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Sample records for aurora kinase inhibitor

  1. Jadomycin B, an Aurora-B kinase inhibitor discovered through virtual screening

    Institute of Scientific and Technical Information of China (English)

    Da Huafu; Jing Zhang; Jian Tingzheng; Yan Li; Ke Qianyang; Yan Changwang; Shu Yisi

    2008-01-01

    @@ Aurora kinases are clearly implicated in tumorgenesis and have emerged as promising targets for cancer therapy in recent years. In a virtual screening attempt, 22 compounds were identified from nearly 15,000 microbial natural products as potential small-molecule inhibitors of human Aurora-B kinase. When tested in yeast models, 2 compounds (one is Jadomycin B) showed preferential inhibition of ipll-321 (yeast Aurora kinase temperature sensitive mutant) than wild type yeast cell, suggesting these compounds are true Aurora kinase inhibitors. Further in vitro biochemical assay using purified recombinant Aurora-B kinase showed Jadomycin B inhibits Aurora-B activity in a dose-dependent fashion while two Jadomycin congeners, Jadomycin S and T, showed no activity.

  2. A Cell Biologist’s Field Guide to Aurora Kinase Inhibitors

    Directory of Open Access Journals (Sweden)

    Christian eDe Groot

    2015-12-01

    Full Text Available Aurora kinases are essential for cell division and are frequently misregulated in human cancers. Based on their potential as cancer therapeutics, a plethora of small molecule Aurora kinase inhibitors have been developed, with a subset having been adopted as tools in cell biology. Here, we fill a gap in the characterization of Aurora kinase inhibitors by using biochemical and cell-based assays to systematically profile a panel of 10 commercially available compounds with reported selectivity for Aurora A (MLN8054, MLN8237, MK-5108, MK-8745, Genentech Aurora Inhibitor 1, Aurora B (Hesperadin, ZM447439, AZD1152-HQPA, GSK1070916 or Aurora A/B (VX-680. We quantify the in vitro effect of each inhibitor on the activity of Aurora A alone, as well as Aurora A and Aurora B bound to fragments of their activators, TPX2 and INCENP, respectively. We also report kinome profiling results for a subset of these compounds to highlight potential off-target effects. In a cellular context, we demonstrate that immunofluorescence-based detection of LATS2 and histone H3 phospho-epitopes provides a facile and reliable means to assess potency and specificity of Aurora A versus Aurora B inhibition, and that G2 duration measured in a live imaging assay is a specific readout of Aurora A activity. Our analysis also highlights variation between HeLa, U2OS and hTERT-RPE1 cells that impacts selective Aurora A inhibition. For Aurora B, all 4 tested compounds exhibit excellent selectivity and do not significantly inhibit Aurora A at effective doses. For Aurora A, MK-5108 and MK-8745 are significantly more selective than the commonly used inhibitors MLN8054 and MLN8237. A crystal structure of an Aurora A/MK-5108 complex that we determined suggests the chemical basis for this higher specificity. Taken together, our quantitative biochemical and cell-based analyses indicate that AZD1152-HQPA and MK-8745 are the best current tools for selectively inhibiting Aurora B and Aurora A

  3. Recent advances in the development of Aurora kinases inhibitors in hematological malignancies

    Science.gov (United States)

    Choudary, Iqra; Barr, Paul M.; Friedberg, Jonathan

    2015-01-01

    Over the last two decades, since the discovery of Drosophila mutants in 1995, much effort has been made to understand Aurora kinase biology. Three mammalian subtypes have been identified thus far which include the Aurora A, B and C kinases. These regulatory proteins specifically work at the cytoskeleton and chromosomal structures between the kinetochores and have vital functions in the early phases of the mitotic cell cycle. Today, there are multiple phase I and phase II clinical trials as well as numerous preclinical studies taking place looking at Aurora kinase inhibitors in both hematologic and solid malignancies. This review focuses on the preclinical and clinical development of Aurora kinase inhibitors in hematological malignancy and discusses their therapeutic potential. PMID:26622997

  4. Identification, SAR studies, and X-ray co-crystallographic analysis of a novel furanopyrimidine aurora kinase A inhibitor.

    Science.gov (United States)

    Coumar, Mohane Selvaraj; Tsai, Ming-Tsung; Chu, Chang-Ying; Uang, Biing-Jiun; Lin, Wen-Hsing; Chang, Chun-Yu; Chang, Teng-Yuan; Leou, Jiun-Shyang; Teng, Chi-Huang; Wu, Jian-Sung; Fang, Ming-Yu; Chen, Chun-Hwa; Hsu, John T-A; Wu, Su-Ying; Chao, Yu-Sheng; Hsieh, Hsing-Pang

    2010-02-01

    Herein we reveal a simple method for the identification of novel Aurora kinase A inhibitors through substructure searching of an in-house compound library to select compounds for testing. A hydrazone fragment conferring Aurora kinase activity and heterocyclic rings most frequently reported in kinase inhibitors were used as substructure queries to filter the in-house compound library collection prior to testing. Five new series of Aurora kinase inhibitors were identified through this strategy, with IC(50) values ranging from approximately 300 nM to approximately 15 microM, by testing only 133 compounds from a database of approximately 125,000 compounds. Structure-activity relationship studies and X-ray co-crystallographic analysis of the most potent compound, a furanopyrimidine derivative with an IC(50) value of 309 nM toward Aurora kinase A, were carried out. The knowledge gained through these studies could help in the future design of potent Aurora kinase inhibitors.

  5. Antineoplastic effects of an Aurora B kinase inhibitor in breast cancer

    Directory of Open Access Journals (Sweden)

    Velazquez-Torres Guermarie

    2010-02-01

    Full Text Available Abstract Background Aurora B kinase is an important mitotic kinase involved in chromosome segregation and cytokinesis. It is overexpressed in many cancers and thus may be an important molecular target for chemotherapy. AZD1152 is the prodrug for AZD1152-HQPA, which is a selective inhibitor of Aurora B kinase activity. Preclinical antineoplastic activity of AZD1152 against acute myelogenous leukemia, multiple myeloma and colorectal cancer has been reported. However, this compound has not been evaluated in breast cancer, the second leading cause of cancer deaths among women. Results The antineoplastic activity of AZD1152-HQPA in six human breast cancer cell lines, three of which overexpress HER2, is demonstrated. AZD1152-HQPA specifically inhibited Aurora B kinase activity in breast cancer cells, thereby causing mitotic catastrophe, polyploidy and apoptosis, which in turn led to apoptotic death. AZD1152 administration efficiently suppressed the tumor growth in a breast cancer cell xenograft model. In addition, AZD1152 also inhibited pulmonary metastatic nodule formation in a metastatic breast cancer model. Notably, it was also found that the protein level of Aurora B kinase declined after inhibition of Aurora B kinase activity by AZD1152-HQPA in a time- and dose-dependent manner. Investigation of the underlying mechanism suggested that AZD1152-HQPA accelerated protein turnover of Aurora B via enhancing its ubiquitination. Conclusions It was shown that AZD1152 is an effective antineoplastic agent for breast cancer, and our results define a novel mechanism for posttranscriptional regulation of Aurora B after AZD1152 treatment and provide insight into dosing regimen design for this kinase inhibitor in metastatic breast cancer treatment.

  6. Aurora kinase inhibitors: which role in the treatment of chronic myelogenous leukemia patients resistant to imatinib?

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    Giovanni Martinelli

    2009-02-01

    Full Text Available At present, there are no compounds in clinical development in the field of chronic myeloid leukemia (CML or Philadelphia-positive (Ph+ acute lymphoblastic leukemia (ALL that have been documented to harbor significant activity against the imatinib-resistant T315I mutation. Recent reports on the pre-clinical activity of some emerging tyrosine kinase inhibitors such as ON012380, VX-680 and PHA-739358 promise possible clinical efficacy against this specific Bcr-Abl mutant form. Here, we focus on the role of aurora kinase inhibitor VX-680 and PHA-739358 in blocking the leukemogenic pathways driven by wild-type and T315I-Bcr-Abl in CML or Ph+ ALL by reviewing recent research evidence. We also discuss the possibility of employing aurora kinase inhibitors as a promising new therapeutic approach in the treatment of CML and Ph+ ALL patients resistant to first and second generation TK inhibitors.

  7. Tripolin A, a novel small-molecule inhibitor of aurora A kinase, reveals new regulation of HURP's distribution on microtubules.

    Directory of Open Access Journals (Sweden)

    Iliana A Kesisova

    Full Text Available Mitotic regulators exhibiting gain of function in tumor cells are considered useful cancer therapeutic targets for the development of small-molecule inhibitors. The human Aurora kinases are a family of such targets. In this study, from a panel of 105 potential small-molecule inhibitors, two compounds Tripolin A and Tripolin B, inhibited Aurora A kinase activity in vitro. In human cells however, only Tripolin A acted as an Aurora A inhibitor. We combined in vitro, in vivo single cell and in silico studies to demonstrate the biological action of Tripolin A, a non-ATP competitive inhibitor. Tripolin A reduced the localization of pAurora A on spindle microtubules (MTs, affected centrosome integrity, spindle formation and length, as well as MT dynamics in interphase, consistent with Aurora A inhibition by RNAi or other specific inhibitors, such as MLN8054 or MLN8237. Interestingly, Tripolin A affected the gradient distribution towards the chromosomes, but not the MT binding of HURP (Hepatoma Up-Regulated Protein, a MT-associated protein (MAP and substrate of the Aurora A kinase. Therefore Tripolin A reveals a new way of regulating mitotic MT stabilizers through Aurora A phosphorylation. Tripolin A is predicted to bind Aurora A similarly but not identical to MLN8054, therefore it could be used to dissect pathways orchestrated by Aurora kinases as well as a scaffold for further inhibitor development.

  8. Tripolin A, a novel small-molecule inhibitor of aurora A kinase, reveals new regulation of HURP's distribution on microtubules.

    Science.gov (United States)

    Kesisova, Iliana A; Nakos, Konstantinos C; Tsolou, Avgi; Angelis, Dimitrios; Lewis, Joe; Chatzaki, Aikaterini; Agianian, Bogos; Giannis, Athanassios; Koffa, Maria D

    2013-01-01

    Mitotic regulators exhibiting gain of function in tumor cells are considered useful cancer therapeutic targets for the development of small-molecule inhibitors. The human Aurora kinases are a family of such targets. In this study, from a panel of 105 potential small-molecule inhibitors, two compounds Tripolin A and Tripolin B, inhibited Aurora A kinase activity in vitro. In human cells however, only Tripolin A acted as an Aurora A inhibitor. We combined in vitro, in vivo single cell and in silico studies to demonstrate the biological action of Tripolin A, a non-ATP competitive inhibitor. Tripolin A reduced the localization of pAurora A on spindle microtubules (MTs), affected centrosome integrity, spindle formation and length, as well as MT dynamics in interphase, consistent with Aurora A inhibition by RNAi or other specific inhibitors, such as MLN8054 or MLN8237. Interestingly, Tripolin A affected the gradient distribution towards the chromosomes, but not the MT binding of HURP (Hepatoma Up-Regulated Protein), a MT-associated protein (MAP) and substrate of the Aurora A kinase. Therefore Tripolin A reveals a new way of regulating mitotic MT stabilizers through Aurora A phosphorylation. Tripolin A is predicted to bind Aurora A similarly but not identical to MLN8054, therefore it could be used to dissect pathways orchestrated by Aurora kinases as well as a scaffold for further inhibitor development.

  9. Benzimidazole-Based Quinazolines: In Vitro Evaluation, Quantitative Structure-Activity Relationship, and Molecular Modeling as Aurora Kinase Inhibitors.

    Science.gov (United States)

    Sharma, Alka; Luxami, Vijay; Saxena, Sanjai; Paul, Kamaldeep

    2016-03-01

    A series of benzimidazole-based quinazoline derivatives with different substitutions of primary and secondary amines at the C2 position (1-12) were evaluated for their Aurora kinase inhibitory activities. All compounds except for 3 and 6 showed good activity against Aurora kinase inhibitors, with IC50 values in the range of 0.035-0.532 μM. The ligand efficiency (LE) of the compounds with Aurora A kinase was also determined. The structure-activity relationship and the quantitative structure-activity relationship revealed that the Aurora inhibitory activities of these derivatives primarily depend on the different substitutions of the amine present at the C2 position of the quinazoline core. Molecular docking studies in the active binding site also provided theoretical support for the experimental biological data acquired. The current study identifies a novel class of Aurora kinase inhibitors, which can further be used for the treatment of cancer.

  10. Discovery of novel inhibitors of Aurora kinases with indazole scaffold: In silico fragment-based and knowledge-based drug design.

    Science.gov (United States)

    Chang, Chun-Feng; Lin, Wen-Hsing; Ke, Yi-Yu; Lin, Yih-Shyan; Wang, Wen-Chieh; Chen, Chun-Hwa; Kuo, Po-Chu; Hsu, John T A; Uang, Biing-Jiun; Hsieh, Hsing-Pang

    2016-11-29

    Aurora kinases have emerged as important anticancer targets so that there are several inhibitors have advanced into clinical study. Herein, we identified novel indazole derivatives as potent Aurora kinases inhibitors by utilizing in silico fragment-based approach and knowledge-based drug design. After intensive hit-to-lead optimization, compounds 17 (dual Aurora A and B), 21 (Aurora B selective) and 30 (Aurora A selective) possessed indazole privileged scaffold with different substituents, which provide sub-type kinase selectivity. Computational modeling helps in understanding that the isoform selectivity could be targeted specific residue in the Aurora kinase binding pocket in particular targeting residues Arg220, Thr217 or Glu177.

  11. Design and synthesis of novel benzoxazole analogs as Aurora B kinase inhibitors.

    Science.gov (United States)

    An, Ying; Lee, Eun; Yu, Yeongji; Yun, Jieun; Lee, Myeong Youl; Kang, Jong Soon; Kim, Woo-Young; Jeon, Raok

    2016-07-01

    A novel series of benzoxazole analogs was designed and synthesized, and their inhibitory activities against Aurora kinases were evaluated. Some of the tested compounds exhibited a promising activity with respect to the inhibition of Aurora B kinase. A structure-activity relationship study indicated that linker length, regiochemistry, and halogen substitution play important roles in kinase inhibitory potency. The binding modes between representative compounds and Aurora kinases were interpreted through a molecular docking study to explain the inhibitory activity and selectivity for Aurora A and B kinases. Compounds 13l and 13q also show an antiproliferative effect on the human tumor cell lines in a dose-dependent manner. The most potent 13q demonstrated good efficacy in the prostate cancer PC-3 tumor xenograft model.

  12. A comparative study of the aneugenic and polyploidy-inducing effects of fisetin and two model Aurora kinase inhibitors.

    Science.gov (United States)

    Gollapudi, P; Hasegawa, L S; Eastmond, D A

    2014-06-01

    Fisetin, a plant flavonol commonly found in fruits, nuts and vegetables, is frequently added to nutritional supplements due to its reported cardioprotective, anti-carcinogenic and antioxidant properties. Earlier reports from our laboratory and others have indicated that fisetin has both aneugenic and clastogenic properties in cultured cells. More recently, fisetin has also been reported to target Aurora B kinase, a Ser/Thr kinase involved in ensuring proper microtubule attachment at the spindle assembly checkpoint, and an enzyme that is overexpressed in several types of cancer. Here we have further characterized the chromosome damage caused by fisetin and compared it with that induced by two known Aurora kinase inhibitors, VX-680 and ZM-447439, in cultured TK6 cells using the micronucleus assay with CREST staining as well as a flow cytometry-based assay that measures multiple types of numerical chromosomal aberrations. The three compounds were highly effective in inducing aneuploidy and polyploidy as evidenced by increases in kinetochore-positive micronuclei, hyperdiploidy, and polyploidy. With fisetin, however, the latter two effects were most significantly observed only after cells were allowed to overcome a cell cycle delay, and occurred at higher concentrations than those induced by the other Aurora kinase inhibitors. Modest increases in kinetochore-negative micronuclei were also seen with the model Aurora kinase inhibitors. These results indicate that fisetin induces multiple types of chromosome abnormalities in human cells, and indicate a need for a thorough investigation of fisetin-augmented dietary supplements.

  13. Dithiocarbamate promoted practical synthesis of N-Aryl-2-aminobenzazoles: Synthesis of novel Aurora-A kinase inhibitor

    Indian Academy of Sciences (India)

    Naresh Kumar Katari; M Venkatanarayana; Kummari Srinivas

    2015-03-01

    Various N-aryl-2-aminobenzoxazoles and N-aryl-2-aminobenzothiazoles were synthesized from o-aminophenol and o-aminothiophenol, respectively, mediated by dithiocarbamate in one step. The salient features of this method include mild reaction condition, high yield and large scale synthesis. Application of this methodology has been demonstrated by synthesizing potent Aurora kinase-A inhibitors.

  14. A kinase inhibitor screen identifies Mcl-1 and Aurora kinase A as novel treatment targets in antiestrogen-resistant breast cancer cells

    DEFF Research Database (Denmark)

    Thrane, S; Pedersen, A M; Thomsen, M B H;

    2015-01-01

    of the anti-apoptotic protein Mcl-1. Mcl-1 expression was found upregulated in the antiestrogen-resistant cell lines and depletion of Mcl-1 in resistant cells caused decreased viability. JNJ-7706621, a dual Aurora kinase and cyclin-dependent kinase inhibitor, specifically inhibited growth and caused G2 phase...

  15. Activity of the Aurora kinase inhibitor VX-680 against Bcr/Abl-positive acute lymphoblastic leukemias.

    Science.gov (United States)

    Fei, Fei; Stoddart, Sonia; Groffen, John; Heisterkamp, Nora

    2010-05-01

    The emergence of resistance to tyrosine kinase inhibitors due to point mutations in Bcr/Abl is a challenging problem for Philadelphia chromosome-positive (Ph-positive) acute lymphoblastic leukemia (ALL) patients, especially for those with the T315I mutation, against which neither nilotinib or dasatinib shows significant activity. VX-680 is a pan-Aurora kinase inhibitor active against all Bcr/Abl proteins but has not been extensively examined in preclinical models of Ph-positive ALL. Here, we have tested VX-680 for the treatment of Bcr/Abl-positive ALL when leukemic cells are protected by the presence of stroma. Under these conditions, VX-680 showed significant effects on primary human Ph-positive ALL cells both with and without the T315I mutation, including ablation of tyrosine phosphorylation downstream of Bcr/Abl, decreased viability, and induction of apoptosis. However, drug treatment of human Ph-positive ALL cells for 3 days followed by drug removal allowed the outgrowth of abnormal cells 21 days later, and on culture of mouse Bcr/Abl ALL cells on stroma with lower concentrations of VX-680, drug-resistant cells emerged. Combined treatment of human ALL cells lacking the T315I mutation with both VX-680 and dasatinib caused significantly more cytotoxicity than each drug alone. We suggest that use of VX-680 together with a second effective drug as first-line treatment for Ph-positive ALL is likely to be safer and more useful than second-line treatment with VX-680 as monotherapy for drug-resistant T315I Ph-positive ALL.

  16. Effect of Human Flavin-Containing Monooxygenase 3 Polymorphism on the Metabolism of Aurora Kinase Inhibitors

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    Sheila J. Sadeghi

    2013-01-01

    Full Text Available Aurora kinases were recently identified as a potential target in anticancer therapy and, amongst their available inhibitors, Tozasertib (VX-680 and Danusertib (PHA-739358 have been indicated as possible substrates of human flavin-containing monooxygenase 3 (hFMO3. Here we report the in vitro rate of oxidation of these drugs by wild-type hFMO3 and its polymorphic variant V257M. The conversion of Tozasertib and Danusertib to their corresponding metabolites, identified by LC-MS, by the purified wild-type and V257M hFMO3 show significant differences. In the case of Tozasertib, the V257M variant shows a catalytic efficiency, expressed as kcat/Km, similar to the wild-type: 0.39 ± 0.06 min−1µM−1 for V257M compared to 0.33 ± 0.04 min−1µM−1 for the wild type. On the other hand, in the case of Danusertib, V257M shows a 3.4× decrease in catalytic efficiency with kcat/Km values of 0.05 ± 0.01 min−1µM−1 for V257M and 0.17 ± 0.03 min−1µM−1 for the wild type. These data reveal how a simple V257M substitution ascribed to a single nucleotide polymorphism affects the N-oxidation of relevant anticancer drugs, with important outcome in their therapeutic effects. These findings demonstrate that codon 257 is important for activity of the hFMO3 gene and the codon change V to M has an effect on the catalytic efficiency of this enzyme.

  17. Phenotypic screening approaches to develop Aurora kinase inhibitors: Drug Discovery perspectives

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    Carlos eMarugán

    2016-01-01

    Full Text Available Targeting mitotic regulators as a strategy to fight cancer implies the development of drugs against key proteins such as Aurora A and B. Current drugs which target mitosis through a general mechanism of action (stabilization/destabilization of microtubules, have several side effects (neutropenia, alopecia, emesis. Pharmaceutical companies aim at avoiding these unwanted effects by generating improved and selective drugs that increase the quality of life of the patients. However, the development of these drugs is an ambitious task that involves testing thousands of compounds through biochemical and cell-based assays. In addition, molecules usually target complex biological processes, involving several proteins and different molecular pathways, further emphasizing the need for high-throughput screening techniques and multiplexing technologies in order to identify drugs with the desired phenotype.We will briefly describe two multiplexing technologies (high-content imaging, microarrays and flow cytometry and two key processes for drug discovery research (assay development and validation following our own published industry quality standards. We will further focus on high-content imaging as a useful tool for phenotypic screening and will provide a concrete example of high-content imaging assay to detect Aurora A or B selective inhibitors discriminating the off-target effects related to inhibition of other cell cycle or non-cell cycle key regulators. Finally, we will describe other assays that can help to characterize the in vitro pharmacology of the inhibitors.

  18. Aurora kinase inhibitors attached to iron oxide nanoparticles enhances inhibition of the growth of liver cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Xiquan [Southeast University, State Key Laboratory of Bioelectronics and Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Science & Medical Engineering (China); Xie, Li [Southeast University, Zhongda Hospital, School of Medicine (China); Zheng, Ming; Yao, Juan [Jiangsu Chai Tai Tianqing Pharmaceutical Co. Ltd. (China); Song, Lina [Southeast University, State Key Laboratory of Bioelectronics and Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Science & Medical Engineering (China); Chang, Weiwei [Jiangsu Chai Tai Tianqing Pharmaceutical Co. Ltd. (China); Zhang, Yu; Ji, Min, E-mail: minji888@hotmail.com; Gu, Ning, E-mail: guning@seu.edu.cn [Southeast University, State Key Laboratory of Bioelectronics and Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Science & Medical Engineering (China); Zhan, Xi, E-mail: zhan01@gmail.com [University of Maryland School of Medicine, The Center of Vascular and Inflammatory Diseases, The Department of Pathology (United States)

    2015-06-15

    We have developed a novel Aurora kinase inhibitor (AKI) AM-005, an analogue of pan-AKI AT-9283. To improve the intracellular efficacy of AM-005 and AT-9283, we utilized magnetite nanoparticles (NPs) to deliver AM-005 and AT-9283 into human SMMC-7721 and HepG2 liver cancer cells. The drug-loaded NPs were prepared through quasi-emulsion solvent diffusion of magnetite NPs with AM-005 or AT-9283. The encapsulated drugs were readily released from NPs, preferentially at low pHs. Upon exposure, cancer cells effectively internalized drug-loaded NPs into lysosome-like vesicles, which triggered a series of cellular changes, including the formation of enlarged cytoplasm, the significant increase of membrane permeability, and the generation of reactive oxygen species (ROS). The increased ROS synthesis sustained over 72 h, whereas that in the cells treated with free-form drugs declined rapidly after 48 h. However, chemical sequestration of the iron core of NPs had a minor influence on the generation of intracellular ROS. On the other hand, uncoupling of AM-005 uptake with NP internalization into cells failed to induce ROS synthesis. Overall, our approach achieved two-fold increase in suppressing the viability of tumor cells in vitro and the growth of tumors in vivo. We conclude that magnetite NPs can be used as pH responsive nanocarriers that are able to improve the efficacy of AKIs.

  19. An exploratory phase 2 study of investigational Aurora A kinase inhibitor alisertib (MLN8237 in acute myelogenous leukemia and myelodysplastic syndromes

    Directory of Open Access Journals (Sweden)

    Stuart L. Goldberg

    2014-01-01

    Full Text Available Alisertib (MLN8237 is an investigational, oral, selective, Aurora A kinase (AAK inhibitor. In this phase 2 trial, 57 patients with acute myeloid leukemia (AML or high-grade myelodysplastic syndrome received alisertib 50 mg BID for 7 days in 21-day cycles. Responses in 6/35 AML patients (17% response rate with an additional 49% stable disease, 34% transfusion independence included 1 complete response lasting >1 year. No responses were observed in MDS patients. Adverse events >30% included diarrhea, fatigue, nausea, febrile neutropenia, and stomatitis. Results suggest modest activity in AML, supporting further research to better understand how AAK inhibition may induce leukemic cell senescence.

  20. Update on Aurora Kinase Targeted Therapeutics in Oncology

    Science.gov (United States)

    Green, Myke R.; Woolery, Joseph E.

    2011-01-01

    Introduction Mammalian cells contain three distinct serine/threonine protein kinases with highly conserved catalytic domains, including aurora A and B kinases that are essential regulators of mitotic entry and progression. Overexpression of aurora A and/or B kinase is associated with high proliferation rates and poor prognosis, making them ideal targets for anti-cancer therapy. Disruption of mitotic machinery is a proven anti-cancer strategy employed by multiple chemotherapeutic agents. Numerous small molecule inhibitors of the aurora kinases have been discovered and tested in vivo and in vitro, with a few currently in phase II testing. Areas covered This review provides the reader with updated results from both preclinical and human studies for each of the aurora kinase inhibitors (AKI) that are currently being investigated. The paper also covers in detail the late breaking and phase I data presented for AKIs thereby allowing the reader to compare and contrast individual and classrelated effects of AKIs. Expert opinion While the successful development and approval of an AKI for anti-cancer therapy remains unresolved, pre-clinical identification of resistant mechanisms would help design better early phase clinical trials where relevant combinations may be evaluated prior to phase II testing. The authors believe that aurora kinases are important anti-cancer targets that operate in collaboration with other oncogenes intimately involved in uncontrolled tumor proliferation and by providing a unique, targeted and complimentary anti-cancer mechanism, expand the available armamentarium against cancer. PMID:21556291

  1. Combining the pan-aurora kinase inhibitor AMG 900 with histone deacetylase inhibitors enhances antitumor activity in prostate cancer

    NARCIS (Netherlands)

    Paller, C.J.; Wissing, M.D.; Mendonca, J.; Sharma, A.; Kim, E.; Kim, H.S.; Kortenhorst, M.S.Q.; Gerber, S.; Rosen, M.; Shaikh, F.; Zahurak, M.L.; Rudek, M.A.; Hammers, H.; Rudin, C.M.; Carducci, M.A.; Kachhap, S.K.

    2014-01-01

    Histone deacetylase inhibitors (HDACIs) are being tested in clinical trials for the treatment of solid tumors. While most studies have focused on the reexpression of silenced tumor suppressor genes, a number of genes/pathways are downregulated by HDACIs. This provides opportunities for combination t

  2. New Treatment of Medullary and Papillary Human Thyroid Cancer: Biological Effects of Hyaluronic Acid Hydrogel Loaded With Quercetin Alone or in Combination to an Inhibitor of Aurora Kinase.

    Science.gov (United States)

    Quagliariello, Vincenzo; Armenia, Emilia; Aurilio, Caterina; Rosso, Francesco; Clemente, Ottavia; de Sena, Gabriele; Barbarisi, Manlio; Barbarisi, Alfonso

    2016-08-01

    The aim of this paper is based on the use of a hyaluronic acid hydrogel of Quercetin tested alone and in combination to an inhibitor of Aurora Kinase type A and B (SNS-314) on human medullary and papillary thyroid cancer cells. Biological investigations were focused on the cellular uptake of the hydrogel, cell viability, antioxidant, and cytokines secretion studies. Quercetin delivered from hydrogel show a time and CD44 dependent interaction with both cell lines with significant anti-inflammatory effects. Combination of Quercetin and SNS-314 leads to a synergistic cytotoxic effect on medullary TT and papillary BCPAP cell lines with a significant reduction of the IC50 value. These results, highlights the importance of synergistic effect of the hyaluronic acid hydrogel of Quercetin with SNS-314 in the regulation of human thyroid cancer cell proliferation and emphasize the anti-tumor activity of these molecules. J. Cell. Physiol. 231: 1784-1795, 2016. © 2015 Wiley Periodicals, Inc.

  3. The aurora kinase inhibitor VX-680 shows anti-cancer effects in primary metastatic cells and the SW13 cell line.

    Science.gov (United States)

    Pezzani, Raffaele; Rubin, Beatrice; Bertazza, Loris; Redaelli, Marco; Barollo, Susi; Monticelli, Halenya; Baldini, Enke; Mian, Caterina; Mucignat, Carla; Scaroni, Carla; Mantero, Franco; Ulisse, Salvatore; Iacobone, Maurizio; Boscaro, Marco

    2016-10-01

    New therapeutic targets are needed to fight cancer. Aurora kinases (AK) were recently identified as vital key regulators of cell mitosis and have consequently been investigated as therapeutic targets in preclinical and clinical studies. Aurora kinase inhibitors (AKI) have been studied in many cancer types, but their potential capacity to limit or delay metastases has rarely been considered, and never in adrenal tissue. Given the lack of an effective pharmacological therapy for adrenal metastasis and adrenocortical carcinoma, we assessed AKI (VX-680, SNS314, ZM447439) in 2 cell lines (H295R and SW13 cells), 3 cell cultures of primary adrenocortical metastases (from lung cancer), and 4 primary adrenocortical tumor cell cultures. We also tested reversan, which is a P-gp inhibitor (a fundamental efflux pump that can extrude drugs), and we measured AK expression levels in 66 adrenocortical tumor tissue samples. Biomolecular and cellular tests were performed (such as MTT, thymidine assay, Wright's staining, cell cycle and apoptosis analysis, Western blot, qRT-PCR, and mutation analysis). Our results are the first to document AK overexpression in adrenocortical carcinoma as well as in H295R and SW13 cell lines, thus proving the efficacy of AKI against adrenal metastases and in the SW13 cancer cell model. We also demonstrated that reversan and AKI Vx-680 are useless in the H295R cell model, and therefore should not be considered as potential treatments for ACC. Serine/threonine AK inhibition, essentially with VX-680, could be a promising, specific therapeutic tool for eradicating metastases in adrenocortical tissue.

  4. Aurora A kinase expression is increased in leukemia stem cells, and a selective Aurora A kinase inhibitor enhances Ara-C-induced apoptosis in acute myeloid leukemia stem cells

    OpenAIRE

    2012-01-01

    Background The overexpression of Aurora A kinase (AurA) has been reported in various malignancies, including acute myeloid leukemia (AML). However, the expression of AurA and the effects of AurA inhibition in cancer stem cells are not yet fully understood. We investigated the expression and inhibition of AurA in AML stem cells (CD34+/CD38-). Methods Expression of AurA was investigated in cell lines (NB4 and KG1) that express high levels of CD34 and low levels of CD38. Primary AML cells were h...

  5. Design, synthesis, quantum chemical studies and biological activity evaluation of pyrazole-benzimidazole derivatives as potent Aurora A/B kinase inhibitors.

    Science.gov (United States)

    Zheng, Youguang; Zheng, Ming; Ling, Xin; Liu, Yi; Xue, Yunsheng; An, Lin; Gu, Ning; Ji, Min; Jin, Min

    2013-06-15

    Novel pyrazole-benzimidazole derivatives have been designed and synthesized. The entire target compounds were determined against cancer cell lines U937, K562, A549, LoVo and HT29 and were screened for Aurora A/B kinase inhibitory activity in vitro. The compounds 7a, 7b, 7i, 7k and 7l demonstrated significant cancer cell lines and Aurora A/B kinase inhibitory activities. Molecular modeling studies suggested the derivatives have bound in the active site of Aurora A kinase through the formation of four hydrogen bonds. Quantum chemical studies were carried out on these compounds to understand the structural features essential for activity. The cellular activity of 7k was also tested by immunofluorescence.

  6. Basal aurora kinase B activity is sufficient for histone H3 phosphorylation in prophase

    Directory of Open Access Journals (Sweden)

    Ly-Thuy-Tram Le

    2013-02-01

    Histone H3 phosphorylation is the hallmark of mitosis deposited by aurora kinase B. Benzo[e]pyridoindoles are a family of potent, broad, ATP-competitive aurora kinase inhibitors. However, benzo[e]pyridoindole C4 only inhibits histone H3 phosphorylation in prophase but not in metaphase. Under the C4 treatment, the cells enter into mitosis with dephosphorylated histone H3, assemble chromosomes normally and progress to metaphase, and then to anaphase. C4 also induces lagging chromosome in anaphase but we demonstrated that these chromosome compaction defects are not related to the absence of H3 phosphorylation in prophase. As a result of C4 action, mitosis lasts longer and the cell cycle is slowed down. We reproduced the mitotic defects with reduced concentrations of potent pan aurora kinase as well as with a specific aurora B ATP-competitive inhibitor; we therefore propose that histone H3 phosphorylation and anaphase chromosome compaction involve the basal activity of aurora kinase B. Our data suggest that aurora kinase B is progressively activated at mitosis entry and at anaphase onset. The full activation of aurora kinase B by its partners, in prometaphase, induces a shift in the catalytic domain of aurora B that modifies its affinity for ATP. These waves of activation/deactivation of aurora B correspond to different conformations of the chromosomal complex revealed by FRAP. The presence of lagging chromosomes may have deleterious consequences on the daughter cells and, unfortunately, the situation may be encountered in patients receiving treatment with aurora kinase inhibitors.

  7. Alisertib, an Aurora kinase A inhibitor, induces apoptosis and autophagy but inhibits epithelial to mesenchymal transition in human epithelial ovarian cancer cells

    Directory of Open Access Journals (Sweden)

    Ding YH

    2015-01-01

    Full Text Available Yong-Hui Ding,1,2 Zhi-Wei Zhou,2,3 Chun-Fang Ha,1 Xue-Yu Zhang,1 Shu-Ting Pan,4 Zhi-Xu He,3 Jeffrey L Edelman,2 Dong Wang,5 Yin-Xue Yang,6 Xueji Zhang,7 Wei Duan,8 Tianxin Yang,9 Jia-Xuan Qiu,4 Shu-Feng Zhou2,3 1Department of Gynecology, General Hospital of Ningxia Medical University, Yinchuan, People’s Republic of China; 2Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA ; 3Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, 4Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, 5Cancer Center, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, 6Department of Colorectal Surgery, General Hospital of Ningxia Medical University, Yinchuan, 7Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 8School of Medicine, Deakin University, Waurn Ponds, Australia; 9Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA Abstract: Ovarian cancer is a leading killer of women, and no cure for advanced ovarian cancer is available. Alisertib (ALS, a selective Aurora kinase A (AURKA inhibitor, has shown potent anticancer effects, and is under clinical investigation for the treatment of advanced solid tumor and hematologic malignancies. However, the role of ALS in the treatment of ovarian cancer remains unclear. This study investigated the effects of ALS on cell growth, apoptosis, autophagy, and epithelial to mesenchymal transition (EMT, and the underlying mechanisms in human epithelial ovarian cancer SKOV3 and OVCAR4 cells. Our docking study showed that ALS, MLN8054, and VX-680 preferentially bound to AURKA

  8. p53 deficiency enhances mitotic arrest and slippage induced by pharmacological inhibition of Aurora kinases.

    Science.gov (United States)

    Marxer, M; Ma, H T; Man, W Y; Poon, R Y C

    2014-07-01

    A number of small-molecule inhibitors of Aurora kinases have been developed and are undergoing clinical trials for anti-cancer therapies. Different Aurora kinases, however, behave as very different targets: while inhibition of Aurora A (AURKA) induces a delay in mitotic exit, inhibition of Aurora B (AURKB) triggers mitotic slippage. Furthermore, while it is evident that p53 is regulated by Aurora kinase-dependent phosphorylation, how p53 may in turn regulate Aurora kinases remains mysterious. To address these issues, isogenic p53-containing and -negative cells were exposed to classic inhibitors that target both AURKA and AURKB (Alisertib and ZM447439), as well as to new generation of inhibitors that target AURKA (MK-5108), AURKB (Barasertib) individually. The fate of individual cells was then tracked with time-lapse microscopy. Remarkably, loss of p53, either by gene disruption or small interfering RNA-mediated depletion, sensitized cells to inhibition of both AURKA and AURKB, promoting mitotic arrest and slippage respectively. As the p53-dependent post-mitotic checkpoint is also important for preventing genome reduplication after mitotic slippage, these studies indicate that the loss of p53 in cancer cells represents a major opportunity for anti-cancer drugs targeting the Aurora kinases.

  9. The pan-inhibitor of Aurora kinases danusertib induces apoptosis and autophagy and suppresses epithelial-to-mesenchymal transition in human breast cancer cells.

    Science.gov (United States)

    Li, Jin-Ping; Yang, Yin-Xue; Liu, Qi-Lun; Zhou, Zhi-Wei; Pan, Shu-Ting; He, Zhi-Xu; Zhang, Xueji; Yang, Tianxin; Pan, Si-Yuan; Duan, Wei; He, Shu-Ming; Chen, Xiao-Wu; Qiu, Jia-Xuan; Zhou, Shu-Feng

    2015-01-01

    Danusertib (Danu) is a pan-inhibitor of Aurora kinases and a third-generation breakpoint cluster region-Abelson murine leukemia viral oncogene homolog 1 (Bcr-Abl) tyrosine kinase inhibitor, but its antitumor effect and underlying mechanisms in the treatment of human breast cancer remain elusive. This study aimed to investigate the effects of Danu on the growth, apoptosis, autophagy, and epithelial-to-mesenchymal transition (EMT) and the molecular mechanisms in human breast cancer MCF7 and MDA-MB-231 cells. The results demonstrated that Danu remarkably inhibited cell proliferation, induced apoptosis and autophagy, and suppressed EMT in both breast cancer cell lines. Danu arrested MCF7 and MDA-MB-231 cells in G2/M phase, accompanied by the downregulation of cyclin-dependent kinase 1 and cyclin B1 and upregulation of p21 Waf1/Cip1, p27 Kip1, and p53. Danu significantly decreased the expression of B-cell lymphoma-extra-large (Bcl-xl) and B-cell lymphoma 2 (Bcl-2), but increased the expression of Bcl-2-associated X protein (Bax) and p53-upregulated modulator of apoptosis (PUMA), and promoted the cleavage of caspases 3 and 9. Furthermore, Danu significantly increased the expression levels of the membrane-bound microtubule-associated protein 1A/1B-light chain 3 (LC3-II) and beclin 1 in breast cancer cells, two markers for autophagy. Danu induced the activation of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinases 1 and 2 (Erk1/2) and inhibited the activation of protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathways in breast cancer cells. Treatment with wortmannin (a phosphatidylinositol 3-kinase inhibitor) markedly inhibited Danu-induced activation of p38 MAPK and conversion of cytosolic LC3-I to membrane-bound LC3-II. Pharmacological inhibition and small interfering RNA-mediated knockdown of p38 MAPK suppressed Akt activation, resulting in LC3-II accumulation and enhanced autophagy. Pharmacological inhibition

  10. Identification of Ustilago maydis Aurora kinase as a novel antifungal target.

    Science.gov (United States)

    Tückmantel, Sandra; Greul, Jörg N; Janning, Petra; Brockmeyer, Andreas; Grütter, Christian; Simard, Jeffrey R; Gutbrod, Oliver; Beck, Michael E; Tietjen, Klaus; Rauh, Daniel; Schreier, Peter H

    2011-09-16

    Infestation of crops by pathogenic fungi has continued to have a major impact by reducing yield and quality, emphasizing the need to identify new targets and develop new agents to improve methods of crop protection. Here we present Aurora kinase from the phytopathogenic fungus Ustilago maydis as a novel target for N-substituted diaminopyrimidines, a class of small-molecule kinase inhibitors. We show that Aurora kinase is essential in U. maydis and that diaminopyrimidines inhibit its activity in vitro. Furthermore, we observed an overall good correlation between in vitro inhibition of Aurora kinase and growth inhibition of diverse fungi in vivo. In vitro inhibition assays with Ustilago and human Aurora kinases indicate that some compounds of the N-substituted diaminopyrimidine class show specificity for the Ustilago enzyme, thus revealing their potential as selective fungicides.

  11. Specificity Rendering ‘Hot-Spots’ for Aurora Kinase Inhibitor Design: The Role of Non-Covalent Interactions and Conformational Transitions

    Science.gov (United States)

    Badrinarayan, Preethi; Sastry, G. Narahari

    2014-01-01

    The present study examines the conformational transitions occurring among the major structural motifs of Aurora kinase (AK) concomitant with the DFG-flip and deciphers the role of non-covalent interactions in rendering specificity. Multiple sequence alignment, docking and structural analysis of a repertoire of 56 crystal structures of AK from Protein Data Bank (PDB) has been carried out. The crystal structures were systematically categorized based on the conformational disposition of the DFG-loop [in (DI) 42, out (DO) 5 and out-up (DOU) 9], G-loop [extended (GE) 53 and folded (GF) 3] and αC-helix [in (CI) 42 and out (CO) 14]. The overlapping subsets on categorization show the inter-dependency among structural motifs. Therefore, the four distinct possibilities a) 2W1C (DI, CI, GE) b) 3E5A (DI, CI, GF) c) 3DJ6 (DI, CO, GF) d) 3UNZ (DOU, CO, GF) along with their co-crystals and apo-forms were subjected to molecular dynamics simulations of 40 ns each to evaluate the variations of individual residues and their impact on forming interactions. The non-covalent interactions formed by the 157 AK co-crystals with different regions of the binding site were initially studied with the docked complexes and structure interaction fingerprints. The frequency of the most prominent interactions was gauged in the AK inhibitors from PDB and the four representative conformations during 40 ns. Based on this study, seven major non-covalent interactions and their complementary sites in AK capable of rendering specificity have been prioritized for the design of different classes of inhibitors. PMID:25485544

  12. Allosteric inhibition of Aurora-A kinase by a synthetic vNAR domain.

    Science.gov (United States)

    Burgess, Selena G; Oleksy, Arkadiusz; Cavazza, Tommaso; Richards, Mark W; Vernos, Isabelle; Matthews, David; Bayliss, Richard

    2016-07-01

    The vast majority of clinically approved protein kinase inhibitors target the ATP-binding pocket directly. Consequently, many inhibitors have broad selectivity profiles and most have significant off-target effects. Allosteric inhibitors are generally more selective, but are difficult to identify because allosteric binding sites are often unknown or poorly characterized. Aurora-A is activated through binding of TPX2 to an allosteric site on the kinase catalytic domain, and this knowledge could be exploited to generate an inhibitor. Here, we generated an allosteric inhibitor of Aurora-A kinase based on a synthetic, vNAR single domain scaffold, vNAR-D01. Biochemical studies and a crystal structure of the Aurora-A/vNAR-D01 complex show that the vNAR domain overlaps with the TPX2 binding site. In contrast with the binding of TPX2, which stabilizes an active conformation of the kinase, binding of the vNAR domain stabilizes an inactive conformation, in which the αC-helix is distorted, the canonical Lys-Glu salt bridge is broken and the regulatory (R-) spine is disrupted by an additional hydrophobic side chain from the activation loop. These studies illustrate how single domain antibodies can be used to characterize the regulatory mechanisms of kinases and provide a rational basis for structure-guided design of allosteric Aurora-A kinase inhibitors.

  13. SMARCA4-inactivating mutations increase sensitivity to Aurora kinase A inhibitor VX-680 in non-small cell lung cancers

    Science.gov (United States)

    Tagal, Vural; Wei, Shuguang; Zhang, Wei; Brekken, Rolf A.; Posner, Bruce A.; Peyton, Michael; Girard, Luc; Hwang, TaeHyun; Wheeler, David A.; Minna, John D.; White, Michael A.; Gazdar, Adi F.; Roth, Michael G.

    2017-01-01

    Mutations in the SMARCA4/BRG1 gene resulting in complete loss of its protein (BRG1) occur frequently in non-small cell lung cancer (NSCLC) cells. Currently, no single therapeutic agent has been identified as synthetically lethal with SMARCA4/BRG1 loss. We identify AURKA activity as essential in NSCLC cells lacking SMARCA4/BRG1. In these cells, RNAi-mediated depletion or chemical inhibition of AURKA induces apoptosis and cell death in vitro and in xenograft mouse models. Disc large homologue-associated protein 5 (HURP/DLGAP5), required for AURKA-dependent, centrosome-independent mitotic spindle assembly is essential for the survival and proliferation of SMARCA4/BRG1 mutant but not of SMARCA4/BRG1 wild-type cells. AURKA inhibitors may provide a therapeutic strategy for biomarker-driven clinical studies to treat the NSCLCs harbouring SMARCA4/BRG1-inactivating mutations. PMID:28102363

  14. The pan-inhibitor of Aurora kinases danusertib induces apoptosis and autophagy and suppresses epithelial-to-mesenchymal transition in human breast cancer cells

    Directory of Open Access Journals (Sweden)

    Li JP

    2015-02-01

    Full Text Available Jin-Ping Li,1,2 Yin-Xue Yang,3 Qi-Lun Liu,1 Zhi-Wei Zhou,2,4 Shu-Ting Pan,2,5 Zhi-Xu He,4 Xueji Zhang,6 Tianxin Yang,7 Si-Yuan Pan,8 Wei Duan,9 Shu-Ming He,10 Xiao-Wu Chen,11 Jia-Xuan Qiu,5 Shu-Feng Zhou2,41Department of Surgical Oncology, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, People’s Republic of China; 2Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA; 3Department of Colorectal Surgery, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, People’s Republic of China; 4Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, Guizhou, People’s Republic of China; 5Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, People’s Republic of China; 6Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 7Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA; 8Department of Pharmacology, School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing, People’s Republic of China; 9School of Medicine, Deakin University, Waurn Ponds, VIC, Australia; 10Department of Obstetrics and Gynecology, Xiaolan People’s Hospital affiliated to Southern Medical University, Zhongshan, Guangdong, People’s Republic of China; 11Department of General Surgery, The First People’s Hospital of Shunde affiliated to Southern Medical University, Shunde, Foshan, Guangdong, People’s Republic of China Abstract: Danusertib (Danu is a pan-inhibitor of Aurora kinases and a third-generation breakpoint cluster region-Abelson murine leukemia viral oncogene homolog 1 (Bcr-Abl tyrosine kinase inhibitor

  15. Aurora kinases in childhood acute leukemia: The promise of aurora B as therapeutic target

    NARCIS (Netherlands)

    S.A. Hartsink-Segers (S.); C.M. Zwaan (Michel); C. Exalto (Carla); M.W.J. Luijendijk (M. W J); V. Calvert (V.); E.F. Petricoin (Emanuel F.); W.E. Evans (William); D. Reinhardt (Dirk); V. de Haas (Valerie); M. Hedtjärn (M.); B.R. Hansen (B.); T. Koch (T.); H.N. Caron (Huib); R. Pieters (Rob); M.L. den Boer (Monique)

    2013-01-01

    textabstractWe investigated the effects of targeting the mitotic regulators aurora kinase A and B in pediatric acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML). Aurora protein expression levels in pediatric ALL and AML patient samples were determined by western blot and reverse ph

  16. Therapeutic potential of mitotic interaction between the nucleoporin Tpr and aurora kinase A.

    Science.gov (United States)

    Kobayashi, Akiko; Hashizume, Chieko; Dowaki, Takayuki; Wong, Richard W

    2015-01-01

    Spindle poles are defined by centrosomes; therefore, an abnormal number or defective structural organization of centrosomes can lead to loss of spindle bipolarity and genetic integrity. Previously, we showed that Tpr (translocated promoter region), a component of the nuclear pore complex (NPC), interacts with Mad1 and dynein to promote proper chromosome segregation during mitosis. Tpr also associates with p53 to induce autophagy. Here, we report that Tpr depletion induces mitotic catastrophe and enhances the rate of tetraploidy and polyploidy. Mechanistically, Tpr interacts, via its central domain, with Aurora A but not Aurora B kinase. In Tpr-depleted cells, the expression levels, centrosomal localization and phosphorylation of Aurora A were all reduced. Surprisingly, an Aurora A inhibitor, Alisertib (MLN8237), also disrupted centrosomal localization of Tpr and induced mitotic catastrophe and cell death in a time- and dose-dependent manner. Strikingly, over-expression of Aurora A disrupted Tpr centrosomal localization only in cells with supernumerary centrosomes but not in bipolar cells. Our results highlight the mutual regulation between Tpr and Aurora A and further confirm the importance of nucleoporin function in spindle pole organization, bipolar spindle assembly, and mitosis; functions that are beyond the conventional nucleocytoplasmic transport and NPC structural roles of nucleoporins. Furthermore, the central coiled-coil domain of Tpr binds to and sequesters extra Aurora A to safeguard bipolarity. This Tpr domain merits further investigation for its ability to inhibit Aurora kinase and as a potential therapeutic agent in cancer treatment.

  17. Combined cisplatin and aurora inhibitor treatment increase neuroblastoma cell death but surviving cells overproduce BDNF.

    Science.gov (United States)

    Polacchini, Alessio; Albani, Clara; Baj, Gabriele; Colliva, Andrea; Carpinelli, Patrizia; Tongiorgi, Enrico

    2016-07-15

    Drug-resistance to chemotherapics in aggressive neuroblastoma (NB) is characterized by enhanced cell survival mediated by TrkB and its ligand, brain-derived neurotrophic factor (BDNF); thus reduction in BDNF levels represent a promising strategy to overcome drug-resistance, but how chemotherapics regulate BDNF is unknown. Here, cisplatin treatment in SK-N-BE neuroblastoma upregulated multiple BDNF transcripts, except exons 5 and 8 variants. Cisplatin increased BDNF mRNA and protein, and enhanced translation of a firefly reporter gene flanked by BDNF 5'UTR exons 1, 2c, 4 or 6 and 3'UTR-long. To block BDNF translation we focused on aurora kinases inhibitors which are proposed as new chemotherapeutics. NB cell survival after 24 h treatment was 43% with cisplatin, and 22% by cisplatin+aurora kinase inhibitor PHA-680632, while the aurora kinases inhibitor alone was less effective; however the combined treatment induced a paradoxical increase of BDNF in surviving cells with strong translational activation of exon6-3'UTR-long transcript, while translation of BDNF transcripts 1, 2C and 4 was suppressed. In conclusion, combined cisplatin and aurora kinase inhibitor treatment increases cell death, but induces BDNF overproduction in surviving cells through an aurora kinase-independent mechanism.

  18. Urochordate ascidians possess a single isoform of Aurora kinase that localizes to the midbody via TPX2 in eggs and cleavage stage embryos.

    Directory of Open Access Journals (Sweden)

    Celine Hebras

    Full Text Available Aurora kinases are key proteins found throughout the eukaryotes that control mitotic progression. Vertebrate Aurora-A and B kinases are thought to have evolved from a single Aurora-kinase isoform closest to that found in present day urochordates. In urochordate ascidians Aurora binds both TPX2 (a vertebrate AURKA partner and INCENP (a vertebrate AURKB partner and localizes to centrosomes and spindle microtubules as well as chromosomes and midbody during both meiosis and mitosis. Ascidian Aurora also displays this localization pattern during mitosis in echinoderms, strengthening the idea that non-vertebrate deuterostomes such as the urochordates and echinoderms possess a single form of Aurora kinase that has properties of vertebrate Aurora-kinase A and B. In the ascidian, TPX2 localizes to the centrosome and the spindle poles also as in vertebrates. However, we were surprised to find that TPX2 also localized strongly to the midbody in ascidian eggs and embryos. We thus examined more closely Aurora localization to the midbody by creating two separate point mutations of ascidian Aurora predicted to perturb binding to TPX2. Both forms of mutated Aurora behaved as predicted: neither localized to spindle poles where TPX2 is enriched. Interestingly, neither form of mutated Aurora localized to the midbody where TPX2 is also enriched, suggesting that ascidian Aurora midbody localization required TPX2 binding in ascidians. Functional analysis revealed that inhibition of Aurora kinase with a pharmacological inhibitor or with a dominant negative kinase dead form of Aurora caused cytokinesis failure and perturbed midbody formation during polar body extrusion. Our data support the view that vertebrate Aurora-A and B kinases evolved from a single non-vertebrate deuterostome ancestor. Moreover, since TPX2 localizes to the midbody in ascidian eggs and cleavage stage embryos it may be worthwhile re-assessing whether Aurora A kinase or TPX2 localize to the midbody

  19. Aurora kinase B is important for antiestrogen resistant cell growth and a potential biomarker for tamoxifen resistant breast cancer

    DEFF Research Database (Denmark)

    Larsen, Sarah L; Yde, Christina W; Laenkholm, Anne-Vibeke

    2015-01-01

    treatment targets. METHODS: Antiestrogen sensitive and resistant T47D breast cancer cell lines were used as model systems. Parental and fulvestrant resistant cell lines were subjected to a kinase inhibitor library. Kinase inhibitors preferentially targeting growth of fulvestrant resistant cells were...... for endocrine resistance, immunohistochemistry was performed on archival primary tumor tissue from breast cancer patients who have received adjuvant endocrine treatment with tamoxifen. RESULTS: The selective Aurora kinase B inhibitor barasertib was identified to preferentially inhibit growth of fulvestrant...... resistant T47D breast cancer cell lines. Compared with parental cells, phosphorylation of Aurora kinase B was higher in the fulvestrant resistant T47D cells. Barasertib induced degradation of Aurora kinase B, caused mitotic errors, and induced apoptotic cell death as measured by accumulation of SubG1 cells...

  20. A phase I trial of the aurora kinase inhibitor, ENMD-2076, in patients with relapsed or refractory acute myeloid leukemia or chronic myelomonocytic leukemia.

    Science.gov (United States)

    Yee, Karen W L; Chen, Hsiao-Wei T; Hedley, David W; Chow, Sue; Brandwein, Joseph; Schuh, Andre C; Schimmer, Aaron D; Gupta, Vikas; Sanfelice, Deborah; Johnson, Tara; Le, Lisa W; Arnott, Jamie; Bray, Mark R; Sidor, Carolyn; Minden, Mark D

    2016-10-01

    ENMD-2076 is a novel, orally-active molecule that inhibits Aurora A kinase, as well as c-Kit, FLT3 and VEGFR2. A phase I study was conducted to determine the maximum tolerated dose (MTD), recommended phase 2 dose (RP2D) and toxicities of ENMD-2076 in patients with acute myeloid leukemia (AML) and chronic myelomonocytic leukemia (CMML). Patients received escalating doses of ENMD-2076 administered orally daily [225 mg (n = 7), 375 mg (n = 6), 325 mg (n = 9), or 275 mg (n = 5)]. Twenty-seven patients were treated (26 AML; 1 CMML-2). The most common non-hematological toxicities of any grade, regardless of association with drug, were fatigue, diarrhea, dysphonia, dyspnea, hypertension, constipation, and abdominal pain. Dose-limiting toxicities (DLTs) consisted of grade 3 fatigue, grade 3 typhilitis, grade 3 syncope and grade 3 QTc prolongation). Of the 16 evaluable patients, one patient achieved a complete remission with incomplete count recovery (CRi), three experienced a morphologic leukemia-free state (MLFS) with a major hematologic improvement in platelets (HI-P), and 5 other patients had a reduction in marrow blast percentage (i.e. 11-65 %). The RP2D in this patient population is 225 mg orally once daily.

