron activation analysis

    CERN Document Server

    Broglio, Eduardo

    2015-01-01

    This work presents the characteristics of the Neutron Activation Analysis (NAA) in the determination of gold in voluminous mineral samples using a linear particle accelerator (Linac) as source of irradiation. The used volumes are one or two orders of magnitude higher than the ones that can be used in nuclear research reactor, which is the source usually used to determine through the NAA. This study was encouraged by a specific legal requirement to determine the gold law in a small Patagonian gold deposit.

  20. Rapid determination of antifungal activity by flow cytometry.

    OpenAIRE

    Green, L.; Petersen, B.; Steimel, L; Haeber, P; Current, W

    1994-01-01

    We have developed a rapid assay of antifungal activity which utilizes flow cytometry to detect accumulation of a vital dye in drug-damaged fungal cells. Results of these studies suggest that flow cytometry may provide an improved, rapid method for determining and comparing the antifungal activities of compounds with differing modes of action.

  1. Determination of activated plasma fibronectin using radioactive labelled collagen I

    DEFF Research Database (Denmark)

    Fenger, M

    1984-01-01

    The plasma concentration of biological active fibronectin was assayed by a protein binding assay using 125I-collagen I as ligand and heparin as activator. The standard curve is linear for a fibronectin range of 1.1-11 pmol (0.5-5.0 micrograms) and the coefficient of variation was less than 10......%. The active or activable fibronectin was compared to the immunoreactive fibronectin in plasma from patients with various bacterial diseases. Similar concentrations were detected by the two assays suggesting that all the circulating fibronectin was functionally active. The assay was also applied to determine...

  2. Effects of the EGFR Inhibitor Erlotinib on Magnesium Handling

    DEFF Research Database (Denmark)

    Dimke, Henrik Anthony; van der Wijst, Jenny; Alexander, Todd R;

    2010-01-01

    A mutation in pro-EGF causes isolated hypomagnesemia, and monoclonal antibodies targeting the extracellular domain of the EGF receptor (EGFR) affect epithelial Mg(2+) transport. The effect of the EGFR tyrosine kinase inhibitor erlotinib on Mg(2+) homeostasis, however, remains unknown. Here, we...... in renal expression of transient receptor potential melastatin 6 (TRPM6) protein, the channel that mediates Mg(2+) reabsorption. Patch clamp analysis in TRPM6-expressing cells demonstrated that 30 muM erlotinib inhibited EGF-induced changes in TRPM6 current density and tyrosine phosphorylation of EGFR; 0...... that observed with antibody-based EGFR inhibitors. These data suggest that typical human dosages of erlotinib are unlikely to severely affect serum Mg(2+) concentrations....

  3. Effects of EGFR Gene Polymorphisms on Efficacy and Prognosis 
in Advanced Non-small Cell Lung Cancer Treated with EGFR-TKIs

    Directory of Open Access Journals (Sweden)

    Liangshan DA

    2013-03-01

    Full Text Available An increasing number of patients with advanced non-small cell lung cancer (NSCLC have been treated with epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs. However, significant differences in response to EGFR-TKIs have been shown among advanced NSCLC patients. Recently, selection of patients was mainly based on EGFR gene mutation detection. Nevertheless, mutation detection is often limited by tumour tissues derivation, technique complexity, high cost, and so on. It is urgent to seek other biological markers to predict efficacy of EGFR-TKIs. Many studies have founded that the EGFR gene polymorphisms are also associated with clinical outcome and prognosis in treatment of advanced NSCLC with EGFR-TKIs. Here, we presented a review discussing the correlation between EGFR gene polymorphisms and the efficacy of EGFR-TKIs in advanced NSCLC.

  4. Physical activity as an important determinant in developing childhood obesity

    OpenAIRE

    Bukara-Radujković Gordana; Zdravković Dragan

    2009-01-01

    The correlation between physical activity and sedentary life style was investigated as a determinant of the body mass index in children and adolescents in Banjaluka region. The study involved 1204 children and adolescents, 6-17 years old, 578 boys, 626 girls. BMI was calculated from their height and weight using standard formula. Each child, together with their parents answered the questions considering their level of involvement in physical versus sedentary activities. Physical activity was ...

  5. Determination of the antioxidant activity of Ficus carica aqueous extract

    OpenAIRE

    Trifunski Svetlana I.; Ardelean Dorina G.

    2012-01-01

    The aim of this study was to examine the antioxidant activity of water extracts from fig leaf. Water extracts were prepared according to traditional medicine. The antioxidant activity of the extracts was spectrophotometrically determined. Using the potassium permanganate colorimetric method it was found that the water extract that was maintained at the refrigerator had lower antioxidant activity than extract that was maintained at the room temperature.

  6. Determination of the antioxidant activity of Ficus carica aqueous extract

    Directory of Open Access Journals (Sweden)

    Trifunski Svetlana I.

    2012-01-01

    Full Text Available The aim of this study was to examine the antioxidant activity of water extracts from fig leaf. Water extracts were prepared according to traditional medicine. The antioxidant activity of the extracts was spectrophotometrically determined. Using the potassium permanganate colorimetric method it was found that the water extract that was maintained at the refrigerator had lower antioxidant activity than extract that was maintained at the room temperature.

  7. A drosophila model for EGFR-Ras and PI3K-dependent human glioma.

    Directory of Open Access Journals (Sweden)

    Renee D Read

    2009-02-01

    Full Text Available Gliomas, the most common malignant tumors of the nervous system, frequently harbor mutations that activate the epidermal growth factor receptor (EGFR and phosphatidylinositol-3 kinase (PI3K signaling pathways. To investigate the genetic basis of this disease, we developed a glioma model in Drosophila. We found that constitutive coactivation of EGFR-Ras and PI3K pathways in Drosophila glia and glial precursors gives rise to neoplastic, invasive glial cells that create transplantable tumor-like growths, mimicking human glioma. Our model represents a robust organotypic and cell-type-specific Drosophila cancer model in which malignant cells are created by mutations in signature genes and pathways thought to be driving forces in a homologous human cancer. Genetic analyses demonstrated that EGFR and PI3K initiate malignant neoplastic transformation via a combinatorial genetic network composed primarily of other pathways commonly mutated or activated in human glioma, including the Tor, Myc, G1 Cyclins-Cdks, and Rb-E2F pathways. This network acts synergistically to coordinately stimulate cell cycle entry and progression, protein translation, and inappropriate cellular growth and migration. In particular, we found that the fly orthologs of CyclinE, Cdc25, and Myc are key rate-limiting genes required for glial neoplasia. Moreover, orthologs of Sin1, Rictor, and Cdk4 are genes required only for abnormal neoplastic glial proliferation but not for glial development. These and other genes within this network may represent important therapeutic targets in human glioma.

  8. Development of a new biosensor for determination of catalase activity.

    Science.gov (United States)

    Teke, Mustafa

    2014-01-01

    Catalase is one of the major antioxidant enzymes that catalyzes the hydrolysis of H2O2. The aim of this study was to suggest a new method for the assay of catalase activity. For this purpose, an amperometric biosensor based on glucose oxidase for determination of catalase activity was developed. Immobilization of glucose oxidase was made by a cross-linking method with glutaraldehyde on a Clark-type electrode (dissolved oxygen probe). Optimization and characterization properties of the biosensor were studied and determination of catalase activity in defined conditions was investigated in artificial serum solution. The results were compared with a reference method. PMID:24499365

  9. Clinical pharmacogenomic testing of KRAS, BRAF and EGFR mutations by high resolution melting analysis and ultra-deep pyrosequencing

    Directory of Open Access Journals (Sweden)

    Agúndez José AG

    2011-09-01

    Full Text Available Abstract Background Epidermal growth factor receptor (EGFR and its downstream factors KRAS and BRAF are mutated in several types of cancer, affecting the clinical response to EGFR inhibitors. Mutations in the EGFR kinase domain predict sensitivity to the tyrosine kinase inhibitors gefitinib and erlotinib in lung adenocarcinoma, while activating point mutations in KRAS and BRAF confer resistance to the anti-EGFR monoclonal antibody cetuximab in colorectal cancer. The development of new generation methods for systematic mutation screening of these genes will allow more appropriate therapeutic choices. Methods We describe a high resolution melting (HRM assay for mutation detection in EGFR exons 19-21, KRAS codon 12/13 and BRAF V600 using formalin-fixed paraffin-embedded samples. Somatic variation of KRAS exon 2 was also analysed by massively parallel pyrosequencing of amplicons with the GS Junior 454 platform. Results We tested 120 routine diagnostic specimens from patients with colorectal or lung cancer. Mutations in KRAS, BRAF and EGFR were observed in 41.9%, 13.0% and 11.1% of the overall samples, respectively, being mutually exclusive. For KRAS, six types of substitutions were detected (17 G12D, 9 G13D, 7 G12C, 2 G12A, 2 G12V, 2 G12S, while V600E accounted for all the BRAF activating mutations. Regarding EGFR, two cases showed exon 19 deletions (delE746-A750 and delE746-T751insA and another two substitutions in exon 21 (one showed L858R with the resistance mutation T590M in exon 20, and the other had P848L mutation. Consistent with earlier reports, our results show that KRAS and BRAF mutation frequencies in colorectal cancer were 44.3% and 13.0%, respectively, while EGFR mutations were detected in 11.1% of the lung cancer specimens. Ultra-deep amplicon pyrosequencing successfully validated the HRM results and allowed detection and quantitation of KRAS somatic mutations. Conclusions HRM is a rapid and sensitive method for moderate

  10. Erlotinib Versus Radiation Therapy for Brain Metastases in Patients With EGFR-Mutant Lung Adenocarcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Gerber, Naamit K.; Yamada, Yoshiya; Rimner, Andreas [Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York (United States); Shi, Weiji [Department of Epidemiology and Biostatistics, Memorial Sloan-Kettering Cancer Center, New York, New York (United States); Riely, Gregory J. [Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York (United States); Beal, Kathryn [Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York (United States); Yu, Helena A. [Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York (United States); Chan, Timothy A. [Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York (United States); Zhang, Zhigang [Department of Epidemiology and Biostatistics, Memorial Sloan-Kettering Cancer Center, New York, New York (United States); Wu, Abraham J., E-mail: wua@mskcc.org [Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York (United States)

    2014-06-01

    Purpose/Objectives: Radiation therapy (RT) is the principal modality in the treatment of patients with brain metastases (BM). However, given the activity of EGFR tyrosine kinase inhibitors in the central nervous system, it is uncertain whether upfront brain RT is necessary for patients with EGFR-mutant lung adenocarcinoma with BM. Methods and Materials: Patients with EGFR-mutant lung adenocarcinoma and newly diagnosed BM were identified. Results: 222 patients were identified. Exclusion criteria included prior erlotinib use, presence of a de novo erlotinib resistance mutation, or incomplete data. Of the remaining 110 patients, 63 were treated with erlotinib, 32 with whole brain RT (WBRT), and 15 with stereotactic radiosurgery (SRS). The median overall survival (OS) for the whole cohort was 33 months. There was no significant difference in OS between the WBRT and erlotinib groups (median, 35 vs 26 months; P=.62), whereas patients treated with SRS had a longer OS than did those in the erlotinib group (median, 64 months; P=.004). The median time to intracranial progression was 17 months. There was a longer time to intracranial progression in patients who received WBRT than in those who received erlotinib upfront (median, 24 vs 16 months, P=.04). Patients in the erlotinib or SRS group were more likely to experience intracranial failure as a component of first failure, whereas WBRT patients were more likely to experience failure outside the brain (P=.004). Conclusions: The survival of patients with EGFR-mutant adenocarcinoma with BM is notably long, whether they receive upfront erlotinib or brain RT. We observed longer intracranial control with WBRT, even though the WBRT patients had a higher burden of intracranial disease. Despite the equivalent survival between the WBRT and erlotinib group, this study underscores the role of WBRT in producing durable intracranial control in comparison with a targeted biologic agent with known central nervous system activity.

  11. Radiolabeled Cetuximab Conjugates for EGFR Targeted Cancer Diagnostics and Therapy

    OpenAIRE

    Wiebke Sihver; Jens Pietzsch; Mechthild Krause; Michael Baumann; Jörg Steinbach; Hans-Jürgen Pietzsch

    2014-01-01

    The epidermal growth factor receptor (EGFR) has evolved over years into a main molecular target for the treatment of different cancer entities. In this regard, the anti-EGFR antibody cetuximab has been approved alone or in combination with: (a) chemotherapy for treatment of colorectal and head and neck squamous cell carcinoma and (b) with external radiotherapy for treatment of head and neck squamous cell carcinoma. The conjugation of radionuclides to cetuximab in combination with the specif...

  12. Detection and Evaluation of EGFR Mutation Status in Serum of Patients with Advanced Non-small Cell Lung Cancer Treated with EGFR-TKIs

    Directory of Open Access Journals (Sweden)

    Ling MA

    2013-06-01

    Full Text Available Background and objective Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs has shown a high response rate in the treatment of lung cancer in patients with (EGFR mutation. The aim of this study is to evaluate the relationship between EGFR mutation status in serum and predicting benefit from EGFR-TKIs therapy in patients with advanced non-small cell lung cancer (NSCLC. Methods We examined EGFR mutation status in serum of 80 patients with advanced, EGFR-TKIs given as first-line therapy NSCLC. All patients were received long-term follow-up, and the drug efficacy were observed and evaluated. Results The EGFR mutation in serum was detected in 33.8% (27/80 of NSCLC patients examined, in which exon 19 deletion mutation was present at a frequency of 44.4% (12/27 and exon 21 point mutation was 55.6% (15/27; The response rate to EGFR-TKI in patients with EGFR mutation in serum was (55.6%, 15/27, which was remarkably higher than that in EGFR wild-type patients (17.0%, 9/53, the difference was statistically significant (χ2=0.370, P<0.001; The median progression free survival (PFS of patients with EGFR mutation in serum was remarkably better than that of EGFR wild-type patients (9.8 months vs 5.7 months, P=0.014. Conclusion In patients with advanced, EGFR-positive in serum NSCLC, EGFR-TKIs given as first-line therapy is associated with improved drug efficacy. The results suggest that it is feasible to use serum to detect EGFR mutation, which can predict a benefit from EGFR-TKIs given as first-line therapy.

  13. Neural cell adhesion molecule-180-mediated homophilic binding induces epidermal growth factor receptor (EGFR) down-regulation and uncouples the inhibitory function of EGFR in neurite outgrowth

    DEFF Research Database (Denmark)

    Povlsen, Gro Klitgaard; Berezin, Vladimir; Bock, Elisabeth

    2008-01-01

    The neural cell adhesion molecule (NCAM) plays important roles in neuronal development, regeneration, and synaptic plasticity. NCAM homophilic binding mediates cell adhesion and induces intracellular signals, in which the fibroblast growth factor receptor plays a prominent role. Recent studies on...... this NCAM-180-induced EGFR down-regulation involves increased EGFR ubiquitination and lysosomal EGFR degradation. Furthermore, NCAM-180-mediated EGFR down-regulation requires NCAM homophilic binding and interactions of the cytoplasmic domain of NCAM-180 with intracellular interaction partners, but does...

  14. Role of [18F]FDG PET in prediction of KRAS and EGFR mutation status in patients with advanced non-small-cell lung cancer

    International Nuclear Information System (INIS)

    The tumour molecular profile predicts the activity of epidermal growth factor receptor (EGFR) inhibitors in non-small-cell lung cancer (NSCLC). However, tissue availability and tumour heterogeneity limit its assessment. We evaluated whether [18F]FDG PET might help predict KRAS and EFGR mutation status in NSCLC. Between January 2005 and October 2011, 340 NSCLC patients were tested for KRAS and EGFR mutation status. We identified patients with stage III and IV disease who had undergone [18F]FDG PET/CT scanning for initial staging. SUVpeak, SUVmax and SUVmean of the single hottest tumour lesions were calculated, and their association with KRAS and EGFR mutation status was assessed. A receiver operator characteristic (ROC) curve analysis and a multivariate analysis (including SUVmean, gender, age and AJCC stage) were performed to identify the potential value of [18F]FDG PET/CT for predicting KRAS mutation. From 102 patients staged using [18F]FDG PET/CT, 28 (27 %) had KRAS mutation (KRAS+), 22 (22 %) had EGFR mutation (EGFR+) and 52 (51 %) had wild-type KRAS and EGFR profiles (WT). KRAS+ patients showed significantly higher [18F]FDG uptake than EGFR+ and WT patients (SUVmean 9.5, 5.7 and 6.6, respectively; p 18F]FDG uptake between EGFR+ patients and WT patients. ROC curve analysis for KRAS mutation status discrimination yielded an area under the curve of 0.740 for SUVmean (p 18F]FDG uptake than WT patients, as assessed in terms of SUVpeak, SUVmax and SUVmean. A multivariate model based on age, gender, AJCC stage and SUVmean might be used as a predictive marker of KRAS mutation status in patients with stage III or IV NSCLC. (orig.)

  15. EGFR signaling in colorectal cancer: a clinical perspective

    Directory of Open Access Journals (Sweden)

    Saletti P

    2015-01-01

    Full Text Available Piercarlo Saletti,1 Francesca Molinari,2 Sara De Dosso,1 Milo Frattini2 1Oncology Institute of Southern Switzerland, Bellinzona, 2Laboratory of Molecular Pathology, Institute of Pathology, Locarno, Switzerland Abstract: Colorectal cancer (CRC remains a formidable health burden worldwide, with up to 50% of patients developing metastases during the course of their disease. This group of CRC patients, characterized by the worst prognosis, has been extensively investigated to improve their life expectancy. Main efforts, focused on the epidermal growth-factor receptor (EGFR, which plays a pivotal role in CRC pathogenesis, have led to the development and introduction in clinical practice of specific targeted therapies (ie, monoclonal antibodies. Subsequently, the scientific community has tried to identify molecular predictors of the efficacy of such therapies. However, it has become clear that EGFR alterations occurring in CRC are difficult to investigate, and therefore their predictive role is unclear. In contrast, the clinical role of two downstream members (KRAS and NRAS has been clearly demonstrated. Currently, EGFR-targeted therapies can be administered only to patients with wild-type KRAS and NRAS genes. Our review addresses the medical management of metastatic CRC. Specifically, we describe in detail the molecular biology of metastatic CRC, focusing on the EGFR signaling pathway, and we discuss the role of current and emerging related biomarkers and therapies in this field. We also summarize the clinical evidence regarding anti-EGFR monoclonal antibodies and examine potential future perspectives. Keywords: colorectal cancer, EGFR, gene mutations, cetuximab, panitumumab

  16. THE EFFECT OF ANTISENSE EPIDERMAL GROWTH FACTOR RECEPTOR (EGFR) RNA ON THE PROLIFERATION OF HUMAN GLIOMA CELLS AND INDUCTION OF CELL APOPTOSIS

    Institute of Scientific and Technical Information of China (English)

    PU Pei-yu; LIU Xu-wen; LIU Ai-xue; WANG Chun-yan; WANG Guang-xiu

    1999-01-01

    Objective: To study the effect of antisense EGFR RNA on the growth of human glioma cells in vitro and evaluate the feasibility of targeting EGFR gene for gene therapy of gliomas. Methods: Southern and Northern blot analysis,in situ hybridization and immunohistochemical staining were used to detect the integration and expression of antisense EGFR constructs. MTT assay and the average number of AgNOR for evaluation of cell proliferation, and the TUNEL method and ultrastructural change for observation of cell apoptosis. Results: Exogenous antisense EGFR cDNA was integrated into the genome of glioma cells and highly expressed, which resulted in a dramatic decrease of endogenous EGFR mRNA and GEPR protein levels.Clones with high expression of the antisense construct showed a lower proliferation activity and the induction of apoptosis in vitro. Conclusion: This study suggests that EGFR plays an important role in the genesis of gliomas; it may be used as a target for antisense gene therapy of gliomas.

  17. Digital PCR analysis of plasma cell-free DNA for non-invasive detection of drug resistance mechanisms in EGFR mutant NSCLC: Correlation with paired tumor samples

    Science.gov (United States)

    Ishii, Hidenobu; Azuma, Koichi; Sakai, Kazuko; Kawahara, Akihiko; Yamada, Kazuhiko; Tokito, Takaaki; Okamoto, Isamu; Nishio, Kazuto; Hoshino, Tomoaki

    2015-01-01

    As the development of resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) has become an issue of concern, identification of the mechanisms responsible has become an urgent priority. However, for research purposes, it is not easy to obtain tumor samples from patients with EGFR mutation-positive non-small-cell lung cancer (NSCLC) that has relapsed after treatment with EGFR-TKIs. Here, using digital PCR assay as an alternative and noninvasive method, we examined plasma and tumor samples from patients with relapsed NSCLC to establish the inter-relationships existing among T790M mutation, activating EGFR mutations, HER2 amplification, and MET amplification. Paired samples of tumor and blood were obtained from a total of 18 patients with NSCLC after they had developed resistance to EGFR-TKI treatment, and the mechanisms of resistance were analyzed by digital PCR. Digital PCR analysis of T790M mutation in plasma had a sensitivity of 81.8% and specificity of 85.7%, the overall concordance between plasma and tissue samples being 83.3%. MET gene copy number gain in tumor DNA was observed by digital PCR in three patients, of whom one exhibited positivity for MET amplification by FISH, whereas no patient demonstrated MET and HER2 copy number gain in plasma DNA. Digital PCR analysis of plasma is feasible and accurate for detection of T790M mutation in NSCLC that becomes resistant to treatment with EGFR-TKIs. PMID:26334838

  18. Antitumor and antiangiogenic effect of the dual EGFR and HER-2 tyrosine kinase inhibitor lapatinib in a lung cancer model

    Directory of Open Access Journals (Sweden)

    Collantes Maria

    2010-05-01

    Full Text Available Abstract Background There is strong evidence demonstrating that activation of epidermal growth factor receptors (EGFRs leads to tumor growth, progression, invasion and metastasis. Erlotinib and gefitinib, two EGFR-targeted agents, have been shown to be relevant drugs for lung cancer treatment. Recent studies demonstrate that lapatinib, a dual tyrosine kinase inhibitor of EGFR and HER-2 receptors, is clinically effective against HER-2-overexpressing metastatic breast cancer. In this report, we investigated the activity of lapatinib against non-small cell lung cancer (NSCLC. Methods We selected the lung cancer cell line A549, which harbors genomic amplification of EGFR and HER-2. Proliferation, cell cycle analysis, clonogenic assays, and signaling cascade analyses (by western blot were performed in vitro. In vivo experiments with A549 cells xenotransplanted into nude mice treated with lapatinib (with or without radiotherapy were also carried out. Results Lapatinib dramatically reduced cell proliferation (P P P in vitro. Furthermore, lapatinib induced G1 cell cycle arrest (P P In vivo experiments revealed that A549 tumor-bearing mice treated with lapatinib had significantly less active tumors (as assessed by PET analysis (P P Conclusion Overall, these data suggest that lapatinib may be a clinically useful agent for the treatment of lung cancer.