  1. Aurora kinase A controls meiosis I progression in mouse oocytes.

    Science.gov (United States)

    Saskova, Adela; Solc, Petr; Baran, Vladimir; Kubelka, Michal; Schultz, Richard M; Motlik, Jan

    2008-08-01

    Aurora kinase A (AURKA), which is a centrosome-localized serine/threonine kinase crucial for cell cycle control, is critically involved in centrosome maturation and spindle assembly in somatic cells. Active T288 phosphorylated AURKA localizes to the centrosome in the late G(2) and also spreads to the minus ends of mitotic spindle microtubules. AURKA activates centrosomal CDC25B and recruits cyclin B1 to centrosomes. We report here functions for AURKA in meiotic maturation of mouse oocytes, which is a model system to study the G(2) to M transition. Whereas AURKA is present throughout the entire GV-stage oocyte with a clear accumulation on microtubule organizing centers (MTOC), active AURKA becomes entirely localized to MTOCs shortly before germinal vesicle breakdown. In contrast to somatic cells in which active AURKA is present at the centrosomes and minus ends of microtubules, active AURKA is mainly located on MTOCs at metaphase I (MI) in oocytes. Inhibitor studies using Roscovitine (CDK1 inhibitor), LY-294002 (PI3K inhibitor) and SH-6 (PKB inhibitor) reveal that activation of AURKA localized on MTOCs is independent on PI3K-PKB and CDK1 signaling pathways and MOTC amplification is observed in roscovitine- and SH-6-treated oocytes that fail to undergo nuclear envelope breakdown. Moreover, microinjection of Aurka mRNA into GV-stage oocytes cultured in 3-isobutyl-1-methyl xanthine (IBMX)-containing medium to prevent maturation also results in MOTC amplification in the absence of CDK1 activation. Overexpression of AURKA also leads to formation of an abnormal MI spindle, whereas RNAi-mediated reduction of AURKA interferes with resumption of meiosis and spindle assembly. Results of these experiments indicate that AURKA is a critical MTOC-associated component involved in resumption of meiosis, MTOC multiplication, proper spindle formation and the metaphase I-metaphase II transition.

  2. Identification of Ski as a target for Aurora A kinase.

    Science.gov (United States)

    Mosquera, Jocelyn; Armisen, Ricardo; Zhao, Hongling; Rojas, Diego A; Maldonado, Edio; Tapia, Julio C; Colombo, Alicia; Hayman, Michael J; Marcelain, Katherine

    2011-06-10

    Ski is a negative regulator of the transforming growth factor-β and other signalling pathways. The absence of SKI in mouse fibroblasts leads to chromosome segregation defects and genomic instability, suggesting a role for Ski during mitosis. At this stage, Ski is phosphorylated but to date little is known about the kinases involved in this process. Here, we show that Aurora A kinase is able to phosphorylate Ski in vitro. In vivo, Aurora A and Ski co-localized at the centrosomes and co-immunoprecipitated. Conversely, a C-terminal truncation mutant of Ski (SkiΔ491-728) lacking a coiled-coil domain, displayed decreased centrosomal localization. This mutant no longer co-immunoprecipitated with Aurora-A in vivo, but was still phosphorylated in vitro, indicating that the Ski-Aurora A interaction takes place at the centrosomes. These data identify Ski as a novel target of Aurora A and contribute to an understanding of the role of these proteins in the mitotic process.

  3. Multifunctional human transcriptional coactivator protein PC4 is a substrate of Aurora kinases and activates the Aurora enzymes.

    Science.gov (United States)

    Dhanasekaran, Karthigeyan; Kumari, Sujata; Boopathi, Ramachandran; Shima, Hiroki; Swaminathan, Amrutha; Bachu, Mahesh; Ranga, Udaykumar; Igarashi, Kazuhiko; Kundu, Tapas K

    2016-03-01

    Positive coactivator 4 (PC4), a human transcriptional coactivator, is involved in diverse processes like chromatin organization and transcription regulation. It is hyperphosphorylated during mitosis, with unknown significance. For the first time, we demonstrate the function of PC4 outside the nucleus upon nuclear envelope breakdown. A fraction of PC4 associates with Aurora A and Aurora B and undergoes phosphorylation, following which PC4 activates both Aurora A and B to sustain optimal kinase activity to maintain the phosphorylation gradient for the proper functioning of the mitotic machinery. This mitotic role is evident in PC4 knockdown cells where the defects are rescued only by the catalytically active Aurora kinases, but not the kinase-dead mutants. Similarly, the PC4 phosphodeficient mutant failed to rescue such defects. Hence, our observations establish a novel mitotic function of PC4 that might be dependent on Aurora kinase-mediated phosphorylation.

  4. Facile identification of dual FLT3-Aurora A inhibitors: a computer-guided drug design approach.

    Science.gov (United States)

    Chang Hsu, Yung; Ke, Yi-Yu; Shiao, Hui-Yi; Lee, Chieh-Chien; Lin, Wen-Hsing; Chen, Chun-Hwa; Yen, Kuei-Jung; Hsu, John T-A; Chang, Chungming; Hsieh, Hsing-Pang

    2014-05-01

    Computer-guided drug design is a powerful tool for drug discovery. Herein we disclose the use of this approach for the discovery of dual FMS-like receptor tyrosine kinase-3 (FLT3)-Aurora A inhibitors against cancer. An Aurora hit compound was selected as a starting point, from which 288 virtual molecules were screened. Subsequently, some of these were synthesized and evaluated for their capacity to inhibit FLT3 and Aurora kinase A. To further enhance FLT3 inhibition, structure-activity relationship studies of the lead compound were conducted through a simplification strategy and bioisosteric replacement, followed by the use of computer-guided drug design to prioritize molecules bearing a variety of different terminal groups in terms of favorable binding energy. Selected compounds were then synthesized, and their bioactivity was evaluated. Of these, one novel inhibitor was found to exhibit excellent inhibition of FLT3 and Aurora kinase A and exert a dramatic antiproliferative effect on MOLM-13 and MV4-11 cells, with an IC50 value of 7 nM. Accordingly, it is considered a highly promising candidate for further development.

  5. A translational regulator, PUM2, promotes both protein stability and kinase activity of Aurora-A.

    Directory of Open Access Journals (Sweden)

    Yei-Hsuan Huang

    Full Text Available Aurora-A, a centrosomal serine-threonine kinase, orchestrates several key aspects of cell division. However, the regulatory pathways for the protein stability and kinase activity of Aurora-A are still not completely understood. In this study, PUM2, an RNA-binding protein, is identified as a novel substrate and interacting protein of Aurora-A. Overexpression of the PUM2 mutant which fails to interact with Aurora-A, and depletion of PUM2 result in a decrease in the amount of Aurora-A. PUM2 physically binds to the D-box of Aurora-A, which is recognized by APC/C(Cdh1. Overexpression of PUM2 prevents ubiquitination and enhances the protein stability of Aurora-A, suggesting that PUM2 protects Aurora-A from APC/C(Cdh1-mediated degradation. Moreover, association of PUM2 with Aurora-A not only makes Aurora-A more stable but also enhances the kinase activity of Aurora-A. Our study suggests that PUM2 plays two different but important roles during cell cycle progression. In interphase, PUM2 localizes in cytoplasm and plays as translational repressor through its RNA binding domain. However, in mitosis, PUM2 physically associates with Aurora-A to ensure enough active Aurora-A at centrosomes for mitotic entry. This is the first time to reveal the moonlight role of PUM2 in mitosis.

  6. Antitumor Activity of KW-2450 Against Triple-Negative Breast Cancer by Inhibiting Aurora A and B Kinases

    Science.gov (United States)

    Kai, Kazuharu; Kondo, Kimie; Wang, Xiaoping; Xie, Xuemei; Pitner, Mary K.; Reyes, Monica E.; Torres-Adorno, Angie M.; Masuda, Hiroko; Hortobagyi, Gabriel N.; Bartholomeusz, Chandra; Saya, Hideyuki; Tripathy, Debu; Sen, Subrata; Ueno, Naoto T.

    2015-01-01

    Currently, no targeted drug is available for triple-negative breast cancer (TNBC), an aggressive breast cancer that does not express estrogen receptor, progesterone receptor, or HER2. TNBC has high mitotic activity, and since Aurora A and B mitotic kinases drive cell division and are overexpressed in tumors with a high mitotic index, we hypothesized that inhibiting Aurora A and B produces a significant antitumor effect in TNBC. We tested this hypothesis by determining the antitumor effects of KW-2450, a multikinase inhibitor of both Aurora A and B kinases. We observed significant inhibitory activities of KW-2450 on cell viability, apoptosis, colony formation in agar, and mammosphere formation in TNBC cells. The growth of TNBC xenografts was significantly inhibited with KW-2450. In cell cycle analysis, KW-2450 induced tetraploid accumulation followed by apoptosis or surviving octaploid (8N) cells, depending on dose. These phenotypes resembled those of Aurora B knockdown and complete pharmaceutical inhibition of Aurora A. We demonstrated that 8N cells resulting from KW-2450 treatment depended on the activation of mitogen-activated protein kinase kinase (MEK) for their survival. When treated with the MEK inhibitor selumetinib combined with KW-2450, compared with KW-2450 alone, the 8N cell population was significantly reduced and apoptosis was increased. Indeed this combination showed synergistic antitumor effect in SUM149 TNBC xenografts. Collectively, Aurora A and B inhibition had a significant antitumor effect against TNBC, and this antitumor effect was maximized by the combination of selumetinib with Aurora A and B inhibition. PMID:26443806

  7. Antitumor Activity of KW-2450 against Triple-Negative Breast Cancer by Inhibiting Aurora A and B Kinases.

    Science.gov (United States)

    Kai, Kazuharu; Kondo, Kimie; Wang, Xiaoping; Xie, Xuemei; Pitner, Mary K; Reyes, Monica E; Torres-Adorno, Angie M; Masuda, Hiroko; Hortobagyi, Gabriel N; Bartholomeusz, Chandra; Saya, Hideyuki; Tripathy, Debu; Sen, Subrata; Ueno, Naoto T

    2015-12-01

    Currently, no targeted drug is available for triple-negative breast cancer (TNBC), an aggressive breast cancer that does not express estrogen receptor, progesterone receptor, or HER2. TNBC has high mitotic activity, and, because Aurora A and B mitotic kinases drive cell division and are overexpressed in tumors with a high mitotic index, we hypothesized that inhibiting Aurora A and B produces a significant antitumor effect in TNBC. We tested this hypothesis by determining the antitumor effects of KW-2450, a multikinase inhibitor of both Aurora A and B kinases. We observed significant inhibitory activities of KW-2450 on cell viability, apoptosis, colony formation in agar, and mammosphere formation in TNBC cells. The growth of TNBC xenografts was significantly inhibited with KW-2450. In cell-cycle analysis, KW-2450 induced tetraploid accumulation followed by apoptosis or surviving octaploid (8N) cells, depending on dose. These phenotypes resembled those of Aurora B knockdown and complete pharmaceutical inhibition of Aurora A. We demonstrated that 8N cells resulting from KW-2450 treatment depended on the activation of mitogen-activated protein kinase kinase (MEK) for their survival. When treated with the MEK inhibitor selumetinib combined with KW-2450, compared with KW-2450 alone, the 8N cell population was significantly reduced and apoptosis was increased. Indeed, this combination showed synergistic antitumor effect in SUM149 TNBC xenografts. Collectively, Aurora A and B inhibition had a significant antitumor effect against TNBC, and this antitumor effect was maximized by the combination of selumetinib with Aurora A and B inhibition.

  8. Deregulated Expression of Aurora Kinases Is Not a Prognostic Biomarker in Papillary Thyroid Cancer Patients

    Science.gov (United States)

    Prinzi, Natalie; Sorrenti, Salvatore; Falvo, Laura; De Vito, Corrado; Catania, Antonio; Tartaglia, Francesco; Mocini, Renzo; Coccaro, Carmela; Alessandrini, Stefania; Barollo, Susi; Mian, Caterina; Antonelli, Alessandro; De Antoni, Enrico; D’Armiento, Massimino; Ulisse, Salvatore

    2015-01-01

    A number of reports indicated that Aurora-A or Aurora-B overexpression represented a negative prognostic factor in several human malignancies. In thyroid cancer tissues a deregulated expression of Aurora kinases has been also demonstrated, butno information regarding its possible prognostic role in differentiated thyroid cancer is available. Here, weevaluated Aurora-A and Aurora-B mRNA expression and its prognostic relevance in a series of 87 papillary thyroid cancers (PTC), with a median follow-up of 63 months. The analysis of Aurora-A and Aurora-B mRNA levels in PTC tissues, compared to normal matched tissues, revealed that their expression was either up- or down-regulatedin the majority of cancer tissues. In particular, Aurora-A and Aurora-B mRNA levels were altered, respectively, in 55 (63.2%) and 79 (90.8%) out of the 87 PTC analyzed.A significant positive correlation between Aurora-A and Aurora-B mRNAswas observed (p=0.001). The expression of both Aurora genes was not affected by the BRAFV600E mutation. Univariate, multivariate and Kaplan-Mayer analyses documented the lack of association between Aurora-A or Aurora-B expression and clinicopathological parameterssuch as gender, age, tumor size, histology, TNM stage, lymph node metastasis and BRAF status as well asdisease recurrences or disease-free interval. Only Aurora-B mRNA was significantly higher in T(3-4) tissues, with respect to T(1-2) PTC tissues. The data reported here demonstrate that the expression of Aurora kinases is deregulated in the majority of PTC tissues, likely contributing to PTC progression. However, differently from other human solid cancers, detection of Aurora-A or Aurora-B mRNAs is not a prognostic biomarker inPTC patients. PMID:25807528

  9. Adaptation of the plasma inhibitory activity assay to detect Aurora, ABL and FLT3 kinase inhibition by AT9283 in pediatric leukemia.

    Science.gov (United States)

    Podesta, Jennifer E; Sugar, Richard; Squires, Matt; Linardopoulos, Spiros; Pearson, Andrew D J; Moore, Andrew S

    2011-09-01

    Non-invasive assessment of biomarker modulation is important for evaluating targeted therapeutics, particularly in pediatrics. The plasma inhibitory activity (PIA) assay is used clinically to assess FLT3 inhibition ex vivo and guide dosing. AT9283 is a novel Aurora kinase inhibitor with secondary activity against FLT3 and ABL. We adapted the PIA assay to simultaneously detect inhibition of Aurora and FLT3 in AML, and Aurora and ABL in CML by AT9283. Furthermore, we optimized the assay for children, where limited blood volumes are available for pharmacodynamic studies. Simultaneously detecting multiple kinase inhibition may identify important mechanisms of action for novel anti-leukemic drugs.

  10. Polo-Like Kinase-1 Controls Aurora A Destruction by Activating APC/C-Cdh1

    NARCIS (Netherlands)

    van Leuken, Renske; Clijsters, Linda; van Zon, Wouter; Lim, Dan; Yao, XueBiao; Wolthuis, Rob M. F.; Yaffe, Michael B.; Medema, Rene H.; van Vugt, Marcel A. T. M.

    2009-01-01

    Polo-like kinase-1 (Plk1) is activated before mitosis by Aurora A and its cofactor Bora. In mitosis, Bora is degraded in a manner dependent on Plk1 kinase activity and the E3 ubiquitin ligase SCF-beta TrCP. Here, we show that Plk1 is also required for the timely destruction of its activator Aurora A

  11. The investigational Aurora kinase A inhibitor alisertib (MLN8237 induces cell cycle G2/M arrest, apoptosis, and autophagy via p38 MAPK and Akt/mTOR signaling pathways in human breast cancer cells

    Directory of Open Access Journals (Sweden)

    Li JP

    2015-03-01

    Full Text Available Jin-Ping Li,1,2 Yin-Xue Yang,3 Qi-Lun Liu,1 Shu-Ting Pan,1,4 Zhi-Xu He,5 Xueji Zhang,6 Tianxin Yang,7 Xiao-Wu Chen,8 Dong Wang,9 Jia-Xuan Qiu,4 Shu-Feng Zhou2,51Department of Surgical Oncology, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, People’s Republic of China; 2Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA; 3Department of Colorectal Surgery, General Hospital of Ningxia Medical University, Yinchuan, Ningxia; 4Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 5Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guizhou Medical University, Guiyang, Guizhou, 6Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 7Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA; 8Department of General Surgery, The First People’s Hospital of Shunde, Southern Medical University, Shunde, Foshan, Guangdong, 9Cancer Center, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, People’s Republic of ChinaAbstract: Alisertib (ALS is an investigational potent Aurora A kinase inhibitor currently undergoing clinical trials for the treatment of hematological and non-hematological malignancies. However, its antitumor activity has not been tested in human breast cancer. This study aimed to investigate the effect of ALS on the growth, apoptosis, and autophagy, and the underlying mechanisms in human breast cancer MCF7 and MDA-MB-231 cells. In the current study, we identified that ALS had potent growth-inhibitory, pro-apoptotic, and pro-autophagic effects in MCF7 and MDA-MB-231 cells. ALS arrested the cells in G2

  12. Interdomain allosteric regulation of Polo kinase by Aurora B and Map205 is required for cytokinesis.

    Science.gov (United States)

    Kachaner, David; Pinson, Xavier; El Kadhi, Khaled Ben; Normandin, Karine; Talje, Lama; Lavoie, Hugo; Lépine, Guillaume; Carréno, Sébastien; Kwok, Benjamin H; Hickson, Gilles R; Archambault, Vincent

    2014-10-27

    Drosophila melanogaster Polo and its human orthologue Polo-like kinase 1 fulfill essential roles during cell division. Members of the Polo-like kinase (Plk) family contain an N-terminal kinase domain (KD) and a C-terminal Polo-Box domain (PBD), which mediates protein interactions. How Plks are regulated in cytokinesis is poorly understood. Here we show that phosphorylation of Polo by Aurora B is required for cytokinesis. This phosphorylation in the activation loop of the KD promotes the dissociation of Polo from the PBD-bound microtubule-associated protein Map205, which acts as an allosteric inhibitor of Polo kinase activity. This mechanism allows the release of active Polo from microtubules of the central spindle and its recruitment to the site of cytokinesis. Failure in Polo phosphorylation results in both early and late cytokinesis defects. Importantly, the antagonistic regulation of Polo by Aurora B and Map205 in cytokinesis reveals that interdomain allosteric mechanisms can play important roles in controlling the cellular functions of Plks.

  13. KNL1 facilitates phosphorylation of outer kinetochore proteins by promoting Aurora B kinase activity.

    Science.gov (United States)

    Caldas, Gina V; DeLuca, Keith F; DeLuca, Jennifer G

    2013-12-23

    Aurora B kinase phosphorylates kinetochore proteins during early mitosis, increasing kinetochore–microtubule (MT) turnover and preventing premature stabilization of kinetochore–MT attachments. Phosphorylation of kinetochore proteins during late mitosis is low, promoting attachment stabilization, which is required for anaphase onset. The kinetochore protein KNL1 recruits Aurora B–counteracting phosphatases and the Aurora B–targeting factor Bub1, yet the consequences of KNL1 depletion on Aurora B phospho-regulation remain unknown. Here, we demonstrate that the KNL1 N terminus is essential for Aurora B activity at kinetochores. This region of KNL1 is also required for Bub1 kinase activity at kinetochores, suggesting that KNL1 promotes Aurora B activity through Bub1-mediated Aurora B targeting. However, ectopic targeting of Aurora B to kinetochores does not fully rescue Aurora B activity in KNL1-depleted cells, suggesting KNL1 influences Aurora B activity through an additional pathway. Our findings establish KNL1 as a requirement for Aurora B activity at kinetochores and for wild-type kinetochore–MT attachment dynamics.

  14. Aurora B kinase is a potent and selective target in MYCN-driven neuroblastoma.

    Science.gov (United States)

    Bogen, Dominik; Wei, Jun S; Azorsa, David O; Ormanoglu, Pinar; Buehler, Eugen; Guha, Rajarshi; Keller, Jonathan M; Mathews Griner, Lesley A; Ferrer, Marc; Song, Young K; Liao, Hongling; Mendoza, Arnulfo; Gryder, Berkley E; Sindri, Sivasish; He, Jianbin; Wen, Xinyu; Zhang, Shile; Shern, John F; Yohe, Marielle E; Taschner-Mandl, Sabine; Shohet, Jason M; Thomas, Craig J; Martin, Scott E; Ambros, Peter F; Khan, Javed

    2015-11-01

    Despite advances in multimodal treatment, neuroblastoma (NB) is often fatal for children with high-risk disease and many survivors need to cope with long-term side effects from high-dose chemotherapy and radiation. To identify new therapeutic targets, we performed an siRNA screen of the druggable genome combined with a small molecule screen of 465 compounds targeting 39 different mechanisms of actions in four NB cell lines. We identified 58 genes as targets, including AURKB, in at least one cell line. In the drug screen, aurora kinase inhibitors (nine molecules) and in particular the AURKB-selective compound, barasertib, were the most discriminatory with regard to sensitivity for MYCN-amplified cell lines. In an expanded panel of ten NB cell lines, those with MYCN-amplification and wild-type TP53 were the most sensitive to low nanomolar concentrations of barasertib. Inhibition of the AURKB kinase activity resulted in decreased phosphorylation of the known target, histone H3, and upregulation of TP53 in MYCN-amplified, TP53 wild-type cells. However, both wild-type and TP53 mutant MYCN-amplified cell lines arrested in G2/M phase upon AURKB inhibition. Additionally, barasertib induced endoreduplication and apoptosis. Treatment of MYCN-amplified/TP53 wild-type neuroblastoma xenografts resulted in profound growth inhibition and tumor regression. Therefore, aurora B kinase inhibition is highly effective in aggressive neuroblastoma and warrants further investigation in clinical trials.

  15. Microtubules accelerate the kinase activity of Aurora-B by a reduction in dimensionality.

    Science.gov (United States)

    Noujaim, Michael; Bechstedt, Susanne; Wieczorek, Michal; Brouhard, Gary J

    2014-01-01

    Aurora-B is the kinase subunit of the Chromosome Passenger Complex (CPC), a key regulator of mitotic progression that corrects improper kinetochore attachments and establishes the spindle midzone. Recent work has demonstrated that the CPC is a microtubule-associated protein complex and that microtubules are able to activate the CPC by contributing to Aurora-B auto-phosphorylation in trans. Aurora-B activation is thought to occur when the local concentration of Aurora-B is high, as occurs when Aurora-B is enriched at centromeres. It is not clear, however, whether distributed binding to large structures such as microtubules would increase the local concentration of Aurora-B. Here we show that microtubules accelerate the kinase activity of Aurora-B by a "reduction in dimensionality." We find that microtubules increase the kinase activity of Aurora-B toward microtubule-associated substrates while reducing the phosphorylation levels of substrates not associated to microtubules. Using the single molecule assay for microtubule-associated proteins, we show that a minimal CPC construct binds to microtubules and diffuses in a one-dimensional (1D) random walk. The binding of Aurora-B to microtubules is salt-dependent and requires the C-terminal tails of tubulin, indicating that the interaction is electrostatic. We show that the rate of Aurora-B auto-activation is faster with increasing concentrations of microtubules. Finally, we demonstrate that microtubules lose their ability to stimulate Aurora-B when their C-terminal tails are removed by proteolysis. We propose a model in which microtubules act as scaffolds for the enzymatic activity of Aurora-B. The scaffolding activity of microtubules enables rapid Aurora-B activation and efficient phosphorylation of microtubule-associated substrates.

  16. The dawn of Aurora kinase research: from fly genetics to the clinic.

    Directory of Open Access Journals (Sweden)

    Mar eCarmena

    2015-11-01

    Full Text Available Aurora kinases comprise a family of highly conserved serine-threonine protein kinases that play a pivotal role in the regulation of cell cycle. Aurora kinases are not only involved in the control of multiple processes during cell division but also coordinate chromosomal and cytoskeletal events, contributing to the regulation of checkpoints and ensuring the smooth progression of the cell cycle.Because of their fundamental contribution to cell cycle regulation, Aurora kinases were originally identified in independent genetic screens designed to find genes involved in the regulation of cell division. The first aurora mutant was part of a collection of mutants isolated in C. Nusslein-Volhard’s laboratory. This collection was screened in D. M. Glover’s laboratory in search for mutations disrupting the centrosome cycle in embryos derived from homozygous mutant mothers. The mutants identified were given names related to the polar regions, and included not only aurora but also the equally famous polo. Ipl1, the only Aurora in yeast, was identified in a genetic screen looking for mutations that caused chromosome segregation defects. The discovery of a second Aurora-like kinase in mammals opened a new chapter in the research of Aurora kinases. The rat kinase AIM was found to be highly homologous to the fly and yeast proteins, but localised at the midzone and midbody and was proposed to have a role in cytokinesis. Homologs of the equatorial Aurora (Aurora B were identified in metazoans ranging from flies to humans. Xenopus Aurora B was found to be in a complex with the chromosomal passenger INCENP, and both proteins were shown to be essential in flies for chromosome structure, segregation, central spindle formation and cytokinesis. Fifteen years on, Aurora kinase research is an active field of research. After the successful introduction of the first anti-mitotic agents in cancer therapy, both Auroras have become the focus of attention as targets for

  17. Aurora kinases are expressed in medullary thyroid carcinoma (MTC and their inhibition suppresses in vitro growth and tumorigenicity of the MTC derived cell line TT

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    Morrone Stefania

    2011-09-01

    Full Text Available Abstract Background The Aurora kinase family members, Aurora-A, -B and -C, are involved in the regulation of mitosis, and alterations in their expression are associated with cell malignant transformation. To date no information on the expression of these proteins in medullary thyroid carcinoma (MTC are available. We here investigated the expression of the Aurora kinases in human MTC tissues and their potential use as therapeutic targets. Methods The expression of the Aurora kinases in 26 MTC tissues at different TNM stages was analyzed at the mRNA level by quantitative RT-PCR. We then evaluated the effects of the Aurora kinase inhibitor MK-0457 on the MTC derived TT cell line proliferation, apoptosis, soft agar colony formation, cell cycle and ploidy. Results The results showed the absence of correlation between tumor tissue levels of any Aurora kinase and tumor stage indicating the lack of prognostic value for these proteins. Treatment with MK-0457 inhibited TT cell proliferation in a time- and dose-dependent manner with IC50 = 49.8 ± 6.6 nM, as well as Aurora kinases phosphorylation of substrates relevant to the mitotic progression. Time-lapse experiments demonstrated that MK-0457-treated cells entered mitosis but were unable to complete it. Cytofluorimetric analysis confirmed that MK-0457 induced accumulation of cells with ≥ 4N DNA content without inducing apoptosis. Finally, MK-0457 prevented the capability of the TT cells to form colonies in soft agar. Conclusions We demonstrate that Aurora kinases inhibition hampered growth and tumorigenicity of TT cells, suggesting its potential therapeutic value for MTC treatment.

  18. Evolutionary relationships of Aurora kinases: Implications for model organism studies and the development of anti-cancer drugs

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    Patrick Denis R

    2004-10-01

    Full Text Available Abstract Background As key regulators of mitotic chromosome segregation, the Aurora family of serine/threonine kinases play an important role in cell division. Abnormalities in Aurora kinases have been strongly linked with cancer, which has lead to the recent development of new classes of anti-cancer drugs that specifically target the ATP-binding domain of these kinases. From an evolutionary perspective, the species distribution of the Aurora kinase family is complex. Mammals uniquely have three Aurora kinases, Aurora-A, Aurora-B, and Aurora-C, while for other metazoans, including the frog, fruitfly and nematode, only Aurora-A and Aurora-B kinases are known. The fungi have a single Aurora-like homolog. Based on the tacit assumption of orthology to human counterparts, model organism studies have been central to the functional characterization of Aurora kinases. However, the ortholog and paralog relationships of these kinases across various species have not been rigorously examined. Here, we present comprehensive evolutionary analyses of the Aurora kinase family. Results Phylogenetic trees suggest that all three vertebrate Auroras evolved from a single urochordate ancestor. Specifically, Aurora-A is an orthologous lineage in cold-blooded vertebrates and mammals, while structurally similar Aurora-B and Aurora-C evolved more recently in mammals from a duplication of an ancestral Aurora-B/C gene found in cold-blooded vertebrates. All so-called Aurora-A and Aurora-B kinases of non-chordates are ancestral to the clade of chordate Auroras and, therefore, are not strictly orthologous to vertebrate counterparts. Comparisons of human Aurora-B and Aurora-C sequences to the resolved 3D structure of human Aurora-A lends further support to the evolutionary scenario that vertebrate Aurora-B and Aurora-C are closely related paralogs. Of the 26 residues lining the ATP-binding active site, only three were variant and all were specific to Aurora-A. Conclusions In

  19. Targeting the interaction of Aurora kinases and SIRT1 mediated by Wnt signaling pathway in colorectal cancer: A critical review.

    Science.gov (United States)

    Subramaniyan, Boopathi; Jagadeesan, Kaviya; Ramakrishnan, Sabitha; Mathan, Ganeshan

    2016-08-01

    The Aurora kinases belong to the family of serine/threonine kinase, a central regulator of mitosis and their expression increased during G2/M phase. It is classified into Aurora A, B and C, each has distinct roles in cellular processes, which includes regulation of spindle assembly, function of centrosomes, cytoskeleton and cytokinesis. During cancer growth, their rapid increase makes most attractive marker for cancer treatment at present. However Aurora A kinase is known to be a marker for cancer therapy, the most important serine/threonine kinase of Aurora B kinase involvement in cancer is still inadequate. Subsequently, the recent findings revealed that the class III histone deacetylase of SIRT1 is a key regulator to activate Aurora kinases from S phase damaged DNA through Wnt signaling pathway. Even if both Aurora A kinase and SIRT1 serve as a marker for cancer therapy, the present review reveals it is interaction in Wnt signaling pathway that solely for colorectal cancer.

  20. Cell cycle-dependent regulation of Aurora kinase B mRNA by the Microprocessor complex.

    Science.gov (United States)

    Jung, Eunsun; Seong, Youngmo; Seo, Jae Hong; Kwon, Young-Soo; Song, Hoseok

    2014-03-28

    Aurora kinase B regulates the segregation of chromosomes and the spindle checkpoint during mitosis. In this study, we showed that the Microprocessor complex, which is responsible for the processing of the primary transcripts during the generation of microRNAs, destabilizes the mRNA of Aurora kinase B in human cells. The Microprocessor-mediated cleavage kept Aurora kinase B at a low level and prevented premature entrance into mitosis. The cleavage was reduced during mitosis leading to the accumulation of Aurora kinase B mRNA and protein. In addition to Aurora kinase B mRNA, the processing of other primary transcripts of miRNAs were also decreased during mitosis. We found that the cleavage was dependent on an RNA helicase, DDX5, and the association of DDX5 and DDX17 with the Microprocessor was reduced during mitosis. Thus, we propose a novel mechanism by which the Microprocessor complex regulates stability of Aurora kinase B mRNA and cell cycle progression.

  1. Aurora kinases as druggable targets in pediatric leukemia: heterogeneity in target modulation activities and cytotoxicity by diverse novel therapeutic agents.

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    Aarthi Jayanthan

    Full Text Available Leukemia is the most common pediatric malignancy, constituting more than 30% of all childhood cancers. Although cure rates have improved greatly, approximately one in five children relapse and poor survival rates post relapse remain a challenge. Given this, more effective and innovative therapeutic strategies are needed in order to improve prognosis. Aurora kinases, a family of serine/threonine kinases essential for the regulation of several mitotic processes, have been identified as potential targets for cancer therapeutics. Elevated expression of Aurora kinases has been demonstrated in several malignancies and is associated with aberrant mitotic activity, aneuploidy and alterations in chromosomal structure and genome instability. Based on this rationale, a number of small molecule inhibitors have been formulated and advanced to human studies in the recent past. A comparative analysis of these agents in cytotoxicity and target modulation analyses against a panel of leukemia cells provides novel insights into the unique mechanisms and codependent activity pathways involved in targeting Aurora kinases, constituting a distinctive preclinical experimental framework to identify appropriate agents and combinations in future clinical studies.

  2. Induction of p21CIP1 protein and cell cycle arrest after inhibition of Aurora B kinase is attributed to aneuploidy and reactive oxygen species.

    Science.gov (United States)

    Kumari, Geeta; Ulrich, Tanja; Krause, Michael; Finkernagel, Florian; Gaubatz, Stefan

    2014-06-01

    Cell cycle progression requires a series of highly coordinated events that ultimately lead to faithful segregation of chromosomes. Aurora B is an essential mitotic kinase, which is involved in regulation of microtubule-kinetochore attachments and cytokinesis. Inhibition of Aurora B results in stabilization of p53 and induction of p53-target genes such as p21 to inhibit proliferation. We have previously demonstrated that induction of p21 by p53 after inhibition of Aurora B is dependent on the p38 MAPK, which promotes transcriptional elongation of p21 by RNA Pol II. In this study, we show that a subset of p53-target genes are induced in a p38-dependent manner upon inhibition of Aurora B. We also demonstrate that inhibition of Aurora B results in down-regulation of E2F-mediated transcription and that the cell cycle arrest after Aurora B inhibition depends on p53 and pRB tumor suppressor pathways. In addition, we report that activation of p21 after inhibition of Aurora B is correlated with increased chromosome missegregation and aneuploidy but not with binucleation or tetraploidy. We provide evidence that p21 is activated in aneuploid cells by reactive oxygen species (ROS) and p38 MAPK. Finally, we demonstrate that certain drugs that act on aneuploid cells synergize with inhibitors of Aurora B to inhibit colony formation and oncogenic transformation. These findings provide an important link between aneuploidy and the stress pathways activated by Aurora B inhibition and also support the use of Aurora B inhibitors in combination therapy for treatment of cancer.

  3. Phosphorylation of MAP65-1 by Arabidopsis Aurora Kinases Is Required for Efficient Cell Cycle Progression1[OPEN

    Science.gov (United States)

    Weimer, Annika K.; Stoppin-Mellet, Virginie; Kosetsu, Ken; Cedeño, Cesyen; Jaquinod, Michel; Njo, Maria; De Milde, Liesbeth; Tompa, Peter; Inzé, Dirk; Beeckman, Tom; Vantard, Marylin

    2017-01-01

    Aurora kinases are key effectors of mitosis. Plant Auroras are functionally divided into two clades. The alpha Auroras (Aurora1 and Aurora2) associate with the spindle and the cell plate and are implicated in controlling formative divisions throughout plant development. The beta Aurora (Aurora3) localizes to centromeres and likely functions in chromosome separation. In contrast to the wealth of data available on the role of Aurora in other kingdoms, knowledge on their function in plants is merely emerging. This is exemplified by the fact that only histone H3 and the plant homolog of TPX2 have been identified as Aurora substrates in plants. Here we provide biochemical, genetic, and cell biological evidence that the microtubule-bundling protein MAP65-1—a member of the MAP65/Ase1/PRC1 protein family, implicated in central spindle formation and cytokinesis in animals, yeasts, and plants—is a genuine substrate of alpha Aurora kinases. MAP65-1 interacts with Aurora1 in vivo and is phosphorylated on two residues at its unfolded tail domain. Its overexpression and down-regulation antagonistically affect the alpha Aurora double mutant phenotypes. Phospho-mutant analysis shows that Aurora contributes to the microtubule bundling capacity of MAP65-1 in concert with other mitotic kinases. PMID:27879390

  4. Indomethacin promotes apoptosis in gastric cancer cells through concomitant degradation of Survivin and Aurora B kinase proteins.

    Science.gov (United States)

    Chiou, Shiun-Kwei; Hoa, Neil; Hodges, Amy; Ge, Lishen; Jadus, Martin R

    2014-09-01

    Regular usage of nonsteroidal anti-inflammatory drugs (NSAIDs) is associated with reduced incidence of a variety of cancers. The molecular mechanisms underlying these chemopreventive effects remain poorly understood. This current investigation showed that in gastric cancer cells: (1) Indomethacin treatment enhanced the degradation of chromosomal passenger proteins, Survivin and Aurora B kinase; (2) Indomethacin treatment down-regulated Aurora B kinase activity in a cell cycle-independent fashion; (3) siRNA knockdown of Survivin level promoted Aurora B kinase protein degradation, and vice versa; (4) ectopic overexpression of Survivin blocked reduction of Aurora B kinase level and activity by indomethacin treatment, and vice versa; (5) siRNA knockdown of Aurora B kinase level and AZD1152 inhibition of its activity induced apoptosis, and overexpression of Aurora B kinase inhibited indomethacin-induced apoptosis; (6) indomethacin treatment reduced Aurora B kinase level, coinciding with reduction of Survivin level and induction of apoptosis, in KATO III and HT-29 cells, and in mouse gastric mucosa. A role for Aurora B kinase function in NSAID-induced apoptosis was not previously explored. Thus this report provides better understanding of the molecular mechanisms underlying the anti-cancer effect of NSAIDs by elucidating a significant role for Aurora B kinase in indomethacin-induced apoptosis.

  5. Specific small-molecule activator of Aurora kinase A induces autophosphorylation in a cell-free system.

    Science.gov (United States)

    Kishore, A Hari; Vedamurthy, B M; Mantelingu, K; Agrawal, Shipra; Reddy, B A Ashok; Roy, Siddhartha; Rangappa, K S; Kundu, Tapas K

    2008-02-28

    Aurora kinases are essential for chromosomal segregation and cell division and thereby important for maintaining the proper genomic integrity. There are three classes of aurora kinases in humans: A, B, and C. Aurora kinase A is frequently overexpressed in various cancers. The link of the overexpression and tumorigenesis is yet to be understood. By employing virtual screening, we have found that anacardic acid, a pentadecane aliphatic chain containing hydroxylcarboxylic acid, from cashew nut shell liquid could be docked in Aurora kinases A and B. Remarkably, we found that anacardic acid could potently activate the Aurora kinase A mediated phosphorylation of histone H3, but at a similar concentration the activity of aurora kinase B remained unaffected in vitro. Mechanistically, anacardic acid induces the structural changes and also the autophosphorylation of the aurora kinase A to enhance the enzyme activity. This data thus indicate anacardic acid as the first small-molecule activator of Aurora kinase, which could be highly useful for probing the function of hyperactive (overexpressed) Aurora kinase A.

  6. Aurora A kinase modulates actin cytoskeleton through phosphorylation of Cofilin: Implication in the mitotic process.

    Science.gov (United States)

    Ritchey, Lisa; Chakrabarti, Ratna

    2014-11-01

    Aurora A kinase regulates early mitotic events through phosphorylation and activation of a variety of proteins. Specifically, Aur-A is involved in centrosomal separation and formation of mitotic spindles in early prophase. The effect of Aur-A on mitotic spindles is mediated by the modulation of microtubule dynamics and association with microtubule binding proteins. In this study we show that Aur-A exerts its effects on spindle organization through the regulation of the actin cytoskeleton. Aurora A phosphorylates Cofilin at multiple sites including S(3) resulting in the inactivation of its actin depolymerizing function. Aur-A interacts with Cofilin in early mitotic phases and regulates its phosphorylation status. Cofilin phosphorylation follows a dynamic pattern during the progression of prophase to metaphase. Inhibition of Aur-A activity induced a delay in the progression of prophase to metaphase. Aur-A inhibitor also disturbed the pattern of Cofilin phosphorylation, which correlated with the mitotic delay. Our results establish a novel function of Aur-A in the regulation of actin cytoskeleton reorganization, through Cofilin phosphorylation during early mitotic stages.

  7. The Aurora B kinase in chromosome biorientation and spindle checkpoint signalling

    Directory of Open Access Journals (Sweden)

    Veronica eKrenn

    2015-10-01

    Full Text Available Aurora B, a member of the Aurora family of serine/threonine protein kinases, is a key player in chromosome segregation. As part of a macromolecular complex known as the chromosome passenger complex, Aurora B concentrates early during mitosis in the proximity of centromeres and kinetochores, the sites of attachment of chromosomes to spindle microtubules. There, it contributes to a number of processes that impart fidelity to cell division, including kinetochore stabilization, kinetochore-microtubule attachment, and the regulation of a surveillance mechanism named the spindle assembly checkpoint. In the regulation of these processes, Aurora B is the fulcrum of a remarkably complex network of interactions that feed back on its localization and activation state. In this review we discuss the multiple roles of Aurora B during mitosis, focusing in particular on its role at centromeres and kinetochores. Many details of the network of interactions at these locations remain poorly understood, and we focus here on several crucial outstanding questions.

  8. Selective disruption of aurora C kinase reveals distinct functions from aurora B kinase during meiosis in mouse oocytes.

    Directory of Open Access Journals (Sweden)

    Ahmed Z Balboula

    2014-02-01

    Full Text Available Aurora B kinase (AURKB is the catalytic subunit of the chromosomal passenger complex (CPC, an essential regulator of chromosome segregation. In mitosis, the CPC is required to regulate kinetochore microtubule (K-MT attachments, the spindle assembly checkpoint, and cytokinesis. Germ cells express an AURKB homolog, AURKC, which can also function in the CPC. Separation of AURKB and AURKC function during meiosis in oocytes by conventional approaches has not been successful. Therefore, the meiotic function of AURKC is still not fully understood. Here, we describe an ATP-binding-pocket-AURKC mutant, that when expressed in mouse oocytes specifically perturbs AURKC-CPC and not AURKB-CPC function. Using this mutant we show for the first time that AURKC has functions that do not overlap with AURKB. These functions include regulating localized CPC activity and regulating chromosome alignment and K-MT attachments at metaphase of meiosis I (Met I. We find that AURKC-CPC is not the sole CPC complex that regulates the spindle assembly checkpoint in meiosis, and as a result most AURKC-perturbed oocytes arrest at Met I. A small subset of oocytes do proceed through cytokinesis normally, suggesting that AURKC-CPC is not the sole CPC complex during telophase I. But, the resulting eggs are aneuploid, indicating that AURKC is a critical regulator of meiotic chromosome segregation in female gametes. Taken together, these data suggest that mammalian oocytes contain AURKC to efficiently execute meiosis I and ensure high-quality eggs necessary for sexual reproduction.

  9. Danusertib, a potent pan-Aurora kinase and ABL kinase inhibitor, induces cell cycle arrest and programmed cell death and inhibits epithelial to mesenchymal transition involving the PI3K/Akt/mTOR-mediated signaling pathway in human gastric cancer AGS and NCI-N78 cells

    Directory of Open Access Journals (Sweden)

    Yuan CX

    2015-03-01

    Full Text Available Chun-Xiu Yuan,1,2 Zhi-Wei Zhou,2,3 Yin-Xue Yang,4 Zhi-Xu He,3 Xueji Zhang,5 Dong Wang,6 Tianxing Yang,7 Si-Yuan Pan,8 Xiao-Wu Chen,9 Shu-Feng Zhou2 1Department of Oncology, General Hospital, Ningxia Medical University, Yinchuan, People’s Republic of China; 2Department of Pharmaceutical Science, College of Pharmacy, University of South Florida, Tampa, FL, USA; 3Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, 4Department of Colorectal Surgery, General Hospital, Ningxia Medical University, Yinchuan, 5Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, 6Cancer Center, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, People’s Republic of China; 7Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA; 8Department of Pharmacology, School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing, 9Department of General Surgery, The First People’s Hospital of Shunde, Southern Medical University, Shunde, People’s Republic of China Abstract: Gastric cancer is the second leading cause of cancer-related death worldwide, with a poor response to current chemotherapy. Danusertib is a pan-inhibitor of the Aurora kinases and a third-generation Bcr-Abl tyrosine kinase inhibitor with potent anticancer effects, but its antitumor effect and underlying mechanisms in the treatment of human gastric cancer are unknown. This study aimed to investigate the effects of danusertib on cell growth, apoptosis, autophagy, and epithelial to mesenchymal transition and the molecular mechanisms involved in human gastric cancer AGS and NCI-N78 cells. The results showed that danusertib had potent growth-inhibitory, apoptosis-inducing, and

  10. Function and regulation of Aurora/Ipl1p kinase family in cell division

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    During mitosis, the parent cell distributes its genetic materials equally into two daughter cells through chromosome segregation, a complex movements orchestrated by mitotic kinases and its effector proteins.Faithful chromosome segregation and cytokinesis ensure that each daughter cell receives a full copy of genetic materials of parent cell. Defects in these processes can lead to aneuploidy or polyploidy. Aurora/Ipl1p fanily,a class of conserved serine/threonine kinases, plays key roles in chromosome segregation and cytokinesis.This article highlights the function and regulation of Aurora/Ipl1p family in mitosis and provides potential links between aberrant regulation of Aurora/Ipl1p kinases and pathogenesis of human cancer.

  11. Aurora A kinase activates YAP signaling in triple-negative breast cancer.

    Science.gov (United States)

    Chang, S-S; Yamaguchi, H; Xia, W; Lim, S-O; Khotskaya, Y; Wu, Y; Chang, W-C; Liu, Q; Hung, M-C

    2017-03-02

    The Yes-associated protein (YAP) is an effector that transduces the output of the Hippo pathway to transcriptional modulation. Considering the role of YAP in cancers, this protein has emerged as a key node in malignancy development. In this study, we determined that Aurora A kinase acts as a positive regulator for YAP-mediated transcriptional machinery. Specifically, YAP associates with Aurora A predominantly in the nucleus. Activation of Aurora A can impinge on YAP activity through direct phosphorylation. Moreover, aberrant expression of YAP and Aurora A signaling is highly correlated with triple-negative breast cancer (TNBC). We herein provide evidence to establish the functional relevance of this newly discovered regulatory axis in TNBC.

  12. Renal targeting of kinase inhibitors

    NARCIS (Netherlands)

    Dolman, M. E. M.; Fretz, M. M.; Segers, Gj. W.; Lacombe, M.; Prakash, J.; Storm, G.; Hennink, W. E.; Kok, R. J.

    2008-01-01

    Activation of proximal tubular cells by fibrotic and inflammatory mediators is an important hallmark of chronic kidney disease. We have developed a novel strategy to intervene in renal fibrosis, by means of locally delivered kinase inhibitors. Such compounds will display enhanced activity within tub

  13. Phosphorylation of multifunctional nucleolar protein nucleophosmin (NPM1) by aurora kinase B is critical for mitotic progression.

    Science.gov (United States)

    Shandilya, Jayasha; Senapati, Parijat; Dhanasekaran, Karthigeyan; Bangalore, Suma S; Kumar, Manoj; Kishore, A Hari; Bhat, Akshay; Kodaganur, Gopinath S; Kundu, Tapas K

    2014-06-27

    The functional association of NPM1 with Aurora kinases is well documented. Surprisingly, although NPM1 is a well characterized phosphoprotein, it is unknown whether it is a substrate of Aurora kinases. We have found that Aurora kinases A and B can phosphorylate NPM1 at a single serine residue, Ser125, in vitro and in vivo. Phosphorylated-S125-NPM1 (pS125-NPM1) localizes to the midbody region during late cytokinesis where it colocalizes with Aurora B. The overexpression of mutant (S125A) NPM1 resulted in the deregulation of centrosome duplication and mitotic defects possibly due to cytokinesis failure. These data suggest that Aurora kinase B-mediated phosphorylation of NPM1 plays a critical role during mitosis, which could have wider implications in oncogenesis.

  14. Inhibitors of protein kinase C

    Institute of Scientific and Technical Information of China (English)

    LIU Shiying; JIANG Yuyang; CAO Jian; LIU Feng; MA Li; ZHAO Yufen

    2005-01-01

    Protein kinase catalyzes the transfer of the γ-phosphoryl group from ATP to the hydroxyl groups of protein side chains, which plays critical roles in signal transduction pathways by transmitting extracellular signals across the plasma membrane and nuclear membrane to the destination sites in the cytoplasm and the nucleus. Protein kinase C (PKC) is a superfamily of phospholipid-dependent Ser/Thr kinase. There are at least 12 isozymes in PKC family. They are distributed in different tissues and play different roles in physiological processes. On account of their concern with a variety of pathophysiologic states, such as cancer, inflammatory conditions, autoimmune disorder, and cardiac diseases, the inhibitors, which can inhibit the activity of PKC and the interaction of cytokine with receptor, and interfere signal transduction pathway, may be candidates of therapeutic drugs. Therefore, intense efforts have been made to develop specific protein kinase inhibitors as biological tools and therapeutic agents. This article reviews the recent development of some of PKC inhibitors based on their interaction with different conserved domains and different inhibition mechanisms.

  15. Aurora B kinase controls the separation of centromeric and telomeric heterochromatin

    OpenAIRE

    Gachet, Yannick; Reyes, Celine; Tournier, Sylvie

    2015-01-01

    The segregation of chromosomes is coordinated at multiple levels to prevent chromosome loss, a phenotype frequently observed in cancers. We recently described an essential role for telomeres in the physical separation of chromosomes and identified Aurora B kinase as a double agent involved in the separation of centromeric and telomeric heterochromatin.

  16. Aurora B kinase controls the separation of centromeric and telomeric heterochromatin.

    Science.gov (United States)

    Gachet, Yannick; Reyes, Celine; Tournier, Sylvie

    2016-03-01

    The segregation of chromosomes is coordinated at multiple levels to prevent chromosome loss, a phenotype frequently observed in cancers. We recently described an essential role for telomeres in the physical separation of chromosomes and identified Aurora B kinase as a double agent involved in the separation of centromeric and telomeric heterochromatin.

  17. Inhibition of mitotic kinase Aurora suppresses Akt-1 activation and induces apoptotic cell death in all-trans retinoid acid-resistant acute promyelocytic leukemia cells

    Directory of Open Access Journals (Sweden)

    Long Zi-Jie

    2011-05-01

    Full Text Available Abstract Background Aurora kinase ensures accurate chromosome segregation during cell cycle, maintaining genetic integrity in cell division. VX-680, a small-molecule Aurora kinase inhibitor, interferes with mitotic entry and formation of bipolar spindles. Here, we evaluated VX-680 as a potential agent for treatment of all-trans retinoid acid (ATRA-resistant acute promyelocytic leukemia (APL in vitro. Methods CD11b expression was utilized to assess cell differentiation by flow cytometry. Immunofluorescence staining was conducted to analyze formation of cell monopolar spindle. Cell proliferation was evaluated by MTT assay. Sub-G1 population and Annexin V/PI staining were used to measure cell apoptosis. Hoechst 33342 staining was applied for identifying morphological changes in nucleus of apoptotic cell. Aurora-A (Aur-A activation and the signaling pathways involved in apoptosis were detected by Western blot. JC-1 probe was employed to measure mitochondrial depolarization. Results VX-680 inhibited Aur-A by reducing autophosphorylation at the activation site, Thr288, accompanied by producing monopolar mitotic spindles in APL cell line NB4-R2 that was resistant to ATRA. In addition, we found that VX-680 inhibited cell proliferation as assessed by MTT assay. Flow cytometry showed that VX-680 led to apoptotic cell death in both dose- and time-dependent manners by either Sub-G1 or Annexin V/PI analysis. Hoechst 33342 staining represented typical apoptotic cells with nuclear fragmentation in VX-680 treated cells. Importantly, VX-680 inhibition of Aurora kinase suppressed Akt-1 activation and induced mitochondrial depolarization, which eventually resulted in apoptosis by activation of caspase pathway, as indicated by increasing proteolytic cleavage of procaspase-3 and poly ADP ribose polymerase (PARP in NB4-R2 cells. Conclusions Our study suggested potential clinical use of mitotic Aurora kinase inhibitor in targeting ATRA-resistant leukemic cells.

  18. Aurora B kinase and protein phosphatase 1 have opposing roles in modulating kinetochore assembly.

    Science.gov (United States)

    Emanuele, Michael J; Lan, Weijie; Jwa, Miri; Miller, Stephanie A; Chan, Clarence S M; Stukenberg, P Todd

    2008-04-21

    The outer kinetochore binds microtubules to control chromosome movement. Outer kinetochore assembly is restricted to mitosis, whereas the inner kinetochore remains tethered to centromeres throughout the cell cycle. The cues that regulate this transient assembly are unknown. We find that inhibition of Aurora B kinase significantly reduces outer kinetochore assembly in Xenopus laevis and human tissue culture cells, frog egg extracts, and budding yeast. In X. leavis M phase extracts, preassembled kinetochores disassemble after inhibiting Aurora B activity with either drugs or antibodies. Kinetochore disassembly, induced by Aurora B inhibition, is rescued by restraining protein phosphatase 1 (PP1) activity. PP1 is necessary for kinetochores to disassemble at the exit from M phase, and purified enzyme is sufficient to cause disassembly on isolated mitotic nuclei. These data demonstrate that Aurora B activity is required for kinetochore maintenance and that PP1 is necessary and sufficient to disassemble kinetochores. We suggest that Aurora B and PP1 coordinate cell cycle-dependent changes in kinetochore assembly though phosphorylation of kinetochore substrates.

  19. Significance of Aurora B overexpression in hepatocellular carcinoma. Aurora B Overexpression in HCC

    Directory of Open Access Journals (Sweden)

    Lin Zhong-Zhe

    2010-08-01

    Full Text Available Abstract Background To investigate the significance of Aurora B expression in hepatocellular carcinoma (HCC. Methods The Aurora B and Aurora A mRNA level was measured in 160 HCCs and the paired nontumorous liver tissues by reverse transcription-polymerase chain reaction. Mutations of the p53 and β-catenin genes were analyzed in 134 and 150 tumors, respectively, by direct sequencing of exon 2 to exon 11 of p53 and exon 3 of β-catenin. Anticancer effects of AZD1152-HQPA, an Aurora B kinase selective inhibitor, were examined in Huh-7 and Hep3B cell lines. Results Aurora B was overexpressed in 98 (61% of 160 HCCs and in all 7 HCC cell lines examined. The overexpression of Aurora B was associated with Aurora A overexpression (P = 0.0003 and p53 mutation (P = 0.002 and was inversely associated with β-catenin mutation (P = 0.002. Aurora B overexpression correlated with worse clinicopathologic characteristics. Multivariate analysis confirmed that Aurora B overexpression was an independent poor prognostic factor, despite its interaction with Aurora A overexpression and mutations of p53 and β-catenin. In Huh-7 and Hep3B cells, AZD1152-HQPA induced proliferation blockade, histone H3 (Ser10 dephosphorylation, cell cycle disturbance, and apoptosis. Conclusion Aurora B overexpression is an independent molecular marker predicting tumor invasiveness and poor prognosis of HCC. Aurora B kinase selective inhibitors are potential therapeutic agents for HCC treatment.