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

    International Nuclear Information System (INIS)

    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

  20. Cardiotoxin III suppresses MDA-MB-231 cell metastasis through the inhibition of EGF/EGFR-mediated signaling pathway.

    Science.gov (United States)

    Tsai, Pei-Chien; Hsieh, Chi-Ying; Chiu, Chien-Chih; Wang, Chih-Kuang; Chang, Long-Sen; Lin, Shinne-Ren

    2012-10-01

    Cardiotoxin III (CTX III), a basic polypeptide isolated from Naja naja atra venom, has been shown to exhibit anticancer activity. Epidermal growth factor (EGF) and its receptor, EGFR, play roles in cancer metastasis in various tumors. We use EGF as a metastatic inducer of MDA-MB-231 cells to investigate the effect of CTX III on cell migration. CTX III inhibited the EGF-induced activation of matrix metalloproteinase-9 (MMP-9), and further suppressed cell invasion and migration without obvious cellular cytotoxicity. CTX III suppressed EGF-induced nuclear factor-kappaB (NF-κB) nuclear translocation and also abrogated the EGF-induced phosphorylation of EGFR, phosphatidylinositol 3-kinase (PI3K)/Akt, and extracellular regulated kinase (ERK)1/2. In addition, CTX III similar to wortmannin (a PI3K inhibitor) and U0126 (an up-stream kinase regulating ERK1/2 inhibitor) attenuated cell migration and invasion induced by EGF. Furthermore, the EGFR inhibitor AG1478 inhibited EGF-induced MMP-9 expression, cell migration and invasion, as well as the activation of ERK1/2 and PI3K/Akt, suggesting that ERK1/2 and PI3K/Akt activation occur downstream of EGFR activation. These findings suggest that CTX III inhibited the EGF-induced invasion and migration of MDA-MB-231 cells via EGFR-dependent PI3K/Akt, ERK1/2, and NF-κB signaling, leading to the down-regulation of MMP-9 expression. These results provide a novel mechanism to explain the role of CTX III as a potent anti-metastatic agent in MDA-MB-231 cells.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  2. Regulation of EGFR signal transduction by analogue-to-digital conversion in endosomes

    Science.gov (United States)

    Villaseñor, Roberto; Nonaka, Hidenori; Del Conte-Zerial, Perla; Kalaidzidis, Yannis; Zerial, Marino

    2015-01-01

    An outstanding question is how receptor tyrosine kinases (RTKs) determine different cell-fate decisions despite sharing the same signalling cascades. Here, we uncovered an unexpected mechanism of RTK trafficking in this process. By quantitative high-resolution FRET microscopy, we found that phosphorylated epidermal growth factor receptor (p-EGFR) is not randomly distributed but packaged at constant mean amounts in endosomes. Cells respond to higher EGF concentrations by increasing the number of endosomes but keeping the mean p-EGFR content per endosome almost constant. By mathematical modelling, we found that this mechanism confers both robustness and regulation to signalling output. Different growth factors caused specific changes in endosome number and size in various cell systems and changing the distribution of p-EGFR between endosomes was sufficient to reprogram cell-fate decision upon EGF stimulation. We propose that the packaging of p-RTKs in endosomes is a general mechanism to ensure the fidelity and specificity of the signalling response. DOI: http://dx.doi.org/10.7554/eLife.06156.001 PMID:25650738

  3. Dynamic Bayesian Network Modeling of the Interplay between EGFR and Hedgehog Signaling.

    Science.gov (United States)

    Fröhlich, Holger; Bahamondez, Gloria; Götschel, Frank; Korf, Ulrike

    2015-01-01

    Aberrant activation of sonic Hegdehog (SHH) signaling has been found to disrupt cellular differentiation in many human cancers and to increase proliferation. The SHH pathway is known to cross-talk with EGFR dependent signaling. Recent studies experimentally addressed this interplay in Daoy cells, which are presumable a model system for medulloblastoma, a highly malignant brain tumor that predominately occurs in children. Currently ongoing are several clinical trials for different solid cancers, which are designed to validate the clinical benefits of targeting the SHH in combination with other pathways. This has motivated us to investigate interactions between EGFR and SHH dependent signaling in greater depth. To our knowledge, there is no mathematical model describing the interplay between EGFR and SHH dependent signaling in medulloblastoma so far. Here we come up with a fully probabilistic approach using Dynamic Bayesian Networks (DBNs). To build our model, we made use of literature based knowledge describing SHH and EGFR signaling and integrated gene expression (Illumina) and cellular location dependent time series protein expression data (Reverse Phase Protein Arrays). We validated our model by sub-sampling training data and making Bayesian predictions on the left out test data. Our predictions focusing on key transcription factors and p70S6K, showed a high level of concordance with experimental data. Furthermore, the stability of our model was tested by a parametric bootstrap approach. Stable network features were in agreement with published data. Altogether we believe that our model improved our understanding of the interplay between two highly oncogenic signaling pathways in Daoy cells. This may open new perspectives for the future therapy of Hedghog/EGF-dependent solid tumors. PMID:26571415

  4. Dynamic Bayesian Network Modeling of the Interplay between EGFR and Hedgehog Signaling.

    Directory of Open Access Journals (Sweden)

    Holger Fröhlich

    Full Text Available Aberrant activation of sonic Hegdehog (SHH signaling has been found to disrupt cellular differentiation in many human cancers and to increase proliferation. The SHH pathway is known to cross-talk with EGFR dependent signaling. Recent studies experimentally addressed this interplay in Daoy cells, which are presumable a model system for medulloblastoma, a highly malignant brain tumor that predominately occurs in children. Currently ongoing are several clinical trials for different solid cancers, which are designed to validate the clinical benefits of targeting the SHH in combination with other pathways. This has motivated us to investigate interactions between EGFR and SHH dependent signaling in greater depth. To our knowledge, there is no mathematical model describing the interplay between EGFR and SHH dependent signaling in medulloblastoma so far. Here we come up with a fully probabilistic approach using Dynamic Bayesian Networks (DBNs. To build our model, we made use of literature based knowledge describing SHH and EGFR signaling and integrated gene expression (Illumina and cellular location dependent time series protein expression data (Reverse Phase Protein Arrays. We validated our model by sub-sampling training data and making Bayesian predictions on the left out test data. Our predictions focusing on key transcription factors and p70S6K, showed a high level of concordance with experimental data. Furthermore, the stability of our model was tested by a parametric bootstrap approach. Stable network features were in agreement with published data. Altogether we believe that our model improved our understanding of the interplay between two highly oncogenic signaling pathways in Daoy cells. This may open new perspectives for the future therapy of Hedghog/EGF-dependent solid tumors.

  5. Lupeol evokes anticancer effects in oral squamous cell carcinoma by inhibiting oncogenic EGFR pathway.

    Science.gov (United States)

    Rauth, Sanchita; Ray, Sudipta; Bhattacharyya, Sayantan; Mehrotra, Debapriya Ghosh; Alam, Neyaz; Mondal, Goutam; Nath, Partha; Roy, Asoke; Biswas, Jaydip; Murmu, Nabendu

    2016-06-01

    Epidermal growth factor receptor (EGFR) pathway is overexpressed in head and neck cancer (HNC). Lupeol, a natural triterpene (phytosterol found in fruits, vegetables, etc.), has been reported to be effective against multiple cancer indications. Here we investigate the antitumor effects of Lupeol and underlying mechanism in oral cancer. Lupeol-induced antitumor response was evaluated in two oral squamous cell carcinoma (OSCC) cell lines (UPCI:SCC131 and UPCI:SCC084) by viability (MTT), proliferation, and colony formation assays. Lupeol-mediated induction of apoptosis was examined by caspase 3/7 assay and flow cytometry. Effect of Lupeol on EGFR in the presence or absence of EGF was delineated by Western blot. The mRNA stability assay was performed to check the role of Lupeol on COX-2 mRNA regulation. Lupeol inhibited proliferation of OSCC cells in vitro by inducing apoptosis 48 h post treatment. Ligand-induced phosphorylation of EGFR and subsequent activation of its downstream molecules such as protein kinase B (PKB or AKT), I kappa B (IκB), and nuclear factor kappa B (NF-κB) was also found to be, in part, suppressed. Interestingly, Lupeol suppressed expression of COX-2 at mRNA and protein level in a time-dependent manner. Primary explants from oral squamous cell carcinoma tissues further confirmed significant inhibition of proliferation (Ki67) in Lupeol-treated explants as compared to untreated control at 48 h. Together these data suggest that Lupeol may act as a potent inhibitor of the EGFR signaling in OSCC and therefore imply its role in triggering antitumor efficacy. PMID:27206736

  6. Molecular docking, MM/GBSA and 3D-QSAR studies on EGFR inhibitors

    Indian Academy of Sciences (India)

    RAJU BATHINI; SREE KANTH SIVAN; SABIHA FATIMA; VIJJULATHA MANGA

    2016-07-01

    Epidermal growth factor receptor (EGFR) is the first growth factor receptor proposed as a target for cancer therapy. Molecular modeling protocols like molecular docking, molecular mechanics/generalized born surface area (MM/GBSA) calculations and three dimensional-quantitative structure activity relationship(3D-QSAR) studies were performed on 45 molecules to understand the structural requirements for EGFR tyrosine kinase inhibitors. Conformation for all the molecules obtained from molecular docking were used as is for 3D-QSAR analysis. Comparative molecular field analysis (CoMFA) and comparative molecular similarityindices analysis (CoMSIA) models were obtained by performing partial least square analysis on 35 training molecules and these models were validated using 10 test moleucles. The models showed good statistical results in terms of r², q² loo and r² pred values. Information rendered from 3D-QSAR model and sitemap analysis was used to optimize lead molecule to design prospective inhibitors. Improvement in EGFR binding affinity can be achieved by substitutional modification on phenyl ring attached to alkynyl group with bulkier hydrogen bond donor and acceptor substituents that can increase favourable interaction with the receptor.

  7. Colon-specific delivery of a probiotic-derived soluble protein ameliorates intestinal inflammation in mice through an EGFR-dependent mechanism.

    Science.gov (United States)

    Yan, Fang; Cao, Hanwei; Cover, Timothy L; Washington, M Kay; Shi, Yan; Liu, LinShu; Chaturvedi, Rupesh; Peek, Richard M; Wilson, Keith T; Polk, D Brent

    2011-06-01

    Probiotic bacteria can potentially have beneficial effects on the clinical course of several intestinal disorders, but our understanding of probiotic action is limited. We have identified a probiotic bacteria-derived soluble protein, p40, from Lactobacillus rhamnosus GG (LGG), which prevents cytokine-induced apoptosis in intestinal epithelial cells. In the current study, we analyzed the mechanisms by which p40 regulates cellular responses in intestinal epithelial cells and p40's effects on experimental colitis using mouse models. We show that the recombinant p40 protein activated EGFR, leading to Akt activation. Activation of EGFR by p40 was required for inhibition of cytokine-induced apoptosis in intestinal epithelial cells in vitro and ex vivo. Furthermore, we developed a pectin/zein hydrogel bead system to specifically deliver p40 to the mouse colon, which activated EGFR in colon epithelial cells. Administration of p40-containing beads reduced intestinal epithelial apoptosis and disruption of barrier function in the colon epithelium in an EGFR-dependent manner, thereby preventing and treating DSS-induced intestinal injury and acute colitis. Furthermore, p40 activation of EGFR was required for ameliorating colon epithelial cell apoptosis and chronic inflammation in oxazolone-induced colitis. These data define what we believe to be a previously unrecognized mechanism of probiotic-derived soluble proteins in protecting the intestine from injury and inflammation.

  8. Ascochlorin suppresses TGF-β1-induced PAI-1 expression through the inhibition of phospho-EGFR in rat kidney fibroblast cells.

    Science.gov (United States)

    Cho, Hyun-Ji; Kang, Jeong-Han; Jeong, Ji-Hak; Jeong, Yun-Jeong; Park, Kwan-Kyu; Park, Yoon-Yub; Moon, Yong-Suk; Kim, Hong-Tae; Chung, Il-Kyung; Kim, Cheorl-Ho; Chang, Hyeun-Wook; Chang, Young-Chae

    2012-04-01

    Fibrosis is induced by the excessive and abnormal deposition of extracellular matrix (ECM) with various growth factors in tissues. Transforming growth factor-β1 (TGF-β1), the growth factor involved in fibrosis, modulates ECM synthesis and accumulation. TGF-β1 enhances the production of stimulators of ECM synthesis such as plasminogen activator inhibitor type 1 (PAI-1). As such, PAI-1 expression directly influences the proteolysis, invasion, and accumulation of ECM. It was shown in this study that ascochlorin, a prenylpenl antiobiotic, prevents the expression of profibrotic factors, such as PAI-1 and collagen type I, and that the TGF-β1-induced PAI-1 promoter activity is inhibited by ascochlorin. Ascochlorin abolishes the phosphorylation of the EGFR-MEK-ERK signaling pathway to regulate the TGF-β1-induced expression of PAI-1 without the inhibition of TβRII phosphorylation. Furthermore, the MEK inhibitor and EGFR siRNA block PAI-1 expression, and the Raf-1, MEK, and ERK signaling pathways for the regulation of PAI-1 expression. Ascochlorin suppresses the matrix metalloproteinases (MMPs) activity to activate the heparin-binding EGF-like growth factor (HB-EGF), to induce the phosphorylation of EGFR, and the MMPs inhibitor suppresses EGFR phosphorylation and the PAI-1 mRNA levels. These results suggest that ascochlorin prevents the expression of PAI-1 via the inhibition of an EGFR-dependent signal transduction pathway activated by MMPs.

  9. Direct potentiometric determination of diastase activity in honey.

    Science.gov (United States)

    Sak-Bosnar, Milan; Sakač, Nikola

    2012-11-15

    A novel method for the determination of diastase activity is reported. The method is based on a direct potentiometric measurement of triiodide ion that is released when a starch-triiodide complex is hydrolysed by honey diastase. The increase of free triiodide ion concentration in a sample is found to be directly proportional to the diastase activity of the sample. A response mechanism of the platinum redox electrode is proposed, allowing a calculation of the diastase activity factor (F). The sensor and analyte parameters, including F, were obtained by least squares fitting of potentiometric data using the optimisation function of the Solver add-in of Microsoft Excel. The values of F obtained by the new direct potentiometric method were compared with those obtained using the standard Phadebas method (DN values), and the two values were found to agree within experimental error. Finally, the diastase activity of nine varieties of honey was determined using the novel method developed here. PMID:22868165

  10. DETERMINANTS OF LEISURE-TIME PHYSICAL ACTIVITY: EVIDENCE FROM MALAYSIA

    OpenAIRE

    YONG KANG CHEAH; Tan, Andrew K. G.

    2014-01-01

    This paper examines how socio-demographic and health-lifestyle factors determine participation and duration of leisure-time physical activity in Malaysia. Based on the Malaysia Non-Communicable Disease Surveillance-1 data, Heckman's sample selection model is employed to estimate the probability to participate and duration on physical activity. Results indicate that gender, age, years of education and family illness history are significant in explaining participation probability in leisure-tim...

  11. EGF stimulates the activation of EGF receptors and the selective activation of major signaling pathways during mitosis.

    Science.gov (United States)

    Wee, Ping; Shi, Huaiping; Jiang, Jennifer; Wang, Yuluan; Wang, Zhixiang

    2015-03-01

    Mitosis and epidermal growth factor (EGF) receptor (EGFR) are both targets for cancer therapy. The role of EGFR signaling in mitosis has been rarely studied and poorly understood. The limited studies indicate that the activation of EGFR and downstream signaling pathways is mostly inhibited during mitosis. However, we recently showed that EGFR is phosphorylated in response to EGF stimulation in mitosis. Here we studied EGF-induced EGFR activation and the activation of major signaling pathways downstream of EGFR during mitosis. We showed that EGFR was strongly activated by EGF during mitosis as all the five major tyrosine residues including Y992, Y1045, Y1068, Y1086, and Y1173 were phosphorylated to a level similar to that in the interphase. We further showed that the activated EGFR is able to selectively activate some downstream signaling pathways while avoiding others. Activated EGFR is able to activate PI3K and AKT2, but not AKT1, which may be responsible for the observed effects of EGF against nocodazole-induced cell death. Activated EGFR is also able to activate c-Src, c-Cbl and PLC-γ1 during mitosis. However, activated EGFR is unable to activate ERK1/2 and their downstream substrates RSK and Elk-1. While it activated Ras, EGFR failed to fully activate Raf-1 in mitosis due to the lack of phosphorylation at Y341 and the lack of dephosphorylation at pS259. We conclude that contrary to the dogma, EGFR is activated by EGF during mitosis. Moreover, EGFR-mediated cell signaling is regulated differently from the interphase to specifically serve the needs of the cell in mitosis.

  12. Differential effects of EGFR ligands on endocytic sorting of the receptor

    DEFF Research Database (Denmark)

    Roepstorff, Kirstine; Grandal, Michael Vibo; Henriksen, Lasse;

    2009-01-01

    Endocytic downregulation is a pivotal mechanism turning off signalling from the EGF receptor (EGFR). It is well established that whereas EGF binding leads to lysosomal degradation of EGFR, transforming growth factor (TGF)-alpha causes receptor recycling. TGF-alpha therefore leads to continuous...... recycling. EGF leads to lysosomal degradation of the majority but not all EGFRs. Amphiregulin does not target EGFR for lysosomal degradation but causes fast as well as slow EGFR recycling. The Cbl ubiquitin ligases, especially c-Cbl, are responsible for EGFR ubiquitination after stimulation with all ligands...

  13. Role of EGFR transactivation in angiotensin II signaling to extracellular regulated kinase in preglomerular smooth muscle cells.

    Science.gov (United States)

    Andresen, Bradley T; Linnoila, Jenny J; Jackson, Edwin K; Romero, Guillermo G

    2003-03-01

    Angiotensin (Ang) II promotes the phosphorylation of extracellular regulated kinase (ERK); however, the mechanisms leading to Ang II-induced ERK phosphorylation are debated. The currently accepted theory involves transactivation of epidermal growth factor receptor (EGFR). We have shown that generation of phosphatidic acid (PA) is required for the recruitment of Raf to membranes and the activation of ERK by multiple agonists, including Ang II. In the present report, we confirm that phospholipase D-dependent generation of PA is required for Ang II-mediated phosphorylation of ERK in Wistar-Kyoto and spontaneously hypertensive rat preglomerular smooth muscle cells (PGSMCs). However, EGF stimulation does not activate phospholipase D or generate PA. These observations indicate that EGF recruits Raf to membranes via a mechanism that does not involve PA, and thus, Ang II-mediated phosphorylation of ERK is partially independent of EGFR-mediated signaling cascades. We hypothesized that phosphoinositide-3-kinase (PI3K) can also act to recruit Raf to membranes; therefore, inhibition of PI3K should inhibit EGF signaling to ERK. Wortmannin, a PI3K inhibitor, inhibited EGF-mediated phosphorylation of ERK (IC50, approximately 14 nmol/L). To examine the role of the EGFR in Ang II-mediated phosphorylation of ERK we utilized 100 nmol/L wortmannin to inhibit EGFR signaling to ERK and T19N RhoA to block Ang II-mediated ERK phosphorylation. Wortmannin treatment inhibited EGF-mediated but not Ang II-mediated phosphorylation of ERK. Furthermore, T19N RhoA inhibited Ang II-mediated ERK phosphorylation, whereas T19N RhoA had significantly less effect on EGF-mediated ERK phosphorylation. We conclude that transactivation of the EGFR is not primarily responsible for Ang II-mediated activation of ERK in PGSMCs.

  14. Role of EGFR transactivation in angiotensin II signaling to extracellular regulated kinase in preglomerular smooth muscle cells.

    Science.gov (United States)

    Andresen, Bradley T; Linnoila, Jenny J; Jackson, Edwin K; Romero, Guillermo G

    2003-03-01

    Angiotensin (Ang) II promotes the phosphorylation of extracellular regulated kinase (ERK); however, the mechanisms leading to Ang II-induced ERK phosphorylation are debated. The currently accepted theory involves transactivation of epidermal growth factor receptor (EGFR). We have shown that generation of phosphatidic acid (PA) is required for the recruitment of Raf to membranes and the activation of ERK by multiple agonists, including Ang II. In the present report, we confirm that phospholipase D-dependent generation of PA is required for Ang II-mediated phosphorylation of ERK in Wistar-Kyoto and spontaneously hypertensive rat preglomerular smooth muscle cells (PGSMCs). However, EGF stimulation does not activate phospholipase D or generate PA. These observations indicate that EGF recruits Raf to membranes via a mechanism that does not involve PA, and thus, Ang II-mediated phosphorylation of ERK is partially independent of EGFR-mediated signaling cascades. We hypothesized that phosphoinositide-3-kinase (PI3K) can also act to recruit Raf to membranes; therefore, inhibition of PI3K should inhibit EGF signaling to ERK. Wortmannin, a PI3K inhibitor, inhibited EGF-mediated phosphorylation of ERK (IC50, approximately 14 nmol/L). To examine the role of the EGFR in Ang II-mediated phosphorylation of ERK we utilized 100 nmol/L wortmannin to inhibit EGFR signaling to ERK and T19N RhoA to block Ang II-mediated ERK phosphorylation. Wortmannin treatment inhibited EGF-mediated but not Ang II-mediated phosphorylation of ERK. Furthermore, T19N RhoA inhibited Ang II-mediated ERK phosphorylation, whereas T19N RhoA had significantly less effect on EGF-mediated ERK phosphorylation. We conclude that transactivation of the EGFR is not primarily responsible for Ang II-mediated activation of ERK in PGSMCs. PMID:12623996

  15. EGFR and KRAS quality assurance schemes in pathology: generating normative data for molecular predictive marker analysis in targeted therapy.

    NARCIS (Netherlands)

    Thunnissen, E.; Bovee, J.V.; Bruinsma, H.; Brule, A.J. van den; Dinjens, W.; Heideman, D.A.; Meulemans, E.; Nederlof, P.; Noesel, C. van; Prinsen, C.F.M.; Scheidel, K.; Ven, P.M. van de; Weger, R. de; Schuuring, E.; Ligtenberg, M.J.

    2011-01-01

    INTRODUCTION: The aim of this study was to compare the reproducibility of epidermal growth factor receptor (EGFR) immunohistochemistry (IHC), EGFR gene amplification analysis, and EGFR and KRAS mutation analysis among different laboratories performing routine diagnostic analyses in pathology in The

  16. Combined EGFR and VEGFR versus single EGFR signaling pathways inhibition therapy for NSCLC: a systematic review and meta-analysis.

    Directory of Open Access Journals (Sweden)

    Xinji Zhang

    Full Text Available BACKGROUND: Lung cancer is a heterogeneous disease with multiple signaling pathways influencing tumor cell survival and proliferation, and it is likely that blocking only one of these pathways allows others to act as salvage or escape mechanisms for cancer cells. Whether combined inhibition therapy has greater anti-tumor activity than single inhibition therapy is a matter of debate. Hence, a meta-analysis comparing therapy inhibiting both VEGFR and EGFR signaling pathways with that inhibiting EGFR signaling pathway alone was performed. METHODOLOGY AND PRINCIPAL FINDINGS: We searched PubMed, EMBASE database and the proceedings of major conferences for relevant clinical trials. Outcomes analyzed were objective tumor response rate (ORR, progression-free survival (PFS, overall survival (OS and toxicity. Besides, subgroup analyses were performed to investigate whether the combined inhibition therapy is best performed using combination of selective agents or a single agent with multiple targets. Six trials recruiting 3,302 patients were included in the analysis. Combined inhibition therapy was associated with a 3% improvement in OS as compared with single-targeted therapy, but this difference was not statistically significant (HR, 0.97; 95% CI, 0.89-1.05; P=0.472. Patients receiving combined inhibition therapy had significant longer PFS than the group with single-targeted therapy (HR, 0.80; 95% CI, 0.67-0.95; P=0.011. There was no difference in the ORR between the groups (OR, 1.44; 95% CI, 0.95-2.18; P=0.085. Subgroup analysis revealed that combined inhibition therapy using combination regimens was associated with statistically significant improvement in both ORR and PFS. Toxicity was greater in combined inhibition therapy. CONCLUSIONS: There is no evidence to support the use of combined inhibition therapy in unselected patients with advanced NSCLC. However, given the significant advantage in ORR and PFS, combined inhibition therapy using combination

  17. Ganoderma lucidum Combined with the EGFR Tyrosine Kinase Inhibitor, Erlotinib Synergize to Reduce Inflammatory Breast Cancer Progression.