  20. Aurora A kinase regulates proper spindle positioning in C. elegans and in human cells.

    Science.gov (United States)

    Kotak, Sachin; Afshar, Katayon; Busso, Coralie; Gönczy, Pierre

    2016-08-01

    Accurate spindle positioning is essential for error-free cell division. The one-cell Caenorhabditis elegans embryo has proven instrumental for dissecting mechanisms governing spindle positioning. Despite important progress, how the cortical forces that act on astral microtubules to properly position the spindle are modulated is incompletely understood. Here, we report that the PP6 phosphatase PPH-6 and its associated subunit SAPS-1, which positively regulate pulling forces acting on spindle poles, associate with the Aurora A kinase AIR-1 in C. elegans embryos. We show that acute inactivation of AIR-1 during mitosis results in excess pulling forces on astral microtubules. Furthermore, we uncover that AIR-1 acts downstream of PPH-6-SAPS-1 in modulating spindle positioning, and that PPH-6-SAPS-1 negatively regulates AIR-1 localization at the cell cortex. Moreover, we show that Aurora A and the PP6 phosphatase subunit PPP6C are also necessary for spindle positioning in human cells. There, Aurora A is needed for the cortical localization of NuMA and dynein during mitosis. Overall, our work demonstrates that Aurora A kinases and PP6 phosphatases have an ancient function in modulating spindle positioning, thus contributing to faithful cell division.

  1. Tension sensing by Aurora B kinase is independent of survivin-based centromere localization.

    Science.gov (United States)

    Campbell, Christopher S; Desai, Arshad

    2013-05-01

    Accurate segregation of the replicated genome requires chromosome biorientation on the spindle. Biorientation is ensured by Aurora B kinase (Ipl1), a member of the four-subunit chromosomal passenger complex (CPC). Localization of the CPC to the inner centromere is central to the current model for how tension ensures chromosome biorientation: kinetochore-spindle attachments that are not under tension remain close to the inner centromere and are destabilized by Aurora B phosphorylation, whereas kinetochores under tension are pulled away from the influence of Aurora B, stabilizing their microtubule attachments. Here we show that an engineered truncation of the Sli15 (known as INCENP in humans) subunit of budding yeast CPC that eliminates association with the inner centromere nevertheless supports proper chromosome segregation during both mitosis and meiosis. Truncated Sli15 suppresses the deletion phenotypes of the inner-centromere-targeting proteins survivin (Bir1), borealin (Nbl1), Bub1 and Sgo1 (ref. 6). Unlike wild-type Sli15, truncated Sli15 localizes to pre-anaphase spindle microtubules. Premature targeting of full-length Sli15 to microtubules by preventing Cdk1 (also known as Cdc28) phosphorylation also suppresses the inviability of Bir1 deletion. These results suggest that activation of Aurora B kinase by clustering either on chromatin or on microtubules is sufficient for chromosome biorientation.

  2. Polo-like kinase-1 controls Aurora A destruction by activating APC/C-Cdh1.

    Directory of Open Access Journals (Sweden)

    Renske van Leuken

    Full Text Available Polo-like kinase-1 (Plk1 is activated before mitosis by Aurora A and its cofactor Bora. In mitosis, Bora is degraded in a manner dependent on Plk1 kinase activity and the E3 ubiquitin ligase SCF-betaTrCP. Here, we show that Plk1 is also required for the timely destruction of its activator Aurora A in late anaphase. It has been shown that Aurora A destruction is controlled by the auxiliary subunit Cdh1 of the Anaphase-Promoting Complex/Cyclosome (APC/C. Remarkably, we found that Plk1-depletion prevented the efficient dephosphorylation of Cdh1 during mitotic exit. Plk1 mediated its effect on Cdh1, at least in part, through direct phosphorylation of the human phosphatase Cdc14A, controlling the phosphorylation state of Cdh1. We conclude that Plk1 facilitates efficient Aurora A degradation through APC/C-Cdh1 activation after mitosis, with a potential role for hCdc14A.

  3. In vitro evaluation of Aurora kinase inhibitor—VX680—in formulation of PLA-TPGS nanoparticles

    Science.gov (United States)

    Thuy Duong Le, Thi; Thu Ha, Phuong; Hai Yen Tran, Thi; Nguyen, Dac Tu; Nguyen, Hoai Nam; Khanh Bui, Van; Nhung Hoang, My

    2016-06-01

    Polymeric nanoparticles prepared from poly(lactide)-tocopheryl polyethylene glycol succinate (PLA-TPGS) were used as potential drug carries with many advantages to overcome the disadvantages of insoluble anticancer drugs and enhance blood circulation time and tissues. VX680 is an Aurora kinase inhibitor and is also the foremost Aurora kinase inhibitor to be studied in clinical trials. In this study, we aimed to investigate whether VX680-loaded PLA-TPGS nanoparticles (VX680-NPs) are able to effectively increase the toxicity of chemotherapy. Accordingly, we first synthesized VX680-loaded nanoparticles and NP characterizations of morphology, mean size, zeta potential, and encapsulation efficiency were spherical shape, 63 nm, -30 mV and 76%, respectively. Then, we investigated the effects on HeLa cells. The cell cytotoxicity was evaluated by the xCELLigence real-time cell analyzer allowing measurement of changes in electrical impedance on the surface of the E-plate. Analysis of nucleus morphology and level of histone H3 phosphorylation was observed by confocal fluorescence scanning microscopy. Cell cycle distribution and apoptosis were analyzed by flow cytometry. Our results showed that VX680-NPs reduced cell viability with IC50 value lower 3.4 times compared to free VX680. Cell proliferation was inhibited by VX680-NPs accompanied by other effects such as high abnormal changes of nucleus, a decrease of phospho-histone H3 at Ser10 level, an increase of polyploid cells and resulted in higher apoptotic cells. These results demonstrated that VX680-NPs had more cytotoxicity than as treated with VX680 alone. Thus, VX680-NPs may be considered as promising drug delivery system for cancer treatment.

  4. The molecular mechanism studies of chirality effect of PHA-739358 on Aurora kinase A by molecular dynamics simulation and free energy calculations

    Science.gov (United States)

    Cheng, Yuanhua; Cui, Wei; Chen, Quan; Tung, Chen-Ho; Ji, Mingjuan; Zhang, Fushi

    2011-02-01

    Aurora kinase family is one of the emerging targets in oncology drug discovery and several small molecules targeting aurora kinases have been discovered and evaluated under early phase I/II trials. Among them, PHA-739358 (compound 1r) is a 3-aminopyrazole derivative with strong activity against Aurora A under early phase II trial. Inhibitory potency of compound 1r (the benzylic substituent at the pro-R position) is 30 times over that of compound 1s (the benzylic substituent at the pro-S position). In present study, the mechanism of how different configurations influence the binding affinity was investigated using molecular dynamics (MD) simulations, free energy calculations and free energy decomposition analysis. The predicted binding free energies of these two complexes are consistent with the experimental data. The analysis of the individual energy terms indicates that although the van der Waals contribution is important for distinguishing the binding affinities of these two inhibitors, the electrostatic contribution plays a more crucial role in that. Moreover, it is observed that different configurations of the benzylic substituent could form different binding patterns with protein, thus leading to variant inhibitory potency of compounds 1r and 1s. The combination of different molecular modeling techniques is an efficient way to interpret the chirality effects of inhibitors and our work gives valuable information for the chiral drug design in the near future.

  5. TYROSINE KINASE INHIBITORS AND PREGNANCY

    Directory of Open Access Journals (Sweden)

    Elisabetta Abruzzese

    2014-04-01

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

  6. Ewing sarcoma EWS protein regulates midzone formation by recruiting Aurora B kinase to the midzone.

    Science.gov (United States)

    Park, Hyewon; Turkalo, Timothy K; Nelson, Kayla; Folmsbee, Stephen Sai; Robb, Caroline; Roper, Brittany; Azuma, Mizuki

    2014-01-01

    Ewing sarcoma is a malignant bone cancer that primarily occurs in children and adolescents. Eighty-five percent of Ewing sarcoma is characterized by the presence of the aberrant chimeric EWS/FLI1 fusion gene. Previously, we demonstrated that an interaction between EWS/FLI1 and wild-type EWS led to the inhibition of EWS activity and mitotic dysfunction. Although defective mitosis is considered to be a critical step in cancer initiation, it is unknown how interference with EWS contributes to Ewing sarcoma formation. Here, we demonstrate that EWS/FLI1- and EWS-knockdown cells display a high incidence of defects in the midzone, a midline structure located between segregating chromatids during anaphase. Defects in the midzone can lead to the failure of cytokinesis and can result in the induction of aneuploidy. The similarity among the phenotypes of EWS/FLI1- and EWS siRNA-transfected HeLa cells points to the inhibition of EWS as the key mechanism for the induction of midzone defects. Supporting this observation, the ectopic expression of EWS rescues the high incidence of midzone defects observed in Ewing sarcoma A673 cells. We discovered that EWS interacts with Aurora B kinase, and that EWS is also required for recruiting Aurora B to the midzone. A domain analysis revealed that the R565 in the RGG3 domain of EWS is essential for both Aurora B interaction and the recruitment of Aurora B to the midzone. Here, we propose that the impairment of EWS-dependent midzone formation via the recruitment of Aurora B is a potential mechanism of Ewing sarcoma development.

  7. Dynamic phosphorylation of Histone Deacetylase 1 by Aurora kinases during mitosis regulates zebrafish embryos development

    Science.gov (United States)

    Loponte, Sara; Segré, Chiara V.; Senese, Silvia; Miccolo, Claudia; Santaguida, Stefano; Deflorian, Gianluca; Citro, Simona; Mattoscio, Domenico; Pisati, Federica; Moser, Mirjam A.; Visintin, Rosella; Seiser, Christian; Chiocca, Susanna

    2016-01-01

    Histone deacetylases (HDACs) catalyze the removal of acetyl molecules from histone and non-histone substrates playing important roles in chromatin remodeling and control of gene expression. Class I HDAC1 is a critical regulator of cell cycle progression, cellular proliferation and differentiation during development; it is also regulated by many post-translational modifications (PTMs). Herein we characterize a new mitosis-specific phosphorylation of HDAC1 driven by Aurora kinases A and B. We show that this phosphorylation affects HDAC1 enzymatic activity and it is critical for the maintenance of a proper proliferative and developmental plan in a complex organism. Notably, we find that Aurora-dependent phosphorylation of HDAC1 regulates histone acetylation by modulating the expression of genes directly involved in the developing zebrafish central nervous system. Our data represent a step towards the comprehension of HDAC1 regulation by its PTM code, with important implications in unravelling its roles both in physiology and pathology. PMID:27458029

  8. Antitumor Activity of KW-2450 Against Triple-Negative Breast Cancer by Inhibiting Aurora A and B Kinases

    OpenAIRE

    Kai, Kazuharu; Kondo, Kimie; Wang, Xiaoping; Xie, Xuemei; Pitner, Mary K.; Reyes, Monica E.; Torres-Adorno, Angie M.; Masuda,Hiroko; Hortobagyi, Gabriel N.; Bartholomeusz, Chandra; Saya, Hideyuki; Tripathy, Debu; Sen, Subrata; Ueno, Naoto T

    2015-01-01

    Currently, no targeted drug is available for triple-negative breast cancer (TNBC), an aggressive breast cancer that does not express estrogen receptor, progesterone receptor, or HER2. TNBC has high mitotic activity, and since Aurora A and B mitotic kinases drive cell division and are overexpressed in tumors with a high mitotic index, we hypothesized that inhibiting Aurora A and B produces a significant antitumor effect in TNBC. We tested this hypothesis by determining the antitumor effects of...

  9. Allosteric small-molecule kinase inhibitors

    DEFF Research Database (Denmark)

    Wu, Peng; Clausen, Mads Hartvig; Nielsen, Thomas E.

    2015-01-01

    current barriers of kinase inhibitors, including poor selectivity and emergence of drug resistance. In spite of the small number of identified allosteric inhibitors in comparison with that of inhibitors targeting the ATP pocket, encouraging results, such as the FDA-approval of the first small...

  10. Scaffold oriented synthesis. Part 3: design, synthesis and biological evaluation of novel 5-substituted indazoles as potent and selective kinase inhibitors employing [2+3] cycloadditions.

    Science.gov (United States)

    Akritopoulou-Zanze, Irini; Wakefield, Brian D; Gasiecki, Alan; Kalvin, Douglas; Johnson, Eric F; Kovar, Peter; Djuric, Stevan W

    2011-03-01

    We report the synthesis and biological evaluation of 5-substituted indazoles and amino indazoles as kinase inhibitors. The compounds were synthesized in a parallel synthesis fashion from readily available starting materials employing [2+3] cycloaddition reactions and were evaluated against a panel of kinase assays. Potent inhibitors were identified for numerous kinases such as Rock2, Gsk3β, Aurora2 and Jak2.

  11. The Aurora Kinase in Trypanosoma brucei plays distinctive roles in metaphase-anaphase transition and cytokinetic initiation.

    Directory of Open Access Journals (Sweden)

    Ziyin Li

    2009-09-01

    Full Text Available Aurora B kinase is an essential regulator of chromosome segregation with the action well characterized in eukaryotes. It is also implicated in cytokinesis, but the detailed mechanism remains less clear, partly due to the difficulty in separating the latter from the former function in a growing cell. A chemical genetic approach with an inhibitor of the enzyme added to a synchronized cell population at different stages of the cell cycle would probably solve this problem. In the deeply branched parasitic protozoan Trypanosoma brucei, an Aurora B homolog, TbAUK1, was found to control both chromosome segregation and cytokinetic initiation by evidence from RNAi and dominant negative mutation. To clearly separate these two functions, VX-680, an inhibitor of TbAUK1, was added to a synchronized T. brucei procyclic cell population at different cell cycle stages. The unique trans-localization pattern of the chromosomal passenger complex (CPC, consisting of TbAUK1 and two novel proteins TbCPC1 and TbCPC2, was monitored during mitosis and cytokinesis by following the migration of the proteins tagged with enhanced yellow fluorescence protein in live cells with time-lapse video microscopy. Inhibition of TbAUK1 function in S-phase, prophase or metaphase invariably arrests the cells in the metaphase, suggesting an action of TbAUK1 in promoting metaphase-anaphase transition. TbAUK1 inhibition in anaphase does not affect mitotic exit, but prevents trans-localization of the CPC from the spindle midzone to the anterior tip of the new flagellum attachment zone for cytokinetic initiation. The CPC in the midzone is dispersed back to the two segregated nuclei, while cytokinesis is inhibited. In and beyond telophase, TbAUK1 inhibition has no effect on the progression of cytokinesis or the subsequent G1, S and G2 phases until a new metaphase is attained. There are thus two clearly distinct points of TbAUK1 action in T. brucei: the metaphase-anaphase transition and

  12. New benzimidazoles and their antitumor effects with Aurora A kinase and KSP inhibitory activities.

    Science.gov (United States)

    Abd El-All, Amira S; Magd-El-Din, Asmaa A; Ragab, Fatma A F; ElHefnawi, Mahmoud; Abdalla, Mohamed M; Galal, Shadia A; El-Rashedy, Ahmed A

    2015-07-01

    A newly synthesized series of anticancer compounds comprising thiazolo[3,2-a]pyrimidine derivatives 6a-q bearing a benzimidazole moiety was produced via a one-pot reaction of N-(4-(1H-benzo[d]imidazol-2-yl)phenyl)-2-cyanoacetamide 5 with 2-aminothiazole and an appropriate aromatic aldehyde. Compound 7 was obtained via the reaction of 4-(1H-benzo[d]imidazol-2yl)benzenamide 1 with carbon disulphide and methyl iodide in the presence of concentrated aqueous solution of NaOH, then treated with o-phenylenediamine to give N-(4-1H-benzo[d]imidazol-2-yl)phenyl)-1H-benzo[d]imidazol-2-amine 8. The structures of the newly synthesized compounds were confirmed by analytical and spectroscopic measurements (IR, MS, and (1) H NMR). The synthesized products were screened and studied for their in vitro antitumor activity against three human cancer cell lines (namely colorectal cancer cell line HCT116, human liver cancer cell line HepG2, and human ovarian cancer cell line A2780) and their Aurora A kinase and KSP inhibitory activities. All newly synthesized compounds revealed marked results comparable with the standard drug CK0106023. The compounds 6e and 6k of the thiazolopyrimidine derivatives were the most active compounds when tested against the three cell lines in comparison with the standard drug CK0106023, and showed potent dual KSP and Aurora A kinase inhibition.

  13. Aurora A Kinase Contributes to a Pole-Based Error Correction Pathway.

    Science.gov (United States)

    Ye, Anna A; Deretic, Jovana; Hoel, Christopher M; Hinman, Albert W; Cimini, Daniela; Welburn, Julie P; Maresca, Thomas J

    2015-07-20

    Chromosome biorientation, where sister kinetochores attach to microtubules (MTs) from opposing spindle poles, is the configuration that best ensures equal partitioning of the genome during cell division. Erroneous kinetochore-MT attachments are commonplace but are often corrected prior to anaphase. Error correction, thought to be mediated primarily by the centromere-enriched Aurora B kinase (ABK), typically occurs near spindle poles; however, the relevance of this locale is unclear. Furthermore, polar ejection forces (PEFs), highest near poles, can stabilize improper attachments by pushing mal-oriented chromosome arms away from spindle poles. Hence, there is a conundrum: erroneous kinetochore-MT attachments are weakened where PEFs are most likely to strengthen them. Here, we report that Aurora A kinase (AAK) opposes the stabilizing effect of PEFs. AAK activity contributes to phosphorylation of kinetochore substrates near poles and its inhibition results in chromosome misalignment and an increased incidence of erroneous kinetochore-MT attachments. Furthermore, AAK directly phosphorylates a site in the N-terminal tail of Ndc80/Hec1 that has been implicated in reducing the affinity of the Ndc80 complex for MTs when phosphorylated. We propose that an AAK activity gradient contributes to correcting mal-oriented kinetochore-MT attachments in the vicinity of spindle poles.

  14. LRD-22, a novel dual dithiocarbamatic acid ester, inhibits Aurora-A kinase and induces apoptosis and cell cycle arrest in HepG2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Huiling; Li, Ridong [State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing (China); Li, Li [Department of Cell Biology, School of Basic Medical Sciences, Peking University, Beijing (China); Ge, Zemei [State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing (China); Zhou, Rouli, E-mail: rlzhou@bjmu.edu.cn [Department of Cell Biology, School of Basic Medical Sciences, Peking University, Beijing (China); Li, Runtao, E-mail: lirt@bjmu.edu.cn [State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing (China)

    2015-02-27

    In this study we investigated the antitumor activity of the novel dual dithiocarbamatic acid ester LRD-22 in vitro and in vivo. Several cancer cell lines were employed to determine the effect of LRD-22 on cell growth, and the MTT assay showed there was a significant decrease in viable tumor cell numbers in the presence of LRD-22, especially in the HepG2 cell line. Colony formation assay also showed LRD-22 strongly inhibits HepG2 cell growth. Evaluation of the mechanism involved showed that inhibitory effects of LRD-22 on cell growth are due to induction of apoptosis and G2/M arrest. LRD-22 inhibited Aurora-A phosphorylation at Thr{sub 288} and subsequently impaired p53 phosphorylation at Ser{sub 315} which was associated with the proteasome degradation pathway. Tumor suppressor protein p53 is stabilized by this mechanism and accumulates through inhibition of Aurora-A kinase activity via treatment with LRD-22. In vivo study of HepG2 xenograft in nude mice also shows LRD-22 suppresses tumor growth at a concentration of 5 mg/kg without animals suffering loss of body weight. In conclusion, our results demonstrate LRD-22 acts as an Aurora-A kinase inhibitor to induce apoptosis and inhibit proliferation in HepG2 cells, and should be considered as a promising targeting agent for HCC therapy. - Highlights: • LRD-22 significantly inhibits cancer cell growth, especially in the HepG2 cell line. • The inhibitory effect of LRD-22 is due to induction of apoptosis and cell cycle arrest. • LRD-22 inhibits Aurora-A phosphorylation which results in subsequent impairment of the p53 pathway. • LRD-22 suppresses tumor growth in xenograft mice without body weight loss.

  15. Conformation-specific inhibitors of Raf kinases.

    Science.gov (United States)

    Wang, Xiaolun; Schleicher, Kristin

    2013-01-01

    Since the discovery linking B-Raf mutations to human tumors in 2002, significant advances in the development of Raf inhibitors have been made, leading to the recent approval of two Raf inhibitor drugs. This chapter includes a brief introduction to B-Raf as a validated target and focuses on the three different binding modes observed with Raf small-molecule inhibitors. These various binding modes lock the Raf kinase in different conformations that impact the toxicity profiles of the inhibitors. Possible solutions to mitigate the side effects caused by inhibitor-induced dimerization are also discussed.

  16. MK615 inhibits pancreatic cancer cell growth by dual inhibition of Aurora A and B kinases

    Institute of Scientific and Technical Information of China (English)

    Toshie Okada; Tokihiko Sawada; Tatsushi Osawa; Masakazu Adachi; Keiichi Kubota

    2008-01-01

    AIM:To investigate the anti-neoplastic effect of MK615,an anti-neoplastic compound isolated from Japanese apricot,against human pancreatic cancer cells in vitro.METHODS:Three human pancreatic cancer cell lines PANC-1,PK-1,and PK45H were cultured with MK615 at concentrations of 600,300,150,and O μg/mL.Growth inhibition was evaluated by cell proliferation assay,and killing activity was determined by lactate dehydrogenase (LDH) assay.Expression of Aurora A and B kinases was detected by real-time polymerase chain reaction (PCR) and Western blotting.Cell cycle stages were evaluated by flow cytometry.RESULTS:The growth inhibitory rates of MK615 at 150,300,and 600 μg/mL were 2.3% ± 0.9%,8.9% ±3.2% and 67.1% ± 8.1% on PANC1 cells,1.3% ± 0.3%,8.7% ± 4.1% and 45.7 ± 7.6% on PK1 cells,and 1.2 ±0.8%,9.1% ± 2.1% and 52.1% ± 5.5% on PK45H cells,respectively (P<0.05).The percentage cytotoxicities of MK615 at 0,150,300,and 600 μg/mL were 19.6% ±1.3%,26.7% ± 1.8%,25.5% ± 0.9% and 26.4% ± 0.9%in PANC1 cells,19.7% ± 1.3%,24.7% ± 0.8%,25.9% ±0.9% and 29.9% ± 1.1% in PK1 cells,and 28.0% ± 0.9%,31.2% ± 0.9%,30.4% ± 1.1% and 35.3 ± 1.0% in PK45H cells,respectively (P<0.05).Real-time PCR and Western blotting showed that MK615 dually inhibited the expression of Aurora A and B kinases.Cell cycle analysis revealed that MK615 increased the population of cells in G2/M phase.CONCLUSION:MK615 exerts an anti-neoplastic effect on human pancreatic cancer cells in vitro by dual inhibition of Aurora A and B kinases.

  17. Aurora kinase A induces papillary thyroid cancer lymph node metastasis by promoting cofilin-1 activity.

    Science.gov (United States)

    Maimaiti, Yusufu; Jie, Tan; Jing, Zhou; Changwen, Wang; Pan, Yu; Chen, Chen; Tao, Huang

    2016-04-22

    Aurora-A (Aur-A), a member of the serine/threonine Aurora kinase family, plays an important role in ensuring genetic stability during cell division. Previous studies indicated that Aur-A possesses oncogenic activity and may be a valuable therapeutic target in cancer therapy. However, the role of Aur-A in the most common thyroid cancer, papillary thyroid cancer (PTC), remains largely unknown. In patients with PTC, cancer cell migration and invasion account for most of the metastasis, recurrence, and cancer-related deaths. Cofilin-1 (CFL-1) is the most important effector of actin polymerization and depolymerization, determining the direction of cell migration. Here, we assessed the correlation between Aur-A and CFL-1 in PTC with lymph node metastasis. Tissue microarray data showed that simultaneous overexpression of Aur-A and CFL-1 correlated with lymph node metastasis in thyroid cancer tissue. Inhibition of Aur-A suppressed thyroid cancer cell migration in vitro and decreased lymph node metastasis in nude mice. Importantly, Aur-A increased the non-phosphorylated, active form of CFL-1 in TPC-1 cells, thus promoting cancer cell migration and thyroid cancer lymph node metastasis. Our findings indicate that the combination of Aur-A and CFL-1 may be useful as a molecular prediction model for lymph node metastasis in thyroid cancer and raise the possibility of targeting Aur-A and CFL-1 for more effective treatment of thyroid cancer.

  18. The Ski Protein is Involved in the Transformation Pathway of Aurora Kinase A.

    Science.gov (United States)

    Rivas, Solange; Armisén, Ricardo; Rojas, Diego A; Maldonado, Edio; Huerta, Hernán; Tapia, Julio C; Espinoza, Jaime; Colombo, Alicia; Michea, Luis; Hayman, Michael J; Marcelain, Katherine

    2016-02-01

    Oncogenic kinase Aurora A (AURKA) has been found to be overexpresed in several tumors including colorectal, breast, and hematological cancers. Overexpression of AURKA induces centrosome amplification and aneuploidy and it is related with cancer progression and poor prognosis. Here we show that AURKA phosphorylates in vitro the transcripcional co-repressor Ski on aminoacids Ser326 and Ser383. Phosphorylations on these aminoacids decreased Ski protein half-life. Reduced levels of Ski resulted in centrosomes amplification and multipolar spindles formation, same as AURKA overexpressing cells. Importantly, overexpression of Ski wild type, but not S326D and S383D mutants inhibited centrosome amplification and cellular transformation induced by AURKA. Altogether, these results suggest that the Ski protein is a target in the transformation pathway mediated by the AURKA oncogene.

  19. Rational design of protein kinase inhibitors

    Directory of Open Access Journals (Sweden)

    Yarmoluk S. M.

    2013-07-01

    Full Text Available Modern methodological approaches to rational design of low molecular weight compounds with specific activity in relation to predetermined biomolecular targets are considered by example of development of high effective protein kinase inhibitors. The application of new computational methods that allow to significantly improve the quality of computational experiments (in, particular, accuracy of low molecular weight compounds activity prediction without increase of computational and time costs are highlighted. The effectiveness of strategy of rational design is demonstrated by examples of several own investigations devoted to development of new inhibitors that are high effective and selective towards protein kinases CK2, FGFR1 and ASK1.

  20. Ipl1/Aurora kinase suppresses S-CDK-driven spindle formation during prophase I to ensure chromosome integrity during meiosis.

    Directory of Open Access Journals (Sweden)

    Louise Newnham

    Full Text Available Cells coordinate spindle formation with DNA repair and morphological modifications to chromosomes prior to their segregation to prevent cell division with damaged chromosomes. Here we uncover a novel and unexpected role for Aurora kinase in preventing the formation of spindles by Clb5-CDK (S-CDK during meiotic prophase I and when the DDR is active in budding yeast. This is critical since S-CDK is essential for replication during premeiotic S-phase as well as double-strand break induction that facilitates meiotic recombination and, ultimately, chromosome segregation. Furthermore, we find that depletion of Cdc5 polo kinase activity delays spindle formation in DDR-arrested cells and that ectopic expression of Cdc5 in prophase I enhances spindle formation, when Ipl1 is depleted. Our findings establish a new paradigm for Aurora kinase function in both negative and positive regulation of spindle dynamics.

  1. Glycogen synthase kinase 3 β activity is required for hBora/Aurora A-mediated mitotic entry.

    Science.gov (United States)

    Lee, Yu-Cheng; Liao, Po-Chi; Liou, Yih-Cherng; Hsiao, Michael; Huang, Chi-Ying; Lu, Pei-Jung

    2013-03-15

    The synthesis and degradation of hBora is important for the regulation of mitotic entry and exist. In G 2 phase, hBora can complex with Aurora A to activate Plk1 and control mitotic entry. However, whether the post-translational modification of hBora is relevant to the mitotic entry still unclear. Here, we used the LC-MS/MS phosphopeptide mapping assay to identify 13 in vivo hBora phosphorylation sites and characterized that GSK3β can interact with hBora and phosphorylate hBora at Ser274 and Ser278. Pharmacological inhibitors of GSK3β reduced the retarded migrating band of hBora in cells and diminished the phosphorylation of hBora by in vitro kinase assay. Moreover, as well as in GSK3β activity-inhibited cells, specific knockdown of GSK3β by shRNA and S274A/S278 hBora mutant-expressing cells also exhibited the reduced Plk1 activation and a delay in mitotic entry. It suggests that GSK3β activity is required for hBora-mediated mitotic entry through Ser274 and Ser278 phosphorylation.

  2. Aurora A Kinase Regulates Mammary Epithelial Cell Fate by Determining Mitotic Spindle Orientation in a Notch-Dependent Manner

    Directory of Open Access Journals (Sweden)

    Joseph L. Regan

    2013-07-01

    Full Text Available Cell fate determination in the progeny of mammary epithelial stem/progenitor cells remains poorly understood. Here, we have examined the role of the mitotic kinase Aurora A (AURKA in regulating the balance between basal and luminal mammary lineages. We find that AURKA is highly expressed in basal stem cells and, to a lesser extent, in luminal progenitors. Wild-type AURKA expression promoted luminal cell fate, but expression of an S155R mutant reduced proliferation, promoted basal fate, and inhibited serial transplantation. The mechanism involved regulation of mitotic spindle orientation by AURKA and the positioning of daughter cells after division. Remarkably, this was NOTCH dependent, as NOTCH inhibitor blocked the effect of wild-type AURKA expression on spindle orientation and instead mimicked the effect of the S155R mutant. These findings directly link AURKA, NOTCH signaling, and mitotic spindle orientation and suggest a mechanism for regulating the balance between luminal and basal lineages in the mammary gland.

  3. Therapeutic Innovations: Tyrosine Kinase Inhibitors in Cancer

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    Nikolaos Dervisis

    2016-01-01

    Full Text Available Conventional cytotoxic chemotherapy involving DNA-interacting agents and indiscriminate cell death is no longer the future of cancer management. While chemotherapy is not likely to completely disappear from the armamentarium; the use of targeted therapies in combination with conventional treatment is becoming the standard of care in human medicine. Tyrosine kinases are pivotal points of functional cellular pathways and have been implicated in malignancy, inflammatory, and immune-mediated diseases. Pharmaceutical interventions targeting aberrant tyrosine kinase signaling has exploded and is the second most important area of drug development. The “Valley of Death” between drug discovery and approval threatens to blunt the enormous strides in cancer management seen thus far. Kinase inhibitors, as targeted small molecules, hold promise in the treatment and diagnosis of cancer. However, there are still many unanswered questions regarding the use of kinase inhibitors in the interpretation and management of cancer. Comparative oncology has the potential to address restrictions and limitations in the advancement in kinase inhibitor therapy.

  4. Discovery of inhibitors of bacterial histidine kinases

    NARCIS (Netherlands)

    Velikova, N.R.

    2014-01-01

    Discovery of Inhibitors of Bacterial Histidine Kinases

    Summary

    The thesis is on novel antibacterial drug discovery (http://youtu.be/NRMWOGgeysM). Using structure-based and fragment-based dru

  5. The transcription factor YY1 is a novel substrate for Aurora B kinase at G2/M transition of the cell cycle.

    Directory of Open Access Journals (Sweden)

    Ari Kassardjian

    Full Text Available Yin Yang 1 (YY1 is a ubiquitously expressed and highly conserved multifunctional transcription factor that is involved in a variety of cellular processes. Many YY1-regulated genes have crucial roles in cell proliferation, differentiation, apoptosis, and cell cycle regulation. Numerous mechanisms have been shown to regulate the function of YY1, such as DNA binding affinity, subcellular localization, and posttranslational modification including phosphorylation. Polo-like kinase 1(Plk1 and Casein kinase 2α (CK2 α were the first two kinases identified to phosphorylate YY1. In this study, we identify a third kinase. We report that YY1 is a novel substrate of the Aurora B kinase both in vitro and in vivo. Serine 184 phosphorylation of YY1 by Aurora B is cell cycle regulated and peaks at G2/M and is rapidly dephosphorylated, likely by protein phosphatase 1 (PP1 as the cells enter G1. Aurora A and Aurora C can also phosphorylate YY1 in vitro, but at serine/threonine residues other than serine 184. We present evidence that phosphorylation of YY1 in the central glycine/alanine (G/A-rich region is important for DNA binding activity, with a potential phosphorylation/acetylation interplay regulating YY1 function. Given their importance in mitosis and overexpression in human cancers, Aurora kinases have been identified as promising therapeutic targets. Increasing our understanding of Aurora substrates will add to the understanding of their signaling pathways.

  6. [Effect of aurora inhibitor VX-680 on proliferation and apoptosis of CML cells].

    Science.gov (United States)

    Yin, Yue; Sun, Hui-Yan; Li, Xiao-Lin; Xiao, Feng-Jun; Wang, Li-Sheng

    2014-12-01

    This study was aimed to explore the effect of VX-680, an aurora inhibitor, on proliferation and apoptosis of K562, KCL22 cell lines and CD34⁺ cells from chronic myeloid leukemia (CML) patients in vitro. The proliferation of K562 and KCL22 cell was detected by CCK-8 method. Apoptosis of cells was detected by Annexin V-PI labeling and flow cytometry. The colony forming ability of bone marrow CD34⁺ cells derived from CML patients and donors was determined by the colony forming test. The results showed that the treatment of K562, KCL22 and CML CD34⁺ cells with VX-680 of 20-100 nmol/L for 3 days could obviously inhibit the cell proliferation in a concentration-dependent manner (P VX-680 treatment significantly induced apoptosis of K562 and KCL22 cells. Compared to bone marrow CD34⁺ cells derived from the healthy donors, the colony forming ability of CML CD34⁺ cells derived from bone marrow of CML patients was remarkably reduced (P VX-680, an aurora inhibitor, can inhibit the proliferation and induce apoptosis of CML cells in vitro.

  7. Inhibitors of p21-activated kinases (PAKs).

    Science.gov (United States)

    Rudolph, Joachim; Crawford, James J; Hoeflich, Klaus P; Wang, Weiru

    2015-01-08

    The p21-activated kinase (PAK) family of serine/threonine protein kinases plays important roles in cytoskeletal organization, cellular morphogenesis, and survival, and members of this family have been implicated in many diseases including cancer, infectious diseases, and neurological disorders. Owing to their large and flexible ATP binding cleft, PAKs, particularly group I PAKs (PAK1, -2, and -3), are difficult to drug; hence, few PAK inhibitors with satisfactory kinase selectivity and druglike properties have been reported to date. Examples are a recently discovered group II PAK (PAK4, -5, -6) selective inhibitor series based on a benzimidazole core, a group I PAK selective series based on a pyrido[2,3-d]pyrimidine-7-one core, and an allosteric dibenzodiazepine PAK1 inhibitor series. Only one compound, an aminopyrazole based pan-PAK inhibitor, entered clinical trials but did not progress beyond phase I trials. Clinical proof of concept for pan-group I, pan-group II, or PAK isoform selective inhibition has yet to be demonstrated.

  8. Delivery of DNAzyme targeting aurora kinase A to inhibit the proliferation and migration of human prostate cancer

    Directory of Open Access Journals (Sweden)

    Xing Z

    2015-09-01

    Full Text Available Zhen Xing,1 Sai Gao,1 Yan Duan,1 Haobo Han,1 Li Li,2 Yan Yang,1 Quanshun Li1 1Key Laboratory for Molecular Enzymology and Engineering of Ministry of Education, School of Life Sciences, Jilin University, 2Department of Clinic Library, Changchun Women and Children’s Health, Changchun, People’s Republic of China Abstract: Herein, a polyethylenimine derivative N-acetyl-L-leucine-polyethylenimine (N-Ac-L-Leu-PEI was employed as a carrier to achieve the delivery of DNAzyme targeting aurora kinase A using PC-3 cell as a model. Flow cytometry and confocal laser scanning microscopy demonstrated that the derivative could realize the cellular uptake of nanoparticles in an energy-dependent and clathrin-mediated pathway and obtain a high DNAzyme concentration in the cytoplasm through further endosomal escape. After DNAzyme transfection, expression level of aurora kinase A would be downregulated at the protein level. Meanwhile, the inhibition of cell proliferation was observed through 3-(4,5-Dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide and cell colony formation assay, attributing to the activation of apoptosis and cell cycle arrest. Through flow cytometric analysis, an early apoptotic ratio of 25.93% and G2 phase of 22.58% has been detected after N-Ac-L-Leu-PEI-mediated DNAzyme transfection. Finally, wound healing and Transwell migration assay showed that DNAzyme transfection could efficiently inhibit the cell migration. These results demonstrated that N-Ac-L-Leu-PEI could successfully mediate the DNAzyme delivery and downregulate the expression level of aurora kinase A, triggering a significant inhibitory effect of excessive proliferation and migration of tumor cells. Keywords: gene therapy, DNAzyme, aurora kinase A, N-acetyl-L-leucine-polyethylenimine, cell proliferation, cell migration

  9. Depletion of Aurora-A in zebrafish causes growth retardation due to mitotic delay and p53-dependent cell death.

    Science.gov (United States)

    Jeon, Hee-Yeon; Lee, Hyunsook

    2013-03-01

    Aurora-A is a serine/threonine mitotic kinase that is required for centrosome maturation. Many cancer cells over-express Aurora-A, and several reports have suggested that Aurora-A has prognostic value in the clinical treatment of cancer. Therefore, inhibitors for Aurora-A kinase have been developed. However, studies on Aurora-A are largely performed in cancer cell lines and are sometimes controversial. For effective evaluation of Aurora-A inhibitors in cancer treatment, it is essential to understand its function at the organism level. Here, we report the crucial functions of Aurora-A in homeostasis of spindle organization in mitosis using zebrafish embryogenesis as a model system. Using morpholino technology, we show that depletion of Aurora-A in zebrafish embryogenesis results in short bent trunks, accompanied by growth retardation and eventual cell death. Live-imaging and immunofluorescence analyses of the embryos revealed that the developmental defects are due to problems in mitosis, manifested through monopolar and disorganized spindle formation. Aurora-A-depleted cells exhibited mitotic arrest with congression failure, leading to activation of the spindle assembly checkpoint. Cell death in the absence of Aurora-A was partially rescued by co-injection of the p53 morpholino, suggesting that apoptosis after Aurora-A depletion is p53-dependent. The clinical implications of these results relate to the indication that Aurora-A inhibitors may be effective towards cancers with intact p53.

  10. Aurora B is regulated by the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway and is a valuable potential target in melanoma cells.

    Science.gov (United States)

    Bonet, Caroline; Giuliano, Sandy; Ohanna, Mickaël; Bille, Karine; Allegra, Maryline; Lacour, Jean-Philippe; Bahadoran, Philippe; Rocchi, Stéphane; Ballotti, Robert; Bertolotto, Corine

    2012-08-24

    Metastatic melanoma is a deadly skin cancer and is resistant to almost all existing treatment. Vemurafenib, which targets the BRAFV600E mutation, is one of the drugs that improves patient outcome, but the patients next develop secondary resistance and a return to cancer. Thus, new therapeutic strategies are needed to treat melanomas and to increase the duration of v-Raf murine sarcoma viral oncogene homolog B1 (BRAF) inhibitor response. The ERK pathway controls cell proliferation, and Aurora B plays a pivotal role in cell division. Here, we confirm that Aurora B is highly expressed in metastatic melanoma cells and that Aurora B inhibition triggers both senescence-like phenotypes and cell death in melanoma cells. Furthermore, we show that the BRAF/ERK axis controls Aurora B expression at the transcriptional level, likely through the transcription factor FOXM1. Our results provide insight into the mechanism of Aurora B regulation and the first molecular basis of Aurora B regulation in melanoma cells. The inhibition of Aurora B expression that we observed in vemurafenib-sensitive melanoma cells was rescued in cells resistant to this drug. Consistently, these latter cells remain sensitive to the effect of the Aurora B inhibitor. Noteworthy, wild-type BRAF melanoma cells are also sensitive to Aurora B inhibition. Collectively, our findings, showing that Aurora B is a potential target in melanoma cells, particularly in those vemurafenib-resistant, may open new avenues to improve the treatment of metastatic melanoma.

  11. Aurora A's functions during mitotic exit: the Guess Who game

    Directory of Open Access Journals (Sweden)

    David eReboutier

    2015-12-01

    Full Text Available Until recently, the knowledge of Aurora A kinase functions during mitosis was limited to pre-metaphase events, particularly centrosome maturation, G2/M transition, and mitotic spindle assembly. However, an involvement of Aurora A in post-metaphase events was also suspected, but not clearly demonstrated due to the technical difficulty to perform the appropriate experiments. Recent developments of both an analog specific version of Aurora A, and of small molecule inhibitors have led to the first demonstration that Aurora A is required for the early steps of cytokinesis. As in pre-metaphase, Aurora A plays diverse functions during anaphase, essentially participating in astral microtubules dynamics and central spindle assembly and functioning. The present review describes the experimental systems used to decipher new functions of Aurora A during late mitosis and situate these functions into the context of cytokinesis mechanisms.

  12. Recent advances in designing substrate-competitive protein kinase inhibitors.

    Science.gov (United States)

    Han, Ki-Cheol; Kim, So Yeon; Yang, Eun Gyeong

    2012-01-01

    Protein kinases play central roles in cellular signaling pathways and their abnormal phosphorylation activity is inseparably linked with various human diseases. Therefore, modulation of kinase activity using potent inhibitors is an attractive strategy for the treatment of human disease. While most protein kinase inhibitors in clinical development are mainly targeted to the highly conserved ATP-binding sites and thus likely promiscuously inhibit multiple kinases including kinases unrelated to diseases, protein substrate-competitive inhibitors are more selective and expected to be promising therapeutic agents. Most substrate-competitive inhibitors mimic peptides derived from substrate proteins, or from inhibitory domains within kinases or inhibitor proteins. In addition, bisubstrate inhibitors are generated by conjugating substrate-competitive peptide inhibitors to ATP-competitive inhibitors to improve affinity and selectivity. Although structural information on protein kinases provides invaluable guidance in designing substrate-competitive inhibitors, other strategies including bioinformatics, computational modeling, and high-throughput screening are often employed for developing specific substrate-competitive kinase inhibitors. This review focuses on recent advances in the design and discovery of substrate-competitive inhibitors of protein kinases.

  13. Occurrence of multipolar mitoses and association with Aurora-A/-B kinases and p53 mutations in aneuploid esophageal carcinoma cells

    Directory of Open Access Journals (Sweden)

    Münch Claudia

    2011-04-01

    Full Text Available Abstract Background Aurora kinases and loss of p53 function are implicated in the carcinogenesis of aneuploid esophageal cancers. Their association with occurrence of multipolar mitoses in the two main histotypes of aneuploid esophageal squamous cell carcinoma (ESCC and Barrett's adenocarcinoma (BAC remains unclear. Here, we investigated the occurrence of multipolar mitoses, Aurora-A/-B gene copy numbers and expression/activation as well as p53 alterations in aneuploid ESCC and BAC cancer cell lines. Results A control esophageal epithelial cell line (EPC-hTERT had normal Aurora-A and -B gene copy numbers and expression, was p53 wild type and displayed bipolar mitoses. In contrast, both ESCC (OE21, Kyse-410 and BAC (OE33, OE19 cell lines were aneuploid and displayed elevated gene copy numbers of Aurora-A (chromosome 20 polysomy: OE21, OE33, OE19; gene amplification: Kyse-410 and Aurora-B (chromosome 17 polysomy: OE21, Kyse-410. Aurora-B gene copy numbers were not elevated in OE19 and OE33 cells despite chromosome 17 polysomy. Aurora-A expression and activity (Aurora-A/phosphoT288 was not directly linked to gene copy numbers and was highest in Kyse-410 and OE33 cells. Aurora-B expression and activity (Aurora-B/phosphoT232 was higher in OE21 and Kyse-410 than in OE33 and OE19 cells. The mitotic index was highest in OE21, followed by OE33 > OE19 > Kyse-410 and EPC-hTERT cells. Multipolar mitoses occurred with high frequency in OE33 (13.8 ± 4.2%, followed by OE21 (7.7 ± 5.0% and Kyse-410 (6.3 ± 2.0% cells. Single multipolar mitoses occurred in OE19 (1.0 ± 1.0% cells. Distinct p53 mutations and p53 protein expression patterns were found in all esophageal cancer cell lines, but complete functional p53 inactivation occurred in OE21 and OE33 only. Conclusions High Aurora-A expression alone is not associated with overt multipolar mitoses in aneuploid ESCC and BAC cancer cells, as specifically shown here for OE21 and OE33 cells, respectively

  14. Bisarylmaleimides & the Corresponding Indolocarbazoles as Kinase Inhibitors

    Institute of Scientific and Technical Information of China (English)

    Zhu Guoxin; Cathy Ogg; Bharvin Patel; Richard M. Schultz; Charles D. Spencer; Beverly Teicher; Scou A. Watkins; Scott E. Conner; Zhou Xun; Chuan Shih; Li Tiechao; Harold B. Brooks; Eileen Considine; Jack A. Dempsey; Margaret M. Faul

    2004-01-01

    Cyclin dependent kinases (CDKs) have recently raised considerable attention because of their central role in the regulation of cell cycle progression. A high incidence of genetic mutation of CDK substrates and deregulaton of CDK modulators were found in a number of disease states,particularly in cancer. A novel series of unsymmetrical substituted indolocarbazoles were synthesized and their kinase inhibitory capability was evaluated in vitro. 6-Substtuted indolocarbazoles were found to be highly potent and selective D1/CDK4 inhibitors. These indolocarbazoles exhibited ATP competitive D1/CDK4 activity and inhibited tumor cell growth,arrested tumor cell at G1 phase. These molecules demonstrated potent anti-tumor activity and inhibited pRb phosphorylation at S780 in the human lung carcinoma (Calu6) and non-small cell lung carcinoma (NCI-H460) xenograft models. The results indicate that these small molecules have potential as therapeutic agents in cancer chemotherapeutc agents.

  15. ENMD-2076 is an orally active kinase inhibitor with antiangiogenic and antiproliferative mechanisms of action.

    Science.gov (United States)

    Fletcher, Graham C; Brokx, Richard D; Denny, Trisha A; Hembrough, Todd A; Plum, Stacy M; Fogler, William E; Sidor, Carolyn F; Bray, Mark R

    2011-01-01

    ENMD-2076 is a novel orally active, small molecule kinase inhibitor with a mechanism of action involving several pathways key to tumor growth and survival: angiogenesis, proliferation, and the cell cycle. ENMD-2076 has selective activity against the mitotic kinase Aurora A, as well as kinases involved in angiogenesis (VEGFRs, FGFRs). ENMD-2076 inhibited the growth in vitro of a wide range of human solid tumor and hematopoietic cancer cell lines with IC(50) values ranging from 0.025 to 0.7 μmol/L. ENMD-2076 was also shown to induce regression or complete inhibition of tumor growth in vivo at well-tolerated doses in tumor xenograft models derived from breast, colon, melanoma, leukemia, and multiple myeloma cell lines. Pharmacodynamic experiments in vivo showed that in addition to inhibiting Aurora A, single doses of ENMD-2076 had sustained inhibitory effects on the activation of Flt3 as well as the angiogenic tyrosine kinases, VEGFR2/KDR and FGFR1 and 2. ENMD-2076 was shown to prevent the formation of new blood vessels and regress formed vessels in vivo at doses equivalent to those that gave substantial activity in tumor xenograft models. These results indicate that ENMD-2076 is a well-tolerated, orally active multitarget kinase inhibitor with a unique antiangiogenic/antiproliferative profile and provides strong preclinical support for use as a therapeutic for human cancers. Several phase 1 studies involving ENMD-2076 have been recently completed, and the compound is currently being evaluated in a phase 2 clinical trial in patients with platinum-resistant ovarian cancer.

  16. Aurora kinase A is not involved in CPEB1 phosphorylation and cyclin B1 mRNA polyadenylation during meiotic maturation of porcine oocytes.

    Science.gov (United States)

    Komrskova, Pavla; Susor, Andrej; Malik, Radek; Prochazkova, Barbora; Liskova, Lucie; Supolikova, Jaroslava; Hladky, Stepan; Kubelka, Michal

    2014-01-01

    Regulation of mRNA translation by cytoplasmic polyadenylation is known to be important for oocyte maturation and further development. This process is generally controlled by phosphorylation of cytoplasmic polyadenylation element binding protein 1 (CPEB1). The aim of this study is to determine the role of Aurora kinase A in CPEB1 phosphorylation and the consequent CPEB1-dependent polyadenylation of maternal mRNAs during mammalian oocyte meiosis. For this purpose, we specifically inhibited Aurora kinase A with MLN8237 during meiotic maturation of porcine oocytes. Using poly(A)-test PCR method, we monitored the effect of Aurora kinase A inhibition on poly(A)-tail extension of long and short cyclin B1 encoding mRNAs as markers of CPEB1-dependent cytoplasmic polyadenylation. Our results show that inhibition of Aurora kinase A activity impairs neither cyclin B1 mRNA polyadenylation nor its translation and that Aurora kinase A is unlikely to be involved in CPEB1 activating phosphorylation.

  17. Aurora kinase A is not involved in CPEB1 phosphorylation and cyclin B1 mRNA polyadenylation during meiotic maturation of porcine oocytes.

    Directory of Open Access Journals (Sweden)

    Pavla Komrskova

    Full Text Available Regulation of mRNA translation by cytoplasmic polyadenylation is known to be important for oocyte maturation and further development. This process is generally controlled by phosphorylation of cytoplasmic polyadenylation element binding protein 1 (CPEB1. The aim of this study is to determine the role of Aurora kinase A in CPEB1 phosphorylation and the consequent CPEB1-dependent polyadenylation of maternal mRNAs during mammalian oocyte meiosis. For this purpose, we specifically inhibited Aurora kinase A with MLN8237 during meiotic maturation of porcine oocytes. Using poly(A-test PCR method, we monitored the effect of Aurora kinase A inhibition on poly(A-tail extension of long and short cyclin B1 encoding mRNAs as markers of CPEB1-dependent cytoplasmic polyadenylation. Our results show that inhibition of Aurora kinase A activity impairs neither cyclin B1 mRNA polyadenylation nor its translation and that Aurora kinase A is unlikely to be involved in CPEB1 activating phosphorylation.

  18. Functional analysis of phosphorylation of the mitotic centromere-associated kinesin by Aurora B kinase in human tumor cells.

    Science.gov (United States)

    Ritter, Andreas; Sanhaji, Mourad; Friemel, Alexandra; Roth, Susanne; Rolle, Udo; Louwen, Frank; Yuan, Juping

    2015-01-01

    Mitotic centromere-associated kinesin (MCAK) is the best characterized member of the kinesin-13 family and plays important roles in microtubule dynamics during mitosis. Its activity and subcellular localization is tightly regulated by an orchestra of mitotic kinases, such as Aurora B. It is well known that serine 196 of MCAK is the major phosphorylation site of Aurora B in Xenopus leavis extracts and that this phosphorylation regulates its catalytic activity and subcellular localization. In the current study, we have addressed the conserved phosphorylation site serine 192 in human MCAK to characterize its function in more depth in human cancer cells. Our data confirm that S192 is the major phosphorylation site of Aurora B in human MCAK and that this phosphorylation has crucial roles in regulating its catalytic activity and localization at the kinetochore/centromere region in mitosis. Interfering with this phosphorylation leads to a delayed progression through prometa- and metaphase associated with mitotic defects in chromosome alignment and segregation. We show further that MCAK is involved in directional migration and invasion of tumor cells, and interestingly, interference with the S192 phosphorylation affects this capability of MCAK. These data provide the first molecular explanation for clinical observation, where an overexpression of MCAK was associated with lymphatic invasion and lymph node metastasis in gastric and colorectal cancer patients.

  19. Stathmin and microtubules regulate mitotic entry in HeLa cells by controlling activation of both Aurora kinase A and Plk1.

    Science.gov (United States)

    Silva, Victoria C; Cassimeris, Lynne

    2013-12-01

    Depletion of stathmin, a microtubule (MT) destabilizer, delays mitotic entry by ∼4 h in HeLa cells. Stathmin depletion reduced the activity of CDC25 and its upstream activators, Aurora A and Plk1. Chemical inhibition of both Aurora A and Plk1 was sufficient to delay mitotic entry by 4 h, while inhibiting either kinase alone did not cause a delay. Aurora A and Plk1 are likely regulated downstream of stathmin, because the combination of stathmin knockdown and inhibition of Aurora A and Plk1 was not additive and again delayed mitotic entry by 4 h. Aurora A localization to the centrosome required MTs, while stathmin depletion spread its localization beyond that of γ-tubulin, indicating an MT-dependent regulation of Aurora A activation. Plk1 was inhibited by excess stathmin, detected in in vitro assays and cells overexpressing stathmin-cyan fluorescent protein. Recruitment of Plk1 to the centrosome was delayed in stathmin-depleted cells, independent of MTs. It has been shown that depolymerizing MTs with nocodazole abrogates the stathmin-depletion induced cell cycle delay; in this study, depolymerization with nocodazole restored Plk1 activity to near normal levels, demonstrating that MTs also contribute to Plk1 activation. These data demonstrate that stathmin regulates mitotic entry, partially via MTs, to control localization and activation of both Aurora A and Plk1.

  20. Inherent formulation issues of kinase inhibitors.

    Science.gov (United States)

    Herbrink, M; Schellens, J H M; Beijnen, J H; Nuijen, B

    2016-10-10

    The small molecular Kinase Inhibitor (smKI) drug class is very promising and rapidly expanding. All of these drugs are administered orally. The clear relationship between structure and function has led to drugs with a general low intrinsic solubility. The majority of the commercial pharmaceutical formulations of the smKIs are physical mixtures that are limited by the low drug solubility of a salt form. This class of drugs is therefore characterized by an impaired and variable bioavailability rendering them costly and their therapies suboptimal. New formulations are sparingly being reported in literature and patents. The presented data suggests that continued research into formulation design can help to develop more efficient and cost-effective smKI formulation. Moreover, it may also be of help in the future design of the formulations of new smKIs.

  1. Differential effects on cell motility, embryonic stem cell self-renewal and senescence by diverse Src kinase family inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Tamm, Christoffer, E-mail: christoffer.tamm@imbim.uu.se; Galito, Sara Pijuan, E-mail: sara.pijuan@imbim.uu.se; Anneren, Cecilia, E-mail: cecilia.anneren@imbim.uu.se

    2012-02-15

    The Src family of non-receptor tyrosine kinases (SFKs) has been shown to play an intricate role in embryonic stem (ES) cell maintenance. In the present study we have focused on the underlying molecular mechanisms responsible for the vastly different effects induced by various commonly used SFK inhibitors. We show that several diverse cell types, including fibroblasts completely lacking SFKs, cannot undergo mitosis in response to SU6656 and that this is caused by an unselective inhibition of Aurora kinases. In contrast, PP2 and PD173952 block motility immediately upon exposure and forces cells to grow in dense colonies. The subsequent halt in proliferation of fibroblast and epithelial cells in the center of the colonies approximately 24 h post-treatment appears to be caused by cell-to-cell contact inhibition rather than a direct effect of SFK kinase inhibition. Interestingly, in addition to generating more homogenous and dense ES cell cultures, without any diverse effect on proliferation, PP2 and PD173652 also promote ES cell self-renewal by reducing the small amount of spontaneous differentiation typically observed under standard ES cell culture conditions. These effects could not be mirrored by the use of Gleevec, a potent inhibitor of c-Abl and PDGFR kinases that are also inhibited by PP2. -- Highlights: Black-Right-Pointing-Pointer SFK inhibitor SU6656 induces senescence in mouse ES cells. Black-Right-Pointing-Pointer SU6656 inhibits mitosis in a SFK-independent manner via cross-selectivity for Aurora kinases. Black-Right-Pointing-Pointer SFK inhibitor PP2 impairs cell motility in various cell lines, including mouse ES cells. Black-Right-Pointing-Pointer Ensuing impeded motility, PP2 inhibits proliferation of various cells lines except for mouse ES cells. Black-Right-Pointing-Pointer SFK inhibitors PP2 and PD173952 impede spontaneous differentiation in standard mouse ES culture maintenance.

  2. Clinical stage EGFR inhibitors irreversibly alkylate Bmx kinase.

    Science.gov (United States)

    Hur, Wooyoung; Velentza, Anastasia; Kim, Sungjoon; Flatauer, Laura; Jiang, Xinnong; Valente, David; Mason, Daniel E; Suzuki, Melissa; Larson, Brad; Zhang, Jianming; Zagorska, Anna; Didonato, Michael; Nagle, Advait; Warmuth, Markus; Balk, Steven P; Peters, Eric C; Gray, Nathanael S

    2008-11-15

    Irreversible HER/erbB inhibitors selectively inhibit HER-family kinases by targeting a unique cysteine residue located within the ATP-binding pocket. Sequence alignment reveals that this rare cysteine is also present in ten other protein kinases including all five Tec-family members. We demonstrate that the Tec-family kinase Bmx is potently inhibited by irreversible modification at Cys496 by clinical stage EGFR inhibitors such as CI-1033. This cross-reactivity may have significant clinical implications.

  3. Skin problems and EGFR-tyrosine kinase inhibitor.

    Science.gov (United States)

    Kozuki, Toshiyuki

    2016-04-01

    Epidermal growth factor receptor inhibition is a good target for the treatment of lung, colon, pancreatic and head and neck cancers. Epidermal growth factor receptor-tyrosine kinase inhibitor was first approved for the treatment of advanced lung cancer in 2002. Epidermal growth factor receptor-tyrosine kinase inhibitor plays an essential role in the treatment of cancer, especially for patients harbouring epidermal growth factor receptor activating mutation. Hence, skin toxicity is the most concerning issue for the epidermal growth factor receptor-tyrosine kinase inhibitor treatment. Skin toxicity is bothersome and sometimes affects the quality of life and treatment compliance. Thus, it is important for physicians to understand the background and how to manage epidermal growth factor receptor-tyrosine kinase inhibitor-associated skin toxicity. Here, the author reviewed the mechanism and upfront preventive and reactive treatments for epidermal growth factor receptor inhibitor-associated skin toxicities.

  4. Inhibiting the Aurora B Kinase Potently Suppresses Repopulation During Fractionated Irradiation of Human Lung Cancer Cell Lines

    Energy Technology Data Exchange (ETDEWEB)

    Sak, Ali, E-mail: ali.sak@uni-due.de [Department of Radiotherapy, West German Cancer Centre (WTZ), University Hospital Essen, University Duisburg-Essen, Essen (Germany); Stuschke, Martin; Groneberg, Michael; Kuebler, Dennis; Poettgen, Christoph [Department of Radiotherapy, West German Cancer Centre (WTZ), University Hospital Essen, University Duisburg-Essen, Essen (Germany); Eberhardt, Wilfried E.E. [Department of Medicine (Cancer Research), West German Cancer Centre (WTZ), University Hospital Essen, University Duisburg-Essen, Essen (Germany)

    2012-10-01

    Purpose: The use of molecular-targeted agents during radiotherapy of non-small-cell lung cancer (NSCLC) is a promising strategy to inhibit repopulation, thereby improving therapeutic outcome. We assessed the combined effectiveness of inhibiting Aurora B kinase and irradiation on human NSCLC cell lines in vitro. Methods and Materials: NSCLC cell lines were exposed to concentrations of AZD1152-hydroxyquinazoline pyrazol anilide (AZD1152-HQPA) inhibiting colony formation by 50% (IC50{sub clone}) in combination with single dose irradiation or different fractionation schedules using multiple 2-Gy fractions per day up to total doses of 4-40 Gy. The total irradiation dose required to control growth of 50% of the plaque monolayers (TCD50) was determined. Apoptosis, G2/M progression, and polyploidization were also analyzed. Results: TCD50 values after single dose irradiation were similar for the H460 and H661 cell lines with 11.4 {+-} 0.2 Gy and 10.7 {+-} 0.3 Gy, respectively. Fractionated irradiation using 3 Multiplication-Sign 2 Gy/day, 2 Multiplication-Sign 2 Gy/day, and 1 Multiplication-Sign 2 Gy/day schedules significantly increased TCD50 values for both cell lines grown as plaque monolayers with increasing radiation treatment time. This could be explained by a repopulation effect per day that counteracts 75 {+-} 8% and 27 {+-} 6% of the effect of a 2-Gy fraction in H460 and H661 cells, respectively. AZD1152-HQPA treatment concomitant to radiotherapy significantly decreased the daily repopulation effect (H460: 28 {+-} 5%, H661: 10 {+-} 4% of a 2-Gy fraction per day). Treatment with IC50{sub clone} AZD1152-HPQA did not induce apoptosis, prolong radiation-induced G2 arrest, or delay cell cycle progression before the spindle check point. However, polyploidization was detected, especially in cell lines without functional p53. Conclusions: Inhibition of Aurora B kinase with low AZD1152-HQPA concentrations during irradiation of NSCLC cell lines affects repopulation during

  5. 有丝分裂关键激酶抑制剂的研究进展%Progress on the research of key kinase inhibitors in mitosis

    Institute of Scientific and Technical Information of China (English)

    陈葆峰; 解鸿波; 沙宇; 程卯生

    2012-01-01

    目的 综述有丝分裂关键激酶抑制剂近年来的研究进展.方法 根据已报道的有丝分裂中关键激酶抑制剂的文献,将对有丝分裂过程中与肿瘤发生联系密切的激酶抑制剂,如Aurora激酶抑制剂、CDK(cyclin-dependent kinase)激酶抑制剂、PLK(Polo-like kinase)激酶抑制剂、CHK(check-point kinase)激酶抑制剂等目前的研究进展进行综述.结果 有丝分裂关键激酶抑制剂已在临床前研究及临床研究中显示出很好的抗肿瘤活性.结论 随着研究的不断深入,有丝分裂关键激酶抑制剂将在肿瘤治疗中发挥更大的作用.%Objective To review the recent progress on the research of key kinase inhibitors in mitosis. Methods According to the references to the key kinase inhibitors in mitosis reported, the kinase inhibitors were introduced closely related to the occurrence of the cancer in the progress of mitosis, such as Aurora kinase in- . hibitors, CDK (cyclin-dependent kinase) inhibitors, PLK (Polo-like kinase) inhibitors and CHK (checkpoint kinase)inhibitors. Results The potent anti-tumor activity of key kinase inhibitors in mitosis had been revealed in the preclinical and clinical research. Conclusions With the further research,key kinase inhibitors in mitosis shows more significant roles in the therapy of cancer.

  6. Identification of aurora kinase B and Wee1-like protein kinase as downstream targets of (V600E)B-RAF in melanoma.

    Science.gov (United States)

    Sharma, Arati; Madhunapantula, SubbaRao V; Gowda, Raghavendra; Berg, Arthur; Neves, Rogerio I; Robertson, Gavin P

    2013-04-01

    BRAF is the most mutated gene in melanoma, with approximately 50% of patients containing V600E mutant protein. (V600E)B-RAF can be targeted using pharmacological agents, but resistance develops in patients by activating other proteins in the signaling pathway. Identifying downstream members in this signaling cascade is important to design strategies to avoid the development of resistance. Unfortunately, downstream proteins remain to be identified and therapeutic potential requires validation. A kinase screen was undertaken to identify downstream targets in the (V600E)B-RAF signaling cascade. Involvement of aurora kinase B (AURKB) and Wee1-like protein kinase (WEE1) as downstream proteins in the (V600E)B-RAF pathway was validated in xenografted tumors, and mechanisms of action were characterized in size- and time-matched tumors. Levels of only AURKB and WEE1 decreased in melanoma cells, when (V600E)B-RAF, mitogen-activated protein kinase 1/2, or extracellular signal-regulated kinase 1/2 protein levels were reduced using siRNA compared with other identified kinases. AURKB and WEE1 were expressed in tumors of patients with melanoma at higher levels than observed in normal human melanocytes. Targeting these proteins reduced tumor development by approximately 70%, similar to that observed when inhibiting (V600E)B-RAF. Furthermore, protein or activity levels of AURKB and WEE1 decreased in melanoma cells when pharmacological agents targeting upstream (V600E)B-RAF or mitogen-activated protein kinase were used to inhibit the (V600E)B-RAF pathway. Thus, AURKB and WEE1 are targets and biomarkers of therapeutic efficacy, lying downstream of (V600E)B-RAF in melanomas.

  7. Analysis of protein phosphatase-1 and aurora protein kinase suppressors reveals new aspects of regulatory protein function in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Anuprita Ghosh

    Full Text Available Protein phosphatase-1 (PP1 controls many processes in eukaryotic cells. Modulation of mitosis by reversing phosphorylation of proteins phosphorylated by aurora protein kinase is a critical function for PP1. Overexpression of the sole PP1, Glc7, in budding yeast, Saccharomyces cerevisiae, is lethal. This work shows that lethality requires the function of Glc7 regulatory proteins Sds22, Reg2, and phosphorylated Glc8. This finding shows that Glc7 overexpression induced cell death requires a specific subset of the many Glc7-interacting proteins and therefore is likely caused by promiscuous dephosphorylation of a variety of substrates. Additionally, suppression can occur by reducing Glc7 protein levels by high-copy Fpr3 without use of its proline isomerase domain. This divulges a novel function of Fpr3. Most suppressors of GLC7 overexpression also suppress aurora protein kinase, ipl1, temperature-sensitive mutations. However, high-copy mutant SDS22 genes show reciprocal suppression of GLC7 overexpression induced cell death or ipl1 temperature sensitivity. Sds22 binds to many proteins besides Glc7. The N-terminal 25 residues of Sds22 are sufficient to bind, directly or indirectly, to seven proteins studied here including the spindle assembly checkpoint protein, Bub3. These data demonstrate that Sds22 organizes several proteins in addition to Glc7 to perform functions that counteract Ipl1 activity or lead to hyper Glc7 induced cell death. These data also emphasize that Sds22 targets Glc7 to nuclear locations distinct from Ipl1 substrates.

  8. Small molecule tyrosine kinase inhibitors in pancreatic cancer

    Directory of Open Access Journals (Sweden)

    Sachin Gupta

    2008-10-01

    Full Text Available Sachin Gupta, Bassel F El-RayesDepartment of Hematology/Oncology, Karmanos Cancer Institute, Wayne State University, MI, USAAbstract: Pancreatic cancer has proven to be chemo-resistant, with gemcitabine being the only cytotoxic agent approved for advanced pancreatic cancer since 1996. Tyrosine kinase inhibitors represent a newer generation of chemotherapeutic agents targeting specific tumor pathways associated with carcinogenesis including cell cycle control, signal transduction, apoptosis and angiogenesis. These agents present a more selective way of treating pancreatic cancer. Erlotinib is the prototype of the tyrosine kinase inhibitors with proven efficacy in advanced pancreatic cancer and has been recently approved in that setting. Multiple other tyrosine kinase inhibitors targeting the VEGFR, PDGFR, and Src kinases are in various phases of clinical trials testing. The preliminary results of these trials have been disappointing. Current challenges in pancreatic cancer clinical trials testing include improving patient selection, identifying effective combinations, improving the predictive value of current preclinical models and better study designs. This review summarizes the present clinical development of tyrosine kinase inhibitors in pancreatic cancer and strategies for future drug development.Keywords: pancreatic cancer, erlotinib, tyrosine kinase inhibitors

  9. Kinase inhibitors: a new class of antirheumatic drugs

    Directory of Open Access Journals (Sweden)

    Kyttaris VC

    2012-09-01

    Full Text Available Vasileios C KyttarisDivision of Rheumatology, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, USAAbstract: The outlook for patients with rheumatoid arthritis has improved significantly over the last three decades with the use of disease-modifying antirheumatic drugs. However, despite the use of methotrexate, cytokine inhibitors, and molecules targeting T and B cells, a percentage of patients do not respond or lose their response over time. The autoimmune process in rheumatoid arthritis depends on activation of immune cells, which utilize intracellular kinases to respond to external stimuli such as cytokines, immune complexes, and antigens. In the past decade, small molecules targeting several kinases, such as p38 MAPK, Syk, and JAK have been developed. Several p38 MAPK inhibitors proved ineffective in treating rheumatoid arthritis. The Syk inhibitor, fostamatinib, proved superior to placebo in Phase II trials and is currently under Phase III investigation. Tofacitinib, a JAK1/3 inhibitor, was shown to be efficacious in two Phase III trials, while VX-509, a JAK3 inhibitor, showed promising results in a Phase II trial. Fostamatinib and tofacitinib were associated with increased rates of infection, elevation of liver enzymes, and neutropenia. Moreover, fostamatinib caused elevations of blood pressure and diarrhea, while tofacitinib was associated with an increase in creatinine and elevation of lipid levels.Keywords: rheumatoid arthritis, kinase inhibitors, mitogen-activated phosphokinase p38, spleen tyrosine kinase, Janus kinases

  10. Essential Roles of mTOR/Akt Pathway in Aurora-A Cell Transformation

    Directory of Open Access Journals (Sweden)

    Makoto Taga, Eiji Hirooka, Toru Ouchi

    2009-01-01

    Full Text Available We have recently demonstrated that Aurora-A kinase is a potential oncogene to develop mammary gland tumors in mice, when expressed under MMTV promoter. These tumors contain phosphorylated forms of Akt and mTOR, suggesting that Akt-mTOR pathway is involved in transformed phenotype induced by Aurora-A. In the present studies, we discovered that stable cell lines expressing Aurora-A contain phosphorylation of Akt Ser473 after prolonged passages of cell culture, not in cells of the early period of cell culture. Levels of PTEN tumor suppressor are significantly reduced in these late passage cells at least in part due to increased poly ubiquitination of the protein. Akt-activated Aurora-A cells formed larger colonies in soft agar and are resistant to UV-induced apoptosis. Aurora-A inhibitor, VX-680, can cause cell death of Aurora-A cells in which Akt is not activated. siRNA-mediated depletion of mTOR in those cells resulted in decreased phosphorylation of Akt Ser473, suggesting that TORC2 complex phosphorylates Akt in Aurora-A cells. Treatment of late-passage Aurora-A cells with mTOR inhibitor reduced colony formation in soft agar. These results strongly suggest that commitment of cell transformation by Aurora-A is determined by at least co-activation of Akt/mTOR pathway.

  11. Pyrrolopyridine inhibitors of mitogen-activated protein kinase-activated protein kinase 2 (MK-2).

    Science.gov (United States)

    Anderson, David R; Meyers, Marvin J; Vernier, William F; Mahoney, Matthew W; Kurumbail, Ravi G; Caspers, Nicole; Poda, Gennadiy I; Schindler, John F; Reitz, David B; Mourey, Robert J

    2007-05-31

    A new class of potent kinase inhibitors selective for mitogen-activated protein kinase-activated protein kinase 2 (MAPKAP-K2 or MK-2) for the treatment of rheumatoid arthritis has been prepared and evaluated. These inhibitors have IC50 values as low as 10 nM against the target and have good selectivity profiles against a number of kinases including CDK2, ERK, JNK, and p38. These MK-2 inhibitors have been shown to suppress TNFalpha production in U397 cells and to be efficacious in an acute inflammation model. The structure-activity relationships of this series, the selectivity for MK-2 and their activity in both in vitro and in vivo models are discussed. The observed selectivity is discussed with the aid of an MK-2/inhibitor crystal structure.

  12. Mini-review: bmx kinase inhibitors for cancer therapy.

    Science.gov (United States)

    Jarboe, John S; Dutta, Shilpa; Velu, Sadanandan E; Willey, Christopher D

    2013-09-01

    Kinase inhibitors are among the fastest growing class of anti-cancer therapies. One family of kinases that has recently gained attention as a target for treating malignant disorders is the Tec kinase family. Evidence has been published that one member of this family; the Bmx kinase, may play a role in the pathogenesis of glioblastoma, prostate, breast and lung cancer. Bmx has also shown potential as an anti-vascular therapy in combination with radiation or as a sensitizer to chemotherapeutic agents. Therefore, several companies such as Pharmacyclics, Avila Therapeutics, Merck and Co., Metaproteomics, IRM, and Moerae Matrix have developed compounds or peptides that function as Bmx kinase inhibitors. These companies have subsequently been issued patents for these inhibitors. Additionally, it has been shown that current clinical stage EGFR inhibitors can irreversibly inhibit Bmx, suggesting these compounds might be rapidly moved to clinical trials for other malignancies. This review will discuss current patents issued since 2009 that contain data specifically on inhibition of the Bmx kinase, and will also discuss the scientific literature that suggests their potential application as therapeutics in the treatment of the aforementioned malignancies.

  13. Two TPX2-dependent switches control the activity of Aurora A.

    Directory of Open Access Journals (Sweden)

    Xue Xu

    Full Text Available Aurora A is an important oncogenic kinase for mitotic spindle assembly and a potentially attractive target for human cancers. Its activation could be regulated by ATP cycle and its activator TPX2. To understand the activation mechanism of Aurora A, a series of 20 ns molecular dynamics (MD simulations were performed on both the wild-type kinase and its mutants. Analyzing the three dynamic trajectories (Aurora A-ATP, Aurora A-ADP, and Aurora A-ADP-TPX2 at the residue level, for the first time we find two TPX2-dependent switches, i.e., switch-1 (Lys-143 and switch-2 (Arg-180, which are tightly associated with Aurora A activation. In the absence of TPX2, Lys-143 exhibits a "closed" state, and becomes hydrogen-bonded to ADP. Once TPX2 binding occurs, switch-1 is forced to "open" the binding site, thus pulling ADP away from Aurora A. Without facilitation of TPX2, switch-2 exits in an "open" conformation which accompanies the outward-flipping movement of P·Thr288 (in an inactive conformation, leading to the crucial phosphothreonine exposed and accessible for deactivation. However, with the binding of TPX2, switch-2 is forced to undergo a "closed" movement, thus capturing P·Thr288 into a buried position and locking its active conformation. Analysis of two Aurora A (K143A and R180A mutants for the two switches further verifies their functionality and reliability in controlling Aurora activity. Our systems therefore suggest two switches determining Aurora A activation, which are important for the development of aurora kinase inhibitors.

  14. Identification of Novel Small Molecule Inhibitors of Oncogenic RET Kinase

    OpenAIRE

    Marialuisa Moccia; Qingsong Liu; Teresa Guida; Giorgia Federico; Annalisa Brescia; Zheng Zhao; Hwan Geun Choi; Xianming Deng; Li Tan; Jinhua Wang; Marc Billaud; Gray, Nathanael S.; Francesca Carlomagno; Massimo Santoro

    2015-01-01

    Oncogenic mutation of the RET receptor tyrosine kinase is observed in several human malignancies. Here, we describe three novel type II RET tyrosine kinase inhibitors (TKI), ALW-II-41-27, XMD15-44 and HG-6-63-01, that inhibit the cellular activity of oncogenic RET mutants at two digit nanomolar concentration. These three compounds shared a 3-trifluoromethyl-4-methylpiperazinephenyl pharmacophore that stabilizes the 'DFG-out' inactive conformation of RET activation loop. They blocked RET-media...

  15. Protein tyrosine phosphatase receptor delta acts as a neuroblastoma tumor suppressor by destabilizing the aurora kinase a oncogene

    Directory of Open Access Journals (Sweden)

    Meehan Maria

    2012-02-01

    Full Text Available Abstract Background Protein tyrosine phosphatase receptor delta (PTPRD is a member of a large family of protein tyrosine phosphatases which negatively regulate tyrosine phosphorylation. Neuroblastoma is a major childhood cancer arising from precursor cells of the sympathetic nervous system which is known to acquire deletions and alterations in the expression patterns of PTPRD, indicating a potential tumor suppressor function for this gene. The molecular mechanism, however, by which PTPRD renders a tumor suppressor effect in neuroblastoma is unknown. Results As a molecular mechanism, we demonstrate that PTPRD interacts with aurora kinase A (AURKA, an oncogenic protein that is over-expressed in multiple forms of cancer, including neuroblastoma. Ectopic up-regulation of PTPRD in neuroblastoma dephosphorylates tyrosine residues in AURKA resulting in a destabilization of this protein culminating in interfering with one of AURKA's primary functions in neuroblastoma, the stabilization of MYCN protein, the gene of which is amplified in approximately 25 to 30% of high risk neuroblastoma. Conclusions PTPRD has a tumor suppressor function in neuroblastoma through AURKA dephosphorylation and destabilization and a downstream destabilization of MYCN protein, representing a novel mechanism for the function of PTPRD in neuroblastoma.

  16. Aurora Kinase A expression predicts platinum-resistance and adverse outcome in high-grade serous ovarian carcinoma patients.

    Science.gov (United States)

    Mignogna, Chiara; Staropoli, Nicoletta; Botta, Cirino; De Marco, Carmela; Rizzuto, Antonia; Morelli, Michele; Di Cello, Annalisa; Franco, Renato; Camastra, Caterina; Presta, Ivan; Malara, Natalia; Salvino, Angela; Tassone, Pierfrancesco; Tagliaferri, Pierosandro; Barni, Tullio; Donato, Giuseppe; Di Vito, Anna

    2016-05-21

    High-Grade Serous Ovarian Carcinoma (HGSOC) is the predominant histotype of epithelial ovarian cancer (EOC), characterized by advanced stage at diagnosis, frequent TP53 mutation, rapid progression, and high responsiveness to platinum-based-chemotherapy. To date, standard first-line-chemotherapy in advanced EOC includes platinum salts and paclitaxel with or without bevacizumab. The major prognostic factor is the response duration from the end of the platinum-based treatment (platinum-free interval) and about 10-0 % of EOC patients bear a platinum-refractory disease or develop early resistance (platinum-free interval shorter than 6 months). On these bases, a careful selection of patients who could benefit from chemotherapy is recommended to avoid unnecessary side effects and for a better disease outcome. In this retrospective study, an immunohistochemical evaluation of Aurora Kinase A (AURKA) was performed on 41 cases of HGSOC according to platinum-status. Taking into account the number and intensity of AURKA positive cells we built a predictive score able to discriminate with high accuracy platinum-sensitive patients from platinum-resistant patients (p platinum-resistance and as prognostic factor, that deserves further investigation in prospective clinical trials. Indeed, in the era of personalized medicine, AURKA could assist the clinicians in selecting the best treatment and represent, at the same time, a promising new therapeutic target in EOC treatment.

  17. Protein tyrosine phosphatase receptor delta acts as a neuroblastoma tumor suppressor by destabilizing the aurora kinase a oncogene

    LENUS (Irish Health Repository)

    Meehan, Maria

    2012-02-05

    Abstract Background Protein tyrosine phosphatase receptor delta (PTPRD) is a member of a large family of protein tyrosine phosphatases which negatively regulate tyrosine phosphorylation. Neuroblastoma is a major childhood cancer arising from precursor cells of the sympathetic nervous system which is known to acquire deletions and alterations in the expression patterns of PTPRD, indicating a potential tumor suppressor function for this gene. The molecular mechanism, however, by which PTPRD renders a tumor suppressor effect in neuroblastoma is unknown. Results As a molecular mechanism, we demonstrate that PTPRD interacts with aurora kinase A (AURKA), an oncogenic protein that is over-expressed in multiple forms of cancer, including neuroblastoma. Ectopic up-regulation of PTPRD in neuroblastoma dephosphorylates tyrosine residues in AURKA resulting in a destabilization of this protein culminating in interfering with one of AURKA\\'s primary functions in neuroblastoma, the stabilization of MYCN protein, the gene of which is amplified in approximately 25 to 30% of high risk neuroblastoma. Conclusions PTPRD has a tumor suppressor function in neuroblastoma through AURKA dephosphorylation and destabilization and a downstream destabilization of MYCN protein, representing a novel mechanism for the function of PTPRD in neuroblastoma.

  18. QM/MD studies of the dynamics of the MTSL spin label in Aurora-A kinase protein activation loop

    CERN Document Server

    Concilio, Maria Grazia; Bayliss, Richard; Burgess, Selena

    2015-01-01

    Molecular dynamics(MD)simulations using a graphics processing unit (GPU) has been employed in order to determine the conformational space of the methane-thiosulfonate spin label (MTSL) attached to the activation loop of the Aurora-A kinase protein and compared with quantum mechanical (QM) methods rooted on density functional theory (DFT). MD provided a wealth of information about interactions between the MTSL and the residues of the protein and on the different motional contributions to the overall dynamics of the MTSL. Data obtained from MD were seen to be in good agreement with those obtained from QM but the dynamics of the system revealed more interactions than those observed from QM methods. A strong correlation between the tumbling of the protein and the transitions of the X4 and X5 dihedral angles of the MTSL, was observed with a consequent effect also the distribution of the nitroxide(NO)group in the space. Theoretical EPR spectra calculated from opportunely selected MD frames showing interactions betw...

  19. Side-effects of protein kinase inhibitors on ion channels

    Indian Academy of Sciences (India)

    Youn Kyoung Son; Hongzoo Park; Amy L Firth; Won Sun Park

    2013-12-01

    Protein kinases are one of the largest gene families and have regulatory roles in all aspects of eukaryotic cell function. Modulation of protein kinase activity is a desirable therapeutic approach for a number of human diseases associated with aberrant kinase activity, including cancers, arthritis and cardiovascular disorders. Several strategies have been used to develop specific and selective protein kinase modulators, primarily via inhibition of phosphorylation and down-regulation of kinase gene expression. These strategies are effective at regulating intracellular signalling pathways, but are unfortunately associated with several undesirable effects, particularly those that modulate ion channel function. In fact, the side-effects have precluded these inhibitors from being both useful experimental tools and therapeutically viable. This review focuses on the ion channel side-effects of several protein kinase inhibitors and specifically on those modulating K+, Na+ and Ca2+ ion channels. It is hoped that the information provided with a detailed summary in this review will assist the future development of novel specific and selective compounds targeting protein kinases both for experimental tools and for therapeutic approaches.

  20. Inhibition of mitotic Aurora kinase A by alisertib induces apoptosis and autophagy of human gastric cancer AGS and NCI-N78 cells

    Directory of Open Access Journals (Sweden)

    Yuan CX

    2015-01-01

    Full Text Available Chun-Xiu Yuan,1,2 Zhi-Wei Zhou,2,3 Yin-Xue Yang,4 Zhi-Xu He,3 Xueji Zhang,5 Dong Wang,6 Tianxing Yang,7 Ning-Ju Wang,1 Ruan Jin Zhao,8 Shu-Feng Zhou21Department of Oncology, General Hospital Ningxia Medical University, Yinchuan, Ningxia, People’s Republic of China; 2Department of Pharmaceutical Science, College of Pharmacy, University of South Florida, Tampa, FL, USA; 3Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino–US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, Guizhou, People’s Republic of China; 4Department of Colorectal Surgery, General Hospital, Ningxia Medical University, Yinchuan, Ningxia, People’s Republic of China; 5Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 6Cancer Center, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, People’s Republic of China; 7Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA; 8Center for Traditional Chinese Medicine, Sarasota, FL, USAAbstract: Gastric cancer is one of the most common cancers and responds poorly to current chemotherapy. Alisertib (ALS is a second-generation, orally bioavailable, highly selective small-molecule inhibitor of the serine/threonine protein kinase Aurora kinase A (AURKA. ALS has been shown to have potent anticancer effects in preclinical and clinical studies, but its role in gastric cancer treatment is unclear. This study aimed to investigate the cancer cell-killing effect of ALS on gastric cancer cell lines AGS and NCI-N78, with a focus on cell proliferation, cell-cycle distribution, apoptosis, and autophagy and the mechanism of action. The results showed that ALS exhibited potent growth-inhibitory, proapoptotic, and proautophagic effects on AGS and

  1. Discovery of benzo[e]pyridoindolones as kinase inhibitors that disrupt mitosis exit while erasing AMPK-Thr172 phosphorylation on the spindle.

    Science.gov (United States)

    Le, Ly-Thuy-Tram; Couvet, Morgane; Favier, Bertrand; Coll, Jean-Luc; Nguyen, Chi-Hung; Molla, Annie

    2015-09-01

    Aurora kinases play an essential role in mitotic progression and are attractive targets in cancer therapy. The first generation of benzo[e]pyridoindole exhibited powerful aurora kinase inhibition but their low solubility limited further development. Grafting a pyperidine-ethoxy group gives rise to a hydrosoluble inhibitor: compound C5M.C5M could efficiently inhibit the proliferation of cells from different origins. C5M prevented cell cycling, induced a strong mitotic arrest then, cells became polyploid and finally died. C5M did not impair the spindle checkpoint, the separation of the sister chromatids and the transfer of aurora B on the mid-zone. C5M prevented histone H3 phosphorylation at mitotic entry and erased AMPK-Thr172 phosphorylation in late mitosis. With this unique profile of inhibition, C5M could be useful for understanding the role of phospho-Thr172-AMPK in abscission and the relationship between the chromosomal complex and the energy sensing machinery.C5M is a multikinase inhibitor with interesting preclinical characteristics: high hydro-solubility and a good stability in plasma. A single dose prevents the expansion of multicellular spheroids. C5M can safely be injected to mice and reduces significantly the development of xenograft. The next step will be to define the protocol of treatment and the cancer therapeutic field of this new anti-proliferative drug.

  2. FDA-approved small-molecule kinase inhibitors

    DEFF Research Database (Denmark)

    Wu, Peng; Nielsen, Thomas E.; Clausen, Mads Hartvig

    2015-01-01

    Kinases have emerged as one of the most intensivelypursued targets in current pharmacological research,especially for cancer, due to their critical roles in cellularsignaling. To date, the US FDA has approved 28 smallmoleculekinase inhibitors, half of which were approvedin the past 3 years. While...

  3. Old Tyrosine Kinase Inhibitors and Newcomers in Gastrointestinal Cancer Treatment.

    Science.gov (United States)

    Giordani, Erika; Zoratto, Federica; Strudel, Martina; Papa, Anselmo; Rossi, Luigi; Minozzi, Marina; Caruso, Davide; Zaccarelli, Eleonora; Verrico, Monica; Tomao, Silverio

    2016-01-01

    Gastrointestinal cancer treatment is based more on molecular biology that has provided increasing knowledge about cancer pathogenesis on which targeted therapy is being developed. Precisely, targeted therapy is defined as a "type of treatment that uses drugs, such as monoclonal antibodies or tyrosine kinase inhibitors, to identify and attack specific cancer cells". Nowadays, the United States Food and Drug Administration has approved many targeted therapies for gastrointestinal cancer treatment, as many are in various phases of development as well. In a previous review we discussed the main monoclonal antibodies used and studied in gastrointestinal cancer. In addition to monoclonal antibodies, tyrosine kinase inhibitors represent another class of targeted therapy and following the approval of imatinib for gastrointestinal stromal tumours, other tyrosine kinase inhibitors have been approved for gastrointestinal cancers treatment such as sunitinib, regoragenib, sorafenib and erlotinib. Moving forward, the purpose of this review is to focus on the efficacy data of main tyrosine kinase inhibitors commonly used in the personalized treatment of each gastrointestinal tumour and to provide a comprehensive overview about experimental targeted therapies ongoing in this setting.

  4. Searching Inhibitors of Adenosine Kinase by Simulation Methods

    Institute of Scientific and Technical Information of China (English)

    ZHU Rui-Xin; ZHANG Xing-Long; DONG Xi-Cheng; CHEN Min-Bo

    2006-01-01

    Searching new inhibitors of adenosine kinase (AK) is still drawing attention of experimental scientists. A better and solid model is here proposed by means of simulation methods from different ways, the direct analysis of receptor itself, the conventional 3D-QSAR methods and the integration of docking method and the conventional QSAR analysis.

  5. Drug-drug interactions with tyrosine-kinase inhibitors

    NARCIS (Netherlands)

    Croes, Sander; de Vries, Frank

    2014-01-01

    In their Review, Roelof van Leeuwen and colleagues1 recommend various dose adjustments during concomitant use of tyrosine-kinase inhibitors and drugs that inhibit or induce cytochrome P450 3A4 (CYP3A4).1 Most information is taken from the US Food and Drug Administration (FDA)'s drug label or the Eur

  6. Identification of a kinase profile that predicts chromosome damage induced by small molecule kinase inhibitors.

    Directory of Open Access Journals (Sweden)

    Andrew J Olaharski

    2009-07-01

    Full Text Available Kinases are heavily pursued pharmaceutical targets because of their mechanistic role in many diseases. Small molecule kinase inhibitors (SMKIs are a compound class that includes marketed drugs and compounds in various stages of drug development. While effective, many SMKIs have been associated with toxicity including chromosomal damage. Screening for kinase-mediated toxicity as early as possible is crucial, as is a better understanding of how off-target kinase inhibition may give rise to chromosomal damage. To that end, we employed a competitive binding assay and an analytical method to predict the toxicity of SMKIs. Specifically, we developed a model based on the binding affinity of SMKIs to a panel of kinases to predict whether a compound tests positive for chromosome damage. As training data, we used the binding affinity of 113 SMKIs against a representative subset of all kinases (290 kinases, yielding a 113x290 data matrix. Additionally, these 113 SMKIs were tested for genotoxicity in an in vitro micronucleus test (MNT. Among a variety of models from our analytical toolbox, we selected using cross-validation a combination of feature selection and pattern recognition techniques: Kolmogorov-Smirnov/T-test hybrid as a univariate filter, followed by Random Forests for feature selection and Support Vector Machines (SVM for pattern recognition. Feature selection identified 21 kinases predictive of MNT. Using the corresponding binding affinities, the SVM could accurately predict MNT results with 85% accuracy (68% sensitivity, 91% specificity. This indicates that kinase inhibition profiles are predictive of SMKI genotoxicity. While in vitro testing is required for regulatory review, our analysis identified a fast and cost-efficient method for screening out compounds earlier in drug development. Equally important, by identifying a panel of kinases predictive of genotoxicity, we provide medicinal chemists a set of kinases to avoid when designing

  7. Endocrine side effects of broad-acting kinase inhibitors.

    Science.gov (United States)

    Lodish, Maya B; Stratakis, Constantine A

    2010-09-01

    Targeted therapy in oncology consists of drugs that specifically interfere with abnormal signaling pathways that are dysregulated in cancer cells. Tyrosine kinase inhibitors (TKIs) take advantage of unique oncogenes that are activated in certain types of cancer, and also target common mechanisms of growth, invasion, metastasis, and angiogenesis. However, many kinase inhibitors for cancer therapy are somewhat nonselective, and most have additional mechanisms of action at the cellular level, which are not completely understood. The use of these agents has increased our knowledge of important side effects, of which the practicing clinician must be aware. Recently, proposed endocrine-related side effects of these agents include alterations in thyroid function, bone metabolism, linear growth, gonadal function, fetal development, and glucose metabolism, and adrenal function. This review summarizes the most recent data on the endocrine side effects of TKIs.

  8. Tyrosine Kinase Inhibitors and Diabetes: A Novel Treatment Paradigm?

    Science.gov (United States)

    Fountas, Athanasios; Diamantopoulos, Leonidas-Nikolaos; Tsatsoulis, Agathocles

    2015-11-01

    Deregulation of protein tyrosine kinase (PTK) activity is implicated in various proliferative conditions. Multi-target tyrosine kinase inhibitors (TKIs) are increasingly used for the treatment of different malignancies. Recently, several clinical cases of the reversal of both type 1 and 2 diabetes mellitus (T1DM, T2DM) during TKI administration have been reported. Experimental in vivo and in vitro studies have elucidated some of the mechanisms behind this effect. For example, inhibition of Abelson tyrosine kinase (c-Abl) results in β cell survival and enhanced insulin secretion, while platelet-derived growth factor receptor (PDGFR) and epidermal growth factor receptor (EGFR) inhibition leads to improvement in insulin sensitivity. In addition, inhibition of vascular endothelial growth factor receptor 2 (VEGFR2) reduces the degree of islet cell inflammation (insulitis). Therefore, targeting several PTKs may provide a novel approach for correcting the pathophysiologic disturbances of diabetes.

  9. Discovery of indazoles as inhibitors of Tpl2 kinase.

    Science.gov (United States)

    Hu, Yonghan; Cole, Derek; Denny, Rajiah Aldrin; Anderson, David R; Ipek, Manus; Ni, Yike; Wang, Xiaolun; Thaisrivongs, Suvit; Chamberlain, Timothy; Hall, J Perry; Liu, Julie; Luong, Michael; Lin, Lih-Ling; Telliez, Jean-Baptiste; Gopalsamy, Ariamala

    2011-08-15

    Synthesis, modeling and structure-activity relationship of indazoles as inhibitors of Tpl2 kinase are described. From a high throughput screening effort, we identified an indazole hit compound 5 that has a single digit micromolar Tpl2 activity. Through SAR modifications at the C3 and C5 positions of the indazole, we discovered compound 31 with good potency in LANCE assay and cell-based p-Erk assay.

  10. Phosphoinositide 3-kinase inhibitors induce DNA damage through nucleoside depletion.

    Science.gov (United States)

    Juvekar, Ashish; Hu, Hai; Yadegarynia, Sina; Lyssiotis, Costas A; Ullas, Soumya; Lien, Evan C; Bellinger, Gary; Son, Jaekyoung; Hok, Rosanna C; Seth, Pankaj; Daly, Michele B; Kim, Baek; Scully, Ralph; Asara, John M; Cantley, Lewis C; Wulf, Gerburg M

    2016-07-26

    We previously reported that combining a phosphoinositide 3-kinase (PI3K) inhibitor with a poly-ADP Rib polymerase (PARP)-inhibitor enhanced DNA damage and cell death in breast cancers that have genetic aberrations in BRCA1 and TP53. Here, we show that enhanced DNA damage induced by PI3K inhibitors in this mutational background is a consequence of impaired production of nucleotides needed for DNA synthesis and DNA repair. Inhibition of PI3K causes a reduction in all four nucleotide triphosphates, whereas inhibition of the protein kinase AKT is less effective than inhibition of PI3K in suppressing nucleotide synthesis and inducing DNA damage. Carbon flux studies reveal that PI3K inhibition disproportionately affects the nonoxidative pentose phosphate pathway that delivers Rib-5-phosphate required for base ribosylation. In vivo in a mouse model of BRCA1-linked triple-negative breast cancer (K14-Cre BRCA1(f/f)p53(f/f)), the PI3K inhibitor BKM120 led to a precipitous drop in DNA synthesis within 8 h of drug treatment, whereas DNA synthesis in normal tissues was less affected. In this mouse model, combined PI3K and PARP inhibition was superior to either agent alone to induce durable remissions of established tumors.

  11. PP2A Inhibitor PME-1 Drives Kinase Inhibitor Resistance in Glioma Cells.

    Science.gov (United States)

    Kaur, Amanpreet; Denisova, Oxana V; Qiao, Xi; Jumppanen, Mikael; Peuhu, Emilia; Ahmed, Shafiq U; Raheem, Olayinka; Haapasalo, Hannu; Eriksson, John; Chalmers, Anthony J; Laakkonen, Pirjo; Westermarck, Jukka

    2016-12-01

    Glioblastoma multiforme lacks effective therapy options. Although deregulated kinase pathways are drivers of malignant progression in glioblastoma multiforme, glioma cells exhibit intrinsic resistance toward many kinase inhibitors, and the molecular basis of this resistance remains poorly understood. Here, we show that overexpression of the protein phosphatase 2A (PP2A) inhibitor protein PME-1 drives resistance of glioma cells to various multikinase inhibitors. The PME-1-elicited resistance was dependent on specific PP2A complexes and was mediated by a decrease in cytoplasmic HDAC4 activity. Importantly, both PME-1 and HDAC4 associated with human glioma progression, supporting clinical relevance of the identified mechanism. Synthetic lethality induced by both PME-1 and HDAC4 inhibition was dependent on the coexpression of proapoptotic protein BAD. Thus, PME-1-mediated PP2A inhibition is a novel mechanistic explanation for multikinase inhibitor resistance in glioma cells. Clinically, these results may inform patient stratification strategies for future clinical trials with selected kinase inhibitors in glioblastoma multiforme. Cancer Res; 76(23); 7001-11. ©2016 AACR.

  12. Photoactivatable Caged Prodrugs of VEGFR-2 Kinase Inhibitors

    Directory of Open Access Journals (Sweden)

    Boris Pinchuk

    2016-04-01

    Full Text Available In this study, we report on the design, synthesis, photokinetic properties and in vitro evaluation of photoactivatable caged prodrugs for the receptor tyrosine kinase VEGFR-2. Highly potent VEGFR-2 inhibitors 1 and 3 were caged by introduction of a photoremovable protecting group (PPG to yield the caged prodrugs 4 and 5. As expected, enzymatic and cellular proliferation assays showed dramatically diminished efficacy of caged prodrugs in vitro. Upon ultraviolet (UV irradiation of the prodrugs original inhibitory activity was completely restored and even distinctly reinforced, as was the case for the prodrug 4. The presented results are a further evidence for caging technique being an interesting approach in the protein kinase field. It could enable spatial and temporal control for the inhibition of VEGFR-2. The described photoactivatable prodrugs might be highly useful as biological probes for studying the VEGFR-2 signal transduction.

  13. What's so Bor(a)ing about Aurora-A activation?

    Science.gov (United States)

    Wiese, Christiane; O'Brien, Lori L

    2006-08-01

    Aurora-A kinases are highly conserved mitotic kinases required for cell division. The regulation of Aurora-A activity is less highly conserved and currently poorly understood. Work by Knoblich and coworkers in this issue of Developmental Cell identifies the conserved protein, Aurora Borealis (Bora), as a key regulator of Aurora-A activity during mitosis.

  14. Cyclin Dependent Kinase 9 Inhibitors for Cancer Therapy.

    Science.gov (United States)

    Sonawane, Yogesh A; Taylor, Margaret A; Napoleon, John Victor; Rana, Sandeep; Contreras, Jacob I; Natarajan, Amarnath

    2016-10-13

    Cyclin dependent kinase (CDK) inhibitors have been the topic of intense research for nearly 2 decades due to their widely varied and critical functions within the cell. Recently CDK9 has emerged as a druggable target for the development of cancer therapeutics. CDK9 plays a crucial role in transcription regulation; specifically, CDK9 mediated transcriptional regulation of short-lived antiapoptotic proteins is critical for the survival of transformed cells. Focused chemical libraries based on a plethora of scaffolds have resulted in mixed success with regard to the development of selective CDK9 inhibitors. Here we review the regulation of CDK9, its cellular functions, and common core structures used to target CDK9, along with their selectivity profile and efficacy in vitro and in vivo.

  15. Development of Selective Covalent Janus Kinase 3 Inhibitors.

    Science.gov (United States)

    Tan, Li; Akahane, Koshi; McNally, Randall; Reyskens, Kathleen M S E; Ficarro, Scott B; Liu, Suhu; Herter-Sprie, Grit S; Koyama, Shohei; Pattison, Michael J; Labella, Katherine; Johannessen, Liv; Akbay, Esra A; Wong, Kwok-Kin; Frank, David A; Marto, Jarrod A; Look, Thomas A; Arthur, J Simon C; Eck, Michael J; Gray, Nathanael S

    2015-08-27

    The Janus kinases (JAKs) and their downstream effectors, signal transducer and activator of transcription proteins (STATs), form a critical immune cell signaling circuit, which is of fundamental importance in innate immunity, inflammation, and hematopoiesis, and dysregulation is frequently observed in immune disease and cancer. The high degree of structural conservation of the JAK ATP binding pockets has posed a considerable challenge to medicinal chemists seeking to develop highly selective inhibitors as pharmacological probes and as clinical drugs. Here we report the discovery and optimization of 2,4-substituted pyrimidines as covalent JAK3 inhibitors that exploit a unique cysteine (Cys909) residue in JAK3. Investigation of structure-activity relationship (SAR) utilizing biochemical and transformed Ba/F3 cellular assays resulted in identification of potent and selective inhibitors such as compounds 9 and 45. A 2.9 Å cocrystal structure of JAK3 in complex with 9 confirms the covalent interaction. Compound 9 exhibited decent pharmacokinetic properties and is suitable for use in vivo. These inhibitors provide a set of useful tools to pharmacologically interrogate JAK3-dependent biology.

  16. Development of covalent inhibitors that can overcome resistance to first-generation FGFR kinase inhibitors.

    Science.gov (United States)

    Tan, Li; Wang, Jun; Tanizaki, Junko; Huang, Zhifeng; Aref, Amir R; Rusan, Maria; Zhu, Su-Jie; Zhang, Yiyun; Ercan, Dalia; Liao, Rachel G; Capelletti, Marzia; Zhou, Wenjun; Hur, Wooyoung; Kim, NamDoo; Sim, Taebo; Gaudet, Suzanne; Barbie, David A; Yeh, Jing-Ruey Joanna; Yun, Cai-Hong; Hammerman, Peter S; Mohammadi, Moosa; Jänne, Pasi A; Gray, Nathanael S

    2014-11-11

    The human FGF receptors (FGFRs) play critical roles in various human cancers, and several FGFR inhibitors are currently under clinical investigation. Resistance usually results from selection for mutant kinases that are impervious to the action of the drug or from up-regulation of compensatory signaling pathways. Preclinical studies have demonstrated that resistance to FGFR inhibitors can be acquired through mutations in the FGFR gatekeeper residue, as clinically observed for FGFR4 in embryonal rhabdomyosarcoma and neuroendocrine breast carcinomas. Here we report on the use of a structure-based drug design to develop two selective, next-generation covalent FGFR inhibitors, the FGFR irreversible inhibitors 2 (FIIN-2) and 3 (FIIN-3). To our knowledge, FIIN-2 and FIIN-3 are the first inhibitors that can potently inhibit the proliferation of cells dependent upon the gatekeeper mutants of FGFR1 or FGFR2, which confer resistance to first-generation clinical FGFR inhibitors such as NVP-BGJ398 and AZD4547. Because of the conformational flexibility of the reactive acrylamide substituent, FIIN-3 has the unprecedented ability to inhibit both the EGF receptor (EGFR) and FGFR covalently by targeting two distinct cysteine residues. We report the cocrystal structure of FGFR4 with FIIN-2, which unexpectedly exhibits a "DFG-out" covalent binding mode. The structural basis for dual FGFR and EGFR targeting by FIIN3 also is illustrated by crystal structures of FIIN-3 bound with FGFR4 V550L and EGFR L858R. These results have important implications for the design of covalent FGFR inhibitors that can overcome clinical resistance and provide the first example, to our knowledge, of a kinase inhibitor that covalently targets cysteines located in different positions within the ATP-binding pocket.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2015-08-15

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

  18. Phosphorylation of microtubule-binding protein Hec1 by mitotic kinase Aurora B specifies spindle checkpoint kinase Mps1 signaling at the kinetochore.

    Science.gov (United States)

    Zhu, Tongge; Dou, Zhen; Qin, Bo; Jin, Changjiang; Wang, Xinghui; Xu, Leilei; Wang, Zhaoyang; Zhu, Lijuan; Liu, Fusheng; Gao, Xinjiao; Ke, Yuwen; Wang, Zhiyong; Aikhionbare, Felix; Fu, Chuanhai; Ding, Xia; Yao, Xuebiao

    2013-12-13

    The spindle assembly checkpoint (SAC) is a quality control device to ensure accurate chromosome attachment to spindle microtubule for equal segregation of sister chromatid. Aurora B is essential for SAC function by sensing chromosome bi-orientation via spatial regulation of kinetochore substrates. However, it has remained elusive as to how Aurora B couples kinetochore-microtubule attachment to SAC signaling. Here, we show that Hec1 interacts with Mps1 and specifies its kinetochore localization via its calponin homology (CH) domain and N-terminal 80 amino acids. Interestingly, phosphorylation of the Hec1 by Aurora B weakens its interaction with microtubules but promotes Hec1 binding to Mps1. Significantly, the temporal regulation of Hec1 phosphorylation orchestrates kinetochore-microtubule attachment and Mps1 loading to the kinetochore. Persistent expression of phosphomimetic Hec1 mutant induces a hyperactivation of SAC, suggesting that phosphorylation-elicited Hec1 conformational change is used as a switch to orchestrate SAC activation to concurrent destabilization of aberrant kinetochore attachment. Taken together, these results define a novel role for Aurora B-Hec1-Mps1 signaling axis in governing accurate chromosome segregation in mitosis.

  19. Crystal structure of inhibitor of ;#954;B kinase [beta

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Guozhou; Lo, Yu-Chih; Li, Qiubai; Napolitano, Gennaro; Wu, Xuefeng; Jiang, Xuliang; Dreano, Michel; Karin, Michael; Wu, Hao (Weill-Med); (Merck-Serono); (UCSD); (EMD Serono)

    2011-07-26

    Inhibitor of {kappa}B (I{kappa}B) kinase (IKK) phosphorylates I{kappa}B proteins, leading to their degradation and the liberation of nuclear factor {kappa}B for gene transcription. Here we report the crystal structure of IKK{beta} in complex with an inhibitor, at a resolution of 3.6 {angstrom}. The structure reveals a trimodular architecture comprising the kinase domain, a ubiquitin-like domain (ULD) and an elongated, {alpha}-helical scaffold/dimerization domain (SDD). Unexpectedly, the predicted leucine zipper and helix-loop-helix motifs do not form these structures but are part of the SDD. The ULD and SDD mediate a critical interaction with I{kappa}B{alpha} that restricts substrate specificity, and the ULD is also required for catalytic activity. The SDD mediates IKK{beta} dimerization, but dimerization per se is not important for maintaining IKK{beta} activity and instead is required for IKK{beta} activation. Other IKK family members, IKK{alpha}, TBK1 and IKK-i, may have a similar trimodular architecture and function.

  20. Identification of Novel Small Molecule Inhibitors of Oncogenic RET Kinase.

    Directory of Open Access Journals (Sweden)

    Marialuisa Moccia

    Full Text Available Oncogenic mutation of the RET receptor tyrosine kinase is observed in several human malignancies. Here, we describe three novel type II RET tyrosine kinase inhibitors (TKI, ALW-II-41-27, XMD15-44 and HG-6-63-01, that inhibit the cellular activity of oncogenic RET mutants at two digit nanomolar concentration. These three compounds shared a 3-trifluoromethyl-4-methylpiperazinephenyl pharmacophore that stabilizes the 'DFG-out' inactive conformation of RET activation loop. They blocked RET-mediated signaling and proliferation with an IC50 in the nM range in fibroblasts transformed by the RET/C634R and RET/M918T oncogenes. They also inhibited autophosphorylation of several additional oncogenic RET-derived point mutants and chimeric oncogenes. At a concentration of 10 nM, ALW-II-41-27, XMD15-44 and HG-6-63-01 inhibited RET kinase and signaling in human thyroid cancer cell lines carrying oncogenic RET alleles; they also inhibited proliferation of cancer, but not non-tumoral Nthy-ori-3-1, thyroid cells, with an IC50 in the nM range. The three compounds were capable of inhibiting the 'gatekeeper' V804M mutant which confers substantial resistance to established RET inhibitors. In conclusion, we have identified a type II TKI scaffold, shared by ALW-II-41-27, XMD15-44 and HG-6-63-01, that may be used as novel lead for the development of novel agents for the treatment of cancers harboring oncogenic activation of RET.

  1. Identification of Novel Small Molecule Inhibitors of Oncogenic RET Kinase.

    Science.gov (United States)

    Moccia, Marialuisa; Liu, Qingsong; Guida, Teresa; Federico, Giorgia; Brescia, Annalisa; Zhao, Zheng; Choi, Hwan Geun; Deng, Xianming; Tan, Li; Wang, Jinhua; Billaud, Marc; Gray, Nathanael S; Carlomagno, Francesca; Santoro, Massimo

    2015-01-01

    Oncogenic mutation of the RET receptor tyrosine kinase is observed in several human malignancies. Here, we describe three novel type II RET tyrosine kinase inhibitors (TKI), ALW-II-41-27, XMD15-44 and HG-6-63-01, that inhibit the cellular activity of oncogenic RET mutants at two digit nanomolar concentration. These three compounds shared a 3-trifluoromethyl-4-methylpiperazinephenyl pharmacophore that stabilizes the 'DFG-out' inactive conformation of RET activation loop. They blocked RET-mediated signaling and proliferation with an IC50 in the nM range in fibroblasts transformed by the RET/C634R and RET/M918T oncogenes. They also inhibited autophosphorylation of several additional oncogenic RET-derived point mutants and chimeric oncogenes. At a concentration of 10 nM, ALW-II-41-27, XMD15-44 and HG-6-63-01 inhibited RET kinase and signaling in human thyroid cancer cell lines carrying oncogenic RET alleles; they also inhibited proliferation of cancer, but not non-tumoral Nthy-ori-3-1, thyroid cells, with an IC50 in the nM range. The three compounds were capable of inhibiting the 'gatekeeper' V804M mutant which confers substantial resistance to established RET inhibitors. In conclusion, we have identified a type II TKI scaffold, shared by ALW-II-41-27, XMD15-44 and HG-6-63-01, that may be used as novel lead for the development of novel agents for the treatment of cancers harboring oncogenic activation of RET.