    Science.gov (United States)

    Suárez-Arroyo, Ivette J; Rios-Fuller, Tiffany J; Feliz-Mosquea, Yismeilin R; Lacourt-Ventura, Mercedes; Leal-Alviarez, Daniel J; Maldonado-Martinez, Gerónimo; Cubano, Luis A; Martínez-Montemayor, Michelle M

    2016-01-01

    The high incidence of resistance to Tyrosine Kinase Inhibitors (TKIs) targeted against EGFR and downstream pathways has increased the necessity to identify agents that may be combined with these therapies to provide a sustained response for breast cancer patients. Here, we investigate the therapeutic potential of Ganoderma lucidum extract (GLE) in breast cancer, focusing on the regulation of the EGFR signaling cascade when treated with the EGFR TKI, Erlotinib. SUM-149, or intrinsic Erlotinib resistant MDA-MB-231 cells, and a successfully developed Erlotinib resistant cell line, rSUM-149 were treated with increasing concentrations of Erlotinib, GLE, or their combination (Erlotinib/GLE) for 72h. Treatment effects were tested on cell viability, cell proliferation, cell migration and invasion. To determine tumor progression, severe combined immunodeficient mice were injected with SUM-149 cells and then treated with Erlotinib/GLE or Erlotinib for 13 weeks. We assessed the protein expression of ERK1/2 and AKT in in vitro and in vivo models. Our results show that GLE synergizes with Erlotinib to sensitize SUM-149 cells to drug treatment, and overcomes intrinsic and developed Erlotinib resistance. Also, Erlotinib/GLE decreases SUM-149 cell viability, proliferation, migration and invasion. GLE increases Erlotinib sensitivity by inactivating AKT and ERK signaling pathways in our models. We conclude that a combinatorial therapeutic approach may be the best way to increase prognosis in breast cancer patients with EGFR overexpressing tumors.

  18. Ganoderma lucidum Combined with the EGFR Tyrosine Kinase Inhibitor, Erlotinib Synergize to Reduce Inflammatory Breast Cancer Progression.

    Science.gov (United States)

    Suárez-Arroyo, Ivette J; Rios-Fuller, Tiffany J; Feliz-Mosquea, Yismeilin R; Lacourt-Ventura, Mercedes; Leal-Alviarez, Daniel J; Maldonado-Martinez, Gerónimo; Cubano, Luis A; Martínez-Montemayor, Michelle M

    2016-01-01

    The high incidence of resistance to Tyrosine Kinase Inhibitors (TKIs) targeted against EGFR and downstream pathways has increased the necessity to identify agents that may be combined with these therapies to provide a sustained response for breast cancer patients. Here, we investigate the therapeutic potential of Ganoderma lucidum extract (GLE) in breast cancer, focusing on the regulation of the EGFR signaling cascade when treated with the EGFR TKI, Erlotinib. SUM-149, or intrinsic Erlotinib resistant MDA-MB-231 cells, and a successfully developed Erlotinib resistant cell line, rSUM-149 were treated with increasing concentrations of Erlotinib, GLE, or their combination (Erlotinib/GLE) for 72h. Treatment effects were tested on cell viability, cell proliferation, cell migration and invasion. To determine tumor progression, severe combined immunodeficient mice were injected with SUM-149 cells and then treated with Erlotinib/GLE or Erlotinib for 13 weeks. We assessed the protein expression of ERK1/2 and AKT in in vitro and in vivo models. Our results show that GLE synergizes with Erlotinib to sensitize SUM-149 cells to drug treatment, and overcomes intrinsic and developed Erlotinib resistance. Also, Erlotinib/GLE decreases SUM-149 cell viability, proliferation, migration and invasion. GLE increases Erlotinib sensitivity by inactivating AKT and ERK signaling pathways in our models. We conclude that a combinatorial therapeutic approach may be the best way to increase prognosis in breast cancer patients with EGFR overexpressing tumors. PMID:26958085

  19. Over-expression of EGFR in Breast Cancer

    Institute of Scientific and Technical Information of China (English)

    BO Ai-hua; HOU Jin-chao; LAN Yong-hao; TIAN Ya-ting; ZHANG Jun-yan

    2008-01-01

    Objective:To explore the relationship of overexpression of epidermal growth factor receptor(EGFR)in occurrence,development and treatment of breast cancer. Methods:Samples of 46 breast adenoma tissues and 86 breast cancer tissues were regularly dehydrate-fixed,embedded in paraffin,sliced in to 5 μm thick,stained with SABC immunohistochemistry and coloured with DAB. Results:The positive staining of EGFR was shown as brown- yellow and distributed in cytoplasm.The positive rates in the tissues of breast adenosis and breast cancer were 17.04%(6/46)and 56.98%(49/86)respectively.The positive rates of EGFR in the tissue of invasive ductal carcinoma was 64.49%(41/59),which was significantly higher than that in in situ carcinoma(P<0.05).The positive rate of lymph metastasis group was higher than that in non-lymph metastasis group (P<0.05). Conclusion:The overexpression of EGFR was related with occurrence,lymph metastasis and pathologic types of breast cancer.The examination of EGFR in the breast cancer can serve as a guidance for target chemotherapy.

  20. Development of personalized treatments in lung cancer: focusing on the EGFR mutations and beyond

    Directory of Open Access Journals (Sweden)

    Suda K

    2013-08-01

    Full Text Available Kenichi Suda,1,2 Tetsuya Mitsudomi1 1Division of Thoracic Surgery, Department of Surgery, Kinki University Faculty of Medicine, Osaka-Sayama, Japan; 2Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan Abstract: Lung cancers with epidermal growth factor receptor (EGFR gene mutation account for ~40% of adenocarcinoma in East Asians and ~15% of that in Caucasians, which makes them one of the most common molecularly defined lung cancer subsets. The role of EGFR mutation as a strong predictive biomarker of response to EGFR-tyrosine kinase inhibitors (TKIs was finally confirmed by the biomarker analysis of Iressa Pan-Asian Study (IPASS. Since the 2004 discovery of EGFR mutation in lung cancer, the EGFR mutation and EGFR-TKI treatment have been widely studied. These include characteristics of lung cancers with EGFR mutations; clinical efficacies and adverse effects of EGFR-TKIs in patients with EGFR-mutated lung cancers; development of novel EGFR-TKIs that may prolong progression-free survival of these patients or overcome resistance to first-generation EGFR-TKIs (gefitinib and erlotinib; optimal treatment schedules for EGFR-TKIs to delay emergence of resistance; molecular mechanisms of acquired resistance to EGFR-TKIs; treatment strategies after patients acquire resistance to EGFR-TKIs; and predictive biomarkers for EGFR-TKIs among patients with EGFR-mutated lung cancers. Some of these results are widely accepted, while others are apparent only in cell line models, preclinical animal models, or retrospective analyses (and sometimes conflict with each other. In this review, we summarize accumulated reports from the past decade, especially focusing on unanswered but important clinical questions in treating patients with EGFR-mutated lung cancers. Keywords: epidermal growth factor receptor mutation, predictive biomarkers, personalized therapy, molecular target, adjuvant therapy, acquired resistance

  1. 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.

  2. Physical activity as an important determinant in developing childhood obesity

    Directory of Open Access Journals (Sweden)

    Bukara-Radujković Gordana

    2009-01-01

    Full Text Available The correlation between physical activity and sedentary life style was investigated as a determinant of the body mass index in children and adolescents in Banjaluka region. The study involved 1204 children and adolescents, 6-17 years old, 578 boys, 626 girls. BMI was calculated from their height and weight using standard formula. Each child, together with their parents answered the questions considering their level of involvement in physical versus sedentary activities. Physical activity was defined as involvement in sports activities, while sedentary life style was defined as time spent on computer, games, video, and TV. The prevalence of overweight and obesity were 12.2% and 6.1% in our study group. Increased physical activity showed strong positive correlation with normal, lower BMI in boys (p<0,05, and girls (p<0.001. Sedentary lifestyle, prolonged TV watching was strongly associated with increased BMI only in girls (p<0.05. However, computer use for 2 hours/day was strongly associated with increased BMI (p<0.05 only in boys, although computer use for more than 3 hours/day was associated with lower BMI in boys. Physical activity and sedentary lifestyle are significant determinants of BMI and risk factors in developing overweight and obesity in childhood, as shown in our study.

  3. Immunotherapy of anti-CD3/anti-EGFR bispecific antibody on the mice borne human gastric cancer%抗EGFR/抗CD3双功能抗体对胃癌荷瘤小鼠的免疫治疗研究

    Institute of Scientific and Technical Information of China (English)

    张林; 侯艳红; 张健; 胡静; 张静

    2012-01-01

    目的 通过抗EGFR/抗CD3双功能抗体(EGFR/CD3 BsAb)治疗SGC7901胃癌细胞移植瘤小鼠模型,初步验证该抗体在体内对胃癌细胞的杀伤能力.方法 采取化学偶联法合成的EGFR/CD3 BsAb联合人外周血淋巴细胞(PBLS)经尾静脉给药注入荷5GC7901胃癌细胞移植瘤小鼠体内.实验裸鼠随机分为抗EGFR单抗联合PBLS组(抗EGFR组)、抗CD3单抗联合PBIS组(抗CD3组)、EGFR/CD3 BsAb联合PBLS组(EGFR/CD3 BsAb组)和空白对照组(生理盐水组).治疗后检测并比较加药各组对胃癌细胞的杀伤能力.结果 EGFR/CD3 BsAb成功制备,分子量为43kD.ECFR/CD3 BsAb组裸鼠的肿瘤抑制率为(57.2±8.6)%,显著高于抗EGFR组的(38.5±6.1)%和抗CD3组的(6.9±7.6)%(P<0.05);治疗结束时EGFR/CD3 BsAb组的瘤重为(517.1±45.4)mg,显著低于抗EGFR组的(737.4±54.3)mg和抗CD3组的(1097.9±167.7)mg(P<0.05).各组移植瘤组织EGFR免疫组化染色均为强阳性.EGFR/CD3 BsAb组裸鼠肿瘤组织CD3免疫组化染色可见部分细胞呈阳性.透射电镜观察显示,EGFR/CD3 BsAb组和抗EGFR组可见肿瘤坏死.结论 EGFR/CD3 BsAb在体内条件下可能对胃癌有治疗作用.%Objective To investigate the effect of the immunotherapy mediated by anti-CD3/anti-EGFR bispecific antibody ( BsAb) on the mice bome human gastric cancer and evaluate the effect of anti-CD3/anti-EGFR BsAb as a targeted therapeutic agent for gastric cancer. Methods The monoclonal antibody(mAb) of anti-CD3 and anti-EGFR were cross-linked to prepare the BsAb by chemical synthesis. The experimental therapy combined with PBLS as effector cells on the mice borne SGC7901 human gastric cancer was performed. Experimental mice were divided in to EGFR mAb group, CD3 mAb group, anti-CD3/anti-EGFR BsAb group and saline group. The comparisons of the curative activity among the anti-CD3/anti-EGFR BsAb, EGFR mAb and CD3 mAb groups were conducted in vivo. Results Anti-CD3/anti-EGFR BsAb was established successfully, and

  4. Correlation between egfr expression and accelerated proliferation during radiotherapy of head and neck squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Pedicini Piernicola