  2. Receptor Tyrosine Kinase and Tyrosine Kinase Inhibitors: New Hope for Success in Multiple Sclerosis Therapy.

    Science.gov (United States)

    Mirshafiey, Abbas; Ghalamfarsa, Ghasem; Asghari, Babak; Azizi, Gholamreza

    2014-07-01

    Receptor tyrosine kinases (RTKs) are essential components of signal transduction pathways that mediate cell-to-cell communication and their function as relay points for signaling pathways. They have a key role in numerous processes that control cellular proliferation and differentiation, regulate cell growth and cellular metabolism, and promote cell survival and apoptosis. Recently, the role of RTKs including TCR, FLT-3, c-Kit, c-Fms, PDGFR, ephrin, neurotrophin receptor, and TAM receptor in autoimmune disorder, especially rheumatoid arthritis and multiple sclerosis has been suggested. In multiple sclerosis pathogenesis, RTKs and their tyrosine kinase enzymes are selective important targets for tyrosine kinase inhibitor (TKI) agents. TKIs, compete with the ATP binding site of the catalytic domain of several tyrosine kinases, and act as small molecules that have a favorable safety profile in disease treatment. Up to now, the efficacy of TKIs in numerous animal models of MS has been demonstrated, but application of these drugs in human diseases should be tested in future clinical trials.

  3. In vivo FRET imaging revealed a regulatory role of RanGTP in kinetochore-microtubule attachments via Aurora B kinase.

    Directory of Open Access Journals (Sweden)

    Yoke-Peng Lee

    Full Text Available Under the fluctuating circumstances provided by the innate dynamics of microtubules and opposing tensions resulted from microtubule-associated motors, it is vital to ensure stable kinetochore-microtubule attachments for accurate segregation. However, a comprehensive understanding of how this regulation is mechanistically achieved remains elusive. Using our newly designed live cell FRET time-lapse imaging, we found that post-metaphase RanGTP is crucial in the maintenance of stable kinetochore-microtubule attachments by regulating Aurora B kinase via the NES-bearing Mst1. More importantly, our study demonstrates that by ensuring stable alignment of metaphase chromosomes prior to segregation, RanGTP is indispensible in governing the genomic integrity and the fidelity of cell cycle progression. Our findings suggest an additional role of RanGTP beyond its known function in mitotic spindle assembly during the prometaphase-metaphase transition.

  4. Identification of small molecule inhibitors of phosphatidylinositol 3-kinase and autophagy

    DEFF Research Database (Denmark)

    Farkas, Thomas; Daugaard, Mads; Jaattela, Marja

    2011-01-01

    by the lack of specific small molecule inhibitors. Thus, we screened two small molecule kinase inhibitor libraries for inhibitors of rapamycin-induced autophagic flux. The three most potent inhibitors identified conferred profound inhibition of autophagic flux by inhibiting the formation of autophagosomes...

  5. Screening of protein kinase inhibitors identifies PKC inhibitors as inhibitors of osteoclastic acid secretion and bone resorption

    Directory of Open Access Journals (Sweden)

    Boutin Jean A

    2010-10-01

    Full Text Available Abstract Background Bone resorption is initiated by osteoclastic acidification of the resorption lacunae. This process is mediated by secretion of protons through the V-ATPase and chloride through the chloride antiporter ClC-7. To shed light on the intracellular signalling controlling extracellular acidification, we screened a protein kinase inhibitor library in human osteoclasts. Methods Human osteoclasts were generated from CD14+ monocytes. The effect of different kinase inhibitors on lysosomal acidification in human osteoclasts was investigated using acridine orange for different incubation times (45 minutes, 4 and 24 hours. The inhibitors were tested in an acid influx assay using microsomes isolated from human osteoclasts. Bone resorption by human osteoclasts on bone slices was measured by calcium release. Cell viability was measured using AlamarBlue. Results Of the 51 compounds investigated only few inhibitors were positive in both acidification and resorption assays. Rottlerin, GF109203X, Hypericin and Ro31-8220 inhibited acid influx in microsomes and bone resorption, while Sphingosine and Palmitoyl-DL-carnitine-Cl showed low levels of inhibition. Rottlerin inhibited lysosomal acidification in human osteoclasts potently. Conclusions In conclusion, a group of inhibitors all indicated to inhibit PKC reduced acidification in human osteoclasts, and thereby bone resorption, indicating that acid secretion by osteoclasts may be specifically regulated by PKC in osteoclasts.

  6. Aurora-A controls cancer cell radio- and chemoresistance via ATM/Chk2-mediated DNA repair networks.

    Science.gov (United States)

    Sun, Huizhen; Wang, Yan; Wang, Ziliang; Meng, Jiao; Qi, Zihao; Yang, Gong

    2014-05-01

    High expression of Aurora kinase A (Aurora-A) has been found to confer cancer cell radio- and chemoresistance, however, the underlying mechanism is unclear. In this study, by using Aurora-A cDNA/shRNA or the specific inhibitor VX680, we show that Aurora-A upregulates cell proliferation, cell cycle progression, and anchorage-independent growth to enhance cell resistance to cisplatin and X-ray irradiation through dysregulation of DNA damage repair networks. Mechanistic studies showed that Aurora-A promoted the expression of ATM/Chk2, but suppressed the expression of BRCA1/2, ATR/Chk1, p53, pp53 (Ser15), H2AX, γH2AX (Ser319), and RAD51. Aurora-A inhibited the focus formation of γH2AX in response to ionizing irradiation. Treatment of cells overexpressing Aurora-A and ATM/Chk2 with the ATM specific inhibitor KU-55933 increased the cell sensitivity to cisplatin and irradiation through increasing the phosphorylation of p53 at Ser15 and inhibiting the expression of Chk2, γH2AX (Ser319), and RAD51. Further study revealed that BRCA1/2 counteracted the function of Aurora-A to suppress the expression of ATM/Chk2, but to activate the expression of ATR/Chk1, pp53, γH2AX, and RAD51, leading to the enhanced cell sensitivity to irradiation and cisplatin, which was also supported by the results from animal assays. Thus, our data provide strong evidences that Aurora-A and BRCA1/2 inversely control the sensitivity of cancer cells to radio- and chemotherapy through the ATM/Chk2-mediated DNA repair networks, indicating that the DNA repair molecules including ATM/Chk2 may be considered for the targeted therapy against cancers with overexpression of Aurora-A.

  7. Tyrosine kinase inhibitors induced immune thrombocytopenia in chronic myeloid leukemia?

    Directory of Open Access Journals (Sweden)

    Avital F. Barak

    2011-12-01

    Full Text Available The outcome and quality of life of chronic myeloid leukemia (CML patients has remarkably changed with the treatment of tyrosine kinase inhibitors (TKIs. Currently, hematopoietic stem cell transplantation (HSCT is considered mainly as a third line salvage therapy in cases of TKIs resistance or intolerance. Here we describe a patient with chronic phase CML who developed both resistance and late occurrence of s severe thrombocytopenia on first and second generation TKIs and eventually underwent HSCT. Although the mechanism of the myelosuppression is not fully understood, we showed for the first time the development of dose dependent platelet antibodies in the presence of TKIs, suggesting the possibility of TKIs induced thrombocytopenia. Our case emphasizes that late development of severe myelosuppression during imatinib treatment is probably an important indication for consideration of early HSCT.

  8. Precision Therapy for Lung Cancer: Tyrosine Kinase Inhibitors and Beyond.

    Science.gov (United States)

    Rajan, Arun; Schrump, David S

    2015-01-01

    For patients with advanced cancers there has been a concerted effort to transition from a generic treatment paradigm to one based on tumor-specific biologic, and patient-specific clinical characteristics. This approach, known as precision therapy has been made possible owing to widespread availability and a reduction in the cost of cutting-edge technologies that are used to study the genomic, proteomic, and metabolic attributes of individual tumors. This review traces the evolution of precision therapy for lung cancer from the identification of molecular subsets of the disease to the development and approval of tyrosine kinase, as well as immune checkpoint inhibitors for lung cancer therapy. Challenges of the precision therapy era including the emergence of acquired resistance, identification of untargetable mutations, and the effect on clinical trial design are discussed. We conclude by highlighting newer applications for the concept of precision therapy.

  9. Exploiting Chemical Libraries, Structure, and Genomics in the Search for Kinase Inhibitors

    NARCIS (Netherlands)

    Gray, Nathanael S.; Wodicka, Lisa; Thunnissen, Andy-Mark W.H.; Norman, Thea C.; Kwon, Soojin; Espinoza, F. Hernan; Morgan, David O.; Barnes, Georjana; LeClerc, Sophie; Meijer, Laurent; Kim, Sung-Hou; Lockhart, David J.; Schultz, Peter G.

    1998-01-01

    Selective protein kinase inhibitors were developed on the basis of the unexpected binding mode of 2,6,9-trisubstituted purines to the adenosine triphosphate-binding site of the human cyclin-dependent kinase 2 (CDK2). By iterating chemical library synthesis and biological screening, potent inhibitors

  10. Nocturnal activation of aurora C in rat pineal gland: its role in the norepinephrine-induced phosphorylation of histone H3 and gene expression.

    Science.gov (United States)

    Price, D M; Kanyo, R; Steinberg, N; Chik, C L; Ho, A K

    2009-05-01

    We have shown previously that Ser10 phosphorylation of histone H3 occurs in rat pinealocytes after stimulation with norepinephrine (NE) and that histone modifications such as acetylation appear to play an important role in pineal gene transcription. Here we report the nocturnal phosphorylation of a Ser10 histone H3 kinase, Aurora C, in the rat pineal gland. The time profile of this phosphorylation parallels the increase in the level of phospho-Ser10 histone H3. Studies with cultured pinealocytes indicate that Aurora C phosphorylation is induced by NE and this induction can be blocked by cotreatment with propranolol or KT5720, a protein kinase A inhibitor. Moreover, only treatment with dibutyryl cAMP, but not other kinase activators, mimics the effect of NE on Aurora C phosphorylation. These results indicate that Aurora C is phosphorylated primarily by a beta-adrenergic/protein kinase A-mediated mechanism. Treatment with an Aurora C inhibitor reduces the NE-induced histone H3 phosphorylation and suppresses the NE-stimulated induction of arylalkylamine N-acetyltransferase (AA-NAT), the rhythm-controlling enzyme of melatonin synthesis, and melatonin production. The effects of Aurora C inhibitors on adrenergic-induced genes in rat pinealocytes are gene specific: inhibitory for Aa-nat and inducible cAMP repressor but stimulatory for c-fos. Together our results support a role for the NE-stimulated phosphorylation of Aurora C and the subsequent remodeling of chromatin in NE-stimulated Aa-nat transcription. This phenomenon suggests that activation of this mitotic kinase can be induced by extracellular signals to participate in the transcriptional induction of a subset of genes in the rat pineal gland.

  11. Structure-based lead discovery for protein kinase C zeta inhibitor design by exploiting kinase-inhibitor complex crystal structure data and potential therapeutics for preterm labour.

    Science.gov (United States)

    Shao, Qing-Chun; Zhang, Cui-Juan; Li, Jie

    2014-10-14

    The protein kinase C (PKC) is a family of serine/threonine kinases with a broad range of cellular targets. Members of the PKC family participate at the diverse biological events involved in cellular proliferation, differentiation and survival. The PKC isoform zeta (PKCζ) is an atypical member that has recently been found to play an essential role in promoting human uterine contractility and thus been raised as a new target for treating preterm labour and other tocolytic diseases. In this study, an integrative protocol was described to graft hundreds of inhibitor ligands from their complex crystal structures with cognate kinases into the active pocket of PKCζ and, based on the modeled structures, to evaluate the binding strength of these inhibitors to the non-cognate PKCζ receptor by using a consensus scoring strategy. A total of 32 inhibitors with top score were compiled, and eight out of them were tested for inhibitory potency against PKCζ. Consequently, five compounds, i.e. CDK6 inhibitor fisetin, PIM1 inhibitor myricetin, CDK9 inhibitor flavopiridol and PknB inhibitor mitoxantrone as well as the promiscuous kinase inhibitor staurosporine showed high or moderate inhibitory activity on PKCζ, with IC50 values of 58 ± 9, 1.7 ± 0.4, 108 ± 17, 280 ± 47 and 0.019 ± 0.004 μM, respectively, while other three compounds, including two marketed drugs dasatinib and sunitinib as well as the Rho inhibitor fasudil, have not been detected to possess observable activity. Next, based on the modeled structure data we modified three flavonoid kinase inhibitors, i.e. fisetin, myricetin and flavopiridol, to generate a number of more potential molecular entities, two of which were found to have a moderately improved activity as compared to their parent compounds.

  12. Benzothiophene inhibitors of MK2. Part 2: improvements in kinase selectivity and cell potency.

    Science.gov (United States)

    Anderson, David R; Meyers, Marvin J; Kurumbail, Ravi G; Caspers, Nicole; Poda, Gennadiy I; Long, Scott A; Pierce, Betsy S; Mahoney, Matthew W; Mourey, Robert J; Parikh, Mihir D

    2009-08-15

    Optimization of kinase selectivity for a set of benzothiophene MK2 inhibitors provided analogs with potencies of less than 500 nM in a cell based assay. The selectivity of the inhibitors can be rationalized by examination of X-ray crystal structures of inhibitors bound to MK2.

  13. Benzothiophene inhibitors of MK2. Part 2: Improvements in kinase selectivity and cell potency

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, David R.; Meyers, Marvin J.; Kurumbail, Ravi G.; Caspers, Nicole; Poda, Gennadiy I.; Long, Scott A.; Pierce, Betsy S.; Mahoney, Matthew W.; Mourey, Robert J.; Parikh, Mihir D.; Pfizer

    2010-10-01

    Optimization of kinase selectivity for a set of benzothiophene MK2 inhibitors provided analogs with potencies of less than 500 nM in a cell based assay. The selectivity of the inhibitors can be rationalized by examination of X-ray crystal structures of inhibitors bound to MK2.

  14. The logic and design of analog-sensitive kinases and their small molecule inhibitors.

    Science.gov (United States)

    Lopez, Michael S; Kliegman, Joseph I; Shokat, Kevan M

    2014-01-01

    Analog-sensitive AS Kinase technology allows for rapid, reversible, and highly specific inhibition of individual engineered kinases in cells and in mouse models of human diseases. The technique consists of two parts: a kinase containing a space-creating mutation in the ATP-binding pocket and a bulky ATP-competitive small molecule inhibitor that complements the shape of the mutant ATP pocket. This strategy enables dissection of phospho-signaling pathways, elucidation of the physiological function of individual kinases, and characterization of the pharmacology of clinical-kinase inhibitors. Here, we present an overview of AS technology and describe a stepwise approach for generating AS Kinase mutants and identifying appropriate small molecule inhibitors. We also describe commonly encountered technical obstacles and provide strategies to overcome them.

  15. Afatinib: emerging next-generation tyrosine kinase inhibitor for NSCLC

    Directory of Open Access Journals (Sweden)

    Nelson V

    2013-03-01

    Full Text Available Valerie Nelson, Jacqueline Ziehr, Mark Agulnik, Melissa JohnsonRobert H Lurie Comprehensive Cancer Center of Northwestern University, Chicago, IL, USAAbstract: The discovery of epidermal growth-factor receptor (EGFR-activating mutations and the introduction of oral EGFR tyrosine kinase inhibitors (EGFR-TKIs have expanded the treatment options for patients with non-small cell lung cancer. The first two reversible EGFR-TKIs, erlotinib and gefitinib, are approved for use in the first-line setting in patients with known EGFR-activating mutations and in the second- and third-line settings for all NSCLC patients. These first-generation EGFR-TKIs improve progression-free survival when compared to chemotherapy in patients with EGFR-activating mutations in the first-line setting. However, nearly all patients develop resistance to EGFR-directed agents. There is a need for further therapy options for patients with disease progression after treatment with reversible EGFR-TKIs. Afatinib is an irreversible ErbB family blocker that inhibits EGFR, HER2, and HER4. In vitro and in vivo, afatinib have shown increased inhibition of the common EGFR-activating mutations as well as the T790M resistance mutation when compared to erlotinib and gefitinib. Clinically, afatinib has been evaluated in the LUX-Lung series of trials, with improvement in progression-free survival reported in patients with EGFR-activating mutations in both first- and second-/third-line settings when compared to chemotherapy. Further investigation is needed to determine the precise role that afatinib will play in the treatment of patients with non-small cell lung cancer and EGFR-activating mutations.Keywords: afatinib, EGFR, irreversible EGFR inhibitor, EGPR-TKIs, LUX lung, resistance mutation, targeted therapy

  16. Fluorescent biosensors for high throughput screening of protein kinase inhibitors.

    Science.gov (United States)

    Prével, Camille; Pellerano, Morgan; Van, Thi Nhu Ngoc; Morris, May C

    2014-02-01

    High throughput screening assays aim to identify small molecules that interfere with protein function, activity, or conformation, which can serve as effective tools for chemical biology studies of targets involved in physiological processes or pathways of interest or disease models, as well as templates for development of therapeutics in medicinal chemistry. Fluorescent biosensors constitute attractive and powerful tools for drug discovery programs, from high throughput screening assays, to postscreen characterization of hits, optimization of lead compounds, and preclinical evaluation of candidate drugs. They provide a means of screening for inhibitors that selectively target enzymatic activity, conformation, and/or function in vitro. Moreover, fluorescent biosensors constitute useful tools for cell- and image-based, multiplex and multiparametric, high-content screening. Application of fluorescence-based sensors to screen large and complex libraries of compounds in vitro, in cell-based formats or whole organisms requires several levels of optimization to establish robust and reproducible assays. In this review, we describe the different fluorescent biosensor technologies which have been applied to high throughput screens, and discuss the prerequisite criteria underlying their successful application. Special emphasis is placed on protein kinase biosensors, since these enzymes constitute one of the most important classes of therapeutic targets in drug discovery.

  17. Structure of Human G Protein-Coupled Receptor Kinase 2 in Complex with the Kinase Inhibitor Balanol

    Energy Technology Data Exchange (ETDEWEB)

    Tesmer, John J.G.; Tesmer, Valerie M.; Lodowski, David T.; Steinhagen, Henning; Huber, Jochen (Sanofi); (Michigan); (Texas)

    2010-07-19

    G protein-coupled receptor kinase 2 (GRK2) is a pharmaceutical target for the treatment of cardiovascular diseases such as congestive heart failure, myocardial infarction, and hypertension. To better understand how nanomolar inhibition and selectivity for GRK2 might be achieved, we have determined crystal structures of human GRK2 in complex with G{beta}{gamma} in the presence and absence of the AGC kinase inhibitor balanol. The selectivity of balanol among human GRKs is assessed.

  18. Regulation of the activity of protein kinases by endogenous heat stable protein inhibitors.

    Science.gov (United States)

    Szmigielski, A

    1985-01-01

    Protein kinase activities are regulated by endogenous thermostable protein inhibitors. Type I inhibitor is a protein of MW 22,000-24,000 which inhibits specifically cyclic AMP-(cAMP) dependent protein kinase (APK) as a competitive inhibitor of catalytic subunits of the enzyme. Type I inhibitor activity changes inversely according to the activation of adenylate cyclase and the changes in cAMP content in tissues. It seems that type I inhibitor serves as a factor preventing spontaneous cAMP-dependent phosphorylation in unstimulated cell. The other thermostable protein which inhibits APK activity has been found in Sertoli cell-enriched testis (testis inhibitor). Physiological role of the testis inhibitor is unknown. Type II inhibitor is a protein of MW 15,000 which blocks phosphorylation mediated by cAMP and cyclic GMP (cGMP) dependent (APK and GPK) and cyclic nucleotide independent protein kinases as a competitive inhibitor of substrate proteins. Activity of this inhibitor specifically changes in reciprocal manner to the changes in cGMP content. It seems that type II inhibitor serves as a factor preventing the phosphorylation catalyzed by GPK when cGMP content is low. Stimulation of guanylate cyclase and activation of GPK is followed by a decrease of type II inhibitor activity. This change in relationship between activities of GPK and type II inhibitor allows for effective phosphorylation catalyzed by this enzyme when cGMP content is increased.

  19. Global effects of kinase inhibitors on signaling networks revealed by quantitative phosphoproteomics

    DEFF Research Database (Denmark)

    Pan, Cuiping; Olsen, Jesper V; Daub, Henrik;

    2009-01-01

    to identify the direct targets of kinase inhibitors upon affinity purification from cellular extracts. Here we introduce a complementary approach to evaluate the effects of kinase inhibitors on the entire cell signaling network. We used triple labeling SILAC (stable isotope labeling by amino acids in cell...... culture) to compare cellular phosphorylation levels for control, epidermal growth factor stimulus, and growth factor combined with kinase inhibitors. Of thousands of phosphopeptides, less than 10% had a response pattern indicative of targets of U0126 and SB202190, two widely used MAPK inhibitors....... Interestingly, 83% of the growth factor-induced phosphorylation events were affected by either or both inhibitors, showing quantitatively that early signaling processes are predominantly transmitted through the MAPK cascades. In contrast to MAPK inhibitors, dasatinib, a clinical drug directed against BCR...

  20. Recent Advances in the Development and Application of Radiolabeled Kinase Inhibitors for PET Imaging

    Directory of Open Access Journals (Sweden)

    Vadim Bernard-Gauthier

    2015-12-01

    Full Text Available Over the last 20 years, intensive investigation and multiple clinical successes targeting protein kinases, mostly for cancer treatment, have identified small molecule kinase inhibitors as a prominent therapeutic class. In the course of those investigations, radiolabeled kinase inhibitors for positron emission tomography (PET imaging have been synthesized and evaluated as diagnostic imaging probes for cancer characterization. Given that inhibitor coverage of the kinome is continuously expanding, in vivo PET imaging will likely find increasing applications for therapy monitoring and receptor density studies both in- and outside of oncological conditions. Early investigated radiolabeled inhibitors, which are mostly based on clinically approved tyrosine kinase inhibitor (TKI isotopologues, have now entered clinical trials. Novel radioligands for cancer and PET neuroimaging originating from novel but relevant target kinases are currently being explored in preclinical studies. This article reviews the literature involving radiotracer design, radiochemistry approaches, biological tracer evaluation and nuclear imaging results of radiolabeled kinase inhibitors for PET reported between 2010 and mid-2015. Aspects regarding the usefulness of pursuing selective vs. promiscuous inhibitor scaffolds and the inherent challenges associated with intracellular enzyme imaging will be discussed.

  1. Enzyme kinetics and distinct modulation of the protein kinase N family of kinases by lipid activators and small molecule inhibitors

    Science.gov (United States)

    Falk, Matthew D.; Liu, Wei; Bolaños, Ben; Unsal-Kacmaz, Keziban; Klippel, Anke; Grant, Stephan; Brooun, Alexei; Timofeevski, Sergei

    2014-01-01

    The PKN (protein kinase N) family of Ser/Thr protein kinases regulates a diverse set of cellular functions, such as cell migration and cytoskeletal organization. Inhibition of tumour PKN activity has been explored as an oncology therapeutic approach, with a PKN3-targeted RNAi (RNA interference)-derived therapeutic agent in Phase I clinical trials. To better understand this important family of kinases, we performed detailed enzymatic characterization, determining the kinetic mechanism and lipid sensitivity of each PKN isoform using full-length enzymes and synthetic peptide substrate. Steady-state kinetic analysis revealed that PKN1–3 follows a sequential ordered Bi–Bi kinetic mechanism, where peptide substrate binding is preceded by ATP binding. This kinetic mechanism was confirmed by additional kinetic studies for product inhibition and affinity of small molecule inhibitors. The known lipid effector, arachidonic acid, increased the catalytic efficiency of each isoform, mainly through an increase in kcat for PKN1 and PKN2, and a decrease in peptide KM for PKN3. In addition, a number of PKN inhibitors with various degrees of isoform selectivity, including potent (Ki<10 nM) and selective PKN3 inhibitors, were identified by testing commercial libraries of small molecule kinase inhibitors. This study provides a kinetic framework and useful chemical probes for understanding PKN biology and the discovery of isoform-selective PKN-targeted inhibitors. PMID:27919031

  2. Current understanding of tyrosine kinase BMX in inflammation and its inhibitors

    Directory of Open Access Journals (Sweden)

    Le Qiu

    2014-07-01

    Full Text Available Tec family kinases, which include tyrosine kinase expressed in hepatocellular carcinoma (TEC, Bruton's tyrosine kinase (BTK, interleukin (IL-2-inducible T-cell kinase (ITK, tyrosine-protein kinase (TXK, and bone marrow tyrosine kinase on chromosome X (BMX, are the second largest group of non-receptor tyrosine kinases and have a highly conserved carboxyl-terminal kinase domain. BMX was identified in human bone marrow cells, and was demonstrated to have been expressed in myeloid hematopoietic lineages cells, endothelial cells, and several types of cancers. Significant progress in this area during the last decade revealed an important role for BMX in inflammation and oncologic disorders. This review focuses on BMX biology, its role in inflammation and possible signaling pathways, and the potential of selective BMX inhibitors.

  3. Current understanding of tyrosine kinase BMX in inflammation and its inhibitors.

    Science.gov (United States)

    Qiu, Le; Wang, Fei; Liu, Sheng; Chen, Xu-Lin

    2014-01-01

    Tec family kinases, which include tyrosine kinase expressed in hepatocellular carcinoma (TEC), Bruton's tyrosine kinase (BTK), interleukin (IL)-2-inducible T-cell kinase (ITK), tyrosine-protein kinase (TXK), and bone marrow tyrosine kinase on chromosome X (BMX), are the second largest group of non-receptor tyrosine kinases and have a highly conserved carboxyl-terminal kinase domain. BMX was identified in human bone marrow cells, and was demonstrated to have been expressed in myeloid hematopoietic lineages cells, endothelial cells, and several types of cancers. Significant progress in this area during the last decade revealed an important role for BMX in inflammation and oncologic disorders. This review focuses on BMX biology, its role in inflammation and possible signaling pathways, and the potential of selective BMX inhibitors.

  4. Possible Role of Aurora-C in Meiosis

    Directory of Open Access Journals (Sweden)

    Kuo-Tai eYang

    2015-08-01

    Full Text Available The meiotic generation of haploid gametes with equal contents of genetic material is important for sexual reproduction in mammals. Errors in the transmission of chromosomes during meiosis may lead to aneuploidy, which is the leading cause of miscarriage and congenital birth defects in humans. The Aurora kinases, which include Aurora-A, Aurora-B, and Aurora-C, are highly conserved serine-threonine kinases that play essential roles in centrosome function, chromosome segregation, and cytokinesis during mitosis and meiosis. While Aurora-A and Aurora-B have been extensively studied in mitosis, the role of Aurora-C in meiosis is only now starting to be revealed. For example, the perturbation of Aurora-C kinase activity by microinjection of Aurora-C-kinase-dead mutant mRNAs into mouse oocytes induced multiple defects, including chromosome misalignment, abnormal kinetochore-microtubule attachment, premature chromosome segregation, and failure of cytokinesis during meiotic division. However, the analysis of such defects is complicated by the possibility that Aurora-B may be present in mammalian germ cells. Interestingly, a homozygous mutation of Aurora-C in humans leads to the production of large-headed polyploid spermatozoa and causes male infertility, but homozygous females are fertile. Mouse studies regarding the roles of Aurora-B and Aurora-C in female meiotic divisions have yielded inconsistent results, and it has proven difficult to explain why homozygous human females have no significant clinical phenotype. In this review, we will discuss the controversial status of Aurora-B in oocytes and the possible role of Aurora-C during meiotic division.

  5. Structural insight into selectivity and resistance profiles of ROS1 tyrosine kinase inhibitors.

    Science.gov (United States)

    Davare, Monika A; Vellore, Nadeem A; Wagner, Jacob P; Eide, Christopher A; Goodman, James R; Drilon, Alexander; Deininger, Michael W; O'Hare, Thomas; Druker, Brian J

    2015-09-29

    Oncogenic ROS1 fusion proteins are molecular drivers in multiple malignancies, including a subset of non-small cell lung cancer (NSCLC). The phylogenetic proximity of the ROS1 and anaplastic lymphoma kinase (ALK) catalytic domains led to the clinical repurposing of the Food and Drug Administration (FDA)-approved ALK inhibitor crizotinib as a ROS1 inhibitor. Despite the antitumor activity of crizotinib observed in both ROS1- and ALK-rearranged NSCLC patients, resistance due to acquisition of ROS1 or ALK kinase domain mutations has been observed clinically, spurring the development of second-generation inhibitors. Here, we profile the sensitivity and selectivity of seven ROS1 and/or ALK inhibitors at various levels of clinical development. In contrast to crizotinib's dual ROS1/ALK activity, cabozantinib (XL-184) and its structural analog foretinib (XL-880) demonstrate a striking selectivity for ROS1 over ALK. Molecular dynamics simulation studies reveal structural features that distinguish the ROS1 and ALK kinase domains and contribute to differences in binding site and kinase selectivity of the inhibitors tested. Cell-based resistance profiling studies demonstrate that the ROS1-selective inhibitors retain efficacy against the recently reported CD74-ROS1(G2032R) mutant whereas the dual ROS1/ALK inhibitors are ineffective. Taken together, inhibitor profiling and stringent characterization of the structure-function differences between the ROS1 and ALK kinase domains will facilitate future rational drug design for ROS1- and ALK-driven NSCLC and other malignancies.

  6. Crystal Structure of the MAP3K TAO2 Kinase Domain Bound by an Inhibitor Staurosporine

    Institute of Scientific and Technical Information of China (English)

    Tian-Jun ZHOU; Li-Guang SUN; Yan GAO; Elizabeth J. GOLDSMITH

    2006-01-01

    Mitogen-activated protein kinase (MAPK) signal transduction pathways are ubiquitous in eukaryotic cells, which transfer signals from the cell surface to the nucleus, controlling multiple cellular programs. MAPKs are activated by MAPK kinases [MAP2Ks or MAP/extracellular signal-regulated kinase (ERK) kinases (MEK)], which in turn are activated by MAPK kinase kinases (MAP3Ks). TAO2 is a MAP3K level kinase that activates the MAP2Ks MEK3 and MEK6 to activate p38 MAPKs. Because p38 MAPKs are key regulators of expression of inflammatory cytokines, they appear to be involved in human diseases such as asthma and autoimmunity. As an upstream activator of p38s, TAO2 represents a potential drug target. Here we report the crystal structure of active TAO2 kinase domain in complex with staurosporine, a broadrange protein kinase inhibitor that inhibits TAO2 with an IC50 of 3 μM. The structure reveals that staurosporine occupies the position where the adenosine of ATP binds in TAO2, and the binding of the inhibitor mimics many features of ATP binding. Both polar and nonpolar interactions contribute to the enzyme-inhibitor recognition. Staurosporine induces conformational changes in TAO2 residues that surround the inhibitor molecule, but causes very limited global changes in the kinase. The structure provides atomic details for TAO2-staurosporine interactions, and explains the relatively low potency of staurosporine against TAO2. The structure presented here should aid in the design of inhibitors specific to TAO2 and related kinases.

  7. Small-molecule kinase inhibitors: an analysis of FDA-approved drugs

    DEFF Research Database (Denmark)

    Wu, Peng; Nielsen, Thomas Eiland; Clausen, Mads Hartvig

    2016-01-01

    Small-molecule kinase inhibitors (SMKIs), 28 of which are approved by the US Food and Drug Administration (FDA), have been actively pursued as promising targeted therapeutics. Here, we assess the key structural and physicochemical properties, target selectivity and mechanism of function, and ther...... to be unreliable. Although previous SMKI research was concentrated on tyrosine kinase inhibitors for cancer treatment, recent progress indicates diversification of SMKI research in terms of new targets, mechanistic types, and therapeutic indications....

  8. Identification of p38α MAP kinase inhibitors by pharmacophore based virtual screening

    DEFF Research Database (Denmark)

    Gangwal, Rahul P; Das, Nihar R; Thanki, Kaushik;

    2014-01-01

    The p38α mitogen-activated protein (MAP) kinase plays a vital role in treating many inflammatory diseases. In the present study, a combined ligand and structure based pharmacophore model was developed to identify potential DFG-in selective p38 MAP kinase inhibitors. Conformations of co-crystallised...

  9. Kinase Inhibitor Screening Identifies Cyclin-Dependent Kinases and Glycogen Synthase Kinase 3 as Potential Modulators of TDP-43 Cytosolic Accumulation during Cell Stress.

    Science.gov (United States)

    Moujalled, Diane; James, Janine L; Parker, Sarah J; Lidgerwood, Grace E; Duncan, Clare; Meyerowitz, Jodi; Nonaka, Takashi; Hasegawa, Masato; Kanninen, Katja M; Grubman, Alexandra; Liddell, Jeffrey R; Crouch, Peter J; White, Anthony R

    2013-01-01

    Abnormal processing of TAR DNA binding protein 43 (TDP-43) has been identified as a major factor in neuronal degeneration during amyotrophic lateral sclerosis (ALS) or frontotemporal lobar degeneration (FTLD). It is unclear how changes to TDP-43, including nuclear to cytosolic translocation and subsequent accumulation, are controlled in these diseases. TDP-43 is a member of the heterogeneous ribonucleoprotein (hnRNP) RNA binding protein family and is known to associate with cytosolic RNA stress granule proteins in ALS and FTLD. hnRNP trafficking and accumulation is controlled by the action of specific kinases including members of the mitogen-activated protein kinase (MAPK) pathway. However, little is known about how kinase pathways control TDP-43 movement and accumulation. In this study, we used an in vitro model of TDP-43-positve stress granule formation to screen for the effect of kinase inhibitors on TDP-43 accumulation. We found that while a number of kinase inhibitors, particularly of the MAPK pathways modulated both TDP-43 and the global stress granule marker, human antigen R (HuR), multiple inhibitors were more specific to TDP-43 accumulation, including inhibitors of cyclin-dependent kinases (CDKs) and glycogen synthase kinase 3 (GSK3). Close correlation was observed between effects of these inhibitors on TDP-43, hnRNP K and TIAR, but often with different effects on HuR accumulation. This may indicate a potential interaction between TDP-43, hnRNP K and TIAR. CDK inhibitors were also found to reverse pre-formed TDP-43-positive stress granules and both CDK and GSK3 inhibitors abrogated the accumulation of C-terminal TDP-43 (219-414) in transfected cells. Further studies are required to confirm the specific kinases involved and whether their action is through phosphorylation of the TDP-43 binding partner hnRNP K. This knowledge provides a valuable insight into the mechanisms controlling abnormal cytoplasmic TDP-43 accumulation and may herald new opportunities

  10. Kinase Inhibitor Screening Identifies Cyclin-Dependent Kinases and Glycogen Synthase Kinase 3 as Potential Modulators of TDP-43 Cytosolic Accumulation during Cell Stress.

    Directory of Open Access Journals (Sweden)

    Diane Moujalled

    Full Text Available Abnormal processing of TAR DNA binding protein 43 (TDP-43 has been identified as a major factor in neuronal degeneration during amyotrophic lateral sclerosis (ALS or frontotemporal lobar degeneration (FTLD. It is unclear how changes to TDP-43, including nuclear to cytosolic translocation and subsequent accumulation, are controlled in these diseases. TDP-43 is a member of the heterogeneous ribonucleoprotein (hnRNP RNA binding protein family and is known to associate with cytosolic RNA stress granule proteins in ALS and FTLD. hnRNP trafficking and accumulation is controlled by the action of specific kinases including members of the mitogen-activated protein kinase (MAPK pathway. However, little is known about how kinase pathways control TDP-43 movement and accumulation. In this study, we used an in vitro model of TDP-43-positve stress granule formation to screen for the effect of kinase inhibitors on TDP-43 accumulation. We found that while a number of kinase inhibitors, particularly of the MAPK pathways modulated both TDP-43 and the global stress granule marker, human antigen R (HuR, multiple inhibitors were more specific to TDP-43 accumulation, including inhibitors of cyclin-dependent kinases (CDKs and glycogen synthase kinase 3 (GSK3. Close correlation was observed between effects of these inhibitors on TDP-43, hnRNP K and TIAR, but often with different effects on HuR accumulation. This may indicate a potential interaction between TDP-43, hnRNP K and TIAR. CDK inhibitors were also found to reverse pre-formed TDP-43-positive stress granules and both CDK and GSK3 inhibitors abrogated the accumulation of C-terminal TDP-43 (219-414 in transfected cells. Further studies are required to confirm the specific kinases involved and whether their action is through phosphorylation of the TDP-43 binding partner hnRNP K. This knowledge provides a valuable insight into the mechanisms controlling abnormal cytoplasmic TDP-43 accumulation and may herald new

  11. Purine inhibitors of protein kinases, G proteins and polymerases

    Energy Technology Data Exchange (ETDEWEB)

    Gray, Nathanael S. (Berkeley, CA); Schultz, Peter (Oakland, CA); Kim, Sung-Hou (Moraga, CA); Meijer, Laurent (Roscoff, FR)

    2001-07-03

    The present invention relates to purine analogs that inhibit, inter alia, protein kinases, G-proteins and polymerases. In addition, the present invention relates to methods of using such purine analogs to inhibit protein kinases, G-proteins, polymerases and other cellular processes and to treat cellular proliferative diseases.

  12. Design, synthesis and characterization of a highly effective inhibitor for analog-sensitive (as) kinases.

    Science.gov (United States)

    Klein, Michael; Morillas, Montse; Vendrell, Alexandre; Brive, Lars; Gebbia, Marinella; Wallace, Iain M; Giaever, Guri; Nislow, Corey; Posas, Francesc; Grøtli, Morten

    2011-01-01

    Highly selective, cell-permeable and fast-acting inhibitors of individual kinases are sought-after as tools for studying the cellular function of kinases in real time. A combination of small molecule synthesis and protein mutagenesis, identified a highly potent inhibitor (1-Isopropyl-3-(phenylethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine) of a rationally engineered Hog1 serine/threonine kinase (Hog1(T100G)). This inhibitor has been successfully used to study various aspects of Hog1 signaling, including a transient cell cycle arrest and gene expression changes mediated by Hog1 in response to stress. This study also underscores that the general applicability of this approach depends, in part, on the selectivity of the designed the inhibitor with respect to activity versus the engineered and wild type kinases. To explore this specificity in detail, we used a validated chemogenetic assay to assess the effect of this inhibitor on all gene products in yeast in parallel. The results from this screen emphasize the need for caution and for case-by-case assessment when using the Analog-Sensitive Kinase Allele technology to assess the physiological roles of kinases.

  13. Opening the door to the development of novel Abl kinase inhibitors.

    Science.gov (United States)

    Bezerra Morais, Pedro Alves; Daltoé, Renata Dalmaschio; Paula, Heberth de

    2016-10-24

    The discovery of the importance of kinase activity and its relationship to the emergence and proliferation of cancer cells, due to changes in normal physiology, opened a remarkable pathway for the treatment of chronic myelogenous leukemia through intense search of drug candidates. Six Abl kinase inhibitors have received the US FDA approval as chronic myelogenous leukemia treatment, and continuous efforts in obtaining new, more effective and selective molecules are being carried out. Herein we discuss the mechanisms of Abl inhibition, structural features and ligand/protein interactions that are important for the design of new Abl kinase inhibitors. This review provides a broad overview of binding mode predictions, through molecular docking, which can be an approach to discover novel Abl kinase inhibitors.

  14. Identification of potent Yes1 kinase inhibitors using a library screening approach.

    Science.gov (United States)

    Patel, Paresma R; Sun, Hongmao; Li, Samuel Q; Shen, Min; Khan, Javed; Thomas, Craig J; Davis, Mindy I

    2013-08-01

    Yes1 kinase has been implicated as a potential therapeutic target in a number of cancers including melanomas, breast cancers, and rhabdomyosarcomas. Described here is the development of a robust and miniaturized biochemical assay for Yes1 kinase that was applied in a high throughput screen (HTS) of kinase-focused small molecule libraries. The HTS provided 144 (17% hit rate) small molecule compounds with IC₅₀ values in the sub-micromolar range. Three of the most potent Yes1 inhibitors were then examined in a cell-based assay for inhibition of cell survival in rhabdomyosarcoma cell lines. Homology models of Yes1 were generated in active and inactive conformations, and docking of inhibitors supports binding to the active conformation (DFG-in) of Yes1. This is the first report of a large high throughput enzymatic activity screen for identification of Yes1 kinase inhibitors, thereby elucidating the polypharmacology of a variety of small molecules and clinical candidates.

  15. Effect of kinase inhibitors on the therapeutic properties of monoclonal antibodies.

    Science.gov (United States)

    Duong, Minh Ngoc; Matera, Eva-Laure; Mathé, Doriane; Evesque, Anne; Valsesia-Wittmann, Sandrine; Clémenceau, Béatrice; Dumontet, Charles

    2015-01-01

    Targeted therapies of malignancies currently consist of therapeutic monoclonal antibodies and small molecule kinase inhibitors. The combination of these novel agents raises the issue of potential antagonisms. We evaluated the potential effect of 4 kinase inhibitors, including the Bruton tyrosine kinase inhibitor ibrutinib, and 3 PI3K inhibitors idelalisib, NVP-BEZ235 and LY294002, on the effects of the 3 monoclonal antibodies, rituximab and obinutuzumab (directed against CD20) and trastuzumab (directed against HER2). We found that ibrutinib potently inhibits antibody-dependent cell-mediated cytotoxicity exerted by all antibodies, with a 50% inhibitory concentration of 0.2 microM for trastuzumab, 0.5 microM for rituximab and 2 microM for obinutuzumab, suggesting a lesser effect in combination with obinutuzumab than with rituximab. The 4 kinase inhibitors were found to inhibit phagocytosis by fresh human neutrophils, as well as antibody-dependent cellular phagocytosis induced by the 3 antibodies. Conversely co-administration of ibrutinib with rituximab, obinutuzumab or trastuzumab did not demonstrate any inhibitory effect of ibrutinib in vivo in murine xenograft models. In conclusion, some kinase inhibitors, in particular, ibrutinib, are likely to exert inhibitory effects on innate immune cells. However, these effects do not compromise the antitumor activity of monoclonal antibodies in vivo in the models that were evaluated.

  16. Structural analysis of ARC-type inhibitor (ARC-1034) binding to protein kinase A catalytic subunit and rational design of bisubstrate analogue inhibitors of basophilic protein kinases.

    Science.gov (United States)

    Lavogina, Darja; Lust, Marje; Viil, Indrek; König, Norbert; Raidaru, Gerda; Rogozina, Jevgenia; Enkvist, Erki; Uri, Asko; Bossemeyer, Dirk

    2009-01-22

    The crystal structure of a complex of the catalytic subunit (type alpha) of cAMP-dependent protein kinase (PKA C alpha) with ARC-type inhibitor (ARC-1034), the presumed lead scaffold of previously reported adenosine-oligo-arginine conjugate-based (ARC-type) inhibitors, was solved. Structural elements important for interaction with the kinase were established with specifically modified derivatives of the lead compound. On the basis of this knowledge, a new generation of inhibitors, conjugates of adenosine-4'-dehydroxymethyl-4'-carboxylic acid moiety and oligo(D-arginine), was developed with inhibitory constants well into the subnanomolar range. The structural determinants of selectivity of the new compounds were established in assays with ROCK-II and PKBgamma.

  17. Structure-based design of isoquinoline-5-sulfonamide inhibitors of protein kinase B.

    Science.gov (United States)

    Collins, Ian; Caldwell, John; Fonseca, Tatiana; Donald, Alastair; Bavetsias, Vassilios; Hunter, Lisa-Jane K; Garrett, Michelle D; Rowlands, Martin G; Aherne, G Wynne; Davies, Thomas G; Berdini, Valerio; Woodhead, Steven J; Davis, Deborah; Seavers, Lisa C A; Wyatt, Paul G; Workman, Paul; McDonald, Edward

    2006-02-15

    Structure-based drug design of novel isoquinoline-5-sulfonamide inhibitors of PKB as potential antitumour agents was investigated. Constrained pyrrolidine analogues that mimicked the bound conformation of linear prototypes were identified and investigated by co-crystal structure determinations with the related protein PKA. Detailed variation in the binding modes between inhibitors with similar overall conformations was observed. Potent PKB inhibitors from this series inhibited GSK3beta phosphorylation in cellular assays, consistent with inhibition of PKB kinase activity in cells.

  18. Exploiting the repertoire of CK2 inhibitors to target DYRK and PIM kinases.

    Science.gov (United States)

    Cozza, Giorgio; Sarno, Stefania; Ruzzene, Maria; Girardi, Cristina; Orzeszko, Andrzej; Kazimierczuk, Zygmunt; Zagotto, Giuseppe; Bonaiuto, Emanuela; Di Paolo, Maria Luisa; Pinna, Lorenzo A

    2013-07-01

    Advantage has been taken of the relative promiscuity of commonly used inhibitors of protein kinase CK2 to develop compounds that can be exploited for the selective inhibition of druggable kinases other than CK2 itself. Here we summarize data obtained by altering the scaffold of CK2 inhibitors to give rise to novel selective inhibitors of DYRK1A and to a powerful cell permeable dual inhibitor of PIM1 and CK2. In the former case one of the new compounds, C624 (naphto [1,2-b]benzofuran-5,9-diol) displays a potency comparable to that of the first-in-class DYRK1A inhibitor, harmine, lacking however the drawback of drastically inhibiting monoamine oxidase-A (MAO-A) as harmine does. On the other hand the promiscuous CK2 inhibitor 4,5,6,7-tetrabromo-1H-benzimidazole (TBI,TBBz) has been derivatized with a sugar moiety to generate a 1-(β-D-2'-deoxyribofuranosyl)-4,5,6,7-tetrabromo-1H-benzimidazole (TDB) compound which inhibits PIM1 and CK2 with comparably high efficacy (IC50 valuescancer cells consistent with concomitant inhibition of both its onco-kinase targets. This article is part of a Special Issue entitled: Inhibitors of Protein Kinases (2012).

  19. Crystal structure of a human cyclin-dependent kinase 6 complexwith a flavonol inhibitor, Fisetin

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Heshu; Chang, Debbie J.; Baratte, Blandine; Meijer, Laurent; Schulze-Gahmen, Ursula

    2005-01-10

    Cyclin-dependent kinases (CDKs) play a central role in cell cycle control, apoptosis, transcription and neuronal functions. They are important targets for the design of drugs with anti-mitotic and/or anti-neurodegenerative effects. CDK4 and CDK6 form a subfamily among the CDKs in mammalian cells, as defined by sequence similarities. Compared to CDK2 and CDK5, structural information on CDK4 and CDK6 is sparse. We describe here the crystal structure of human CDK6 in complex with a viral cyclin and a flavonol inhibitor, fisetin. Fisetin binds to the active form of CDK6, forming hydrogen bonds with the side chains of residues in the binding pocket that undergo large conformational changes during CDK activation by cyclin binding. The 4-keto group and the 3-hydroxyl group of fisetin are hydrogen bonded with the backbone in the hinge region between the N-terminal and C-terminal kinase domain, as has been observed for many CDK inhibitors. However, CDK2 and HCK kinase in complex with other flavone inhibitors such as quercetin and flavopiridol showed a different binding mode with the inhibitor rotated by about 180. The structural information of the CDK6-fisetin complex is correlated with the binding affinities of different flavone inhibitors for CDK6. This complex structure is the first description of an inhibitor complex with a kinase from the CDK4/6 subfamily and can provide a basis for selecting and designing inhibitor compounds with higher affinity and specificity.

  20. Ability of the Met kinase inhibitor crizotinib and new generation EGFR inhibitors to overcome resistance to EGFR inhibitors.

    Directory of Open Access Journals (Sweden)

    Shigeki Nanjo

    Full Text Available PURPOSE: Although EGF receptor tyrosine kinase inhibitors (EGFR-TKI have shown dramatic effects against EGFR mutant lung cancer, patients ultimately develop resistance by multiple mechanisms. We therefore assessed the ability of combined treatment with the Met inhibitor crizotinib and new generation EGFR-TKIs to overcome resistance to first-generation EGFR-TKIs. EXPERIMENTAL DESIGN: Lung cancer cell lines made resistant to EGFR-TKIs by the gatekeeper EGFR-T790M mutation, Met amplification, and HGF overexpression and mice with tumors induced by these cells were treated with crizotinib and a new generation EGFR-TKI. RESULTS: The new generation EGFR-TKI inhibited the growth of lung cancer cells containing the gatekeeper EGFR-T790M mutation, but did not inhibit the growth of cells with Met amplification or HGF overexpression. In contrast, combined therapy with crizotinib plus afatinib or WZ4002 was effective against all three types of cells, inhibiting EGFR and Met phosphorylation and their downstream molecules. Crizotinib combined with afatinib or WZ4002 potently inhibited the growth of mouse tumors induced by these lung cancer cell lines. However, the combination of high dose crizotinib and afatinib, but not WZ4002, triggered severe adverse events. CONCLUSIONS: Our results suggest that the dual blockade of mutant EGFR and Met by crizotinib and a new generation EGFR-TKI may be promising for overcoming resistance to reversible EGFR-TKIs but careful assessment is warranted clinically.

  1. The maternal-effect gene cellular island encodes aurora B kinase and is essential for furrow formation in the early zebrafish embryo.

    Directory of Open Access Journals (Sweden)

    Taijiro Yabe

    2009-06-01

    Full Text Available Females homozygous for a mutation in cellular island (cei produce embryos with defects in cytokinesis during early development. Analysis of the cytoskeletal events associated with furrow formation reveal that these defects include a general delay in furrow initiation as well as a complete failure to form furrow-associated structures in distal regions of the blastodisc. A linkage mapping-based candidate gene approach, including transgenic rescue, shows that cei encodes the zebrafish Aurora B kinase homologue. Genetic complementation analysis between the cei mutation and aurB zygotic lethal mutations corroborate gene assignment and reveal a complex nature of the maternal-effect cei allele, which appears to preferentially affect a function important for cytokinesis in the early blastomeres. Surprisingly, in cei mutant embryos a short yet otherwise normal furrow forms in the center of the blastodisc. Furrow formation is absent throughout the width of the blastodisc in cei mutant embryos additionally mutant for futile cycle, which lack a spindle apparatus, showing that the residual furrow signal present in cei mutants is derived from the mitotic spindle. Our analysis suggests that partially redundant signals derived from the spindle and astral apparatus mediate furrow formation in medial and distal regions of the early embryonic blastomeres, respectively, possibly as a spatial specialization to achieve furrow formation in these large cells. In addition, our data also suggest a role for Cei/AurB function in the reorganization of the furrow-associated microtubules in both early cleavage- and somite-stage embryos. In accordance with the requirement for cei/aurB in furrow induction in the early cleavage embryo, germ plasm recruitment to the forming furrow is also affected in embryos lacking normal cei/aurB function.

  2. Mutations of the aurora kinase C gene causing macrozoospermia are the most frequent genetic cause of male infertility in Algerian men

    Directory of Open Access Journals (Sweden)

    Leyla Ounis

    2015-02-01

    Full Text Available Klinefelter syndrome and Y-chromosomal microdeletion analyses were once the only two genetic tests offered to infertile men. Analyses of aurora kinase C (AURKC and DPY19L2 are now recommended for patients presenting macrozoospermia and globozoospermia, respectively, two rare forms of teratozoospermia particularly frequent among North African men. We carried out genetic analyses on Algerian patients, to evaluate the prevalence of these syndromes in this population and to compare it with the expected frequency of Klinefelter syndrome and Y-microdeletions. We carried out a retrospective study on 599 consecutive patients consulting for couple infertility at the assisted reproduction unit of the Ibn Rochd Clinique, Constantine, Algeria. Abnormal sperm parameters were observed in 404 men. Fourteen and seven men had typical macrozoospermia and globozoospermia profiles, respectively. Molecular diagnosis was carried out for these patients, for the AURKC and DPY19L2 genes. Eleven men with macrozoospermia had a homozygous AURKC mutation (79%, corresponding to 2.7% of all patients with abnormal spermograms. All the men with globozoospermia studied (n = 5, corresponding to 1.2% of all infertile men, presented a homozygous DPY19L2 deletion. By comparison, we would expect 1.6% of the patients in this cohort to have Klinefelter syndrome and 0.23% to have Y-microdeletion. Our findings thus indicate that AURKC mutations are more frequent than Klinefelter syndrome and constitute the leading genetic cause of infertility in North African men. Furthermore, we estimate that AURKC and DPY19L2 molecular defects are 10 and 5 times more frequent, respectively, than Y-microdeletions.

  3. Condensin phosphorylated by the Aurora-B-like kinase Ark1 is continuously required until telophase in a mode distinct from Top2.

    Science.gov (United States)

    Nakazawa, Norihiko; Mehrotra, Rajesh; Ebe, Masahiro; Yanagida, Mitsuhiro

    2011-06-01

    Condensin is a conserved protein complex that functions in chromosome condensation and segregation. It has not been previously unequivocally determined whether condensin is required throughout mitosis. Here, we examined whether Schizosaccharomyces pombe condensin continuously acts on chromosomes during mitosis and compared its role with that of DNA topoisomerase II (Top2). Using double mutants containing a temperature-sensitive allele of the condensin SMC2 subunit cut14 (cut14-208) or of top2, together with the cold-sensitive nda3-KM311 mutation (in β-tubulin), temperature-shift experiments were performed. These experiments allowed inactivation of condensin or Top2 at various stages throughout mitosis, even after late anaphase. The results established that mitotic chromosomes require condensin and Top2 throughout mitosis, even in telophase. We then showed that the Cnd2 subunit of condensin (also known as Barren) is the target subunit of Aurora-B-like kinase Ark1 and that Ark1-mediated phosphorylation of Cnd2 occurred throughout mitosis. The phosphorylation sites in Cnd2 were determined by mass spectrometry, and alanine and glutamate residue replacement mutant constructs for these sites were constructed. Alanine substitution mutants of Cnd2, which mimic the unphosphorylated protein, exhibited broad mitotic defects, including at telophase, and overexpression of these constructs caused a severe dominant-negative effect. By contrast, glutamate substitution mutants, which mimic the phosphorylated protein, alleviated the segregation defect in Ark1-inhibited cells. In telophase, the condensin subunits in cut14-208 mutant accumulated in lumps that contained telomeric DNA and proteins that failed to segregate. Condensin might thus serve to keep the segregated chromosomes apart during telophase.

  4. Reduced Proteolytic Shedding of Receptor Tyrosine Kinases is a Post-Translational Mechanism of Kinase Inhibitor Resistance

    Science.gov (United States)

    Miller, Miles A.; Oudin, Madeleine J.; Sullivan, Ryan J.; Wang, Stephanie J.; Meyer, Aaron S.; Im, Hyungsoon; Frederick, Dennie T.; Tadros, Jenny; Griffith, Linda G.; Lee, Hakho; Weissleder, Ralph; Flaherty, Keith T.; Gertler, Frank B.; Lauffenburger, Douglas A.

    2016-01-01

    Kinase inhibitor resistance often involves upregulation of poorly understood “bypass” signaling pathways. Here, we show that extracellular proteomic adaptation is one path to bypass signaling and drug resistance. Proteolytic shedding of surface receptors, which can provide negative feedback on signaling activity, is blocked by kinase inhibitor treatment and enhances bypass signaling. In particular, MEK inhibition broadly decreases shedding of multiple receptor tyrosine kinases (RTKs) including HER4, MET, and most prominently AXL, an ADAM10 and ADAM17 substrate, thus increasing surface RTK levels and mitogenic signaling. Progression-free survival of melanoma patients treated with clinical BRAF/MEK inhibitors inversely correlates with RTK shedding reduction following treatment, as measured non-invasively in blood plasma. Disrupting protease inhibition by neutralizing TIMP1 improves MAPK inhibitor efficacy, and combined MAPK/AXL inhibition synergistically reduces tumor growth and metastasis in xenograft models. Altogether, extracellular proteomic rewiring through reduced RTK shedding represents a surprising mechanism for bypass signaling in cancer drug resistance. PMID:26984351

  5. MOLECULAR MODELING AND DRUG DISCOVERY OF POTENTIAL INHIBITORS FOR ANTICANCER TARGET GENE MELK (MATERNAL EMBRYONIC LEUCINE ZIPPER KINASE

    Directory of Open Access Journals (Sweden)

    Sabitha. K

    2011-12-01

    Full Text Available Maternal embryonic leucine zipper kinase (MELK, a member of the AMP serine/threonine kinase family, exhibits multiple features consistent with the potential utility of this gene as an anticancer target. Reports show that MELK functions as a cancer-specific protein kinase, and that down-regulation of MELK results in growth suppression of breast cancer cells. There are many inhibitors which bind to kinases and are in clinical trials too. In our study we have taken a library of different inhibitors and docked those using GLIDE Induced Fit. From docking result we can conclude that Syk inhibitor II, Rho kinase inhibitor IV, p38 MAP Kinase Inhibitor III, HA 1004, Dihydrochloride and IKK -2 inhibitor VI have good binding affinity towards MELK and may have anticancer activity.

  6. Purine inhibitors of protein kinases, G proteins and polymerases

    Energy Technology Data Exchange (ETDEWEB)

    Gray, Nathanael S.; Schultz, Peter; Kim, Sung-Hou; Meijer, Laurent

    2004-10-12

    The present invention relates to 2-N-substituted 6-(4-methoxybenzylamino)-9-isopropylpurines that inhibit, inter alia, protein kinases, G-proteins and polymerases. In addition, the present invention relates to methods of using such 2-N-substituted 6-(4-methoxybenzylamino)-9-isopropylpurines to inhibit protein kinases, G-proteins, polymerases and other cellular processes and to treat cellular proliferative diseases.

  7. Crystal structures of human RIP2 kinase catalytic domain complexed with ATP-competitive inhibitors: Foundations for understanding inhibitor selectivity.

    Science.gov (United States)

    Charnley, Adam K; Convery, Máire A; Lakdawala Shah, Ami; Jones, Emma; Hardwicke, Philip; Bridges, Angela; Ouellette, Michael; Totoritis, Rachel; Schwartz, Benjamin; King, Bryan W; Wisnoski, David D; Kang, James; Eidam, Patrick M; Votta, Bartholomew J; Gough, Peter J; Marquis, Robert W; Bertin, John; Casillas, Linda

    2015-11-01

    Receptor interacting protein 2 (RIP2) is an intracellular kinase and key signaling partner for the pattern recognition receptors NOD1 and NOD2 (nucleotide-binding oligomerization domain-containing proteins 1 and 2). As such, RIP2 represents an attractive target to probe the role of these pathways in disease. In an effort to design potent and selective inhibitors of RIP2 we established a crystallographic system and determined the structure of the RIP2 kinase domain in an apo form and also in complex with multiple inhibitors including AMP-PCP (β,γ-Methyleneadenosine 5'-triphosphate, a non-hydrolysable adenosine triphosphate mimic) and structurally diverse ATP competitive chemotypes identified via a high-throughput screening campaign. These structures represent the first set of diverse RIP2-inhibitor co-crystal structures and demonstrate that the protein possesses the ability to adopt multiple DFG-in as well as DFG-out and C-helix out conformations. These structures reveal key protein-inhibitor structural insights and serve as the foundation for establishing a robust structure-based drug design effort to identify both potent and highly selective inhibitors of RIP2 kinase.

  8. Targeting Cyclin-Dependent Kinases in Human Cancers: From Small Molecules to Peptide Inhibitors

    Directory of Open Access Journals (Sweden)

    Marion Peyressatre

    2015-01-01

    Full Text Available Cyclin-dependent kinases (CDK/Cyclins form a family of heterodimeric kinases that play central roles in regulation of cell cycle progression, transcription and other major biological processes including neuronal differentiation and metabolism. Constitutive or deregulated hyperactivity of these kinases due to amplification, overexpression or mutation of cyclins or CDK, contributes to proliferation of cancer cells, and aberrant activity of these kinases has been reported in a wide variety of human cancers. These kinases therefore constitute biomarkers of proliferation and attractive pharmacological targets for development of anticancer therapeutics. The structural features of several of these kinases have been elucidated and their molecular mechanisms of regulation characterized in depth, providing clues for development of drugs and inhibitors to disrupt their function. However, like most other kinases, they constitute a challenging class of therapeutic targets due to their highly conserved structural features and ATP-binding pocket. Notwithstanding, several classes of inhibitors have been discovered from natural sources, and small molecule derivatives have been synthesized through rational, structure-guided approaches or identified in high throughput screens. The larger part of these inhibitors target ATP pockets, but a growing number of peptides targeting protein/protein interfaces are being proposed, and a small number of compounds targeting allosteric sites have been reported.

  9. Structure-guided inhibitor design expands the scope of analog-sensitive kinase technology.

    Science.gov (United States)

    Zhang, Chao; Lopez, Michael S; Dar, Arvin C; Ladow, Eva; Finkbeiner, Steven; Yun, Cai-Hong; Eck, Michael J; Shokat, Kevan M

    2013-09-20

    Engineered analog-sensitive (AS) protein kinases have emerged as powerful tools for dissecting phospho-signaling pathways, for elucidating the cellular function of individual kinases, and for deciphering unanticipated effects of clinical therapeutics. A crucial and necessary feature of this technology is a bioorthogonal small molecule that is innocuous toward native cellular systems but potently inhibits the engineered kinase. In order to generalize this method, we sought a molecule capable of targeting divergent AS-kinases. Here we employ X-ray crystallography and medicinal chemistry to unravel the mechanism of current inhibitors and use these insights to design the most potent, selective, and general AS-kinase inhibitors reported to date. We use large-scale kinase inhibitor profiling to characterize the selectivity of these molecules as well as examine the consequences of potential off-target effects in chemical genetic experiments. The molecules reported here will serve as powerful tools in efforts to extend AS-kinase technology to the entire kinome and the principles discovered may help in the design of other engineered enzyme/ligand pairs.

  10. Protein Kinase Inhibitors CK59 and CID755673 Alter Primary Human NK Cell Effector Functions

    Science.gov (United States)

    Scheiter, Maxi; Bulitta, Björn; van Ham, Marco; Klawonn, Frank; König, Sebastian; Jänsch, Lothar

    2013-01-01

    Natural killer (NK) cells are part of the innate immune response and play a crucial role in the defense against tumors and virus-infected cells. Their effector functions include the specific killing of target cells, as well as the modulation of other immune cells by cytokine release. Kinases constitute a relevant part in signaling, are prime targets in drug research and the protein kinase inhibitor Dasatinib is already used for immune-modulatory therapies. In this study, we tested the effects of the kinase inhibitors CK59 and CID755673. These inhibitors are directed against calmodulin kinase II (CaMKII; CK59) and PKD family kinases (CID755673) that were previously suggested as novel components of NK activation pathways. Here, we use a multi-parameter, FACS-based assay to validate the influence of CK59 and CID755673 on the effector functions of primary NK cells. Treatment with CK59 and CID755673 indeed resulted in a significant dose-dependent reduction of NK cell degranulation markers and cytokine release in freshly isolated Peripheral blood mononuclear cell populations from healthy blood donors. These results underline the importance of CaMKII for NK cell signaling and suggest protein kinase D2 as a novel signaling component in NK cell activation. Notably, kinase inhibition studies on pure NK cell populations indicate significant donor variations. PMID:23508354

  11. Aurora B interacts with NIR-p53, leading to p53 phosphorylation in its DNA-binding domain and subsequent functional suppression.

    Science.gov (United States)

    Wu, Liming; Ma, Chi A; Zhao, Yongge; Jain, Ashish

    2011-01-21

    NIR (novel INHAT repressor) is a transcriptional co-repressor with inhibitor of histone acetyltransferase (INHAT) activity and has previously been shown to physically interact with and suppress p53 transcriptional activity and function. However, the mechanism by which NIR suppresses p53 is not completely understood. Using a proteomic approach, we have identified the Aurora kinase B as a novel binding partner of NIR. We show that Aurora B, NIR and p53 exist in a protein complex in which Aurora B binds to NIR, thus also indirectly associates with p53. Functionally, overexpression of Aurora B or NIR suppresses p53 transcriptional activity, and depletion of Aurora B or NIR causes p53-dependent apoptosis and cell growth arrest, due to the up-regulation of p21 and Bax. We then demonstrate that Aurora B phosphorylates multiple sites in the p53 DNA-binding domain in vitro, and this phosphorylation probably also occurs in cells. Importantly, the Aurora B-mediated phosphorylation on Ser(269) or Thr(284) significantly compromises p53 transcriptional activity. Taken together, these results provide novel insight into NIR-mediated p53 suppression and also suggest an additional way for p53 regulation.

  12. Structure and inhibitor specificity of the PCTAIRE-family kinase CDK16.

    Science.gov (United States)

    Dixon-Clarke, Sarah E; Shehata, Saifeldin N; Krojer, Tobias; Sharpe, Timothy D; von Delft, Frank; Sakamoto, Kei; Bullock, Alex N

    2017-02-20

    CDK16 (also known as PCTAIRE1 or PCTK1) is an atypical member of the cyclin-dependent kinase (CDK) family that has emerged as a key regulator of neurite outgrowth, vesicle trafficking and cancer cell proliferation. CDK16 is activated through binding to cyclin Y via a phosphorylation-dependent 14-3-3 interaction and has a unique consensus substrate phosphorylation motif compared with conventional CDKs. To elucidate the structure and inhibitor-binding properties of this atypical CDK, we screened the CDK16 kinase domain against different inhibitor libraries and determined the co-structures of identified hits. We discovered that the ATP-binding pocket of CDK16 can accommodate both type I and type II kinase inhibitors. The most potent CDK16 inhibitors revealed by cell-free and cell-based assays were the multitargeted cancer drugs dabrafenib and rebastinib. An inactive DFG-out binding conformation was confirmed by the first crystal structures of CDK16 in separate complexes with the inhibitors indirubin E804 and rebastinib, respectively. The structures revealed considerable conformational plasticity, suggesting that the isolated CDK16 kinase domain was relatively unstable in the absence of a cyclin partner. The unusual structural features and chemical scaffolds identified here hold promise for the development of more selective CDK16 inhibitors and provide opportunity to better characterise the role of CDK16 and its related CDK family members in various physiological and pathological contexts.

  13. Structure and inhibitor specificity of the PCTAIRE-family kinase CDK16

    Science.gov (United States)

    Dixon-Clarke, Sarah E.; Shehata, Saifeldin N.; Krojer, Tobias; Sharpe, Timothy D.; vonDelft, Frank; Sakamoto, Kei

    2017-01-01

    CDK16 (also known as PCTAIRE1 or PCTK1) is an atypical member of the cyclin-dependent kinase (CDK) family that has emerged as a key regulator of neurite outgrowth, vesicle trafficking and cancer cell proliferation. CDK16 is activated through binding to cyclin Y via a phosphorylation-dependent 14-3-3 interaction and has a unique consensus substrate phosphorylation motif compared with conventional CDKs. To elucidate the structure and inhibitor-binding properties of this atypical CDK, we screened the CDK16 kinase domain against different inhibitor libraries and determined the co-structures of identified hits. We discovered that the ATP-binding pocket of CDK16 can accommodate both type I and type II kinase inhibitors. The most potent CDK16 inhibitors revealed by cell-free and cell-based assays were the multitargeted cancer drugs dabrafenib and rebastinib. An inactive DFG-out binding conformation was confirmed by the first crystal structures of CDK16 in separate complexes with the inhibitors indirubin E804 and rebastinib, respectively. The structures revealed considerable conformational plasticity, suggesting that the isolated CDK16 kinase domain was relatively unstable in the absence of a cyclin partner. The unusual structural features and chemical scaffolds identified here hold promise for the development of more selective CDK16 inhibitors and provide opportunity to better characterise the role of CDK16 and its related CDK family members in various physiological and pathological contexts. PMID:28057719

  14. Control of centrin stability by Aurora A.

    Directory of Open Access Journals (Sweden)

    Kara B Lukasiewicz

    Full Text Available Aurora A is an oncogenic serine/threonine kinase which can cause cell transformation and centrosome amplification when over-expressed. Human breast tumors show excess Aurora A and phospho-centrin in amplified centrosomes. Here, we show that Aurora A mediates the phosphorylation of and localizes with centrin at the centrosome, with both proteins reaching maximum abundance from prophase through metaphase, followed by their precipitous loss in late stages of mitosis. Over-expression of Aurora A results in excess phospho-centrin and centrosome amplification. In contrast, centrosome amplification is not seen in cells over-expressing Aurora A in the presence of a recombinant centrin mutant lacking the serine phosphorylation site at residue 170. Expression of a kinase dead Aurora A results in a decrease in mitotic index and abrogation of centrin phosphorylation. Finally, a recombinant centrin mutation that mimics centrin phosphorylation increases centrin's stability against APC/C-mediated proteasomal degradation. Taken together, these results suggest that the stability of centrin is regulated in part by Aurora A, and that excess phosphorylated centrin may promote centrosome amplification in cancer.

  15. Recent advances in protein kinase inhibition: current molecular scaffolds used for inhibitor synthesis.

    Science.gov (United States)

    Stover, D R; Lydon, N B; Nunes, J J

    1999-07-01

    Early efforts to discover and develop protein kinase inhibitors have focused largely on a small group of oncology targets such as the EGFR and PKC enzymes. More recently, hundreds of protein kinases have been identified at the genetic level, many of which are now being assigned functions in a variety of signaling pathways. Additionally, mutagenesis and X-ray crystallographic studies have further defined common structural features associated with binding of the ATP cofactor within a conserved ATP binding cleft. These studies have also demonstrated significant differences in the ATP binding cleft between individual kinases, providing a molecular basis for understanding and exploiting inhibitor specificity. The current review will focus on recent developments in the field of ATP site-directed inhibitors with particular emphasis on the major scaffolds being derivatized to take advantage of variable regions of the active site.

  16. Science Signaling Podcast for 15 November 2016: A new type of kinase inhibitor.

    Science.gov (United States)

    Eldar-Finkelman, Hagit; VanHook, Annalisa M

    2016-11-15

    This Podcast features an interview with Hagit Eldar-Finkelman, author of a Research Article that appears in the 15 November 2016 issue of Science Signaling, about a newly developed inhibitor of glycogen synthase kinase 3 (GSK-3). GSK-3 participates in several signaling networks and has been implicated in various pathologies, including neurodegenerative diseases, cognitive impairments, and cancer. Licht-Murava et al developed L807mts, a substrate-competitive peptide inhibitor that blocks GSK-3 activity through an unusual mechanism. L807mts not only bound to the substrate recognition domain of GSK-3, it was also phosphorylated by the kinase. This phosphorylated form of L807mts remained associated with GSK-3 and inhibited GSK-3 activity. L807mts treatment reduced cellular, cognitive, and behavioral symptoms in a mouse model of Alzheimer's disease. L807mts is an advance in kinase inhibitor development because it is both highly specific and very potent.Listen to Podcast.

  17. Lichen planopilaris-like eruption during treatment with tyrosine kinase inhibitor nilotinib*

    Science.gov (United States)

    Leitão, Juliana Ribeiro; Valente, Neusa Yuriko Sakai; Kakizaki, Priscila; Veronez, Isis Suga; Pires, Mario Cezar

    2016-01-01

    Tyrosine kinase inhibitors are effective as a target therapy for malignant neoplasms. Imatinib was the first tyrosine kinase inhibitor used. After its introduction, several other drugs have appeared with a similar mechanism of action, but less prone to causing resistance. Even though these drugs are selective, their toxicity does not exclusively target cancer cells, and skin toxicity is the most common non-hematologic adverse effect. We report an eruption similar to lichen planopilaris that developed during therapy with nilotinib, a second generation tyrosine kinase inhibitor, in a patient with chronic myeloid leukemia resistant to imatinib. In a literature review, we found only one report of non-scarring alopecia due to the use of nilotinib.

  18. Pyrazolo[3,4-b]quinoxalines. A new class of cyclin-dependent kinases inhibitors.

    Science.gov (United States)

    Ortega, Miguel A; Montoya, María E; Zarranz, Belén; Jaso, Andrés; Aldana, Ignacio; Leclerc, Sophie; Meijer, Laurent; Monge, Antonio

    2002-07-01

    Protein kinases are involved in most physiological processes and in numerous diseases. Therefore, inhibitors of protein kinases have therefore a wide therapeutic potential. While screening for inhibitors of cyclin-dependent kinases (CDK's) and glycogen synthase kinase-3 (GSK-3), we identified pyrazolo[3,4-b]quinoxalines as sub-micromolar inhibitors of CDK1/cyclin B. A preliminary structure-activity relationship study suggests that this family of compounds can be optimized to inhibit CDK's and GSK-3. Compounds were tested for their anti-proliferative activity and the results show that several of them displayed a significant inhibitory effect on CDK1/cyclin B. The most active compound (1) was also tested against the brain kinases CDK5/p25 and GSK-3, and proved to be a good inhibitor of both of them. On the contrary, none of the compounds showed any activity in the CDC25 phosphatase assay. As an additional approach, affinity chromatography on immobilized pyrazolo[3,4-b]quinoxalines will be used to identify the intracellular targets of this family of compounds.

  19. Structural Bioinformatics-Based Prediction of Exceptional Selectivity of p38 MAP Kinase Inhibitor PH-797804

    Energy Technology Data Exchange (ETDEWEB)

    Xing, Li; Shieh, Huey S.; Selness, Shaun R.; Devraj, Rajesh V.; Walker, John K.; Devadas, Balekudru; Hope, Heidi R.; Compton, Robert P.; Schindler, John F.; Hirsch, Jeffrey L.; Benson, Alan G.; Kurumbail, Ravi G.; Stegeman, Roderick A.; Williams, Jennifer M.; Broadus, Richard M.; Walden, Zara; Monahan, Joseph B.; Pfizer

    2009-07-24

    PH-797804 is a diarylpyridinone inhibitor of p38{alpha} mitogen-activated protein (MAP) kinase derived from a racemic mixture as the more potent atropisomer (aS), first proposed by molecular modeling and subsequently confirmed by experiments. On the basis of structural comparison with a different biaryl pyrazole template and supported by dozens of high-resolution crystal structures of p38{alpha} inhibitor complexes, PH-797804 is predicted to possess a high level of specificity across the broad human kinase genome. We used a structural bioinformatics approach to identify two selectivity elements encoded by the TXXXG sequence motif on the p38{alpha} kinase hinge: (i) Thr106 that serves as the gatekeeper to the buried hydrophobic pocket occupied by 2,4-difluorophenyl of PH-797804 and (ii) the bidentate hydrogen bonds formed by the pyridinone moiety with the kinase hinge requiring an induced 180{sup o} rotation of the Met109-Gly110 peptide bond. The peptide flip occurs in p38{alpha} kinase due to the critical glycine residue marked by its conformational flexibility. Kinome-wide sequence mining revealed rare presentation of the selectivity motif. Corroboratively, PH-797804 exhibited exceptionally high specificity against MAP kinases and the related kinases. No cross-reactivity was observed in large panels of kinase screens (selectivity ratio of >500-fold). In cellular assays, PH-797804 demonstrated superior potency and selectivity consistent with the biochemical measurements. PH-797804 has met safety criteria in human phase I studies and is under clinical development for several inflammatory conditions. Understanding the rationale for selectivity at the molecular level helps elucidate the biological function and design of specific p38{alpha} kinase inhibitors.

  20. Unique Cyclin-Dependent Kinase (CDK) Inhibitors at the ATP-site

    Institute of Scientific and Technical Information of China (English)

    LI Lin; LUNDGREN Karen; ESCOBAR Jorge; MINNICK Sharon price; HUBER Andrea; KOUDRIAKOVA Tatiana; ARRUDA Jeannie; SISSON Wes; AUST Robert M.; VERKHIVKER Gennady M.; SCHAFFER Lana; CHONG Wesley K. M.; ROSE Peter w.; LEWIS Cristrina T; DUVADIE Rohit K.; CHU Shao Song; YANG Y. Michelle; NONOMIYA Jim; TUCKER Kadthleen D.; KNIGHTON Daniel R.; FERRE RoseAnn

    2001-01-01

    @@ Control of the cell cycle could be applicable in new approaches for cancer chemotherapy. The cyclin-dependent kinases (CDK's) and their corresponding complexes with cyclins are regulatory enzymes for which we have discovered a novel small molecule series of inhibitors, with potencies in the nanomolar range and good selectivity for the CDK's versus other kinases. We will discuss structure-based drug design efforts with crystal structures of complexes with certain CDK's. Cellular effects and some preliminary examination of in vivo cancer efficacy by these inhibitors will also be discussed.

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

    Directory of Open Access Journals (Sweden)

    Havelange V

    2013-08-01

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

  2. Application of multiplexed kinase inhibitor beads to study kinome adaptations in drug-resistant leukemia.

    Directory of Open Access Journals (Sweden)

    Matthew J Cooper

    Full Text Available Protein kinases play key roles in oncogenic signaling and are a major focus in the development of targeted cancer therapies. Imatinib, a BCR-Abl tyrosine kinase inhibitor, is a successful front-line treatment for chronic myelogenous leukemia (CML. However, resistance to imatinib may be acquired by BCR-Abl mutations or hyperactivation of Src family kinases such as Lyn. We have used multiplexed kinase inhibitor beads (MIBs and quantitative mass spectrometry (MS to compare kinase expression and activity in an imatinib-resistant (MYL-R and -sensitive (MYL cell model of CML. Using MIB/MS, expression and activity changes of over 150 kinases were quantitatively measured from various protein kinase families. Statistical analysis of experimental replicates assigned significance to 35 of these kinases, referred to as the MYL-R kinome profile. MIB/MS and immunoblotting confirmed the over-expression and activation of Lyn in MYL-R cells and identified additional kinases with increased (MEK, ERK, IKKα, PKCβ, NEK9 or decreased (Abl, Kit, JNK, ATM, Yes abundance or activity. Inhibiting Lyn with dasatinib or by shRNA-mediated knockdown reduced the phosphorylation of MEK and IKKα. Because MYL-R cells showed elevated NF-κB signaling relative to MYL cells, as demonstrated by increased IκBα and IL-6 mRNA expression, we tested the effects of an IKK inhibitor (BAY 65-1942. MIB/MS and immunoblotting revealed that BAY 65-1942 increased MEK/ERK signaling and that this increase was prevented by co-treatment with a MEK inhibitor (AZD6244. Furthermore, the combined inhibition of MEK and IKKα resulted in reduced IL-6 mRNA expression, synergistic loss of cell viability and increased apoptosis. Thus, MIB/MS analysis identified MEK and IKKα as important downstream targets of Lyn, suggesting that co-targeting these kinases may provide a unique strategy to inhibit Lyn-dependent imatinib-resistant CML. These results demonstrate the utility of MIB/MS as a tool to identify

  3. Radiolabeled Small Molecule Protein Kinase Inhibitors for Imaging with PET or SPECT

    Directory of Open Access Journals (Sweden)

    Justin W. Hicks

    2010-11-01

    Full Text Available Imaging protein kinase expression with radiolabeled small molecule inhibitors has been actively pursued to monitor the clinical potential of targeted therapeutics and treatments as well as to determine kinase receptor density changes related to disease progression. The goal of the present review is to provide an overview of the breadth of radiolabeled small molecules that have been synthesized to target intracellular protein kinases, not only for imaging in oncology, but also for other areas of interest, particularly the central nervous system.  Considerable radiotracer development has focused on imaging receptor tyrosine kinases of growth factors, protein kinases A, B and C, and glycogen synthase kinase–3β. Design considerations, structural attributes and relevant biological results are summarized.

  4. Tyrosine Kinase Inhibitors and Vascular Toxicity: Impetus for a Classification System?

    Science.gov (United States)

    Herrmann, Joerg

    2016-06-01

    The introduction of molecularly targeted therapies with tyrosine kinase inhibitors has revolutionized cancer therapy and has contributed to a steady decline in cancer-related mortality since the late 1990s. However, not only cardiac but also vascular toxicity has been reported for these agents, some as expected on-target effects (e.g., VEGF receptor inhibitors) and others as unanticipated events (e.g., BCR-Abl inhibitors). A sound understanding of these cardiovascular toxic effects is critical to advance mechanistic insight into vascular disease and clinical care. From a conceptual standpoint, there might be value in defining type I (permanent) and type II (transient) vascular toxicity. This review will focus on the tyrosine kinase inhibitors in current clinical use and their associated vascular side effects.

  5. Selective Inhibitors of Cyclin G Associated Kinase (GAK) as Anti-Hepatitis C Agents.

    Science.gov (United States)

    Kovackova, Sona; Chang, Lei; Bekerman, Elena; Neveu, Gregory; Barouch-Bentov, Rina; Chaikuad, Apirat; Heroven, Christina; Šála, Michal; De Jonghe, Steven; Knapp, Stefan; Einav, Shirit; Herdewijn, Piet

    2015-04-23

    Cyclin G associated kinase (GAK) emerged as a promising drug target for the treatment of viral infections. However, no potent and selective GAK inhibitors have been reported in the literature to date. This paper describes the discovery of isothiazolo[5,4-b]pyridines as selective GAK inhibitors, with the most potent congeners displaying low nanomolar binding affinity for GAK. Cocrystallization experiments revealed that these compounds behaved as classic type I ATP-competitive kinase inhibitors. In addition, we have demonstrated that these compounds exhibit a potent activity against hepatitis C virus (HCV) by inhibiting two temporally distinct steps in the HCV life cycle (i.e., viral entry and assembly). Hence, these GAK inhibitors represent chemical probes to study GAK function in different disease areas where GAK has been implicated (including viral infection, cancer, and Parkinson's disease).

  6. Methods Of Using Chemical Libraries To Search For New Kinase Inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Gray, Nathanael S. (Berkeley, CA), Schultz, Peter (Oakland, CA), Wodicka, Lisa (Santa Clara, CA), Meijer, Laurent (Roscoff, FR), Lockhart, David J. (Mountain View, CA)

    2003-06-03

    The generation of selective inhibitors for specific protein kinases would provide new tools for analyzing signal transduction pathways and possibly new therapeutic agents. We have invented an approach to the development of selective protein kinase inhibitors based on the unexpected binding mode of 2,6,9-trisubstituted purines to the ATP binding site of human CDK2. The most potent inhibitor, purvalanol B (IC.sub.50 =6 nM), binds with a 30-fold greater affinity than the known CDK2 inhibitor, flavopiridol. The cellular effects of this class of compounds were examined and compared to those of flavopiridol by monitoring changes in mRNA expression levels for all genes in treated cells of Saccharomyces cerevisiae using high-density oligonucleotide probe arrays.

  7. Clinical development of phosphatidylinositol-3 kinase pathway inhibitors.

    Science.gov (United States)

    Arteaga, Carlos L

    2010-01-01

    The PI3K pathway is the most commonly altered in human cancer. Several recent phase I studies with therapeutic inhibitors of this pathway have shown that pharmacological inhibition of PI3K in humans is feasible and overall well tolerated. Furthermore, there has already been clinical evidence of anti-tumor activity in patients with advanced cancer. The intensity and duration of PI3K inhibition required for an antitumor effect and the optimal pharmacodynamic biomarker(s) of pathway inactivation remain to be established. Preclinical and early clinical data support focusing on trials with PI3K inhibitors that are at a minimum enriched with patients with alterations in this signaling pathway. These inhibitors are likely to be more effective in combination with established and other novel molecular therapies.

  8. Structural Mechanism of the Pan-BCR-ABL Inhibitor Ponatinib (AP24534): Lessons for Overcoming Kinase Inhibitor Resistance

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Tianjun; Commodore, Lois; Huang, Wei-Sheng; Wang, Yihan; Thomas, Mathew; Keats, Jeff; Xu, Qihong; Rivera, Victor M.; Shakespeare, William C.; Clackson, Tim; Dalgarno, David C.; Zhu, Xiaotian (ARIAD)

    2012-01-20

    The BCR-ABL inhibitor imatinib has revolutionized the treatment of chronic myeloid leukemia. However, drug resistance caused by kinase domain mutations has necessitated the development of new mutation-resistant inhibitors, most recently against the T315I gatekeeper residue mutation. Ponatinib (AP24534) inhibits both native and mutant BCR-ABL, including T315I, acting as a pan-BCR-ABL inhibitor. Here, we undertook a combined crystallographic and structure-activity relationship analysis on ponatinib to understand this unique profile. While the ethynyl linker is a key inhibitor functionality that interacts with the gatekeeper, virtually all other components of ponatinib play an essential role in its T315I inhibitory activity. The extensive network of optimized molecular contacts found in the DFG-out binding mode leads to high potency and renders binding less susceptible to disruption by single point mutations. The inhibitory mechanism exemplified by ponatinib may have broad relevance to designing inhibitors against other kinases with mutated gatekeeper residues.

  9. Virtual Screening for Potential Allosteric Inhibitors of Cyclin-Dependent Kinase 2 from Traditional Chinese Medicine

    Directory of Open Access Journals (Sweden)

    Fang Lu

    2016-09-01

    Full Text Available Cyclin-dependent kinase 2 (CDK2, a member of Cyclin-dependent kinases (CDKs, plays an important role in cell division and DNA replication. It is regarded as a desired target to treat cancer and tumor by interrupting aberrant cell proliferation. Compared to lower subtype selectivity of CDK2 ATP-competitive inhibitors, CDK2 allosteric inhibitor with higher subtype selectivity has been used to treat CDK2-related diseases. Recently, the first crystal structure of CDK2 with allosteric inhibitor has been reported, which provides new opportunities to design pure allosteric inhibitors of CDK2. The binding site of the ATP-competition inhibitors and the allosteric inhibitors are partially overlapped in space position, so the same compound might interact with the two binding sites. Thus a novel screening strategy was essential for the discovery of pure CDK2 allosteric inhibitors. In this study, pharmacophore and molecular docking were used to screen potential CDK2 allosteric inhibitors and ATP-competition inhibitors from Traditional Chinese Medicine (TCM. In the docking result of the allosteric site, the compounds which can act with the CDK2 ATP site were discarded, and the remaining compounds were regarded as the potential pure allosteric inhibitors. Among the results, prostaglandin E1 and nordihydroguaiaretic acid (NDGA were available and their growth inhibitory effect on human HepG2 cell lines was determined by MTT assay. The two compounds could substantially inhibit the growth of HepG2 cell lines with an estimated IC50 of 41.223 μmol/L and 45.646 μmol/L. This study provides virtual screening strategy of allosteric compounds and a reliable method to discover potential pure CDK2 allosteric inhibitors from TCM. Prostaglandin E1 and NDGA could be regarded as promising candidates for CDK2 allosteric inhibitors.

  10. [Effect of inhibitors serine/threonine protein kinases and protein phosphatases on mitosis progression of synchronized tobacco by-2 cells].

    Science.gov (United States)

    Sheremet, Ia A; Emets, A I; Azmi, A; Vissenberg, K; Verbelen, J-P; Blium, Ia B

    2012-01-01

    In order to investigate the role of various serine/ threonine protein kinases and protein phosphatases in the regulation of mitosis progression in plant cells the influence of cyclin-dependent (olomoucine) and Ca2+ -calmodulin-dependent (W7) protein kinases inhibitors, as well as protein kinase C inhibitors (H7 and staurosporine) and protein phosphatases inhibitor (okadaic acid) on mitosis progression in synchronized tobacco BY-2 cells has been studied. It was found that BY-2 culture treatment with inhibitors of cyclin dependent protein kinases and protein kinase C causes prophase delay, reduces the mitotic index and displaces of mitotic peak as compare with control cells. Inhibition of Ca2+ -calmodulin dependent protein kinases enhances the cell entry into prophase and delays their exit from mitosis. Meanwhile inhibition of serine/threonine protein phosphatases insignificantly enhances of synchronized BY-2 cells entering into all phases of mitosis.

  11. Protein kinase inhibitors CK59 and CID755673 alter primary human NK cell effector functions

    Directory of Open Access Journals (Sweden)

    Maxi eScheiter

    2013-03-01

    Full Text Available Natural killer (NK cells are part of the innate immune response and play a crucial role in the defense against tumors and virus-infected cells. Their effector functions include the specific killing of target cells, as well as the modulation of other immune cells by cytokine release. Kinases constitute a relevant part in signaling, are prime targets in drug research and the protein kinase inhibitor Dasatinib is already used for immune-modulatory theraphies. In this study, we have tested the effects of the kinase inhibitors CK59 and CID755673. These inhibitors are directed against CaMKII (CK59 and PKD family kinases (CID755673 that were previously suggested as novel components of NK activation pathways. Here, we use a multi-parameter, FACS-based assay to validate the influence of CK59 and CID755673 on the effector functions of primary NK cells. Dose dependent treatment with CK59 and CID755673 indeed results in a significant reduction of NK cell degranulation markers and cytokine release in freshly isolated PBMC populations from healthy blood donors. These results underline the importance of CaMKII for NK cell signaling and suggest PKD2 as a novel signaling component in NK cell activation. Notably, kinase inhibition studies on pure NK cell populations indicate significant donor variations.

  12. Receptor-Based Virtual Screening of EGFR Kinase Inhibitors from the NCI Diversity Database

    Directory of Open Access Journals (Sweden)

    Kiattawee Choowongkomon

    2010-06-01

    Full Text Available Epidermal growth factor receptor (EGFR abnormalities have been associated with several types of human cancer. The crystal structures of its tyrosine kinase domain (EGFR-TK complexed with small molecule inhibitors revealed the kinase inhibition modes, prompting us to search for novel anti-cancer drugs. A total of 1,990 compounds from the National Cancer Institute (NCI diversity set with nonredundant structures have been tested to inhibit cancer cell lines with unknown mechanism. Cancer inhibition through EGFR-TK is one of the mechanisms of these compounds. In this work, we performed receptor-based virtual screening against the NCI diversity database. Using two different docking algorithms, AutoDock and Gold, combined with subsequent post-docking analyses, we found eight candidate compounds with high scoring functions that all bind to the ATP-competitive site of the kinase. None of these compounds belongs to the main group of the currently known EGFR-TK inhibitors. Binding mode analyses revealed that the way these compounds complexed with EGFR-TK differs from quinazoline inhibitor binding and the interaction mainly involves hydrophobic interactions. Also, the common kinase-inhibitor (NH---N and CO---HC hydrogen bonds between the hinge region and the hit compounds are rarely observed. Our results suggest that these molecules could be developed as novel lead compounds in anti-cancer drug design.

  13. Metabolite profiling of the multiple tyrosine kinase inhibitor lenvatinib : a cross-species comparison

    NARCIS (Netherlands)

    Dubbelman, Anne Charlotte; Nijenhuis, Cynthia M.; Jansen, Robert S; Rosing, Hilde; Mizuo, Hitoshi; Kawaguchi, Shinki; Critchley, David; Shumaker, Robert; Schellens, Jan H M; Beijnen, Jos H.

    2016-01-01

    Lenvatinib is an oral, multiple receptor tyrosine kinase inhibitor. Preclinical drug metabolism studies showed unique metabolic pathways for lenvatinib in monkeys and rats. A human mass balance study demonstrated that lenvatinib related material is mainly excreted via feces with a small fraction as

  14. Crystal structure of the PIM2 kinase in complex with an organoruthenium inhibitor.

    Directory of Open Access Journals (Sweden)

    Alex N Bullock

    Full Text Available BACKGROUND: The serine/threonine kinase PIM2 is highly expressed in human leukemia and lymphomas and has been shown to positively regulate survival and proliferation of tumor cells. Its diverse ATP site makes PIM2 a promising target for the development of anticancer agents. To date our knowledge of catalytic domain structures of the PIM kinase family is limited to PIM1 which has been extensively studied and which shares about 50% sequence identity with PIM2. PRINCIPAL FINDINGS: Here we determined the crystal structure of PIM2 in complex with an organoruthenium complex (inhibition in sub-nanomolar level. Due to its extraordinary shape complementarity this stable organometallic compound is a highly potent inhibitor of PIM kinases. SIGNIFICANCE: The structure of PIM2 revealed several differences to PIM1 which may be explored further to generate isoform selective inhibitors. It has also demonstrated how an organometallic inhibitor can be adapted to the binding site of protein kinases to generate highly potent inhibitors. ENHANCED VERSION: This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.

  15. COMPARATIVE PATHOGENESIS OF HALOACETIC ACID AND PROTEIN KINASE INHIBITOR EMBRYOTOXICITY IN MOUSE WHOLE EMBRYO CULTURE

    Science.gov (United States)

    Comparative pathogenesis of haloacetic acid and protein kinase inhibitor embryotoxicity in mouse whole embryo culture.Ward KW, Rogers EH, Hunter ES 3rd.Curriculum in Toxicology, University of North Carolina at Chapel Hill, 27599-7270, USA.Haloacetic acids ...

  16. Synthesis of Benzofuran Analogue of Go6976, an Isoform Selective Protein Kinase C Inhibitor

    Institute of Scientific and Technical Information of China (English)

    MA, Da-Wei; ZHANG, Xin-Rong; WU, Shi-Hui; TAO, Feng-Gang

    2001-01-01

    Based on the structure of Go6976, a known isoform-selective protein kinase C inhibitor, a benzofuran analogue (1) was designed. This analogue was synthesized by coupling of benzofuran 3-acetic acid and 8-oxo-tryptamine and subsequent intramolecular Dieckmann condensation, alkylation, oxidative photocyclization and cyanation reaction of mesylate.

  17. Tyrosine kinase inhibitors target cancer stem cells in renal cell cancer.

    Science.gov (United States)

    Czarnecka, Anna M; Solarek, Wojciech; Kornakiewicz, Anna; Szczylik, Cezary

    2016-03-01

    This study was designed to analyze the impact of multi-targeted tyrosine kinase inhibitors on the cancer stem cell subpopulation in renal cell cancer. The second objective was to evaluate the effect of tumor growth inhibition related to a tumor niche factor - oxygen deprivation - as hypoxia develops along with the anti-angiogenic activity of tyrosine kinase inhibitors in renal tumors. Cells were treated with tyrosine kinase inhibitors, sunitinib, sorafenib and axitinib, in 2D and 3D culture conditions. Cell proliferation along with drug toxicity were evaluated. It was shown that the proliferation rate of cancer stem cells was decreased by the tyrosine kinase inhibitors. The efficacy of the growth inhibition was limited by hypoxic conditions and 3D intratumoral cell-cell interactions. We conclude that understanding the complex molecular interaction feedback loops between differentiated cancer cells, cancer stem cells and the tumor microenvironment in 3D culture should aid the identification of novel treatment targets and to evalute the efficacy of renal cancer therapies. Cell-cell interaction may represent a critical microenvironmental factor regulating cancer stem cell self-renewal potential, enhancing the stem cell phenotype and limiting drug toxicity. At the same time the role of hypoxia in renal cancer stem cell biology is also significant.

  18. Discovery of a Selective Inhibitor of Oncogenic B-Raf Kinase With Potent Antimelanoma Activity

    Energy Technology Data Exchange (ETDEWEB)

    Tsai, J.; Lee, J.T.; Wang, W.; Zhang, J.; Cho, H.; Mamo, S.; Bremer, R.; Gillette, S.; Kong, J.; Haass, N.K.; Sproesser, K.; Li, L.; Smalley, K.S.M.; Fong, D.; Zhu, Y.-L.; Marimuthu, A.; Nguyen, H.; Lam, B.; Liu, J.; Cheung, I.; Rice, J.

    2009-05-26

    BRAF{sup V600E} is the most frequent oncogenic protein kinase mutation known. Furthermore, inhibitors targeting 'active' protein kinases have demonstrated significant utility in the therapeutic repertoire against cancer. Therefore, we pursued the development of specific kinase inhibitors targeting B-Raf, and the V600E allele in particular. By using a structure-guided discovery approach, a potent and selective inhibitor of active B-Raf has been discovered. PLX4720, a 7-azaindole derivative that inhibits B-Raf{sup V600E} with an IC{sub 50} of 13 nM, defines a class of kinase inhibitor with marked selectivity in both biochemical and cellular assays. PLX4720 preferentially inhibits the active B-Raf{sup V600E} kinase compared with a broad spectrum of other kinases, and potent cytotoxic effects are also exclusive to cells bearing the V600E allele. Consistent with the high degree of selectivity, ERK phosphorylation is potently inhibited by PLX4720 in B-Raf{sup V600E}-bearing tumor cell lines but not in cells lacking oncogenic B-Raf. In melanoma models, PLX4720 induces cell cycle arrest and apoptosis exclusively in B-Raf{sup V600E}-positive cells. In B-Raf{sup V600E}-dependent tumor xenograft models, orally dosed PLX4720 causes significant tumor growth delays, including tumor regressions, without evidence of toxicity. The work described here represents the entire discovery process, from initial identification through structural and biological studies in animal models to a promising therapeutic for testing in cancer patients bearing B-Raf{sup V600E}-driven tumors.

  19. QSAR, molecular docking studies of thiophene and imidazopyridine derivatives as polo-like kinase 1 inhibitors

    Science.gov (United States)

    Cao, Shandong

    2012-08-01

    The purpose of the present study was to develop in silico models allowing for a reliable prediction of polo-like kinase inhibitors based on a large diverse dataset of 136 compounds. As an effective method, quantitative structure activity relationship (QSAR) was applied using the comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA). The proposed QSAR models showed reasonable predictivity of thiophene analogs (Rcv2=0.533, Rpred2=0.845) and included four molecular descriptors, namely IC3, RDF075m, Mor02m and R4e+. The optimal model for imidazopyridine derivatives (Rcv2=0.776, Rpred2=0.876) was shown to perform good in prediction accuracy, using GATS2m and BEHe1 descriptors. Analysis of the contour maps helped to identify structural requirements for the inhibitors and served as a basis for the design of the next generation of the inhibitor analogues. Docking studies were also employed to position the inhibitors into the polo-like kinase active site to determine the most probable binding mode. These studies may help to understand the factors influencing the binding affinity of chemicals and to develop alternative methods for prescreening and designing of polo-like kinase inhibitors.

  20. A targeted library screen reveals a new inhibitor scaffold for protein kinase D.

    Directory of Open Access Journals (Sweden)

    Manuj Tandon

    Full Text Available Protein kinase D (PKD has emerged as a potential therapeutic target in multiple pathological conditions, including cancer and heart diseases. Potent and selective small molecule inhibitors of PKD are valuable for dissecting PKD-mediated cellular signaling pathways and for therapeutic application. In this study, we evaluated a targeted library of 235 small organic kinase inhibitors for PKD1 inhibitory activity at a single concentration. Twenty-eight PKD inhibitory chemotypes were identified and six exhibited excellent PKD1 selectivity. Five of the six lead structures share a common scaffold, with compound 139 being the most potent and selective for PKD vs PKC and CAMK. Compound 139 was an ATP-competitive PKD1 inhibitor with a low double-digit nanomolar potency and was also cell-active. Kinase profiling analysis identified this class of small molecules as pan-PKD inhibitors, confirmed their selectivity again PKC and CAMK, and demonstrated an overall favorable selectivity profile that could be further enhanced through structural modification. Furthermore, using a PKD homology model based on similar protein kinase structures, docking modes for compound 139 were explored and compared to literature examples of PKD inhibition. Modeling of these compounds at the ATP-binding site of PKD was used to rationalize its high potency and provide the foundation for future further optimization. Accordingly, using biochemical screening of a small number of privileged scaffolds and computational modeling, we have identified a new core structure for highly potent PKD inhibition with promising selectivity against closely related kinases. These lead structures represent an excellent starting point for the further optimization and the design of selective and therapeutically effective small molecule inhibitors of PKD.

  1. Key Structures and Interactions for Binding of Mycobacterium tuberculosis Protein Kinase B Inhibitors from Molecular Dynamics Simulation.

    Science.gov (United States)

    Punkvang, Auradee; Kamsri, Pharit; Saparpakorn, Patchreenart; Hannongbua, Supa; Wolschann, Peter; Irle, Stephan; Pungpo, Pornpan

    2015-07-01

    Substituted aminopyrimidine inhibitors have recently been introduced as antituberculosis agents. These inhibitors show impressive activity against protein kinase B, a Ser/Thr protein kinase that is essential for cell growth of M. tuberculosis. However, up to now, X-ray structures of the protein kinase B enzyme complexes with the substituted aminopyrimidine inhibitors are currently unavailable. Consequently, structural details of their binding modes are questionable, prohibiting the structural-based design of more potent protein kinase B inhibitors in the future. Here, molecular dynamics simulations, in conjunction with molecular mechanics/Poisson-Boltzmann surface area binding free-energy analysis, were employed to gain insight into the complex structures of the protein kinase B inhibitors and their binding energetics. The complex structures obtained by the molecular dynamics simulations show binding free energies in good agreement with experiment. The detailed analysis of molecular dynamics results shows that Glu93, Val95, and Leu17 are key residues responsible to the binding of the protein kinase B inhibitors. The aminopyrazole group and the pyrimidine core are the crucial moieties of substituted aminopyrimidine inhibitors for interaction with the key residues. Our results provide a structural concept that can be used as a guide for the future design of protein kinase B inhibitors with highly increased antagonistic activity.

  2. San Huang Decoction Promotes Apoptosis of Breast Cancer Cells Through Regulating Aurora Kinase A%三黄煎剂调节Aurora激酶A促进乳腺癌细胞凋亡的实验研究

    Institute of Scientific and Technical Information of China (English)

    许岩磊; 陈曦琰; 陈绪; 卞卫和; 姚昶; 朱小纾; 陈佳静

    2015-01-01

    目的:探讨三黄煎剂对乳腺癌 MCF?7、MDA?MB?231细胞凋亡的影响及 Aurora 激酶 A(Aurora A)蛋白表达及功能的影响,并探讨其内在机制。方法采用 CCK?8法检测三黄煎剂对乳腺癌细胞 MCF?7、MDA?MB?231增殖的影响。AnnexinV? FITC/PI 法检测 MCF?7、MDA?MB?231细胞凋亡率。q?PCR 法检测 MCF?7、MDA?MB?231细胞 Aurora A、p53的 mRNA 表达水平。Western Blot 法检测 MCF?7、MDA?MB?231细胞凋亡相关蛋白的表达及 Aurora A 蛋白的表达。结果三黄煎剂对MCF?7、MDA?MB?231细胞增殖抑制率呈浓度梯度依赖增长(P <0.05),给药48 h 疗效好于24 h(P <0.05),与给药72 h 无统计学差异(P >0.05)。三黄煎剂能够诱导 MCF?7、MDA?MB?231细胞凋亡,并上调 c?PARP、c?Caspase 3、Bax 蛋白的表达,下调 Bcl?2蛋白的表达,呈浓度梯度依赖。三黄煎剂能够下调 Aurora A 蛋白及 mRNA 的表达、上调 p53蛋白及 mRNA 的表达。结论三黄煎剂能够通过下调 Aurora A 蛋白及 mRNA 的表达,抑制 Aurora A 的生物活性,抑制 MCF?7、MDA?MB?231细胞增殖,诱导其凋亡。%ABSTRACTOBJECTIVE To explore the effect of San Huang decoction on the apoptosis of breast cancer cells and the effect on the mRNA and protein expression and function of Aurora kinase A and discuss the underlying mechanism of San Huang in-duced apoptosis.METHODS The inhibition of breast cancer cells proliferation was determined by CCK?8 assay.The apoptosis of breast cancer cells was detected by AnnexinV?FITC/PI Staining.The expression of mRNA of Aurora A was examined by q ?PCR analysis.The expression of apoptosis?related proteins and Aurora A were determined by Western Blot analysis.RE-SULTS San Huang decoction inhibited the proliferation of breast cancer cells in a does?dependent mannerP 0.05.San Huang decoction was also found to induce apoptosis in both MCF?7 and

  3. From the cyclooxygenase-2 inhibitor celecoxib to a novel class of 3-phosphoinositide-dependent protein kinase-1 inhibitors.

    Science.gov (United States)

    Zhu, Jiuxiang; Huang, Jui-Wen; Tseng, Ping-Hui; Yang, Ya-Ting; Fowble, Joseph; Shiau, Chung-Wai; Shaw, Yeng-Jeng; Kulp, Samuel K; Chen, Ching-Shih

    2004-06-15

    The blockade of Akt activation through the inhibition of 3-phosphoinositide-dependent kinase-1 (PDK-1) represents a major signaling mechanism whereby celecoxib mediates apoptosis. Celecoxib, however, is a weak PDK-1 inhibitor (IC(50), 48 microM), requiring at least 30 microM to exhibit discernable effects on the growth of tumor cells in vitro. Here, we report the structure-based optimization of celecoxib to develop PDK-1 inhibitors with greater potency in enzyme inhibition and growth inhibition. Kinetics of PDK-1 inhibition by celecoxib with respect to ATP suggest that celecoxib derivatives inhibit PDK-1 by competing with ATP for binding, a mechanism reminiscent to that of many kinase inhibitors. Structure-activity analysis together with molecular modeling was used to generate compounds that were tested for their potency in inhibiting PDK-1 kinase activity and in inducing apoptosis in PC-3 prostate cancer cells. Docking of potent compounds into the ATP-binding site of PDK-1 was performed for lead optimization, leading to two compounds, OSU-03012 and OSU-03013, with IC(50) values in PDK-1 inhibition and apoptosis induction in the low microM range. Exposure of PC-3 cells to these agents led to Akt dephosphorylation and inhibition of p70 S6 kinase activity. Moreover, overexpression of constitutively active forms of PDK-1 and Akt partially protected OSU-03012-induced apoptosis. Screening in a panel of 60 cell lines and more extensive testing in PC-3 cells indicated that the mean concentration for total growth inhibition was approximately 3 microM for both agents. Considering the conserved role of PDK-1/Akt signaling in promoting tumorigenesis, these celecoxib analogs are of translational relevance for cancer prevention and therapy.

  4. On-Chip Peptide Mass Spectrometry Imaging for Protein Kinase Inhibitor Screening.

    Science.gov (United States)

    Cho, Young-Lai; Kim, Young-Pil; Son, Jin Gyeong; Son, Miyoung; Lee, Tae Geol

    2017-01-03

    Protein kinases are enzymes that are important targets for drug discovery because of their involvement in regulating the essential cellular processes. For this reason, the changes in protein kinase activity induced by each drug candidate (the inhibitor in this case) need to be accurately determined. Here, an on-chip secondary ion mass spectrometry (SIMS) imaging technique of the peptides was developed for determining protein kinase activity and inhibitor screening without a matrix. In our method, cysteine-tethered peptides adsorbed onto a gold surface produced changes in the relative peak intensities of the phosphorylated and unphosphorylated substrate peptides, which were quantitatively dependent on protein kinase activity. Using mass spectrometry imaging of multiple compartments on the gold surface in the presence of a peptide substrate, we screened 13,727 inhibitors, of which seven were initially found to have inhibitor efficiencies that surpassed 50%. Of these, we were able to identify a new breakpoint cluster region-abelson (BCR-ABL)(T315I) kinase inhibitor, henceforth referred to as KR135861. KR135861 showed no cytotoxicity and was subsequently confirmed to be superior to imatinib, a commercial drug marketed as Gleevec. Moreover, KR135861 exhibited a greater inhibitory effect on the BCR-ABL(T315I) tyrosine kinase, with an IC50 value as low as 1.3 μM. In in vitro experiments, KR135861 reduced the viability of both Ba/F3 cells expressing wild-type BCR-ABL and BCR-ABL(T315I), in contrast to imatinib's inhibitory effects only on Ba/F3 cells expressing wild-type BCR-ABL. Due to the surface sensitivity and selectivity of SIMS imaging, it is anticipated that our approach will make it easier to validate the small modifications of a substrate in relation to enzyme activity as well as for drug discovery. This mass spectrometry imaging analysis enables efficient screening for protein kinase inhibitors, thus permitting high-throughput drug screening with high accuracy

  5. Development of a cell-based, high-throughput screening assay for ATM kinase inhibitors.

    Science.gov (United States)

    Guo, Kexiao; Shelat, Anang A; Guy, R Kiplin; Kastan, Michael B

    2014-04-01

    The ATM (ataxia-telangiectasia, mutated) protein kinase is a major regulator of cellular responses to DNA double-strand breaks (DSBs), DNA lesions that can be caused by ionizing irradiation (IR), oxidative damage, or exposure to certain chemical agents. In response to DSBs, the ATM kinase is activated and subsequently phosphorylates numerous downstream substrates, including p53, Chk2, BRCA1, and KAP1, which affect processes such as cell cycle progression and DNA repair. Numerous studies have demonstrated that loss of ATM function results in enhanced sensitivity to ionizing irradiation in clinically relevant dose ranges, suggesting that ATM kinase is an attractive therapeutic target for enhancing tumor cell kill with radiotherapy. Previously identified small-molecule ATM kinase inhibitors, such as CP466722 and Ku55933, were identified using in vitro kinase assays carried out with recombinant ATM kinase isolated from mammalian cells. Since it has not been feasible to express full-length recombinant ATM in bacterial or baculovirus systems, a robust in vitro screening tool has been lacking. We have developed a cell-based assay that is robust, straightforward, and sensitive. Using this high-throughput assay, we screened more than 7000 compounds and discovered additional small molecules that inhibit the ATM kinase and further validated these hits by secondary assays.

  6. Identification and Structure-Function Analysis of Subfamily Selective G Protein-Coupled Receptor Kinase Inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Homan, Kristoff T.; Larimore, Kelly M.; Elkins, Jonathan M.; Szklarz, Marta; Knapp, Stefan; Tesmer, John J.G. [Michigan; (Oxford)

    2015-02-13

    Selective inhibitors of individual subfamilies of G protein-coupled receptor kinases (GRKs) would serve as useful chemical probes as well as leads for therapeutic applications ranging from heart failure to Parkinson’s disease. To identify such inhibitors, differential scanning fluorimetry was used to screen a collection of known protein kinase inhibitors that could increase the melting points of the two most ubiquitously expressed GRKs: GRK2 and GRK5. Enzymatic assays on 14 of the most stabilizing hits revealed that three exhibit nanomolar potency of inhibition for individual GRKs, some of which exhibiting orders of magnitude selectivity. Most of the identified compounds can be clustered into two chemical classes: indazole/dihydropyrimidine-containing compounds that are selective for GRK2 and pyrrolopyrimidine-containing compounds that potently inhibit GRK1 and GRK5 but with more modest selectivity. The two most potent inhibitors representing each class, GSK180736A and GSK2163632A, were cocrystallized with GRK2 and GRK1, and their atomic structures were determined to 2.6 and 1.85 Å spacings, respectively. GSK180736A, developed as a Rho-associated, coiled-coil-containing protein kinase inhibitor, binds to GRK2 in a manner analogous to that of paroxetine, whereas GSK2163632A, developed as an insulin-like growth factor 1 receptor inhibitor, occupies a novel region of the GRK active site cleft that could likely be exploited to achieve more selectivity. However, neither compound inhibits GRKs more potently than their initial targets. This data provides the foundation for future efforts to rationally design even more potent and selective GRK inhibitors.