    2012-08-01

    Full Text Available Abstract Purpose To investigate the correlation between the expression of Epidermal Growth Factor receptor (EGFr and the reduction of the effective doubling time (TD during radiotherapy treatment and also to determine the dose per fraction to be taken into account when the overall treatment time (OTT is reduced in accelerated radiotherapy of head and neck squamous cell carcinoma (HNSCC. Methods A survey of the published papers comparing 3-years of local regional control rate (LCR for a total of 2162 patients treated with conventional and accelerated radiotherapy and with a pretreatment assessment of EGFr expression, was made. Different values of TD were obtained by a model incorporating the overall time corrected biologically effective dose (BED and a 3-year clinical LCR for high and low EGFr groups of patients (HEGFr and LEGFr, respectively. By obtaining the TD from the above analysis and the sub-sites’ potential doubling time (Tpot from flow cytometry and immunohistochemical methods, we were able to estimate the average TD for each sub-site included in the analysis. Moreover, the dose that would be required to offset the modified proliferation occurring in one day (Dprolif, was estimated. Results The averages of TD were 77 (27-9095% days in LEGFr and 8.8 (7.3-11.095% days in HEGFr, if an onset of accelerated proliferation TK at day 21 was assumed. The correspondent HEGFr sub-sites’ TD were 5.9 (6.6, 5.9 (6.6, 4.6 (6.1, 14.3 (12.9 days, with respect to literature immunohistochemical (flow cytometry data of Tpot for Oral-Cavity, Oro-pharynx, Hypo-pharynx, and Larynx respectively. The Dprolif for the HEGFr groups were 0.33 (0.29, 0.33 (0.29, 0.42 (0.31, 0.14 (0.15 Gy/day if α = 0.3 Gy-1 and α/β = 10 Gy were assumed. Conclusions A higher expression of the EGFr leads to enhanced proliferation. This study allowed to quantify the extent of the effect which EGFr expression has in terms of reduced TD and Dprolif for each head and neck

  5. Correlation between egfr expression and accelerated proliferation during radiotherapy of head and neck squamous cell carcinoma

    International Nuclear Information System (INIS)

    To investigate the correlation between the expression of Epidermal Growth Factor receptor (EGFr) and the reduction of the effective doubling time (TD) during radiotherapy treatment and also to determine the dose per fraction to be taken into account when the overall treatment time (OTT) is reduced in accelerated radiotherapy of head and neck squamous cell carcinoma (HNSCC). A survey of the published papers comparing 3-years of local regional control rate (LCR) for a total of 2162 patients treated with conventional and accelerated radiotherapy and with a pretreatment assessment of EGFr expression, was made. Different values of TD were obtained by a model incorporating the overall time corrected biologically effective dose (BED) and a 3-year clinical LCR for high and low EGFr groups of patients (HEGFr and LEGFr), respectively. By obtaining the TD from the above analysis and the sub-sites’ potential doubling time (Tpot) from flow cytometry and immunohistochemical methods, we were able to estimate the average TD for each sub-site included in the analysis. Moreover, the dose that would be required to offset the modified proliferation occurring in one day (Dprolif), was estimated. The averages of TD were 77 (27-90)95% days in LEGFr and 8.8 (7.3-11.0)95% days in HEGFr, if an onset of accelerated proliferation TK at day 21 was assumed. The correspondent HEGFr sub-sites’ TD were 5.9 (6.6), 5.9 (6.6), 4.6 (6.1), 14.3 (12.9) days, with respect to literature immunohistochemical (flow cytometry) data of Tpot for Oral-Cavity, Oro-pharynx, Hypo-pharynx, and Larynx respectively. The Dprolif for the HEGFr groups were 0.33 (0.29), 0.33 (0.29), 0.42 (0.31), 0.14 (0.15) Gy/day if α = 0.3 Gy-1 and α/β = 10 Gy were assumed. A higher expression of the EGFr leads to enhanced proliferation. This study allowed to quantify the extent of the effect which EGFr expression has in terms of reduced TD and Dprolif for each head and neck sub-site

  6. EGFR-targeted plasmonic magnetic nanoparticles suppress lung tumor growth by abrogating G2/M cell-cycle arrest and inducing DNA damage

    Directory of Open Access Journals (Sweden)

    Kuroda S

    2014-08-01

    Full Text Available Shinji Kuroda,1 Justina Tam,2 Jack A Roth,1 Konstantin Sokolov,2 Rajagopal Ramesh3–5 1Department of Thoracic and Cardiovascular Surgery, 2Department of Imaging Physics, The University of Texas MD Anderson Cancer Center, Houston, TX, USA; 3Department of Pathology, 4Graduate Program in Biomedical Sciences, 5Stephenson Cancer Center, University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA Background: We have previously demonstrated the epidermal growth factor receptor (EGFR-targeted hybrid plasmonic magnetic nanoparticles (225-NP produce a therapeutic effect in human lung cancer cell lines in vitro. In the present study, we investigated the molecular mechanism of 225-NP-mediated antitumor activity both in vitro and in vivo using the EGFR-mutant HCC827 cell line. Methods: The growth inhibitory effect of 225-NP on lung tumor cells was determined by cell viability and cell-cycle analysis. Protein expression related to autophagy, apoptosis, and DNA-damage were determined by Western blotting and immunofluorescence. An in vivo efficacy study was conducted using a human lung tumor xenograft mouse model. Results: The 225-NP treatment markedly reduced tumor cell viability at 72 hours compared with the cell viability in control treatment groups. Cell-cycle analysis showed the percentage of cells in the G2/M phase was reduced when treated with 225-NP, with a concomitant increase in the number of cells in Sub-G1 phase, indicative of cell death. Western blotting showed LC3B and PARP cleavage, indicating 225-NP-treatment activated both autophagy- and apoptosis-mediated cell death. The 225-NP strongly induced γH2AX and phosphorylated histone H3, markers indicative of DNA damage and mitosis, respectively. Additionally, significant γH2AX foci formation was observed in 225-NP-treated cells compared with control treatment groups, suggesting 225-NP induced cell death by triggering DNA damage. The 225-NP-mediated DNA damage involved abrogation of the

  7. Nicotinamide attenuates aquaporin 3 overexpression induced by retinoic acid through inhibition of EGFR/ERK in cultured human skin keratinocytes.

    Science.gov (United States)

    Song, Xiuzu; Xu, Aie; Pan, Wei; Wallin, Brittany; Kivlin, Rebecca; Lu, Shan; Cao, Cong; Bi, Zhigang; Wan, Yinsheng

    2008-08-01

    The most common adverse effects that are related to all-trans retinoic acid (atRA) treatment are irritation and dryness of the skin. atRA therapy is reported to impair barrier function as achieved by trans-epidermal water loss (TEWL). Treatment with nicotinamide prior to initiation of atRA therapy provides additional barrier protection and thus reduces susceptibility of retinoic acid. Our previous studies showed that atRA upregulates aquaporin 3 (AQP3) in cultured human skin keratinocytes and fibroblasts. Others have demonstrated that in atopic dermatitis, overexpression of AQP3 is linked to elevated TEWL and that nicotinamide treatment reduces skin TEWL. In this study, we observed that while atRA upregulates AQP3 expression in cultured human skin keratinocytes (HaCaT cells), nicotinamide attenuates the effect of atRA in a concentration-dependent manner. atRA treatment induces EGFR and ERK activation. PD153035, an EGFR inhibitor, and U0126, an ERK inhibitor, inhibit atRA-induced upregulation of AQP3. Nicotinamide also inhibits atRA-induced activation of EGFR/ERK signal transduction and decreases water permeability by downregulating AQP3 expression. Collectively, our results indicate that the effect of atRA on AQP3 expression is at least partly mediated by EGFR/ERK signaling in cultured human skin keratinocytes. Nicotinamide attenuates atRA-induced AQP3 expression through inhibition of EGFR/ERK signal transduction and eventually decreases water permeability and water loss. Our study provides insights into the molecular mechanism through which nicotinamide reverses the side effects of dryness in human skin after treatment with atRA.

  8. Role of epidermal growth factor receptor activation in regulating mucin synthesis

    Directory of Open Access Journals (Sweden)

    Nadel Jay A

    2001-02-01

    Full Text Available Abstract Healthy individuals have few goblet cells in their airways, but in patients with hypersecretory diseases goblet-cell upregulation results in mucus hypersecretion, airway plugging, and death. Multiple stimuli produce hypersecretion via epidermal growth factor receptor (EGFR expression and activation, causing goblet-cell metaplasia from Clara cells by a process of cell differentiation. These cells are also believed to be the cells of origin of non-small-cell lung cancer, but this occurs via cell multiplication. The mechanisms that determine which pathway is chosen are critical but largely unknown. Although no effective therapy exists for hypersecretion at present, the EGFR cascade suggests methods for effective therapeutic intervention.

  9. Colourimetric Determination of Phospholipase Activities in Balamuthia mandrillaris

    Directory of Open Access Journals (Sweden)

    Syed Razi Haider

    2007-01-01

    Full Text Available Balamuthia mandrillaris is a recently identified protozoan pathogen that can cause fatal granulomatous encephalitis however the pathogenesis and pathophysiology associated with Balamuthia encephalitis remain unclear. We have recently isolated B. mandrillaris from a 33-years old male who died of encephalitis. Using this isolate, we demonstrated for the first time that B. mandrillaris exhibited phospholipase activities. More specifically, B. mandrillaris exhibited phospholipase A2 and phospholipase D activities. For the first time we used colourimetric technique based on spectrophotometer and designed phospholipases assays to determine these phospholipase activities. The functional role of phospholipases was determined in in vitro assays using human brain microvascular endothelial cells (HBMEC. We observed that PLA2-specific inhibitor i.e., cytidine 5'-diphosphocholine significantly inhibited B. mandrillaris binding to HBMEC. Similarly PLD inhibitor i.e., compound 48/80 inhibited B. mandrillaris binding to HBMEC. Moreover, both inhibitors inhibited B. mandrillaris-mediated HBMEC cytotoxicity. Overall these results clearly demonstrate that phospholipases are important virulence determinants in B. mandrillaris. Further studies will identify the precise role of phospholipases in the pathogenesis of B. mandrillaris, which may help develop therapeutic interventions. Using a novel spectrophotometric-based assay we demonstrated for the first time that B. mandrillaris exhibit phospholipase activities.

  10. Factor H-related proteins determine complement-activating surfaces.

    Science.gov (United States)

    Józsi, Mihály; Tortajada, Agustin; Uzonyi, Barbara; Goicoechea de Jorge, Elena; Rodríguez de Córdoba, Santiago

    2015-06-01

    Complement factor H-related proteins (FHRs) are strongly associated with different diseases involving complement dysregulation, which suggests a major role for these proteins regulating complement activation. Because FHRs are evolutionarily and structurally related to complement inhibitor factor H (FH), the initial assumption was that the FHRs are also negative complement regulators. Whereas weak complement inhibiting activities were originally reported for these molecules, recent developments indicate that FHRs may enhance complement activation, with important implications for the role of these proteins in health and disease. We review these findings here, and propose that FHRs represent a complex set of surface recognition molecules that, by competing with FH, provide improved discrimination of self and non-self surfaces and play a central role in determining appropriate activation of the complement pathway.

  11. Human monoamine oxidase A gene determines levels of enzyme activity.

    OpenAIRE

    Hotamisligil, G S; Breakefield, X O

    1991-01-01

    Monoamine oxidase (MAO) is a critical enzyme in the degradative deamination of biogenic amines throughout the body. Two biochemically distinct forms of the enzyme, A and B, are encoded in separate genes on the human X chromosome. In these studies we investigated the role of the structural gene for MAO-A in determining levels of activity in humans, as measured in cultured skin fibroblasts. The coding sequence of the mRNA for MAO-A was determined by first-strand cDNA synthesis, PCR amplificatio...

  12. Determination of mercury in dentists through Neutron activation analysis

    International Nuclear Information System (INIS)

    It was determined the mercury levels in urine through Neutron activation analysis to 25 dentists who have been exposed to mercury by several time periods, because of the routine manipulations of amalgams. The determined concentrations of mercury were less to 10 μ g Hg/l of urine. The results were founded inside the safety limits reported in the literature. The mercury levels in the dentists are associated with a wide variety of factors that contribute to their exposure as: number of years of dental practice, number of amalgams manipulated between others. (Author)

  13. Determinants of clean development mechanism activity: Evidence from China

    International Nuclear Information System (INIS)

    As regards the dual goals of additionality and sustainability, some clean development mechanism (CDM) projects were successfully registered while some were rejected by the Executive Board (EB). This paper focuses on two final statuses of these projects and aims to explore the potential determinants of CDM activities. Based on the Chinese samples of the rejections and registrations, we find that (1) according to the financial barrier analysis with the internal rate of return without CDM, the additionality is the most important and robust rule of CDM activities and the CDM projects successfully registered are more additional than the rejected ones in China; (2) the sustainability is another important determinant of CDM activities in China and Chinese CDM activities are sustainable in terms of size, while the EB is criticized for its selection of unsustainable CDM projects in terms of cost-effectiveness on emission reductions within a long period or in terms of hydropower plants; (3) some other factors including the prices of sale electricity and certified emission reductions, the generating capacity and installed capacity all tend to pose significant impacts on the final status of each project that features the CDM activities in local areas of China. - Highlights: • The final status of each project features the CDM activities in local areas. • Microeconomic factors have significant impacts on the final statuses. • The additionality is the most important and robust rule of CDM activities in China. • Chinese CDM activities are sustainable in terms of size. • Chinese unsustainable CDM projects were selected by the EB in terms of cost-effectiveness

  14. EGFR-Targeted Therapeutics: Focus on SCCHN and NSCLC

    Directory of Open Access Journals (Sweden)

    Martin Sattler

    2008-01-01

    Full Text Available Cancers of the head and neck and of the lung are associated with high morbidity and mortality rates that have remained relatively unchanged for more than 3 decades, despite advances in radiation therapies and chemotherapies over the same time. It is generally believed that the efficacy of standard therapy regimens has reached a plateau for these cancers. The discovery of specific aberrant molecular signaling pathways in solid tumors has afforded promising new directions for newer “targeted” cancer therapeutics. Among these, the epidermal growth factor receptor (EGFR shows promise as a therapeutic target. Clinical studies have demonstrated that this targeted approach provides clinically meaningful benefit. This article reviews EGFR-targeted therapies in use and in development, with a focus on the role of EGFR in the pathophysiology of head and neck and lung cancer, and new concepts being investigated to improve outcomes with these agents.

  15. Multiple myeloma acquires resistance to EGFR inhibitor via induction of pentose phosphate pathway.

    Science.gov (United States)

    Chen, Yan; Huang, Ruibin; Ding, Jianghua; Ji, Dexiang; Song, Bing; Yuan, Liya; Chang, Hong; Chen, Guoan

    2015-04-20

    Multiple myeloma (MM) was characterized by frequent mutations in KRAS/NRAS/BRAF within the EGFR pathway that could induce resistance to EGFR inhibitors. We here report that EGFR inhibition solely exhibited moderate inhibition in KRAS/NRAS/BRAF wildtype (triple-WT) MM cells, whilst had no effect in myeloma cells with any of the mutated genes. The moderate inhibitory effect was conferred by induction of pentose phosphate pathway (PPP) when cells were treated with Gefitinib, the EGFR inhibitor. Combination of Gefitinib with PPP inhibitor 6AN effected synergistically in triple-WT cells. The inhibition could be restored by addition of NADPH. Dual EGFR/ERBB2 inhibitor Afatinib also exhibited similar effects. Further genetic silencing of EGFR, ERBB2 and mTOR indicated that major effect conferred by ERBB2 was via convergence to EGFR pathway in MM. Our results contributed to the individualized targeted therapy with EGFR inhibitors in MM.

  16. Wogonin inhibits the proliferation and invasion, and induces the apoptosis of HepG2 and Bel7402 HCC cells through NF‑κB/Bcl-2, EGFR and EGFR downstream ERK/AKT signaling.

    Science.gov (United States)

    Liu, Xiaodong; Tian, Shuo; Liu, Mei; Jian, Lingyan; Zhao, Limei

    2016-10-01

    The anticancer effects of the natural flavonoid, wogonin, have been reported. However, its molecular mechanisms of action have not yet been fully explored. In the present study, we aimed to examine the molecular mechanisms of action of wogonin and its effects on the biological behavior of the HepG2 and Bel7402 hepatocellular carcinoma (HCC) cell lines. We also examined the effects of wogonin on nuclear factor-κB (NF-κB)/Bcl-2 and epidermal growth factor receptor (EGFR) signaling, as well as on downstream pathways of EGFR, namely extracellular signal-regulated kinase (ERK)/AKT signaling. We found that treatment with wogonin inhibited the proliferation and invasion, and induced the apoptosis of the HepG2 and Bel7402 cells. In addition, treatment with wogonin decreased cyclin D1, cyclin E, CDK4/6, Bcl-2 and matrix metalloproteinase 2 (MMP2) expression, and promoted the cleavage of caspase-3 and caspase-9 in a concentration-dependent manner. Further experiments revealed that wogonin inhibited NF-κB/Bcl-2 signaling by decreasing the IκB and p65 phosphorylation levels. Wogonin also inhibited the activation of the EGFR (Tyr845) signaling pathway, and that of downstream pathways of EGFR, namely ERK/AKT/MMP2 signaling. The depletion of EGFR by siRNA partly abolished the inhibitory effects of wogonin on cyclin D1, MMP2 expression. On the whole, our our findings demonstrate that wogonin effectively suppresses the proliferation, invasion and survival of HCC cells through the modulation of the NF-κB and EGFR signaling pathways.

  17. Determinants of Regional Entrepreneurial Activity in the Czech Republic

    Directory of Open Access Journals (Sweden)

    Ondřej Dvouletý

    2016-07-01

    Full Text Available The following study is focused on analysis of registered businesses in the 14 regions of the Czech Republic during the period of years 1995-2013. The aim of the study was to quantify factors that affect entrepreneurial activity expressed as rate of registered businesses per capita. Based on the previous empirical studies, the determinants were selected and hypothesis stated. Formed hypothesis investigated positive impact of GDP per capita, unemployment rate and R&D institutions on rate of registered business activity. To evaluate them, data were obtained from the Czech Statistical Office and formed into dataset. Firstly, panel regressions estimated with fixed effects method were employed and secondly, Granger causality tests to evaluate the relationship between entrepreneurial activity and GDP per capita were used. Regression estimates proved positive relationship between entrepreneurial activity in Czech regions and GDP per capita, unemployment rate and support activities of R&D institutions. Positive impact was also confirmed for population density, average age, share of tertiary educated population and real R&D expenditures. Testing Granger causality proved dual causality between entrepreneurial activity and GDP per capita confirming that GDP per capita as good predictor of economic development of Czech regions. Finally, economic growth motivates Czech individuals to enter entrepreneurial activity.

  18. Multielement determination in soil extracts by instrumental neutron activation analysis

    International Nuclear Information System (INIS)

    Forest Swiss soils from 39 locations, each sampled at three depths, were extracted with a buffered solution of pH 4.65 containing EDTA. Nearly 30 elements were determined by instrumental neutron activation analysis. The results obtained are discussed in terms of efficiency of the extractant, precision and accuracy. Summarized results are presented for the quantity fractions of the extracted elements. (author) 18 refs.; 4 figs.; 3 tabs

  19. Determination Of Antioxidant Activities In Freshliver (Salvia Officinalis) Plant

    OpenAIRE

    Arıduru, Rana; Arabacı, Gülnur

    2013-01-01

    In this study, we determined the antioxidant activities of four different solvent fractions (ethanol, methanol, acetone and ethyl acetate) obtained from Freshliver plant leaves (Salvia officinalis) by employing two different assays such as 2,2-diphenyl-1-picrylhydrazyl hydrate (DPPH) and Folin-Ciocaltaeu method. The results showed that ethanolextract of freshliver plant exhibited the highest total phenolic contents (43.55 mg GAE/g extract), followed by methanol-extract of freshliver plant (23...

  20. Which biomarker predicts benefit from EGFR-TKI treatment for patients with lung cancer?

    OpenAIRE

    Uramoto, H; Mitsudomi, T.

    2007-01-01

    Subsets of patients with non-small cell lung cancer respond remarkably well to small molecule tyrosine kinase inhibitors (TKI) specific for epidermal growth factor receptor (EGFR) such as gefitinib or erlotinib. In 2004, it was found that EGFR mutations occurring in the kinase domain are strongly associated with EGFR-TKI sensitivity. However, subsequent studies revealed that this relationship was not perfect and various predictive markers have been reported. These include EGFR gene copy numbe...

  1. EGFR-inhibitors, radiotherapy and normal tissue toxicity

    DEFF Research Database (Denmark)

    Eriksen, J. G.

    2015-01-01

    EGFR-inhibitors have been used in several clinical settings during the last decade and side-effects related to normal tissues like the skin, mucosa and kidney has been well described. However, when EGFR-inhibitors are combined with radiotherapy, then different skin and mucosa toxicity profiles can...... variations that are shared between tumour and normal tissue like skin and mucosa. At present, it is not possible to predict which patients that will develop severe skin toxicity and thereby potentially benefit from EGFRinhibition in terms of tumour response. However, emerging data suggests that certain...

  2. Quantum dot-based quantification revealed differences in subcellular localization of EGFR and E-cadherin between EGFR-TKI sensitive and insensitive cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Huang Donghai; Su Ling; Peng Xianghong; Zhang Hongzheng; Khuri, Fadlo R; Shin, Dong M; Chen Zhuo [Department of Hematology and Medical Oncology, Winship Cancer Institute, Emory University School of Medicine, Atlanta, GA (United States)], E-mail: gzchen@emory.edu

    2009-06-03

    Nanoparticle quantum dots (QDs) provide sharper and more photostable fluorescent signals than organic dyes, allowing quantification of multiple biomarkers simultaneously. In this study, we quantified the expression of epidermal growth factor receptor (EGFR) and E-cadherin (E-cad) in the same cells simultaneously by using secondary antibody-conjugated QDs with two different emission wavelengths (QD605 and QD565) and compared the cellular distribution of EGFR and E-cad between EGFR-tyrosine kinase inhibitor (TKI)-insensitive and -sensitive lung and head and neck cancer cell lines. Relocalization of EGFR and E-cad upon treatment with the EGFR-TKI erlotinib in the presence of EGF was visualized and analyzed quantitatively. Our results showed that QD-immunocytochemistry (ICC)-based technology can not only quantify basal levels of multiple biomarkers but also track the localization of the biomarkers upon biostimulation. With this new technology we found that in EGFR-TKI-insensitive cells, EGFR and E-cad were located mainly in the cytoplasm; while in sensitive cells, they were found mainly on the cell membrane. After induction with EGF, both EGFR and E-cad internalized to the cytoplasm, but the internalization capability in sensitive cells was greater than that in insensitive cells. Quantification also showed that inhibition of EGF-induced EGFR and E-cad internalization by erlotinib in the sensitive cells was stronger than that in the insensitive cells. These studies demonstrate substantial differences between EGFR-TKI-insensitive and -sensitive cancer cells in EGFR and E-cad expression and localization both at the basal level and in response to EGF and erlotinib. QD-based analysis facilitates the understanding of the features of EGFR-TKI-insensitive versus -sensitive cancer cells and may be used in the prediction of patient response to EGFR-targeted therapy.

  3. 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.