  7. Comparative VEGF receptor tyrosine kinase modeling for the development of highly specific inhibitors of tumor angiogenesis.

    Science.gov (United States)

    Schmidt, Ulrike; Ahmed, Jessica; Michalsky, Elke; Hoepfner, Michael; Preissner, Robert

    2008-01-01

    The Vascular Endothelial Growth Factor receptors (VEGF-Rs) play a significant role in tumor development and tumor angiogenesis and are therefore interesting targets in cancer therapy. Targeting the VEGF-R is of special importance as the feed of the tumor has to be reduced. In general, this can be carried out by inhibiting the tyrosine kinase function of the VEGF-R. Nevertheless, there arise some problems with the specificity of known kinase inhibitors: they bind to the ATP-binding site and inhibit a number of kinases, moreover the so far most specific inhibitors act at least on these three major types of VEGF-Rs: Flt-1, Flk-1/KDR, Flt-4. The goal is a selective VEGF-R-2 (Flk-1/KDR) inhibitor, because this receptor triggers rather unspecific signals from VEGF-A, -C, -D and -E. Here, we describe a protocol starting from an established inhibitor (Vatalanib) with 2D-/3D-searching and property filtering of the in silico screening hits and the "negative docking approach". With this approach we were able to identify a compound, which shows a fourfold higher reduction of the proliferation rate of endothelial cells compared to the reduction effect of the lead structure.

  8. A class of selective antibacterials derived from a protein kinase inhibitor pharmacophore.

    Science.gov (United States)

    Miller, J Richard; Dunham, Steve; Mochalkin, Igor; Banotai, Craig; Bowman, Matthew; Buist, Susan; Dunkle, Bill; Hanna, Debra; Harwood, H James; Huband, Michael D; Karnovsky, Alla; Kuhn, Michael; Limberakis, Chris; Liu, Jia Y; Mehrens, Shawn; Mueller, W Thomas; Narasimhan, Lakshmi; Ogden, Adam; Ohren, Jeff; Prasad, J V N Vara; Shelly, John A; Skerlos, Laura; Sulavik, Mark; Thomas, V Hayden; VanderRoest, Steve; Wang, LiAnn; Wang, Zhigang; Whitton, Amy; Zhu, Tong; Stover, C Kendall

    2009-02-10

    As the need for novel antibiotic classes to combat bacterial drug resistance increases, the paucity of leads resulting from target-based antibacterial screening of pharmaceutical compound libraries is of major concern. One explanation for this lack of success is that antibacterial screening efforts have not leveraged the eukaryotic bias resulting from more extensive chemistry efforts targeting eukaryotic gene families such as G protein-coupled receptors and protein kinases. Consistent with a focus on antibacterial target space resembling these eukaryotic targets, we used whole-cell screening to identify a series of antibacterial pyridopyrimidines derived from a protein kinase inhibitor pharmacophore. In bacteria, the pyridopyrimidines target the ATP-binding site of biotin carboxylase (BC), which catalyzes the first enzymatic step of fatty acid biosynthesis. These inhibitors are effective in vitro and in vivo against fastidious gram-negative pathogens including Haemophilus influenzae. Although the BC active site has architectural similarity to those of eukaryotic protein kinases, inhibitor binding to the BC ATP-binding site is distinct from the protein kinase-binding mode, such that the inhibitors are selective for bacterial BC. In summary, we have discovered a promising class of potent antibacterials with a previously undescribed mechanism of action. In consideration of the eukaryotic bias of pharmaceutical libraries, our findings also suggest that pursuit of a novel inhibitor leads for antibacterial targets with active-site structural similarity to known human targets will likely be more fruitful than the traditional focus on unique bacterial target space, particularly when structure-based and computational methodologies are applied to ensure bacterial selectivity.

  9. A class of selective antibacterials derived from a protein kinase inhibitor pharmacophore

    Energy Technology Data Exchange (ETDEWEB)

    Miller, J. Richard; Dunham, Steve; Mochalkin, Igor; Banotai, Craig; Bowman, Matthew; Buist, Susan; Dunkle, Bill; Hanna, Debra; Harwood, H. James; Huband, Michael D.; Karnovsky, Alla; Kuhn, Michael; Limberakis, Chris; Liu, Jia Y.; Mehrens, Shawn; Mueller, W. Thomas; Narasimhan, Lakshmi; Ogden, Adam; Ohren, Jeff; Prasad, J.V.N. Vara; Shelly, John A.; Skerlos, Laura; Sulavik, Mark; Thomas, V. Hayden; VanderRoest, Steve; Wang, LiAnn; Wang, Zhigang; Whitton, Amy; Zhu, Tong; Stover, C. Kendall; (Pfizer)

    2009-06-25

    As the need for novel antibiotic classes to combat bacterial drug resistance increases, the paucity of leads resulting from target-based antibacterial screening of pharmaceutical compound libraries is of major concern. One explanation for this lack of success is that antibacterial screening efforts have not leveraged the eukaryotic bias resulting from more extensive chemistry efforts targeting eukaryotic gene families such as G protein-coupled receptors and protein kinases. Consistent with a focus on antibacterial target space resembling these eukaryotic targets, we used whole-cell screening to identify a series of antibacterial pyridopyrimidines derived from a protein kinase inhibitor pharmacophore. In bacteria, the pyridopyrimidines target the ATP-binding site of biotin carboxylase (BC), which catalyzes the first enzymatic step of fatty acid biosynthesis. These inhibitors are effective in vitro and in vivo against fastidious Gram-negative pathogens including Haemophilus influenzae. Although the BC active site has architectural similarity to those of eukaryotic protein kinases, inhibitor binding to the BC ATP-binding site is distinct from the protein kinase-binding mode, such that the inhibitors are selective for bacterial BC. In summary, we have discovered a promising class of potent antibacterials with a previously undescribed mechanism of action. In consideration of the eukaryotic bias of pharmaceutical libraries, our findings also suggest that pursuit of a novel inhibitor leads for antibacterial targets with active-site structural similarity to known human targets will likely be more fruitful than the traditional focus on unique bacterial target space, particularly when structure-based and computational methodologies are applied to ensure bacterial selectivity.

  10. Using ovality to predict nonmutagenic, orally efficacious pyridazine amides as cell specific spleen tyrosine kinase inhibitors.

    Science.gov (United States)

    Lucas, Matthew C; Bhagirath, Niala; Chiao, Eric; Goldstein, David M; Hermann, Johannes C; Hsu, Pei-Yuan; Kirchner, Stephan; Kennedy-Smith, Joshua J; Kuglstatter, Andreas; Lukacs, Christine; Menke, John; Niu, Linghao; Padilla, Fernando; Peng, Ying; Polonchuk, Liudmila; Railkar, Aruna; Slade, Michelle; Soth, Michael; Xu, Daigen; Yadava, Preeti; Yee, Calvin; Zhou, Mingyan; Liao, Cheng

    2014-03-27

    Inhibition of spleen tyrosine kinase has attracted much attention as a mechanism for the treatment of cancers and autoimmune diseases such as asthma, rheumatoid arthritis, and systemic lupus erythematous. We report the structure-guided optimization of pyridazine amide spleen tyrosine kinase inhibitors. Early representatives of this scaffold were highly potent and selective but mutagenic in an Ames assay. An approach that led to the successful identification of nonmutagenic examples, as well as further optimization to compounds with reduced cardiovascular liabilities is described. Select pharmacokinetic and in vivo efficacy data are presented.

  11. Anti-Angiogenic Therapy: Strategies to Develop Potent VEGFR-2 Tyrosine Kinase Inhibitors and Future Prospect.

    Science.gov (United States)

    Shi, Leilei; Zhou, Jianfeng; Wu, Jifeng; Shen, Yuemao; Li, Xun

    2016-01-01

    Tumor angiogenesis has always been a major gap for effective cancer therapy. Interruption of aberrant angiogenesis by specific inhibitors targeting receptor tyrosine kinases (RTKs) has been of great interests to medicinal chemists. Among the factors that are involved in tumor angiogenesis, vascular endothelial growth factor receptor-2 (VEGFR-2) is validated as the most closely related factor which can drive angiogenesis through binding with its natural ligand VEGF. The well-validated VEGF-driven VEGFR-2 signaling pathway can stimulate many endothelial responses, including increasing vessel permeability and enhancing endothelial cell proliferation, migration and differentiation. Consequently, circumventing angiogenesis by VEGFR-2 inhibitors represents a promising strategy for counteracting various VEGFR-2-mediated disorders as well as drug resistance. Over the past decades, a considerable number of novel small molecular VEGFR-2 inhibitors have been exploited with diverse chemical scaffolds. Especially, recent frequently launched inhibitors have declared their research values and therapeutic potentials in oncology. Still, the antiangiogenesis based treatment remains an ongoing challenge. In this review, a comprehensive retrospective of newly emerged VEGFR-2 inhibitors have been summarized, with the emphasis on the structure-activity relationship (SAR) investigation, and also binding patterns of representative inhibitors with biotargets. On the basis of all of this information, varied strategies for developing potent VEGFR-2 inhibitors and the future prospect of the clinical application of antiangiogenic inhibitors are discussed hereby.

  12. Ligand-protein interactions of selective casein kinaseinhibitors.

    Science.gov (United States)

    Mente, Scot; Arnold, Eric; Butler, Todd; Chakrapani, Subramanyam; Chandrasekaran, Ramalakshmi; Cherry, Kevin; DiRico, Ken; Doran, Angela; Fisher, Katherine; Galatsis, Paul; Green, Michael; Hayward, Matthew; Humphrey, John; Knafels, John; Li, Jianke; Liu, Shenping; Marconi, Michael; McDonald, Scott; Ohren, Jeff; Paradis, Vanessa; Sneed, Blossom; Walton, Kevin; Wager, Travis

    2013-09-12

    Casein kinase 1δ (CK1δ) and 1ε (CK1ε) are believed to be necessary enzymes for the regulation of circadian rhythms in all mammals. On the basis of our previously published work demonstrating a CK1ε-preferring compound to be an ineffective circadian clock modulator, we have synthesized a series of pyrazole-substitued pyridine inhibitors, selective for the CK1δ isoform. Additionally, using structure-based drug design, we have been able to exploit differences in the hinge region between CK1δ and p38 to find selective inhibitors that have minimal p38 activity. The SAR, brain exposure, and the effect of these inhibitors on mouse circadian rhythms are described. The in vivo evaluation of these inhibitors demonstrates that selective inhibition of CK1δ at sufficient central exposure levels is capable of modulating circadian rhythms.

  13. An overview of the binding models of FGFR tyrosine kinases in complex with small molecule inhibitors.

    Science.gov (United States)

    Cheng, Weiyan; Wang, Mixiang; Tian, Xin; Zhang, Xiaojian

    2017-01-27

    The fibroblast growth factor receptor (FGFR) family receptor tyrosine kinase (RTK) includes four structurally related members, termed as FGFR1, FGFR2, FGFR3, and FGFR4. Given its intimate role in the progression of several solid tumors, excessive FGFR signaling provides an opportunity for anticancer therapy. Along with extensive pharmacological studies validating the therapeutic potential of targeting the FGFRs for cancer treatment, co-crystal structures of FGFRs/inhibitors are continuously coming up to study the mechanism of actions and explore new inhibitors. Herein, we review the reported co-crystals of FGFRs in complex with the corresponding inhibitors, main focusing our attention on the binding models and the pharmacological activities of the inhibitors.

  14. Excess F-actin mechanically impedes mitosis leading to cytokinesis failure in X-linked neutropenia by exceeding Aurora B kinase error correction capacity.

    Science.gov (United States)

    Moulding, Dale A; Moeendarbary, Emad; Valon, Leo; Record, Julien; Charras, Guillaume T; Thrasher, Adrian J

    2012-11-01

    The constitutively active mutant of the Wiskott-Aldrich Syndrome protein (CA-WASp) is the cause of X-linked neutropenia and is linked with genomic instability and myelodysplasia. CA-WASp generates abnormally high levels of cytoplasmic F-actin through dysregulated activation of the Arp2/3 complex leading to defects in cell division. As WASp has no reported role in cell division, we hypothesized that alteration of cell mechanics because of increased F-actin may indirectly disrupt dynamic events during mitosis. Inhibition of the Arp2/3 complex revealed that excess cytoplasmic F-actin caused increased cellular viscosity, slowed all phases of mitosis, and perturbed mitotic mechanics. Comparison of chromosome velocity to the cytoplasmic viscosity revealed that cells compensated for increased viscosity by up-regulating force applied to chromosomes and increased the density of microtubules at kinetochores. Mitotic abnormalities were because of overload of the aurora signaling pathway as subcritical inhibition of Aurora in CA-WASp cells caused increased cytokinesis failure, while overexpression reduced defects. These findings demonstrate that changes in cell mechanics can cause significant mitotic abnormalities leading to genomic instability, and highlight the importance of mechanical sensors such as Aurora B in maintaining the fidelity of hematopoietic cell division.

  15. Inflammatory Signaling by NOD-RIPK2 Is Inhibited by Clinically Relevant Type II Kinase Inhibitors.

    Science.gov (United States)

    Canning, Peter; Ruan, Qui; Schwerd, Tobias; Hrdinka, Matous; Maki, Jenny L; Saleh, Danish; Suebsuwong, Chalada; Ray, Soumya; Brennan, Paul E; Cuny, Gregory D; Uhlig, Holm H; Gyrd-Hansen, Mads; Degterev, Alexei; Bullock, Alex N

    2015-09-17

    RIPK2 mediates pro-inflammatory signaling from the bacterial sensors NOD1 and NOD2, and is an emerging therapeutic target in autoimmune and inflammatory diseases. We observed that cellular RIPK2 can be potently inhibited by type II inhibitors that displace the kinase activation segment, whereas ATP-competitive type I inhibition was only poorly effective. The most potent RIPK2 inhibitors were the US Food and Drug Administration-approved drugs ponatinib and regorafenib. Their mechanism of action was independent of NOD2 interaction and involved loss of downstream kinase activation as evidenced by lack of RIPK2 autophosphorylation. Notably, these molecules also blocked RIPK2 ubiquitination and, consequently, inflammatory nuclear factor κB signaling. In monocytes, the inhibitors selectively blocked NOD-dependent tumor necrosis factor production without affecting lipopolysaccharide-dependent pathways. We also determined the first crystal structure of RIPK2 bound to ponatinib, and identified an allosteric site for inhibitor development. These results highlight the potential for type II inhibitors to treat indications of RIPK2 activation as well as inflammation-associated cancers.

  16. Combining RNA interference and kinase inhibitors against cell signalling components involved in cancer

    Directory of Open Access Journals (Sweden)

    Hanson Bonnie J

    2005-10-01

    Full Text Available Abstract Background The transcription factor activator protein-1 (AP-1 has been implicated in a large variety of biological processes including oncogenic transformation. The tyrosine kinases of the epidermal growth factor receptor (EGFR constitute the beginning of one signal transduction cascade leading to AP-1 activation and are known to control cell proliferation and differentiation. Drug discovery efforts targeting this receptor and other pathway components have centred on monoclonal antibodies and small molecule inhibitors. Resistance to such inhibitors has already been observed, guiding the prediction of their use in combination therapies with other targeted agents such as RNA interference (RNAi. This study examines the use of RNAi and kinase inhibitors for qualification of components involved in the EGFR/AP-1 pathway of ME180 cells, and their inhibitory effects when evaluated individually or in tandem against multiple components of this important disease-related pathway. Methods AP-1 activation was assessed using an ME180 cell line stably transfected with a beta-lactamase reporter gene under the control of AP-1 response element following epidermal growth factor (EGF stimulation. Immunocytochemistry allowed for further quantification of small molecule inhibition on a cellular protein level. RNAi and RT-qPCR experiments were performed to assess the amount of knockdown on an mRNA level, and immunocytochemistry was used to reveal cellular protein levels for the targeted pathway components. Results Increased potency of kinase inhibitors was shown by combining RNAi directed towards EGFR and small molecule inhibitors acting at proximal or distal points in the pathway. After cellular stimulation with EGF and analysis at the level of AP-1 activation using a β-lactamase reporter gene, a 10–12 fold shift or 2.5–3 fold shift toward greater potency in the IC50 was observed for EGFR and MEK-1 inhibitors, respectively, in the presence of RNAi

  17. Structural basis for isoform selectivity in a class of benzothiazole inhibitors of phosphoinositide 3-kinase γ.

    Science.gov (United States)

    Collier, Philip N; Martinez-Botella, Gabriel; Cornebise, Mark; Cottrell, Kevin M; Doran, John D; Griffith, James P; Mahajan, Sudipta; Maltais, François; Moody, Cameron S; Huck, Emilie Porter; Wang, Tiansheng; Aronov, Alex M

    2015-01-08

    Phosphoinositide 3-kinase γ (PI3Kγ) is an attractive target to potentially treat a range of disease states. Herein, we describe the evolution of a reported phenylthiazole pan-PI3K inhibitor into a family of potent and selective benzothiazole inhibitors. Using X-ray crystallography, we discovered that compound 22 occupies a previously unreported hydrophobic binding cleft adjacent to the ATP binding site of PI3Kγ, and achieves its selectivity by exploiting natural sequence differences among PI3K isoforms in this region.

  18. Role of tyrosine-kinase inhibitors in myeloproliferative neoplasms: comparative lessons learned

    Directory of Open Access Journals (Sweden)

    Pinilla-Ibarz J

    2016-08-01

    Full Text Available Javier Pinilla-Ibarz, Kendra L Sweet, Gabriela M Corrales-Yepez, Rami S Komrokji Department of Malignant Hematology, H Lee Moffitt Cancer Center and Research Institute, Tampa, FL, USA Abstract: An important pathogenetic distinction in the classification of myeloproliferative neoplasms (MPNs is the presence or absence of the BCR–ABL fusion gene, which encodes a unique oncogenic tyrosine kinase. The BCR–ABL fusion, caused by the formation of the Philadelphia chromosome (Ph through translocation, constitutes the disease-initiating event in chronic myeloid leukemia. The development of successive BCR–ABL-targeted tyrosine-kinase inhibitors has led to greatly improved outcomes in patients with chronic myeloid leukemia, including high rates of complete hematologic, cytogenetic, and molecular responses. Such levels of treatment success have long been elusive for patients with Ph-negative MPNs, because of the difficulties in identifying specific driver proteins suitable as drug targets. However, in recent years an improved understanding of the complex pathobiology of classic Ph-negative MPNs, characterized by variable, overlapping multimutation profiles, has prompted the development of better and more broadly targeted (to pathway rather than protein treatment options, particularly JAK inhibitors. In classic Ph-negative MPNs, overactivation of JAK-dependent signaling pathways is a central pathogenic mechanism, and mutually exclusive mutations in JAK2, MPL, and CALR linked to aberrant JAK activation are now recognized as key drivers of disease progression in myelofibrosis (MF. In clinical trials, the JAK1/JAK2 inhibitor ruxolitinib – the first therapy approved for MF worldwide – improved disease-related splenomegaly and symptoms independent of JAK2V617F mutational status, and prolonged survival compared with placebo or standard therapy in patients with advanced MF. In separate trials, ruxolitinib also provided comprehensive hematologic control in

  19. Evaluating the predictivity of virtual screening for ABL kinase inhibitors to hinder drug resistance

    DEFF Research Database (Denmark)

    Gani, Osman A B S M; Narayanan, Dilip; Engh, Richard A

    2013-01-01

    Virtual screening methods are now widely used in early stages of drug discovery, aiming to rank potential inhibitors. However, any practical ligand set (of active or inactive compounds) chosen for deriving new virtual screening approaches cannot fully represent all relevant chemical space for pot...... inactive ligands, even without assuming knowledge of the binding mode. We believe that this study can be extended to other therapeutically important kinases in prospective virtual screening studies....... for potential new compounds. In this study, we have taken a retrospective approach to evaluate virtual screening methods for the leukemia target kinase ABL1 and its drug-resistant mutant ABL1-T315I. 'Dual active' inhibitors against both targets were grouped together with inactive ligands chosen from different...

  20. Combination treatment of prostate cancer with FGF receptor and AKT kinase inhibitors

    Science.gov (United States)

    Feng, Shu; Shao, Longjiang; Castro, Patricia; Coleman, Ilsa; Nelson, Peter S; Smith, Paul D; Davies, Barry R; Ittmann, Michael

    2017-01-01

    Activation of the PI3K/AKT pathway occurs in the vast majority of advanced prostate cancers (PCas). Activation of fibroblast growth factor receptor (FGFR) signaling occurs in a wide variety of malignancies, including PCa. RNA-Seq of castration resistant PCa revealed expression of multiple FGFR signaling components compatible with FGFR signaling in all cases, with multiple FGF ligands expressed in 90% of cases. Immunohistochemistry confirmed FGFR signaling in the majority of xenografts and advanced PCas. AZD5363, an AKT kinase inhibitor and AZD4547, a FGFR kinase inhibitor are under active clinical development. We therefore sought to determine if these two drugs have additive effects in PCa models. The effect of both agents, singly and in combination was evaluated in a variety of PCa cell lines in vitro and in vivo. All cell lines tested responded to both drugs with decreased invasion, soft agar colony formation and growth in vivo, with additive effects seen with combination treatment. Activation of the FGFR, AKT, ERK and STAT3 pathways was examined in treated cells. AZD5363 inhibited AKT signaling and increased FGFR1 signaling, which partially compensated for decreased AKT kinase activity. While AZD4547 could effectively block the ERK pathway, combination treatment was needed to completely block STAT3 activation. Thus combination treatment with AKT and FGFR kinase inhibitors have additive effects on malignant phenotypes in vitro and in vivo by inhibiting multiple signaling pathways and mitigating the compensatory upregulation of FGFR signaling induced by AKT kinase inhibition. Our studies suggest that co-targeting these pathways may be efficacious in advanced PCa. PMID:28008155

  1. Masitinib (AB1010, a potent and selective tyrosine kinase inhibitor targeting KIT.

    Directory of Open Access Journals (Sweden)

    Patrice Dubreuil

    Full Text Available BACKGROUND: The stem cell factor receptor, KIT, is a target for the treatment of cancer, mastocytosis, and inflammatory diseases. Here, we characterise the in vitro and in vivo profiles of masitinib (AB1010, a novel phenylaminothiazole-type tyrosine kinase inhibitor that targets KIT. METHODOLOGY/PRINCIPAL FINDINGS: In vitro, masitinib had greater activity and selectivity against KIT than imatinib, inhibiting recombinant human wild-type KIT with an half inhibitory concentration (IC(50 of 200+/-40 nM and blocking stem cell factor-induced proliferation and KIT tyrosine phosphorylation with an IC(50 of 150+/-80 nM in Ba/F3 cells expressing human or mouse wild-type KIT. Masitinib also potently inhibited recombinant PDGFR and the intracellular kinase Lyn, and to a lesser extent, fibroblast growth factor receptor 3. In contrast, masitinib demonstrated weak inhibition of ABL and c-Fms and was inactive against a variety of other tyrosine and serine/threonine kinases. This highly selective nature of masitinib suggests that it will exhibit a better safety profile than other tyrosine kinase inhibitors; indeed, masitinib-induced cardiotoxicity or genotoxicity has not been observed in animal studies. Molecular modelling and kinetic analysis suggest a different mode of binding than imatinib, and masitinib more strongly inhibited degranulation, cytokine production, and bone marrow mast cell migration than imatinib. Furthermore, masitinib potently inhibited human and murine KIT with activating mutations in the juxtamembrane domain. In vivo, masitinib blocked tumour growth in mice with subcutaneous grafts of Ba/F3 cells expressing a juxtamembrane KIT mutant. CONCLUSIONS: Masitinib is a potent and selective tyrosine kinase inhibitor targeting KIT that is active, orally bioavailable in vivo, and has low toxicity.

  2. Identification and Characterization of Amlexanox as a G Protein-Coupled Receptor Kinase 5 Inhibitor

    Directory of Open Access Journals (Sweden)

    Kristoff T. Homan

    2014-10-01

    Full Text Available G protein-coupled receptor kinases (GRKs have been implicated in human diseases ranging from heart failure to diabetes. Previous studies have identified several compounds that selectively inhibit GRK2, such as paroxetine and balanol. Far fewer selective inhibitors have been reported for GRK5, a target for the treatment of cardiac hypertrophy, and the mechanism of action of reported compounds is unknown. To identify novel scaffolds that selectively inhibit GRK5, a differential scanning fluorometry screen was used to probe a library of 4480 compounds. The best hit was amlexanox, an FDA-approved anti-inflammatory, anti-allergic immunomodulator. The crystal structure of amlexanox in complex with GRK1 demonstrates that its tricyclic aromatic ring system forms ATP-like interactions with the hinge of the kinase domain, which is likely similar to how this drug binds to IκB kinase ε (IKKε, another kinase known to be inhibited by this compound. Amlexanox was also able to inhibit myocyte enhancer factor 2 transcriptional activity in neonatal rat ventricular myocytes in a manner consistent with GRK5 inhibition. The GRK1 amlexanox structure thus serves as a springboard for the rational design of inhibitors with improved potency and selectivity for GRK5 and IKKε.

  3. Casein kinase 1δ/ε inhibitor PF-5006739 attenuates opioid drug-seeking behavior.

    Science.gov (United States)

    Wager, Travis T; Chandrasekaran, Ramalakshmi Y; Bradley, Jenifer; Rubitski, David; Berke, Helen; Mente, Scot; Butler, Todd; Doran, Angela; Chang, Cheng; Fisher, Katherine; Knafels, John; Liu, Shenping; Ohren, Jeff; Marconi, Michael; DeMarco, George; Sneed, Blossom; Walton, Kevin; Horton, David; Rosado, Amy; Mead, Andy

    2014-12-17

    Casein kinase 1 delta (CK1δ) and casein kinase 1 epsilon (CK1ε) inhibitors are potential therapeutic agents for a range of psychiatric disorders. The feasibility of developing a CNS kinase inhibitor has been limited by an inability to identify safe brain-penetrant compounds with high kinome selectivity. Guided by structure-based drug design, potent and selective CK1δ/ε inhibitors have now been identified that address this gap, through the design and synthesis of novel 4-[4-(4-fluorophenyl)-1-(piperidin-4-yl)-1H-imidazol-5-yl]pyrimidin-2-amine derivatives. PF-5006739 (6) possesses a desirable profile, with low nanomolar in vitro potency for CK1δ/ε (IC50 = 3.9 and 17.0 nM, respectively) and high kinome selectivity. In vivo, 6 demonstrated robust centrally mediated circadian rhythm phase-delaying effects in both nocturnal and diurnal animal models. Further, 6 dose-dependently attenuated opioid drug-seeking behavior in a rodent operant reinstatement model in animals trained to self-administer fentanyl. Collectively, our data supports further development of 6 as a promising candidate to test the hypothesis of CK1δ/ε inhibition in treating multiple indications in the clinic.

  4. Mesenchymal stem cells are sensitive to treatment with kinase inhibitors and ionizing radiation

    Energy Technology Data Exchange (ETDEWEB)

    Nicolay, Nils H.; Debus, Juergen; Huber, Peter E. [Heidelberg University Hospital, Department of Radiation Oncology, Heidelberg (Germany); German Cancer Research Center (dkfz), Department of Molecular and Radiation Oncology, Heidelberg (Germany); Sommer, Eva; Lopez Perez, Ramon; Wirkner, Ute [German Cancer Research Center (dkfz), Department of Molecular and Radiation Oncology, Heidelberg (Germany); Bostel, Tilman [Heidelberg University Hospital, Department of Radiation Oncology, Heidelberg (Germany); Ho, Anthony D.; Saffrich, Rainer [Heidelberg University Hospital, Department of Hematology, Oncology and Rheumatology, Heidelberg (Germany); Lahn, Michael [Lilly Research Laboratories, Oncology Early Clinical Investigation, Indianapolis, IN (United States)

    2014-11-15

    Mesenchymal stem cells (MSCs) can regenerate damaged tissues and may therefore be of importance for normal tissue repair after cancer treatment. Small molecule receptor kinase inhibitors (RKIs) have recently been introduced into cancer treatment. However, the influence of these drugs - particularly in combination with radiotherapy - on the survival of MSCs is largely unknown. The sensitivity of human primary MSCs from healthy volunteers and primary human fibroblast cells to small molecule kinase inhibitors of the vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF) and transforming growth factor β (TGFβ) receptors, as well to inhibitors of c-Kit, was examined in combination with ionizing radiation (IR); cell survival and proliferation were assessed. Expression patterns of different kinase receptors and ligands were investigated using gene arrays. MSCs were highly sensitive to the tyrosine kinase inhibitors SU14816 (imatinib) and SU11657 (sunitinib), but showed only moderate sensitivity to the selective TGFβ receptor 1 inhibitor LY2109761. Primary adult human fibroblasts were comparably resistant to all three inhibitors. The addition of IR had an additive or supra-additive effect in the MSCs, but this was not the case for differentiated fibroblasts. Proliferation was markedly reduced in MSCs following kinase inhibition, both with and without IR. Gene expression analysis revealed high levels of the PDGF α and β receptors, and lower levels of the TGFβ receptor 2 and Abl kinase. IR did not alter the expression of kinase receptors or their respective ligands in either MSCs or adult fibroblasts. These data show that MSCs are highly sensitive to RKIs and combination treatments incorporating IR. Expression analyses suggest that high levels of PDGF receptors may contribute to this effect. (orig.) [German] Mesenchymale Stammzellen (MSCs) koennen die Geweberegeneration unterstuetzen und haben daher moeglicherweise eine Rolle bei der Reparatur

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

    Science.gov (United States)

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

    2012-01-01

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

  6. To fast or not to fast? How food could impact on the absorption of kinase inhibitors and its economical value

    Directory of Open Access Journals (Sweden)

    Camillo Porta

    2011-12-01

    Full Text Available Presently, cancer patients receiving oral kinase inhibitors are suggested to take the drug with an empty stomach or, in the case of drugs such as Sorafenib which are administered twice daily, at least following a lowfat meal...

  7. EBNA3C-mediated regulation of aurora kinase B contributes to Epstein-Barr virus-induced B-cell proliferation through modulation of the activities of the retinoblastoma protein and apoptotic caspases.

    Science.gov (United States)

    Jha, Hem Chandra; Lu, Jie; Saha, Abhik; Cai, Qiliang; Banerjee, Shuvomoy; Prasad, Mahadesh A J; Robertson, Erle S

    2013-11-01

    Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that is implicated in several human malignancies, including Burkitt's lymphoma (BL), posttransplant lymphoproliferative disease (PTLD), nasopharyngeal carcinoma (NPC), and AIDS-associated lymphomas. Epstein-Barr nuclear antigen 3C (EBNA3C), one of the essential EBV latent antigens, can induce mammalian cell cycle progression through its interaction with cell cycle regulators. Aurora kinase B (AK-B) is important for cell division, and deregulation of AK-B is associated with aneuploidy, incomplete mitotic exit, and cell death. Our present study shows that EBNA3C contributes to upregulation of AK-B transcript levels by enhancing the activity of its promoter. Further, EBNA3C also increased the stability of the AK-B protein, and the presence of EBNA3C leads to reduced ubiquitination of AK-B. Importantly, EBNA3C in association with wild-type AK-B but not with its kinase-dead mutant led to enhanced cell proliferation, and AK-B knockdown can induce nuclear blebbing and cell death. This phenomenon was rescued in the presence of EBNA3C. Knockdown of AK-B resulted in activation of caspase 3 and caspase 9, along with poly(ADP-ribose) polymerase 1 (PARP1) cleavage, which is known to be an important contributor to apoptotic signaling. Importantly, EBNA3C failed to stabilize the kinase-dead mutant of AK-B compared to wild-type AK-B, which suggests a role for the kinase domain in AK-B stabilization and downstream phosphorylation of the cell cycle regulator retinoblastoma protein (Rb). This study demonstrates the functional relevance of AK-B kinase activity in EBNA3C-regulated B-cell proliferation and apoptosis.

  8. Structure-based design and synthesis of potent benzothiazole inhibitors of interleukin-2 inducible T cell kinase (ITK).

    Science.gov (United States)

    MacKinnon, Colin H; Lau, Kevin; Burch, Jason D; Chen, Yuan; Dines, Jonathon; Ding, Xiao; Eigenbrot, Charles; Heifetz, Alexander; Jaochico, Allan; Johnson, Adam; Kraemer, Joachim; Kruger, Susanne; Krülle, Thomas M; Liimatta, Marya; Ly, Justin; Maghames, Rosemary; Montalbetti, Christian A G N; Ortwine, Daniel F; Pérez-Fuertes, Yolanda; Shia, Steven; Stein, Daniel B; Trani, Giancarlo; Vaidya, Darshan G; Wang, Xiaolu; Bromidge, Steven M; Wu, Lawren C; Pei, Zhonghua

    2013-12-01

    Inhibition of the non-receptor tyrosine kinase ITK, a component of the T-cell receptor signalling cascade, may represent a novel treatment for allergic asthma. Here we report the structure-based optimization of a series of benzothiazole amides that demonstrate sub-nanomolar inhibitory potency against ITK with good cellular activity and kinase selectivity. We also elucidate the binding mode of these inhibitors by solving the X-ray crystal structures of several inhibitor-ITK complexes.

  9. Structural characterization of nonactive site, TrkA-selective kinase inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Su, Hua-Poo; Rickert, Keith; Burlein, Christine; Narayan, Kartik; Bukhtiyarova, Marina; Hurzy, Danielle M.; Stump, Craig A.; Zhang, Xufang; Reid, John; Krasowska-Zoladek, Alicja; Tummala, Srivanya; Shipman, Jennifer M.; Kornienko, Maria; Lemaire, Peter A.; Krosky, Daniel; Heller, Amanda; Achab, Abdelghani; Chamberlin, Chad; Saradjian, Peter; Sauvagnat, Berengere; Yang, Xianshu; Ziebell, Michael R.; Nickbarg, Elliott; Sanders, John M.; Bilodeau, Mark T.; Carroll, Steven S.; Lumb, Kevin J.; Soisson, Stephen M.; Henze, Darrell A.; Cooke, Andrew J. (Merck)

    2016-12-30

    Current therapies for chronic pain can have insufficient efficacy and lead to side effects, necessitating research of novel targets against pain. Although originally identified as an oncogene, Tropomyosin-related kinase A (TrkA) is linked to pain and elevated levels of NGF (the ligand for TrkA) are associated with chronic pain. Antibodies that block TrkA interaction with its ligand, NGF, are in clinical trials for pain relief. Here, we describe the identification of TrkA-specific inhibitors and the structural basis for their selectivity over other Trk family kinases. The X-ray structures reveal a binding site outside the kinase active site that uses residues from the kinase domain and the juxtamembrane region. Three modes of binding with the juxtamembrane region are characterized through a series of ligand-bound complexes. The structures indicate a critical pharmacophore on the compounds that leads to the distinct binding modes. The mode of interaction can allow TrkA selectivity over TrkB and TrkC or promiscuous, pan-Trk inhibition. This finding highlights the difficulty in characterizing the structure-activity relationship of a chemical series in the absence of structural information because of substantial differences in the interacting residues. These structures illustrate the flexibility of binding to sequences outside of—but adjacent to—the kinase domain of TrkA. This knowledge allows development of compounds with specificity for TrkA or the family of Trk proteins.

  10. Small-Molecule Inhibitors of the Receptor Tyrosine Kinases: Promising Tools for Targeted Cancer Therapies

    Directory of Open Access Journals (Sweden)

    Mohammad Hojjat-Farsangi

    2014-08-01

    Full Text Available Chemotherapeutic and cytotoxic drugs are widely used in the treatment of cancer. In spite of the improvements in the life quality of patients, their effectiveness is compromised by several disadvantages. This represents a demand for developing new effective strategies with focusing on tumor cells and minimum side effects. Targeted cancer therapies and personalized medicine have been defined as a new type of emerging treatments. Small molecule inhibitors (SMIs are among the most effective drugs for targeted cancer therapy. The growing number of approved SMIs of receptor tyrosine kinases (RTKs i.e., tyrosine kinase inhibitors (TKIs in the clinical oncology imply the increasing attention and application of these therapeutic tools. Most of the current approved RTK–TKIs in preclinical and clinical settings are multi-targeted inhibitors with several side effects. Only a few specific/selective RTK–TKIs have been developed for the treatment of cancer patients. Specific/selective RTK–TKIs have shown less deleterious effects compared to multi-targeted inhibitors. This review intends to highlight the importance of specific/selective TKIs for future development with less side effects and more manageable agents. This article provides an overview of: (1 the characteristics and function of RTKs and TKIs; (2 the recent advances in the improvement of specific/selective RTK–TKIs in preclinical or clinical settings; and (3 emerging RTKs for targeted cancer therapies by TKIs.

  11. Inhibition of dihydroceramide desaturase activity by the sphingosine kinase inhibitor SKI II.

    Science.gov (United States)

    Cingolani, Francesca; Casasampere, Mireia; Sanllehí, Pol; Casas, Josefina; Bujons, Jordi; Fabrias, Gemma

    2014-08-01

    Sphingosine kinase inhibitor (SKI) II has been reported as a dual inhibitor of sphingosine kinases (SKs) 1 and 2 and has been extensively used to prove the involvement of SKs and sphingosine-1-phosphate (S1P) in cellular processes. Dihydroceramide desaturase (Des1), the last enzyme in the de novo synthesis of ceramide (Cer), regulates the balance between dihydroceramides (dhCers) and Cers. Both SKs and Des1 have interest as therapeutic targets. Here we show that SKI II is a noncompetitive inhibitor (Ki = 0.3 μM) of Des1 activity with effect also in intact cells without modifying Des1 protein levels. Molecular modeling studies support that the SKI II-induced decrease in Des1 activity could result from inhibition of NADH-cytochrome b5 reductase. SKI II, but not the SK1-specific inhibitor PF-543, provoked a remarkable accumulation of dhCers and their metabolites, while both SKI II and PF-543 reduced S1P to almost undetectable levels. SKI II, but not PF543, reduced cell proliferation with accumulation of cells in the G0/G1 phase. SKI II, but not PF543, induced autophagy. These overall findings should be taken into account when using SKI II as a pharmacological tool, as some of the effects attributed to decreased S1P may actually be caused by augmented dhCers and/or their metabolites.

  12. Novel small molecule inhibitors of 3-phosphoinositide-dependent kinase-1.

    Science.gov (United States)

    Feldman, Richard I; Wu, James M; Polokoff, Mark A; Kochanny, Monica J; Dinter, Harald; Zhu, Daguang; Biroc, Sandra L; Alicke, Bruno; Bryant, Judi; Yuan, Shendong; Buckman, Brad O; Lentz, Dao; Ferrer, Mike; Whitlow, Marc; Adler, Marc; Finster, Silke; Chang, Zheng; Arnaiz, Damian O

    2005-05-20

    The phosphoinositide 3-kinase/3-phosphoinositide-dependent kinase 1 (PDK1)/Akt signaling pathway plays a key role in cancer cell growth, survival, and tumor angiogenesis and represents a promising target for anticancer drugs. Here, we describe three potent PDK1 inhibitors, BX-795, BX-912, and BX-320 (IC(50) = 11-30 nm) and their initial biological characterization. The inhibitors blocked PDK1/Akt signaling in tumor cells and inhibited the anchorage-dependent growth of a variety of tumor cell lines in culture or induced apoptosis. A number of cancer cell lines with elevated Akt activity were >30-fold more sensitive to growth inhibition by PDK1 inhibitors in soft agar than on tissue culture plastic, consistent with the cell survival function of the PDK1/Akt signaling pathway, which is particularly important for unattached cells. BX-320 inhibited the growth of LOX melanoma tumors in the lungs of nude mice after injection of tumor cells into the tail vein. The effect of BX-320 on cancer cell growth in vitro and in vivo indicates that PDK1 inhibitors may have clinical utility as anticancer agents.

  13. (Secondary) solid tumors in thyroid cancer patients treated with the multi-kinase inhibitor sorafenib may present diagnostic challenges

    NARCIS (Netherlands)

    Schneider, T.C.; Kapiteijn, E.; Wezel, T. van; Smit, J.W.A.; Hoeven, J.J.M. van der; Morreau, H.

    2016-01-01

    BACKGROUND: Sorafenib is an orally active multikinase tyrosine kinase inhibitor (TKI) that targets B-type Raf kinase (BRAF), vascular endothelial growth factor receptors (VEGFR) 1 and 2, and rearranged during transfection (RET), inducing anti-angiogenic and pro-apoptotic actions in a wide range of s

  14. Identification Of Akn-032, A Novel 2-Aminopyrazine Tyrosine Kinase Inhibitor, With Significant Preclinical Activity In Acute Myeloid Leukemia

    OpenAIRE

    Eriksson, Anna; Höglund, Martin; Lindhagen, Elin; Åleskog, Anna; Hassan, Sadia Bashir; Ekholm, Carina; Fhölenhag, Karin; Jensen, Annika Jenmalm; Löthgren, Agneta; Scobie, Martin; Larsson, Rolf; Parrow, Vendela

    2010-01-01

    Abstract Aberrant signal transduction by mutant or overexpressed protein kinases has emerged as a promising target for treatment of acute myeloid leukemia (AML). We here present a novel low molecular weight kinase inhibitor, AKN-032, targeting the FMS-like tyrosine kinase 3 (FLT3) and discovered in a new type of screening funnel combining the target therapy approach with sequential cellular screens. AKN-032 was identified among 150 selected hits from three different high throughput...

  15. Implications of promiscuous Pim-1 kinase fragment inhibitor hydrophobic interactions for fragment-based drug design.

    Science.gov (United States)

    Good, Andrew C; Liu, Jinyu; Hirth, Bradford; Asmussen, Gary; Xiang, Yibin; Biemann, Hans-Peter; Bishop, Kimberly A; Fremgen, Trisha; Fitzgerald, Maria; Gladysheva, Tatiana; Jain, Annuradha; Jancsics, Katherine; Metz, Markus; Papoulis, Andrew; Skerlj, Renato; Stepp, J David; Wei, Ronnie R

    2012-03-22

    We have studied the subtleties of fragment docking and binding using data generated in a Pim-1 kinase inhibitor program. Crystallographic and docking data analyses have been undertaken using inhibitor complexes derived from an in-house surface plasmon resonance (SPR) fragment screen, a virtual needle screen, and a de novo designed fragment inhibitor hybrid. These investigations highlight that fragments that do not fill their binding pocket can exhibit promiscuous hydrophobic interactions due to the lack of steric constraints imposed on them by the boundaries of said pocket. As a result, docking modes that disagree with an observed crystal structure but maintain key crystallographically observed hydrogen bonds still have potential value in ligand design and optimization. This observation runs counter to the lore in fragment-based drug design that all fragment elaboration must be based on the parent crystal structure alone.

  16. Skp2 is required for Aurora B activation in cell mitosis and spindle checkpoint.

    Science.gov (United States)

    Wu, Juan; Huang, Yu-Fan; Zhou, Xin-Ke; Zhang, Wei; Lian, Yi-Fan; Lv, Xiao-Bin; Gao, Xiu-Rong; Lin, Hui-Kuan; Zeng, Yi-Xin; Huang, Jian-Qing

    2015-01-01

    The Aurora B kinase plays a critical role in cell mitosis and spindle checkpoint. Here, we showed that the ubiquitin E3-ligase protein Skp2, also as a cell-cycle regulatory protein, was required for the activation of Aurora B and its downstream protein. When we restored Skp2 knockdown Hela cells with Skp2 and Skp2-LRR E3 ligase dead mutant we found that Skp2 could rescue the defect in the activation of Aurora B, but the mutant failed to do so. Furthermore, we discovered that Skp2 could interact with Aurora B and trigger Aurora B Lysine (K) 63-linked ubiquitination. Finally, we demonstrated the essential role of Skp2 in cell mitosis progression and spindle checkpoint, which was Aurora B dependent. Our results identified a novel ubiquitinated substrate of Skp2, and also indicated that Aurora B ubiquitination might serve as an important event for Aurora B activation in cell mitosis and spindle checkpoint.

  17. The phosphoinositide-dependent protein kinase 1 inhibitor, UCN-01, induces fragmentation: possible role of metalloproteinases.

    Science.gov (United States)

    Alcántara-Hernández, Rocío; Hernández-Méndez, Aurelio; García-Sáinz, J Adolfo

    2014-10-05

    Phosphoinositide-dependent protein kinase 1 (PDK1) is a key enzyme, master regulator of cellular proliferation and metabolism; it is considered a key target for pharmacological intervention. Using membranes obtained from DDT1 MF-2 cells, phospho-PDK1 was identified by Western blotting, as two major protein bands of Mr 58-68 kDa. Cell incubation with the PDK1 inhibitor, UCN-01, induced a time- and concentration-dependent decrease in the amount of phospho-PDK1 with a concomitant appearance of a ≈42 kDa phosphorylated fragment. Knocking down PDK1 diminished the amount of phospho-PDK1 detected in membranes, accompanied by similarly decreased fragment generation. UCN-01-induced fragment generation was also observed in membranes from cells stably expressing a myc-tagged PDK1 construct. Other PDK1 inhibitors were also tested: OSU-03012 induced a clear decrease in phospho-PDK1 and increased the presence of the phosphorylated fragment in membrane preparations; in contrast, GSK2334470 and staurosporine induced only marginal increases in the amount of PDK1 fragment. Galardin and batimastat, two metalloproteinase inhibitors, markedly attenuated inhibitor-induced PDK1 fragment generation. Metalloproteinases 2, 3, and 9 co-immunoprecipitated with myc-PDK1 under baseline conditions and this interaction was stimulated by UCN-01; batimastat also markedly diminished this effect of the PDK1 inhibitor. Our results indicate that a series of protein kinase inhibitors, namely UCN-01 and OSU-03012 and to a lesser extent GSK2334470 and staurosporine induce PDK1 fragmentation and suggest that metalloproteinases could participate in this effect.

  18. Comparison of the cancer gene targeting and biochemical selectivities of all targeted kinase inhibitors approved for clinical use.

    Directory of Open Access Journals (Sweden)

    Joost C M Uitdehaag

    Full Text Available The anti-proliferative activities of all twenty-five targeted kinase inhibitor drugs that are in clinical use were measured in two large assay panels: (1 a panel of proliferation assays of forty-four human cancer cell lines from diverse tumour tissue origins; and (2 a panel of more than 300 kinase enzyme activity assays. This study provides a head-on comparison of all kinase inhibitor drugs in use (status Nov. 2013, and for six of these drugs, the first kinome profiling data in the public domain. Correlation of drug activities with cancer gene mutations revealed novel drug sensitivity markers, suggesting that cancers dependent on mutant CTNNB1 will respond to trametinib and other MEK inhibitors, and cancers dependent on SMAD4 to small molecule EGFR inhibitor drugs. Comparison of cellular targeting efficacies reveals the most targeted inhibitors for EGFR, ABL1 and BRAF(V600E-driven cell growth, and demonstrates that the best targeted agents combine high biochemical potency with good selectivity. For ABL1 inhibitors, we computationally deduce optimized kinase profiles for use in a next generation of drugs. Our study shows the power of combining biochemical and cellular profiling data in the evaluation of kinase inhibitor drug action.

  19. The potent Cdc7-Dbf4 (DDK kinase inhibitor XL413 has limited activity in many cancer cell lines and discovery of potential new DDK inhibitor scaffolds.

    Directory of Open Access Journals (Sweden)

    Nanda Kumar Sasi

    Full Text Available Cdc7-Dbf4 kinase or DDK (Dbf4-dependent kinase is required to initiate DNA replication by phosphorylating and activating the replicative Mcm2-7 DNA helicase. DDK is overexpressed in many tumor cells and is an emerging chemotherapeutic target since DDK inhibition causes apoptosis of diverse cancer cell types but not of normal cells. PHA-767491 and XL413 are among a number of potent DDK inhibitors with low nanomolar IC50 values against the purified kinase. Although XL413 is highly selective for DDK, its activity has not been extensively characterized on cell lines. We measured anti-proliferative and apoptotic effects of XL413 on a panel of tumor cell lines compared to PHA-767491, whose activity is well characterized. Both compounds were effective biochemical DDK inhibitors but surprisingly, their activities in cell lines were highly divergent. Unlike PHA-767491, XL413 had significant anti-proliferative activity against only one of the ten cell lines tested. Since XL413 did not effectively inhibit DDK in multiple cell lines, this compound likely has limited bioavailability. To identify potential leads for additional DDK inhibitors, we also tested the cross-reactivity of ∼400 known kinase inhibitors against DDK using a DDK thermal stability shift assay (TSA. We identified 11 compounds that significantly stabilized DDK. Several inhibited DDK with comparable potency to PHA-767491, including Chk1 and PKR kinase inhibitors, but had divergent chemical scaffolds from known DDK inhibitors. Taken together, these data show that several well-known kinase inhibitors cross-react with DDK and also highlight the opportunity to design additional specific, biologically active DDK inhibitors for use as chemotherapeutic agents.

  20. The potent Cdc7-Dbf4 (DDK) kinase inhibitor XL413 has limited activity in many cancer cell lines and discovery of potential new DDK inhibitor scaffolds.

    Science.gov (United States)

    Sasi, Nanda Kumar; Tiwari, Kanchan; Soon, Fen-Fen; Bonte, Dorine; Wang, Tong; Melcher, Karsten; Xu, H Eric; Weinreich, Michael

    2014-01-01

    Cdc7-Dbf4 kinase or DDK (Dbf4-dependent kinase) is required to initiate DNA replication by phosphorylating and activating the replicative Mcm2-7 DNA helicase. DDK is overexpressed in many tumor cells and is an emerging chemotherapeutic target since DDK inhibition causes apoptosis of diverse cancer cell types but not of normal cells. PHA-767491 and XL413 are among a number of potent DDK inhibitors with low nanomolar IC50 values against the purified kinase. Although XL413 is highly selective for DDK, its activity has not been extensively characterized on cell lines. We measured anti-proliferative and apoptotic effects of XL413 on a panel of tumor cell lines compared to PHA-767491, whose activity is well characterized. Both compounds were effective biochemical DDK inhibitors but surprisingly, their activities in cell lines were highly divergent. Unlike PHA-767491, XL413 had significant anti-proliferative activity against only one of the ten cell lines tested. Since XL413 did not effectively inhibit DDK in multiple cell lines, this compound likely has limited bioavailability. To identify potential leads for additional DDK inhibitors, we also tested the cross-reactivity of ∼400 known kinase inhibitors against DDK using a DDK thermal stability shift assay (TSA). We identified 11 compounds that significantly stabilized DDK. Several inhibited DDK with comparable potency to PHA-767491, including Chk1 and PKR kinase inhibitors, but had divergent chemical scaffolds from known DDK inhibitors. Taken together, these data show that several well-known kinase inhibitors cross-react with DDK and also highlight the opportunity to design additional specific, biologically active DDK inhibitors for use as chemotherapeutic agents.

  1. New pyrazolopyrimidine inhibitors of protein kinase d as potent anticancer agents for prostate cancer cells.

    Directory of Open Access Journals (Sweden)

    Manuj Tandon

    Full Text Available The emergence of protein kinase D (PKD as a potential therapeutic target for several diseases including cancer has triggered the search for potent, selective, and cell-permeable small molecule inhibitors. In this study, we describe the identification, in vitro characterization, structure-activity analysis, and biological evaluation of a novel PKD inhibitory scaffold exemplified by 1-naphthyl PP1 (1-NA-PP1. 1-NA-PP1 and IKK-16 were identified as pan-PKD inhibitors in a small-scale targeted kinase inhibitor library assay. Both screening hits inhibited PKD isoforms at about 100 nM and were ATP-competitive inhibitors. Analysis of several related kinases indicated that 1-NA-PP1 was highly selective for PKD as compared to IKK-16. SAR analysis showed that 1-NA-PP1 was considerably more potent and showed distinct substituent effects at the pyrazolopyrimidine core. 1-NA-PP1 was cell-active, and potently blocked prostate cancer cell proliferation by inducing G2/M arrest. It also potently blocked the migration and invasion of prostate cancer cells, demonstrating promising anticancer activities on multiple fronts. Overexpression of PKD1 or PKD3 almost completely reversed the growth arrest and the inhibition of tumor cell invasion caused by 1-NA-PP1, indicating that its anti-proliferative and anti-invasive activities were mediated through the inhibition of PKD. Interestingly, a 12-fold increase in sensitivity to 1-NA-PP1 could be achieved by engineering a gatekeeper mutation in the active site of PKD1, suggesting that 1-NA-PP1 could be paired with the analog-sensitive PKD1(M659G for dissecting PKD-specific functions and signaling pathways in various biological systems.

  2. The discovery of novel 3-(pyrazin-2-yl)-1H-indazoles as potent pan-Pim kinase inhibitors.

    Science.gov (United States)

    Wang, Hui-Ling; Cee, Victor J; Chavez, Frank; Lanman, Brian A; Reed, Anthony B; Wu, Bin; Guerrero, Nadia; Lipford, J Russell; Sastri, Christine; Winston, Jeff; Andrews, Kristin L; Huang, Xin; Lee, Matthew R; Mohr, Christopher; Xu, Yang; Zhou, Yihong; Tasker, Andrew S

    2015-02-15

    The three Pim kinases are a small family of serine/threonine kinases regulating several signaling pathways that are fundamental to tumorigenesis. As such, the Pim kinases are a very attractive target for pharmacological inhibition in cancer therapy. Herein, we describe our efforts toward the development of a potent, pan-Pim inhibitor. The synthesis and hit-to-lead SAR development from a 3-(pyrazin-2-yl)-1H-indazole derived hit 2 to the identification of a series of potent, pan-Pim inhibitors such as 13o are described.