  4. Antiproliferative effect of growth hormone-releasing hormone (GHRH antagonist on ovarian cancer cells through the EGFR-Akt pathway

    Directory of Open Access Journals (Sweden)

    Varga Jozsef

    2010-05-01

    Full Text Available Abstract Background Antagonists of growth hormone-releasing hormone (GHRH are being developed for the treatment of various human cancers. Methods MTT assay was used to test the proliferation of SKOV3 and CaOV3. The splice variant expression of GHRH receptors was examined by RT-PCR. The expression of protein in signal pathway was examined by Western blotting. siRNA was used to block the effect of EGFR. Results In this study, we investigated the effects of a new GHRH antagonist JMR-132, in ovarian cancer cell lines SKOV3 and CaOV3 expressing splice variant (SV1 of GHRH receptors. MTT assay showed that JMR-132 had strong antiproliferative effects on SKOV3 and CaOV3 cells in both a time-dependent and dose-dependent fashion. JMR-132 also induced the activation and increased cleaved caspase3 in a time- and dose-dependent manner in both cell lines. In addition, JMR-132 treatments decreased significantly the epidermal growth factor receptor (EGFR level and the phosphorylation of Akt (p-Akt, suggesting that JMR-132 inhibits the EGFR-Akt pathway in ovarian cancer cells. More importantly, treatment of SKOV3 and CaOV3 cells with 100 nM JMR-132 attenuated proliferation and the antiapoptotic effect induced by EGF in both cell lines. After the knockdown of the expression of EGFR by siRNA, the antiproliferative effect of JMR-132 was abolished in SKOV3 and CaOV3 cells. Conclusions The present study demonstrates that the inhibitory effect of the GHRH antagonist JMR-132 on proliferation is due, in part, to an interference with the EGFR-Akt pathway in ovarian cancer cells.

  5. Emerging Roles of MicroRNAs in EGFR-Targeted Therapies for Lung Cancer

    Directory of Open Access Journals (Sweden)

    Fei Han

    2015-01-01

    Full Text Available Lung cancer is a leading cause of cancer mortality worldwide. Several molecular pathways underlying mechanisms of this disease have been partly elucidated, among which the epidermal growth factor receptor (EGFR pathway is one of the well-known signaling cascades that plays a critical role in tumorigenesis. Dysregulation of the EGFR signaling is frequently found in lung cancer. The strategies to effectively inhibit EGFR signaling pathway have been mounted for developing anticancer therapeutic agents. However, most anti-EGFR-targeted agents fail to repress cancer progression because of developing drug-resistance. Therefore, studies of the mechanisms underpinning the resistance toward anti-EGFR agents may provide important findings for lung cancer treatment using anti-EGFR therapies. Recently, increasing numbers of miRNAs are correlated with the drug resistance of lung cancer cells to anti-EGFR agents, indicating that miRNAs may serve as novel targets and/or promising predictive biomarkers for anti-EGFR therapy. In this paper, we summarize the emerging role of miRNAs as regulators to modulate the EGFR signaling and the resistance of lung cancer cells to anti-EGFR therapy. We also highlight the evidence supporting the use of miRNAs as biomarkers for response to anti-EGFR agents and as novel therapeutic targets to circumvent the resistance of lung cancer cells to EGFR inhibitors.

  6. File list: Oth.ALL.20.EGFR.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.20.EGFR.AllCell hg19 TFs and others EGFR All cell types SRX853217,SRX853214...,SRX853216,SRX853215 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.ALL.20.EGFR.AllCell.bed ...

  7. File list: Oth.ALL.10.EGFR.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.10.EGFR.AllCell hg19 TFs and others EGFR All cell types SRX853214,SRX853217...,SRX853216,SRX853215 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.ALL.10.EGFR.AllCell.bed ...

  8. File list: Oth.Utr.10.EGFR.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Utr.10.EGFR.AllCell hg19 TFs and others EGFR Uterus SRX853214,SRX853217,SRX8532...16,SRX853215 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Utr.10.EGFR.AllCell.bed ...

  9. File list: Oth.ALL.50.EGFR.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.50.EGFR.AllCell hg19 TFs and others EGFR All cell types SRX853217,SRX853215...,SRX853214,SRX853216 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.ALL.50.EGFR.AllCell.bed ...

  10. File list: Oth.Utr.20.EGFR.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  11. File list: Oth.Utr.05.EGFR.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  12. File list: Oth.Utr.50.EGFR.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  13. File list: Oth.ALL.05.EGFR.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.05.EGFR.AllCell hg19 TFs and others EGFR All cell types SRX853214,SRX853217...,SRX853215,SRX853216 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.ALL.05.EGFR.AllCell.bed ...

  14. EGFR Signaling in the Brain Is Necessary for Olfactory Learning in "Drosophila" Larvae

    Science.gov (United States)

    Rahn, Tasja; Leippe, Matthias; Roeder, Thomas; Fedders, Henning

    2013-01-01

    Signaling via the epidermal growth factor receptor (EGFR) pathway has emerged as one of the key mechanisms in the development of the central nervous system in "Drosophila melanogaster." By contrast, little is known about the functions of EGFR signaling in the differentiated larval brain. Here, promoter-reporter lines of EGFR and its most prominent…

  15. Ofloxacin induces apoptosis via β1 integrin-EGFR-Rac1-Nox2 pathway in microencapsulated chondrocytes

    Energy Technology Data Exchange (ETDEWEB)

    Sheng, Zhi-Guo [State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Science, Chinese Academy of Sciences, 18 Shuangqing Road, Beijing 100085 (China); Huang, Wei [Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University Health Science Center, Beijing 1000191 (China); Liu, Yu-Xiang [State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Science, Chinese Academy of Sciences, 18 Shuangqing Road, Beijing 100085 (China); Yuan, Ye [Beijing Institute of Pharmacology and Toxicology, 27 Taiping Road, Beijing 100850 (China); Zhu, Ben-Zhan, E-mail: bzhu@rcees.ac.cn [State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Science, Chinese Academy of Sciences, 18 Shuangqing Road, Beijing 100085 (China); Linus Pauling Institute, Oregon State University, Corvallis, OR 97331 (United States)

    2013-02-15

    Quinolones (QNs)-induced arthropathy is an important toxic side-effect in immature animals leading to the restriction of their therapeutic use in pediatrics. Ofloxacin, a typical QN, was found to induce the chondrocytes apoptosis in the early phase (12–48 h) of arthropathy in our previous study. However, the exact mechanism(s) is unclear. Microencapsulated juvenile rabbit joint chondrocytes, a three-dimensional culture system, is utilized to perform the present study. Ofloxacin, at a therapeutically relevant concentration (10 μg/ml), disturbs the interaction between β1 integrin and activated intracellular signaling proteins at 12 h, which is inhibited when supplementing Mg{sup 2+}. Intracellular reactive oxygen species (ROS) significantly increases in a time-dependent manner after exposure to ofloxacin for 12–48 h. Furthermore, ofloxacin markedly enhances the level of activated Rac1 and epidermal growth factor receptor (EGFR) phosphorylation, and its inhibition in turn reduces the ROS production, apoptosis and Rac1 activation. Silencing Nox2, Rac1 or supplementing Mg{sup 2+} inhibits ROS accumulation, apoptosis occurrence and EGFR phosphorylation induced by ofloxacin. However, depletion of Nox2, Rac1 and inhibition of EGFR do not affect ofloxacin-mediated loss of interaction between β1 integrin and activated intracellular signaling proteins. In addition, ofloxacin also induces Vav2 phosphorylation, which is markedly suppressed after inactivating EGFR or supplementing Mg{sup 2+}. These results suggest that ofloxacin causes Nox2-mediated intracellular ROS production by disrupting the β1 integrin function and then activating the EGFR-Vav2-Rac1 pathway, finally resulting in apoptosis within 12–48 h exposure. The present study provides a novel insight regarding the potential role of Nox-driven ROS in QNs-induced arthropathy. - Highlights: ► Ofloxacin induces Nox2-driven ROS in encapsulated chondrocyte at 12–48 h. ► Ofloxacin stimulates ROS production via

  16. Structural Determinants of Clostridium difficile Toxin A Glucosyltransferase Activity

    Energy Technology Data Exchange (ETDEWEB)

    Pruitt, Rory N.; Chumbler, Nicole M.; Rutherford, Stacey A.; Farrow, Melissa A.; Friedman, David B.; Spiller, Ben; Lacy, D. Borden (Vanderbilt)

    2012-03-28

    The principle virulence factors in Clostridium difficile pathogenesis are TcdA and TcdB, homologous glucosyltransferases capable of inactivating small GTPases within the host cell. We present crystal structures of the TcdA glucosyltransferase domain in the presence and absence of the co-substrate UDP-glucose. Although the enzymatic core is similar to that of TcdB, the proposed GTPase-binding surface differs significantly. We show that TcdA is comparable with TcdB in its modification of Rho family substrates and that, unlike TcdB, TcdA is also capable of modifying Rap family GTPases both in vitro and in cells. The glucosyltransferase activities of both toxins are reduced in the context of the holotoxin but can be restored with autoproteolytic activation and glucosyltransferase domain release. These studies highlight the importance of cellular activation in determining the array of substrates available to the toxins once delivered into the cell.

  17. The structural determinants of insulin-like peptide 3 activity

    Directory of Open Access Journals (Sweden)

    Ross AD Bathgate

    2012-02-01

    Full Text Available INSL3 is a hormone and/or paracrine factor which is a member of the relaxin peptide family. It has key roles as a fertility regulator in both males and females. The receptor for INSL3 is the leucine rich repeat (LRR containing G-protein coupled receptor 8 (LGR8 which is now known as relaxin family peptide receptor 2 (RXFP2. Receptor activation by INSL3 involves binding to the LRRs in the large ectodomain of RXFP2 by residues within the B-chain of INSL3 as well as an interaction with the transmembrane exoloops of the receptor. Although the binding to the LRRs is well characterized the features of the peptide and receptor involved in the exoloop interaction are currently unknown. This study was designed to determine the key INSL3 determinants for RXFP2 activation. A chimeric peptide approach was first utilized to demonstrate that the A-chain is critical for receptor activation. Replacement of the INSL3 A-chain with that from the related peptides INSL5 and INSL6 resulted in complete loss of activity despite only minor changes in binding affinity. Subsequent replacement of specific A-chain residues with those from the INSL5 peptide highlighted that the N-terminus of the A-chain of INSL3 is critical for its activity. Remarkably, replacement of the entire N-terminus with four or five alanine residues resulted in peptides with near native activity suggesting that specific residues are not necessary for activity. Additionally removal of two amino acids at the C-terminus of the A-chain and mutation of Lys-8 in the B-chain also resulted in minor decreases in peptide activity. Therefore we have demonstrated that the activity of the INSL3 peptide is driven predominantly by residues 5-9 in the A-chain, with minor additional contributions from the two C-terminal A-chain residues and Lys-8 in the B-chain. Using this new knowledge, we were able to produce a truncated INSL3 peptide structure which retained native activity, despite having 14 fewer residues than

  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

    Institute of Scientific and Technical Information of China (English)

    Jie-Ying Chen; Ya-Nan Cheng; Lei Han; Feng Wei; Wen-Wen Yu; Xin-Wei Zhang; Shui Cao; Jin-Pu Yu

    2015-01-01

    Objective:A meta-analysis was performed to augment the insuffcient data on the impact of mutative EGFR downstream phosphatidylinositol-3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) pathways on the clinical effciency of epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) treatment of non-small cell lung cancer (NSCLC) patients. Methods: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. Results:Mutation in K-ras signiifcantly predicted poor ORR [OR=0.22;95%conifdence 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 signiifcantly 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. Conclusion: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 beneift from EGFR-TKI treatment.

  19. Predictive value of K-ras and PIK3CA in non-small cell lung cancer patients treated with EGFR-TKIs: a systemic review and meta-analysis

    Science.gov (United States)

    Chen, Jie-Ying; Cheng, Ya-Nan; Han, Lei; Wei, Feng; Yu, Wen-Wen; Zhang, Xin-Wei; Cao, Shui; Yu, Jin-Pu

    2015-01-01

    Objective 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. Methods 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. Results 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. Conclusion 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. PMID:26175928

  20. 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 (