  3. A high-throughput, nonisotopic, competitive binding assay for kinases using nonselective inhibitor probes (ED-NSIP).

    Science.gov (United States)

    Vainshtein, Inna; Silveria, Scott; Kaul, Poonam; Rouhani, Riaz; Eglen, Richard M; Wang, John

    2002-12-01

    A novel competitive binding assay for protein kinase inhibitors has been developed for high-throughput screening (HTS). Unlike functional kinase assays, which are based on detection of substrate phosphorylation by the enzyme, this novel method directly measures the binding potency of compounds to the kinase ATP binding site through competition with a conjugated binding probe. The binding interaction is coupled to a signal amplification system based on complementation of beta-galactosidase enzyme fragments, a homogeneous, nonisotopic assay technology platform developed by DiscoveRx Corp. In the present study, staurosporine, a potent, nonselective kinase inhibitor, was chemically conjugated to a small fragment of beta-galactosidase (termed ED-SS). This was used as the binding probe to the kinase ATP binding pocket. The binding potencies of several inhibitors with diverse structures were assessed by displacement of ED-SS from the kinase. The assay format was specifically evaluated with GSK3alpha, an enzyme previously screened in a radioactive kinase assay (i.e., measurement of [(33)P]-gamma-ATP incorporation into the kinase peptide substrate). Under optimized assay conditions, nonconjugated staurosporine inhibited ED-SS binding in a concentration-dependent manner with an apparent potency (IC(50)) of 11 nM, which was similar to the IC(50) value determined in a radioactive assay. Furthermore, 9 kinase inhibitors with diverse structures, previously identified from chemical compound library screening, were screened using the competitive binding assay. The potencies in the binding assay were in very good agreement with those obtained previously in the isotopic functional activity assay. The binding assay was adapted for automated HTS using selected compound libraries in a 384-well microtiter plate format. The HTS assay was observed to be highly robust and reproducible (Z' factors > 0.7) with high interassay precision (R(2) > 0.96). Interference of compounds with the beta

  4. Determination of human serum alpha1-acid glycoprotein and albumin binding of various marketed and preclinical kinase inhibitors.

    Science.gov (United States)

    Zsila, Ferenc; Fitos, Ilona; Bencze, Gyula; Kéri, György; Orfi, László

    2009-01-01

    There are about 380 protein kinase inhibitors in drug development as of today and 15 drugs have been marketed already for the treatment of cancer. This time 139 validated kinase targets are in the focus of drug research of pharmaceutical companies and big efforts are made for the development of new, druglike kinase inhibitors. Plasma protein binding is an important factor of the ADME profiling of a drug compound. Human serum albumin (HSA) and alpha(1)-acid glycoprotein (AAG) are the most relevant drug carriers in blood plasma. Since previous literature data indicated that AAG is the principal plasma binding component of some kinase inhibitors the present work focuses on the comprehensive evaluation of AAG binding of a series of marketed and experimental kinase inhibitors by using circular dichroism (CD) spectroscopy approach. HSA binding was also evaluated by affinity chromatography. Protein binding interactions of twenty-six kinase inhibitors are characterized. The contribution of AAG and HSA binding data to the pharmacokinetic profiles of the investigated therapeutic agents is discussed. Structural, biological and drug binding properties of AAG as well as the applicability of the CD method in studying drug-protein binding interactions are also briefly reviewed.

  5. QSAR based docking studies of marine algal anticancer compounds as inhibitors of protein kinase B (PKBβ).

    Science.gov (United States)

    Davis, G Dicky John; Vasanthi, A Hannah Rachel

    2015-08-30

    Marine algae are prolific source of bioactive secondary metabolites and are found to be active against different cancer cell lines. QSAR studies will explicate the significance of a particular class of descriptor in eliciting anticancer activity against a cancer type. Marine algal compounds showing anticancer activity against six different cancer cell lines namely MCF-7, A431, HeLa, HT-29, P388 and A549 taken from Seaweed metabolite database were subjected to comprehensive QSAR modeling studies. A hybrid-GA (genetic algorithm) optimization technique for descriptor space reduction and multiple linear regression analysis (MLR) approach was used as fitness functions. Cell lines HeLa and MCF-7 showed good statistical quality (R(2)∼0.75, Q(2)∼0.65) followed by A431, HT29 and P388 cell lines with reasonable statistical values (R(2)∼0.70, Q(2)∼0.60). The models developed were interpretable, with good statistical and predictive significance. Molecular descriptor analyses revealed that Baumann's alignment-independent topological descriptors had a major role in variation of activity along with other descriptors. Incidentally, earlier QSAR analysis on a variety of chemically diverse PKBα inhibitors revealed Baumann's alignment-independent topological descriptors that differentiated the molecules binding to Protein kinase B (PKBα) kinase or PH domain, hence a docking study of two crystal structures of PKBβ was performed for identification of novel ATP-competitive inhibitors of PKBβ. Five compounds had a good docking score and Callophycin A showed better ligand efficiency than other PKBβ inhibitors. Furthermore in silico pharmacokinetic and toxicity studies also showed that Callophycin A had a high drug score (0.85) compared to the other inhibitors. These results encourages discovering novel inhibitors for cancer therapeutic targets by screening metabolites from marine algae.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2009-11-20

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

  7. Effects of 4 multitargeted receptor tyrosine kinase inhibitors on regional hemodynamics in conscious, freely moving rats

    Science.gov (United States)

    Carter, Joanne J.; Fretwell, Laurice V.; Woolard, Jeanette

    2017-01-01

    VEGF inhibitors, including receptor tyrosine kinase inhibitors, are used as adjunct therapies in a number of cancer treatments. An emerging issue with these drugs is that most cause hypertension. To gain insight into the physiological mechanisms involved, we evaluated their regional hemodynamic effects in conscious rats. Male Sprague Dawley rats (350–450 g) were chronically implanted with pulsed Doppler flow probes (renal and mesenteric arteries, and the descending abdominal aorta) and catheters (jugular vein, peritoneal cavity, and distal abdominal aorta). Regional hemodynamics were measured over 4 d, before and after daily administration of cediranib (3 and 6 mg/kg, 3 and 6 mg/kg/h for 1 h, i.v.), sorafenib (10 and 20 mg/kg, 10 and 20 mg kg/h for 1 h, i.v.), pazopanib (30 and100 mg/kg, i.p.), or vandetanib (12.5 and 25 mg/kg, i.p.). All drugs evoked significant increases (P phentolamine and propranolol (each 1 mg/kg/h), suggesting a need for new strategies to overcome them.—Carter, J. J., Fretwell, L. V., Woolard, J. Effects of 4 multitargeted receptor tyrosine kinase inhibitors on regional hemodynamics in conscious, freely moving rats. PMID:27986807

  8. Molecular docking and NMR binding studies to identify novel inhibitors of human phosphomevalonate kinase

    Energy Technology Data Exchange (ETDEWEB)

    Boonsri, Pornthip [Chemical Proteomics Facility at Marquette, Department of Chemistry, Marquette University, Milwaukee, WI 53201 (United States); Department of Chemistry, NANOTEC Center of Nanotechnology, National Nanotechnology Center, Faculty of Science, Kasetsart University, Bangkok 10900 (Thailand); Neumann, Terrence S.; Olson, Andrew L.; Cai, Sheng [Chemical Proteomics Facility at Marquette, Department of Chemistry, Marquette University, Milwaukee, WI 53201 (United States); Herdendorf, Timothy J.; Miziorko, Henry M. [Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri-Kansas City, Kansas City, MO 64110 (United States); Hannongbua, Supa [Department of Chemistry, NANOTEC Center of Nanotechnology, National Nanotechnology Center, Faculty of Science, Kasetsart University, Bangkok 10900 (Thailand); Sem, Daniel S., E-mail: daniel.sem@cuw.edu [Chemical Proteomics Facility at Marquette, Department of Chemistry, Marquette University, Milwaukee, WI 53201 (United States)

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer Natural and synthetic inhibitors of human phosphomevalonate kinase identified. Black-Right-Pointing-Pointer Virtual screening yielded a hit rate of 15%, with inhibitor K{sub d}'s of 10-60 {mu}M. Black-Right-Pointing-Pointer NMR studies indicate significant protein conformational changes upon binding. -- Abstract: Phosphomevalonate kinase (PMK) phosphorylates mevalonate-5-phosphate (M5P) in the mevalonate pathway, which is the sole source of isoprenoids and steroids in humans. We have identified new PMK inhibitors with virtual screening, using autodock. Promising hits were verified and their affinity measured using NMR-based {sup 1}H-{sup 15}N heteronuclear single quantum coherence (HSQC) chemical shift perturbation and fluorescence titrations. Chemical shift changes were monitored, plotted, and fitted to obtain dissociation constants (K{sub d}). Tight binding compounds with K{sub d}'s ranging from 6-60 {mu}M were identified. These compounds tended to have significant polarity and negative charge, similar to the natural substrates (M5P and ATP). HSQC cross peak changes suggest that binding induces a global conformational change, such as domain closure. Compounds identified in this study serve as chemical genetic probes of human PMK, to explore pharmacology of the mevalonate pathway, as well as starting points for further drug development.

  9. De novo design of protein kinase inhibitors by in silico identification of hinge region-binding fragments.

    Science.gov (United States)

    Urich, Robert; Wishart, Grant; Kiczun, Michael; Richters, André; Tidten-Luksch, Naomi; Rauh, Daniel; Sherborne, Brad; Wyatt, Paul G; Brenk, Ruth

    2013-05-17

    Protein kinases constitute an attractive family of enzyme targets with high relevance to cell and disease biology. Small molecule inhibitors are powerful tools to dissect and elucidate the function of kinases in chemical biology research and to serve as potential starting points for drug discovery. However, the discovery and development of novel inhibitors remains challenging. Here, we describe a structure-based de novo design approach that generates novel, hinge-binding fragments that are synthetically feasible and can be elaborated to small molecule libraries. Starting from commercially available compounds, core fragments were extracted, filtered for pharmacophoric properties compatible with hinge-region binding, and docked into a panel of protein kinases. Fragments with a high consensus score were subsequently short-listed for synthesis. Application of this strategy led to a number of core fragments with no previously reported activity against kinases. Small libraries around the core fragments were synthesized, and representative compounds were tested against a large panel of protein kinases and subjected to co-crystallization experiments. Each of the tested compounds was active against at least one kinase, but not all kinases in the panel were inhibited. A number of compounds showed high ligand efficiencies for therapeutically relevant kinases; among them were MAPKAP-K3, SRPK1, SGK1, TAK1, and GCK for which only few inhibitors are reported in the literature.

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

    Directory of Open Access Journals (Sweden)

    Richard L. Schroeder

    2014-09-01

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

  11. Inhibitor of apoptosis signal-regulating kinase 1 protects against acetaminophen-induced liver injury

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Yuchao; Ramachandran, Anup [Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Breckenridge, David G.; Liles, John T. [Department of Biology, Gilead Sciences, Inc., Foster City, CA (United States); Lebofsky, Margitta [Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Farhood, Anwar [Department of Pathology, St. David' s North Austin Medical Center, Austin, TX 78756 (United States); Jaeschke, Hartmut, E-mail: hjaeschke@kumc.edu [Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States)

    2015-07-01

    Metabolic activation and oxidant stress are key events in the pathophysiology of acetaminophen (APAP) hepatotoxicity. The initial mitochondrial oxidative stress triggered by protein adduct formation is amplified by c-jun-N-terminal kinase (JNK), resulting in mitochondrial dysfunction and ultimately cell necrosis. Apoptosis signal-regulating kinase 1 (ASK1) is considered the link between oxidant stress and JNK activation. The objective of the current study was to assess the efficacy and mechanism of action of the small-molecule ASK1 inhibitor GS-459679 in a murine model of APAP hepatotoxicity. APAP (300 mg/kg) caused extensive glutathione depletion, JNK activation and translocation to the mitochondria, oxidant stress and liver injury as indicated by plasma ALT activities and area of necrosis over a 24 h observation period. Pretreatment with 30 mg/kg of GS-459679 almost completely prevented JNK activation, oxidant stress and injury without affecting the metabolic activation of APAP. To evaluate the therapeutic potential of GS-459679, mice were treated with APAP and then with the inhibitor. Given 1.5 h after APAP, GS-459679 was still protective, which was paralleled by reduced JNK activation and p-JNK translocation to mitochondria. However, GS-459679 treatment was not more effective than N-acetylcysteine, and the combination of GS-459679 and N-acetylcysteine exhibited similar efficacy as N-acetylcysteine monotherapy, suggesting that GS-459769 and N-acetylcysteine affect the same pathway. Importantly, inhibition of ASK1 did not impair liver regeneration as indicated by PCNA staining. In conclusion, the ASK1 inhibitor GS-459679 protected against APAP toxicity by attenuating JNK activation and oxidant stress in mice and may have therapeutic potential for APAP overdose patients. - Highlights: • Two ASK1 inhibitors protected against acetaminophen-induced liver injury. • The ASK1 inhibitors protect when used as pre- or post-treatment. • Protection by ASK1 inhibitor is

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

    Science.gov (United States)

    Breccia, Massimo; Molica, Matteo; Alimena, Giuliana

    2014-12-01

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

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

    DEFF Research Database (Denmark)

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

    2008-01-01

    There is strong clinical evidence that trastuzumab, a monoclonal antibody targeting the human epidermal growth factor receptor (HER) two tyrosine kinase receptor, is an important component of first-line treatment of patients with HER2-positive metastatic breast cancer. In particular the combination...... with taxanes and vinorelbine has been established. In the preoperative setting inclusion of trastuzumab has significantly increased the pathological complete response rate. Results from large phase III trials evaluating adjuvant therapy in HER2-positive early breast cancer indicate that the addition...... of trastuzumab to chemotherapy improves disease-free and overall survival. The use of lapatinib, a dual tyrosine kinase inhibitor of both HER1 and HER2, in combination with capecitabine in the second-line treatment of HER2-positive patients with metastatic breast cancer previously treated with trastuzumab has...

  14. Clinical pharmacology of tyrosine kinase inhibitors becoming generic drugs: the regulatory perspective.

    Science.gov (United States)

    Eckstein, Niels; Röper, Lea; Haas, Bodo; Potthast, Henrike; Hermes, Ulrike; Unkrig, Christoph; Naumann-Winter, Frauke; Enzmann, Harald

    2014-02-07

    Over the last decades, billions have been spent and huge efforts have been taken in basic and clinical cancer research [CA Cancer J Clin63:11-30]. About a decade ago, the arms race between drugs and cancer cells reached a new level by introduction of tyrosine kinase inhibitors (TKI) into pharmacological anti-cancer therapy. According to their molecular mechanism of action, TKI in contrast to so-called "classic" or "conventional" cytostatics belong to the group of targeted cancer medicines, characterized by accurately fitting with biological structures (i.e. active centers of kinases). Numerous (partly orphan) indications are covered by this new class of substances. Approximately ten years after the first substances of this class of medicines were authorized, patent protection will end within the next years. The following article covers clinical meaning and regulatory status of anti-cancer TKI and gives an outlook to what is expected from the introduction of generic anti-cancer TKI.

  15. (R)-roscovitine, a cyclin-dependent kinase inhibitor, enhances tonic GABA inhibition in rat hippocampus.

    Science.gov (United States)

    Ivanov, A; Tyzio, R; Zilberter, Y; Ben-Ari, Yehezkel

    2008-10-02

    Pharmacological agents that mediate a persistent GABAergic conductance are of considerable interest for treatment of epilepsy. (R)-roscovitine is a membrane permeable cyclin-dependent kinase inhibitor, designed to block cell division. It is currently undergoing a phase II clinical trial as an anticancer drug. We show that (R)-roscovitine increases a tonic GABA-mediated current in rat hippocampal neurons. This enhanced tonic current appears independent of synaptic GABA release and requires functional transmembrane GABA transport. The effect of (R)-roscovitine is associated with neither modification of GABAA receptors nor protein kinase activity, but is associated with a significant increase in intracellular GABA concentration in hippocampal GABAergic neurons. (R)-roscovitine-induced tonic inhibition significantly suppresses spontaneous spiking activity of hippocampal pyramidal cells. Therefore, (R)-roscovitine is a potent modulator of neuronal activity in rat hippocampus and may provide a tool for preventing paroxysmal activity.

  16. Discovery and optimization of pyrrolo[1,2-a]pyrazinones leads to novel and selective inhibitors of PIM kinases.

    Science.gov (United States)

    Casuscelli, Francesco; Ardini, Elena; Avanzi, Nilla; Casale, Elena; Cervi, Giovanni; D'Anello, Matteo; Donati, Daniele; Faiardi, Daniela; Ferguson, Ronald D; Fogliatto, Gianpaolo; Galvani, Arturo; Marsiglio, Aurelio; Mirizzi, Danilo G; Montemartini, Marisa; Orrenius, Christian; Papeo, Gianluca; Piutti, Claudia; Salom, Barbara; Felder, Eduard R

    2013-12-01

    A novel series of PIM inhibitors was derived from a combined effort in natural product-inspired library generation and screening. The novel pyrrolo[1,2-a]pyrazinones initial hits are inhibitors of PIM isoforms with IC50 values in the low micromolar range. The application of a rational optimization strategy, guided by the determination of the crystal structure of the complex in the kinase domain of PIM1 with compound 1, led to the discovery of compound 15a, which is a potent PIM kinases inhibitor exhibiting excellent selectivity against a large panel of kinases, representative of each family. The synthesis, structure-activity relationship studies, and pharmacokinetic data of compounds from this inhibitor class are presented herein. Furthermore, the cellular activities including inhibition of cell growth and modulation of downstream targets are also described.

  17. Discovery of a novel class of targeted kinase inhibitors that blocks protein kinase C signaling and ameliorates retinal vascular leakage in a diabetic rat model.

    Science.gov (United States)

    Grant, Stephan; Tran, Phong; Zhang, Qin; Zou, Aihua; Dinh, Dac; Jensen, Jordan; Zhou, Sue; Kang, Xiaolin; Zachwieja, Joseph; Lippincott, John; Liu, Kevin; Johnson, Sarah Ludlum; Scales, Stephanie; Yin, Chunfeng; Nukui, Seiji; Stoner, Chad; Prasanna, Ganesh; Lafontaine, Jennifer; Wells, Peter; Li, Hui

    2010-02-10

    Protein kinase C (PKC) family members such as PKCbetaII may become activated in the hyperglycemic state associated with diabetes. Preclinical and clinical data implicate aberrant PKC activity in the development of diabetic microvasculature abnormalities. Based on this potential etiological role for PKC in diabetic complications, several therapeutic PKC inhibitors have been investigated in clinical trials for the treatment of diabetic patients. In this report, we present the discovery and preclinical evaluation of a novel class of 3-amino-pyrrolo[3,4-c]pyrazole derivatives as inhibitors of PKC that are structurally distinct from the prototypical indolocarbazole and bisindolylmaleimide PKC inhibitors. From this pyrrolo-pyrazole series, several compounds were identified from biochemical assays as potent, ATP-competitive inhibitors of PKC activity with high specificity for PKC over other protein kinases. These compounds were also found to block PKC signaling activity in multiple cellular functional assays. PF-04577806, a representative from this series, inhibited PKC activity in retinal lysates from diabetic rats stimulated with phorbol myristate acetate. When orally administered, PF-04577806 showed good exposure in the retina of diabetic Long-Evans rats and ameliorated retinal vascular leakage in a streptozotocin-induced diabetic rat model. These novel PKC inhibitors represent a promising new class of targeted protein kinase inhibitors with potential as therapeutic agents for the treatment of patients with diabetic microvascular complications.

  18. Protein Kinase C Inhibitors as Modulators of Vascular Function and Their Application in Vascular Disease

    Directory of Open Access Journals (Sweden)

    Raouf A. Khalil

    2013-03-01

    Full Text Available Blood pressure (BP is regulated by multiple neuronal, hormonal, renal and vascular control mechanisms. Changes in signaling mechanisms in the endothelium, vascular smooth muscle (VSM and extracellular matrix cause alterations in vascular tone and blood vessel remodeling and may lead to persistent increases in vascular resistance and hypertension (HTN. In VSM, activation of surface receptors by vasoconstrictor stimuli causes an increase in intracellular free Ca2+ concentration ([Ca2+]i, which forms a complex with calmodulin, activates myosin light chain (MLC kinase and leads to MLC phosphorylation, actin-myosin interaction and VSM contraction. Vasoconstrictor agonists could also increase the production of diacylglycerol which activates protein kinase C (PKC. PKC is a family of Ca2+-dependent and Ca2+-independent isozymes that have different distributions in various blood vessels, and undergo translocation from the cytosol to the plasma membrane, cytoskeleton or the nucleus during cell activation. In VSM, PKC translocation to the cell surface may trigger a cascade of biochemical events leading to activation of mitogen-activated protein kinase (MAPK and MAPK kinase (MEK, a pathway that ultimately increases the myofilament force sensitivity to [Ca2+]i, and enhances actin-myosin interaction and VSM contraction. PKC translocation to the nucleus may induce transactivation of various genes and promote VSM growth and proliferation. PKC could also affect endothelium-derived relaxing and contracting factors as well as matrix metalloproteinases (MMPs in the extracellular matrix further affecting vascular reactivity and remodeling. In addition to vasoactive factors, reactive oxygen species, inflammatory cytokines and other metabolic factors could affect PKC activity. Increased PKC expression and activity have been observed in vascular disease and in certain forms of experimental and human HTN. Targeting of vascular PKC using PKC inhibitors may function in

  19. Spleen tyrosine kinase inhibitors for rheumatoid arthritis: where are we now?

    Science.gov (United States)

    Scott, Ian C; Scott, David L

    2014-03-01

    The development of small-molecule inhibitors of inflammatory cascade signaling kinases offers a potential approach to treating rheumatoid arthritis (RA). Spleen tyrosine kinase is one such tyrosine kinase. Recent research efforts have focussed on the development and testing of a spleen tyrosine kinase inhibitor, fostamatinib. We reviewed the results of the clinical trials of fostamatinib in RA with the aim of outlining its clinical efficacy and the nature and frequency of its main adverse events. To date, this drug has been evaluated in over 3,200 RA patients enrolled in three phase II, one phase IIb and three phase III trials. These studies showed fostamatinib was effective. In four trials in which patients received 100 mg twice daily, fostamatinib reduced inflammatory synovitis; the relative risks of achieving American College of Rheumatology Responder rates compared with placebo in the combined studies ranged from 1.6 for 20 % of responders to 3.7 for 70 % of responders. There was a similar relative risk of achieving a clinically meaningful reduction in disability of 1.6 for the chance of patients achieving a reduction in health assessment questionnaire scores of 0.22 or more. Three of the trials examined the impact of fostamatinib on erosive radiographic damage using changes in the modified total Sharp score. None of them provided any evidence for a significant effect of fostamatinib on erosive damage over 6 months. All the trials included descriptions of adverse events. Hypertension was common, involving over 40 % of patients treated. Other common adverse events included diarrhoea, neutropenia and increases in hepatic enzyme levels. Some patients developed infections. On the conclusion of the phase III trials, one of the main pharmaceutical sponsors decided not to further develop fostamatinib for RA.

  20. ALK kinase domain mutations in primary anaplastic large cell lymphoma: consequences on NPM-ALK activity and sensitivity to tyrosine kinase inhibitors.

    Directory of Open Access Journals (Sweden)

    Federica Lovisa

    Full Text Available ALK inhibitor crizotinib has shown potent antitumor activity in children with refractory Anaplastic Large Cell Lymphoma (ALCL and the opportunity to include ALK inhibitors in first-line therapies is oncoming. However, recent studies suggest that crizotinib-resistance mutations may emerge in ALCL patients. In the present study, we analyzed ALK kinase domain mutational status of 36 paediatric ALCL patients at diagnosis to identify point mutations and gene aberrations that could impact on NPM-ALK gene expression, activity and sensitivity to small-molecule inhibitors. Amplicon ultra-deep sequencing of ALK kinase domain detected 2 single point mutations, R335Q and R291Q, in 2 cases, 2 common deletions of exon 23 and 25 in all the patients, and 7 splicing-related INDELs in a variable number of them. The functional impact of missense mutations and INDELs was evaluated. Point mutations were shown to affect protein kinase activity, signalling output and drug sensitivity. INDELs, instead, generated kinase-dead variants with dominant negative effect on NPM-ALK kinase, in virtue of their capacity of forming non-functional heterocomplexes. Consistently, when co-expressed, INDELs increased crizotinib inhibitory activity on NPM-ALK signal processing, as demonstrated by the significant reduction of STAT3 phosphorylation. Functional changes in ALK kinase activity induced by both point mutations and structural rearrangements were resolved by molecular modelling and dynamic simulation analysis, providing novel insights into ALK kinase domain folding and regulation. Therefore, these data suggest that NPM-ALK pre-therapeutic mutations may be found at low frequency in ALCL patients. These mutations occur randomly within the ALK kinase domain and affect protein activity, while preserving responsiveness to crizotinib.

  1. ALK kinase domain mutations in primary anaplastic large cell lymphoma: consequences on NPM-ALK activity and sensitivity to tyrosine kinase inhibitors.

    Science.gov (United States)

    Lovisa, Federica; Cozza, Giorgio; Cristiani, Andrea; Cuzzolin, Alberto; Albiero, Alessandro; Mussolin, Lara; Pillon, Marta; Moro, Stefano; Basso, Giuseppe; Rosolen, Angelo; Bonvini, Paolo

    2015-01-01

    ALK inhibitor crizotinib has shown potent antitumor activity in children with refractory Anaplastic Large Cell Lymphoma (ALCL) and the opportunity to include ALK inhibitors in first-line therapies is oncoming. However, recent studies suggest that crizotinib-resistance mutations may emerge in ALCL patients. In the present study, we analyzed ALK kinase domain mutational status of 36 paediatric ALCL patients at diagnosis to identify point mutations and gene aberrations that could impact on NPM-ALK gene expression, activity and sensitivity to small-molecule inhibitors. Amplicon ultra-deep sequencing of ALK kinase domain detected 2 single point mutations, R335Q and R291Q, in 2 cases, 2 common deletions of exon 23 and 25 in all the patients, and 7 splicing-related INDELs in a variable number of them. The functional impact of missense mutations and INDELs was evaluated. Point mutations were shown to affect protein kinase activity, signalling output and drug sensitivity. INDELs, instead, generated kinase-dead variants with dominant negative effect on NPM-ALK kinase, in virtue of their capacity of forming non-functional heterocomplexes. Consistently, when co-expressed, INDELs increased crizotinib inhibitory activity on NPM-ALK signal processing, as demonstrated by the significant reduction of STAT3 phosphorylation. Functional changes in ALK kinase activity induced by both point mutations and structural rearrangements were resolved by molecular modelling and dynamic simulation analysis, providing novel insights into ALK kinase domain folding and regulation. Therefore, these data suggest that NPM-ALK pre-therapeutic mutations may be found at low frequency in ALCL patients. These mutations occur randomly within the ALK kinase domain and affect protein activity, while preserving responsiveness to crizotinib.

  2. Switch control pocket inhibitors of p38-MAP kinase. Durable type II inhibitors that do not require binding into the canonical ATP hinge region

    Energy Technology Data Exchange (ETDEWEB)

    Ahn, Yu Mi; Clare, Michael; Ensinger, Carol L.; Hood, Molly M.; Lord, John W.; Lu, Wei-Ping; Miller, David F.; Patt, William C.; Smith, Bryan D.; Vogeti, Lakshminarayana; Kaufman, Michael D.; Petillo, Peter A.; Wise, Scott C.; Abendroth, Jan; Chun, Lawrence; Clark, Robin; Feese, Michael; Kim, Hidong; Stewart, Lance; Flynn, Daniel L. (Deciphera); (Emerald); (Cocrystal)

    2012-01-20

    Switch control pocket inhibitors of p38-alpha kinase are described. Durable type II inhibitors were designed which bind to arginines (Arg67 or Arg70) that function as key residues for mediating phospho-threonine 180 dependant conformational fluxing of p38-alpha from an inactive type II state to an active type I state. Binding to Arg70 in particular led to potent inhibitors, exemplified by DP-802, which also exhibited high kinase selectivity. Binding to Arg70 obviated the requirement for binding into the ATP Hinge region. X-ray crystallography revealed that DP-802 and analogs induce an enhanced type II conformation upon binding to either the unphosphorylated or the doubly phosphorylated form of p38-alpha kinase.

  3. Switch control pocket inhibitors of p38-MAP kinase. Durable type II inhibitors that do not require binding into the canonical ATP hinge region.

    Science.gov (United States)

    Ahn, Yu Mi; Clare, Michael; Ensinger, Carol L; Hood, Molly M; Lord, John W; Lu, Wei-Ping; Miller, David F; Patt, William C; Smith, Bryan D; Vogeti, Lakshminarayana; Kaufman, Michael D; Petillo, Peter A; Wise, Scott C; Abendroth, Jan; Chun, Lawrence; Clark, Robin; Feese, Michael; Kim, Hidong; Stewart, Lance; Flynn, Daniel L

    2010-10-01

    Switch control pocket inhibitors of p38-alpha kinase are described. Durable type II inhibitors were designed which bind to arginines (Arg67 or Arg70) that function as key residues for mediating phospho-threonine 180 dependant conformational fluxing of p38-alpha from an inactive type II state to an active type I state. Binding to Arg70 in particular led to potent inhibitors, exemplified by DP-802, which also exhibited high kinase selectivity. Binding to Arg70 obviated the requirement for binding into the ATP Hinge region. X-ray crystallography revealed that DP-802 and analogs induce an enhanced type II conformation upon binding to either the unphosphorylated or the doubly phosphorylated form of p38-alpha kinase.

  4. Efficacy of HER2-targeted therapy in metastatic breast cancer. Monoclonal antibodies and tyrosine kinase inhibitors

    DEFF Research Database (Denmark)

    Nielsen, Dorte L; Kümler, Iben; Palshof, Jesper Andreas;

    2013-01-01

    Therapies targeting the human epidermal growth factor receptor (HER) 2 are effective in metastatic breast cancer (MBC). We review the efficacy of HER2-directed therapies, focussing on monoclonal antibodies and tyrosine kinase inhibitors targeting HER2 that have been tested in phase II-III studies...... to those obtained for capecitabine plus lapatinib (48%), continuing trastuzumab in combination with capecitabine (48%), pertuzumab plus trastuzumab (24%), and neratinib (24%). Strategies combining multiple HER2-directed therapies might yield additive or synergistic effects and lead to improved outcome...

  5. Pregnancies in patients with chronic myeloid leukemia treated with tyrosine kinase inhibitor.

    Science.gov (United States)

    Zhou, Li; You, Jian-Hua; Wu, Wen; Li, Jun-Min; Shen, Zhi-Xiang; Wang, Ai-Hua

    2013-10-01

    We presented our experience in chronic myeloid leukemia (CML) patients who conceived children and/or became pregnant while receiving tyrosine kinase inhibitor (TKI). Among 7 male patients, 7 pregnancies resulted in the birth of 7 healthy babies. Among 18 female patients, 8 ended in elective abortion; 3 had spontaneous abortion, and 7 carried to term, resulting in the birth of 8 healthy babies. All children have normal growth and development. All patients remain in TKI therapy and in good response. It is suggested that female patients are advised to practice adequate contraception. No special precautions apply for male patients receiving TKI.

  6. Influence of protein kinase C inhibitor in phagocytosis activity toward Candida sp

    Directory of Open Access Journals (Sweden)

    Adiprayitno Adiprayitno

    2001-09-01

    Full Text Available Protein kinase C isoenzyme family that expresses in all of cells plays a pivotal role in the signal transduction pathway of a variety of hormones, cytokines, neurotransmitter, and growth factors. The immunity against Candida sp is mainly mediated and performed by the T cells and macrophages. The objective of this experiment is to know the influence the protein kinase C inhibitor - bisindolylmaleimides in phagocytosis activity toward Candida sp. The culture of peritoneal macrophage derived from BALB/c mice are treated with bisindolylmaleimides as a protein kinase C inhibitor concentration varied from 5 ng/ml to 100 ng/ml for as long as 10 minute. Then the Candida sp added is observed after every 30 minute for as long as 120 minute. As the experimental design is used the method of factorial and orthogonal polynomial. The data consisting the length of pseudopodia and the number of Candida sp which are phagocytosed are analyzed applying the Anova. One Way Anova to show the differences of each manipulation, the Two Way Anova to show the interaction of manipulations and the Student's t Test to show the differences with control. Statistical test show significant differences on the length of pseudopodia, and phagocytosed Candida sp, at different bisindolylmaleimides concentration (p<0.001 and different observed time (p<0.001. The data show a significant interaction between the bisindolylmaleimides concentration and observed time (p<0.001. The higher the bisindolylmaleimides concentration, the earlier the observed time, the much number the protein kinase C are going inactive and the shorter the length of pseudopodia or the lower the macrophages phagocytic activity toward Candida sp. The result of this experiment indicates that bisindolylmaleimides can inhibit the macrophage mobility and phagocytic activity toward Candida sp. Further experiment in protein kinase C, especially in macrophage, is suggested. (Med J Indones 2001; 10: 150

  7. Molecular Shape Analysis-Guided Virtual Screening Platform for Adenosine Kinase Inhibitors

    Science.gov (United States)

    Bhutoria, Savita; Das, Ballari; Ghoshal, Nanda

    2016-01-01

    We propose a new application of molecular shape descriptors in hierarchical selection during virtual screening (VS). Here, a structure-based pharmacophore and docking-guided VS protocol have been evolved to identify inhibitors against adenosine kinase (AK). The knowledge gained on the shape requirements has been extrapolated in classifying active and inactive molecules against this target. This classification enabled us to pick the appropriate ligand conformation in the binding site. We have suggested a set of hierarchical filters for VS, from a simple molecular shape analysis (MSA) descriptor-based recursive models to docking scores. This approach permits a systematic study to understand the importance of spatial requirements and limitations for inhibitors against AK. Finally, the guidelines on how to select compounds for AK to achieve success have been highlighted. The utility of this approach has been suggested by giving an example of database screening for plausible active compounds. PMID:27478367

  8. Further investigation of inhibitors of MRSA pyruvate kinase: Towards the conception of novel antimicrobial agents.

    Science.gov (United States)

    Labrière, Christophe; Gong, Huansheng; Finlay, B Brett; Reiner, Neil E; Young, Robert N

    2017-01-05

    Several novel series of compounds were synthesized and evaluated as inhibitors of methicillin-resistant Staphylococcus aureus (MRSA) pyruvate kinase (PK). PK has been identified as a highly interconnected essential 'hub' protein in MRSA, with structural features distinct from the human homologs which makes it a novel antimicrobial target. Several MRSA PK inhibitors (including the hydrazide 1) were identified using in silico screening combined with enzyme assays and were found to be selective for bacterial enzyme compared to human PK isoforms. Structure-activity relationship (SAR) studies were carried out on the replacement of the hydrazide linker with 3-atoms, 2-atoms and 0-atom linkers and led us to discover more potent compounds with enzyme inhibiting activities in the low nanomolar range and some were found to effectively inhibit bacteria growth in culture with minimum inhibitory concentrations (MIC) as low as 1 μg/mL.

  9. Fragment-Based and Structure-Guided Discovery and Optimization of Rho Kinase Inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Li, Rongshi; Martin, Mathew P.; Liu, Yan; Wang, Binglin; Patel, Ronil A.; Zhu, Jin-Yi; Sun, Nan; Pireddu, Roberta; Lawrence, Nicholas J.; Li, Jiannong; Haura, Eric B.; Sung, Shen-Shu; Guida, Wayne C.; Schonbrunn, Ernst; Sebti, Said M. (Moffitt)

    2012-05-14

    Using high concentration biochemical assays and fragment-based screening assisted by structure-guided design, we discovered a novel class of Rho-kinase inhibitors. Compound 18 was equipotent for ROCK1 (IC{sub 50} = 650 nM) and ROCK2 (IC{sub 50} = 670 nM), whereas compound 24 was more selective for ROCK2 (IC{sub 50} = 100 nM) over ROCK1 (IC{sub 50} = 1690 nM). The crystal structure of the compound 18-ROCK1 complex revealed that 18 is a type 1 inhibitor that binds the hinge region in the ATP binding site. Compounds 18 and 24 inhibited potently the phosphorylation of the ROCK substrate MLC2 in intact human breast cancer cells.

  10. Pyrrolo[2,3-b]pyridine derivatives as potent Bruton's tyrosine kinase inhibitors.

    Science.gov (United States)

    Zhao, Xinge; Huang, Wei; Wang, Yazhou; Xin, Minhang; Jin, Qiu; Cai, Jianfeng; Tang, Feng; Zhao, Yong; Xiang, Hua

    2015-08-01

    A series of pyrrolo[2,3-b]pyridine-based derivatives were designed as potent Bruton's tyrosine kinase (BTK) inhibitors by using a scaffold-hopping strategy. Structure-activity relationship studies identified five compounds (3n, 3p, 3q, 3r, and 3s) with IC50 of less than 10nM in BTK enzyme assay and five compounds (3m, 3n, 3o, 3p, and 3t) with IC50 of less than 20 nM in Ramos cell assay. As one of the most potent inhibitors, compound 3p exhibited superior activity to that of compound 1 (RN486) and pyrrolo[2,3-d]pyrimidine derivative 2 in both BTK enzymatic (IC50=6.0 nM) and cellular inhibition (IC50=14 nM) assays. In addition, 3p displayed favorable overall pharmacokinetic profiles compared with 1 and 2.

  11. Attenuation of endothelin-1-induced calcium response by tyrosine kinase inhibitors in vascular smooth muscle cells.

    Science.gov (United States)

    Liu, C Y; Sturek, M

    1996-06-01

    Although tyrosine kinases play an important role in cell growth and have been implicated in regulation of smooth muscle contraction, their role in agonist-induced myoplasmic Ca2+ responses is unclear. We examined effects of the tyrosine kinase inhibitors genistein and methyl 2,5-dihydroxycinnamate (MDHC) on the endothelin-1 (ET-1)-induced Ca2+ response and determined underlying mechanisms for the effects. Freshly isolated smooth muscle cells from porcine coronary arteries were loaded with fura 2 ester, and myoplasmic free Ca2+ (Ca2+ (m)) concentration was estimated with fura 2 microfluorometry. Both genistein and MDHC inhibited the initial transient Cam2+ response to ET by 54 and 81%, respectively (P latent period from ET-1 application to the beginning of the Cam2+ response being increased from 1.08 +/- 0.17 to 2.65 +/- 0.52 min (P < 0.05). In the absence of extracellular Ca2+, genistein inhibited the ET-1-induced Cam2+ response by 93% (P < 0.05). The Cam2+ responses to caffeine (5 mM) or inositol trisphosphate (IP3) applied intracellularly via a patch-clamp pipette were not affected by genistein. Both genistein and MDHC also abolished the sustained Cam2+ response to ET-1. However, the Cam2+ response to depolarization by 80 mM K+ was not inhibited by MDHC and only inhibited 22% by genistein (P < 0.05). These results indicate that 1) activation of tyrosine kinases is an important regulatory mechanism for the ET-1-induced Cam2+ response in vascular smooth muscle and 2) tyrosine kinases mediate ET-1-induced Ca2+ release with no direct effect on IP3-mediated Ca2+ release. Thus ET-1-mediated signaling upstream of IP3 interaction with the Ca2+ stores is regulated by tyrosine kinases.

  12. The Potent Cdc7-Dbf4 (DDK) Kinase Inhibitor XL413 Has Limited Activity in Many Cancer Cell Lines and Discovery of Potential New DDK Inhibitor Scaffolds

    OpenAIRE

    Nanda Kumar Sasi; Kanchan Tiwari; Fen-Fen Soon; Dorine Bonte; Tong Wang; Karsten Melcher; H. Eric Xu; Michael Weinreich

    2014-01-01

    Cdc7-Dbf4 kinase or DDK (Dbf4-dependent kinase) is required to initiate DNA replication by phosphorylating and activating the replicative Mcm2-7 DNA helicase. DDK is overexpressed in many tumor cells and is an emerging chemotherapeutic target since DDK inhibition causes apoptosis of diverse cancer cell types but not of normal cells. PHA-767491 and XL413 are among a number of potent DDK inhibitors with low nanomolar IC50 values against the purified kinase. Although XL413 is highly selective fo...

  13. Protein Kinase CK2α Maintains Extracellular Signal-regulated Kinase (ERK) Activity in a CK2α Kinase-independent Manner to Promote Resistance to Inhibitors of RAF and MEK but Not ERK in BRAF Mutant Melanoma.

    Science.gov (United States)

    Zhou, Bingying; Ritt, Daniel A; Morrison, Deborah K; Der, Channing J; Cox, Adrienne D

    2016-08-19

    The protein kinase casein kinase 2 (CK2) is a pleiotropic and constitutively active kinase that plays crucial roles in cellular proliferation and survival. Overexpression of CK2, particularly the α catalytic subunit (CK2α, CSNK2A1), has been implicated in a wide variety of cancers and is associated with poorer survival and resistance to both conventional and targeted anticancer therapies. Here, we found that CK2α protein is elevated in melanoma cell lines compared with normal human melanocytes. We then tested the involvement of CK2α in drug resistance to Food and Drug Administration-approved single agent targeted therapies for melanoma. In BRAF mutant melanoma cells, ectopic CK2α decreased sensitivity to vemurafenib (BRAF inhibitor), dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor) by a mechanism distinct from that of mutant NRAS. Conversely, knockdown of CK2α sensitized cells to inhibitor treatment. CK2α-mediated RAF-MEK kinase inhibitor resistance was tightly linked to its maintenance of ERK phosphorylation. We found that CK2α post-translationally regulates the ERK-specific phosphatase dual specificity phosphatase 6 (DUSP6) in a kinase dependent-manner, decreasing its abundance. However, we unexpectedly showed, by using a kinase-inactive mutant of CK2α, that RAF-MEK inhibitor resistance did not rely on CK2α kinase catalytic function, and both wild-type and kinase-inactive CK2α maintained ERK phosphorylation upon inhibition of BRAF or MEK. That both wild-type and kinase-inactive CK2α bound equally well to the RAF-MEK-ERK scaffold kinase suppressor of Ras 1 (KSR1) suggested that CK2α increases KSR facilitation of ERK phosphorylation. Accordingly, CK2α did not cause resistance to direct inhibition of ERK by the ERK1/2-selective inhibitor SCH772984. Our findings support a kinase-independent scaffolding function of CK2α that promotes resistance to RAF- and MEK-targeted therapies.

  14. The pan-HER family tyrosine kinase inhibitor afatinib overcomes HER3 ligand heregulin-mediated resistance to EGFR inhibitors in non-small cell lung cancer

    OpenAIRE

    Yonesaka, Kimio; KUDO, KEITA; Nishida, Satomi; Takahama, Takayuki; Iwasa, Tsutomu; Yoshida, Takeshi; Tanaka, Kaoru; Takeda, Masayuki; Kaneda, Hiroyasu; Okamoto, Isamu; Nishio, Kazuto; Nakagawa, Kazuhiko

    2015-01-01

    Afatinib is a second generation epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) characterized as an irreversible pan-human EGFR (HER) family inhibitor. Afatinib remains effective for a subpopulation of patients with non-small cell lung cancer (NSCLC) with acquired resistance to first generation EGFF-TKIs such as erlotinib. Heregulin activates HER3 in an autocrine fashion and causes erlotinib resistance in NSCLC. Here we examine whether afatinib is effective against hereg...

  15. Monte Carlo method based QSAR modeling of maleimide derivatives as glycogen synthase kinase-3β inhibitors.

    Science.gov (United States)

    Živković, Jelena V; Trutić, Nataša V; Veselinović, Jovana B; Nikolić, Goran M; Veselinović, Aleksandar M

    2015-09-01

    The Monte Carlo method was used for QSAR modeling of maleimide derivatives as glycogen synthase kinase-3β inhibitors. The first QSAR model was developed for a series of 74 3-anilino-4-arylmaleimide derivatives. The second QSAR model was developed for a series of 177 maleimide derivatives. QSAR models were calculated with the representation of the molecular structure by the simplified molecular input-line entry system. Two splits have been examined: one split into the training and test set for the first QSAR model, and one split into the training, test and validation set for the second. The statistical quality of the developed model is very good. The calculated model for 3-anilino-4-arylmaleimide derivatives had following statistical parameters: r(2)=0.8617 for the training set; r(2)=0.8659, and r(m)(2)=0.7361 for the test set. The calculated model for maleimide derivatives had following statistical parameters: r(2)=0.9435, for the training, r(2)=0.9262 and r(m)(2)=0.8199 for the test and r(2)=0.8418, r(av)(m)(2)=0.7469 and ∆r(m)(2)=0.1476 for the validation set. Structural indicators considered as molecular fragments responsible for the increase and decrease in the inhibition activity have been defined. The computer-aided design of new potential glycogen synthase kinase-3β inhibitors has been presented by using defined structural alerts.

  16. Anaplastic lymphoma kinase (ALK) inhibitors in the treatment of ALK-driven lung cancers.

    Science.gov (United States)

    Roskoski, Robert

    2017-03-01

    Anaplastic lymphoma kinase is expressed in two-thirds of the anaplastic large-cell lymphomas as an NPM-ALK fusion protein. Physiological ALK is a receptor protein-tyrosine kinase within the insulin receptor superfamily of proteins that participates in nervous system development. The EML4-ALK fusion protein and four other ALK-fusion proteins play a fundamental role in the development in about 5% of non-small cell lung cancers. The amino-terminal portions of the ALK fusion proteins result in dimerization and subsequent activation of the ALK protein kinase domain that plays a key role in the pathogenesis of various tumors. Downstream signaling from the ALK fusion protein leads to the activation of the Ras/Raf/MEK/ERK1/2 cell proliferation module and the JAK/STAT cell survival pathways. Moreover, nearly two dozen ALK activating mutations are involved in the pathogenesis of childhood neuroblastomas. The occurrence of oncogenic ALK-fusion proteins, particularly in non-small cell lung cancer, has fostered considerable interest in the development of ALK inhibitors. Crizotinib was the first such inhibitor approved by the US Food and Drug Administration for the treatment of ALK-positive non-small cell lung cancer in 2011. The median time for the emergence of crizotinib drug resistance is 10.5 months after the initiation of therapy. Such resistance prompted the development of second-generation drugs including ceritinib and alectinib, which are approved for the treatment of non-small cell lung cancer. Unlike the single gatekeeper mutation that occurs in drug-resistant epidermal growth factor receptor in lung cancer, nearly a dozen different mutations in the catalytic domain of ALK fusion proteins have been discovered that result in crizotinib resistance. Crizotinib, ceritinib, and alectinib form a complex within the front cleft between the small and large lobes of an inactive ALK protein-kinase domain with a compact activation segment. These drugs are classified as type I½ B

  17. A Receptor Tyrosine Kinase Inhibitor, Dovitinib (TKI-258), Enhances BMP-2-Induced Osteoblast Differentiation In Vitro

    Science.gov (United States)

    Lee, Yura; Bae, Kyoung Jun; Chon, Hae Jung; Kim, Seong Hwan; Kim, Soon Ae; Kim, Jiyeon

    2016-01-01

    Dovitinib (TKI258) is a small molecule multi-kinase inhibitor currently in clinical phase I/II/III development for the treatment of various types of cancers. This drug has a safe and effective pharmacokinetic/pharmacodynamic profile. Although dovitinib can bind several kinases at nanomolar concentrations, there are no reports relating to osteoporosis or osteoblast differentiation. Herein, we investigated the effect of dovitinib on human recombinant bone morphogenetic protein (BMP)-2-induced osteoblast differentiation in a cell culture model. Dovitinib enhanced the BMP-2-induced alkaline phosphatase (ALP) induction, which is a representative marker of osteoblast differentiation. Dovitinib also stimulated the translocation of phosphorylated Smad1/5/8 into the nucleus and phosphorylation of mitogen-activated protein kinases, including ERK1/2 and p38. In addition, the mRNA expression of BMP-4, BMP-7, ALP, and OCN increased with dovitinib treatment. Our results suggest that dovitinib has a potent stimulating effect on BMP-2-induced osteoblast differentiation and this existing drug has potential for repositioning in the treatment of bone-related disorders. PMID:27025387

  18. The c-Met receptor tyrosine kinase inhibitor MP470 radiosensitizes glioblastoma cells

    Directory of Open Access Journals (Sweden)

    Bearss David

    2009-12-01

    Full Text Available Abstract Purpose Glioblastoma multiforme (GBM is resistant to current cytotoxic therapies, in part because of enhanced DNA repair. Activation of the receptor tyrosine kinase c-Met has been shown to protect cancer cells from DNA damage. We hypothesized that inhibiting c-Met would decrease this protection and thus sensitize resistant tumor cells to the effects of radiation therapy. Materials and methods Eight human GBM cell lines were screened for radiosensitivity to the small-molecule c-Met inhibitor MP470 with colony-count assays. Double-strand (ds DNA breaks was quantified by using antibodies to gamma H2AX. Western blotting demonstrate expression of RAD51, glycogen synthase kinase (GSK-3β, and other proteins. A murine xenograft tumor flank model was used for in vivo radiosensitization studies. Results MP470 reduced c-Met phosphorylation and enhanced radiation-induced cell kill by 0.4 logs in SF767 cells. Cells pretreated with MP470 had more ds DNA damage than cells treated with radiation alone. Mechanistically, MP470 was shown to inhibit dsDNA break repair and increase apoptosis. MP470 influences various survival and DNA repair related proteins such as pAKT, RAD51 and GSK3β. In vivo, the addition of MP470 to radiation resulted in a tumor-growth-delay enhancement ratio of 2.9 over radiation alone and extended survival time. Conclusions GBM is a disease site where radiation is often used to address both macroscopic and microscopic disease. Despite attempts at dose escalation outcomes remain poor. MP470, a potent small-molecule tyrosine kinase inhibitor of c-Met, radiosensitized several GBM cell lines both in vitro and in vivo, and may help to improve outcomes for patients with GBM.

  19. Investigations on Binding Pattern of Kinase Inhibitors with PPARγ: Molecular Docking, Molecular Dynamic Simulations, and Free Energy Calculation Studies

    Science.gov (United States)

    Mazumder, Mohit; Das, Umashankar; Gourinath, Samudrala

    2017-01-01

    Peroxisome proliferator-activated receptor gamma (PPARγ) is a potential target for the treatment of several disorders. In view of several FDA approved kinase inhibitors, in the current study, we have investigated the interaction of selected kinase inhibitors with PPARγ using computational modeling, docking, and molecular dynamics simulations (MDS). The docked conformations and MDS studies suggest that the selected KIs interact with PPARγ in the ligand binding domain (LBD) with high positive predictive values. Hence, we have for the first time shown the plausible binding of KIs in the PPARγ ligand binding site. The results obtained from these in silico investigations warrant further evaluation of kinase inhibitors as PPARγ ligands in vitro and in vivo.

  20. Treatment of Experimental Candida Sepsis with a Janus Kinase Inhibitor Controls Inflammation and Prolongs Survival.

    Science.gov (United States)

    Tsirigotis, P; Papanikolaou, N; Elefanti, A; Konstantinou, P; Gkirkas, K; Rontogianni, D; Siafakas, N; Karakitsos, P; Roilides, E; Dimitriadis, G; Zerva, L; Meletiadis, J

    2015-12-01

    Janus kinases (JAK) are intracellular tyrosine kinases that transduce cytokine-mediated signals to the nucleus, promoting gene expression. Cytokines play a major role in microbial sepsis, which is often associated with uncontrolled inflammation leading to death. JAK inhibitors have been used for the treatment of several autoimmune diseases by modulating immune response, but they have never been tested against microbial sepsis. Ruxolitinib is a small-molecule inhibitor of JAK1/2 proteins, which are involved in the downstream signaling pathway of the vast majority of proinflammatory and anti-inflammatory cytokines. We therefore studied the effect of ruxolitinib in a mouse model of sepsis due to Candida albicans. When ruxolitinib therapy (50 mg/kg [of body weight]/day) was started 1 day before infection, the median survival time was reduced by 3 days, the fungal loads in all organs were higher, the inflammation was significantly less, and serum tumor necrosis factor alpha (TNF-α) and interleukin 10 (IL-10) levels and IL-10/TNF-α ratios were higher than in controls. When ruxolitinib therapy (50 to 1.5 mg/kg/day) was started 1 day after infection, an inverted-U relationship was found, with 6.25 mg/kg/day prolonging median survival time by 6 days, resulting in similar fungal loads, less inflammation, and similar cytokine levels but higher IL-10/TNF-α ratios than the controls. The optimal dose of ruxolitinib controlled infection and prolonged survival with less inflammation than in control animals. Administration of JAK inhibitors may be a promising therapeutic adjunct that needs further investigation.

  1. Inhibitor of Apoptosis Signal-Regulating Kinase 1 Protects Against Acetaminophen-induced Liver Injury

    Science.gov (United States)

    Xie, Yuchao; Ramachandran, Anup; Breckenridge, David G.; Liles, John T.; Lebofsky, Margitta; Farhood, Anwar; Jaeschke, Hartmut

    2015-01-01

    Metabolic activation and oxidant stress are key events in the pathophysiology of acetaminophen (APAP) hepatotoxicity. The initial mitochondrial oxidative stress triggered by protein adduct formation is amplified by c-jun-N-terminal kinase (JNK), resulting in mitochondrial dysfunction and ultimately cell necrosis. Apoptosis signal-regulating kinase 1 (ASK1) is considered the link between oxidant stress and JNK activation. The objective of the current study was to assess the efficacy and mechanism of action of the small-molecule ASK1 inhibitor GS-459679 in a murine model of APAP hepatotoxicity. APAP (300 mg/kg) caused extensive glutathione depletion, JNK activation and translocation to the mitochondria, oxidant stress and liver injury as indicated by plasma ALT activities and area of necrosis over a 24h observation period. Pretreatment with 30 mg/kg of GS-459679 almost completely prevented JNK activation, oxidant stress and injury without affected the metabolic activation of APAP. To evaluate the therapeutic potential of GS-459679, mice were treated with APAP and then with the inhibitor. Given 1.5h after APAP, GS-459679 was still protective, which was paralleled by reduced JNK activation and p-JNK translocation to mitochondria. However, GS-459679 treatment was not more effective than N-acetylcysteine, and the combination of GS-459679 and N-acetylcysteine exhibited similar efficacy as N-acetylcysteine monotherapy, suggesting that GS-459769 and N-acetylcysteine affect the same pathway. Importantly, inhibition of ASK1 did not impair liver regeneration as indicated by PCNA staining. In conclusion, the ASK1 inhibitor GS-459679 protected against APAP toxicity by attenuating JNK activation and oxidant stress in mice and may have therapeutic potential for APAP overdose patients. PMID:25818599

  2. The Phosphoinositide 3-OH Kinase/AKT2 Pathway as a Critical Target for Farnesyltransferase Inhibitor-Induced Apoptosis

    OpenAIRE

    Jiang, Kun; Coppola, Domenico; Crespo, Nichole C.; Nicosia, Santo V.; Hamilton, Andrew D.; Sebti, Said M.; Cheng, Jin Q.

    2000-01-01

    Farnesyltransferase inhibitors (FTIs) represent a novel class of anticancer drugs that exhibit a remarkable ability to inhibit malignant transformation without toxicity to normal cells. However, the mechanism by which FTIs inhibit tumor growth is not well understood. Here, we demonstrate that FTI-277 inhibits phosphatidylinositol 3-OH kinase (PI 3-kinase)/AKT2-mediated growth factor- and adhesion-dependent survival pathways and induces apoptosis in human cancer cells that overexpress AKT2. Fu...

  3. Investigation into the Role of PI3K and JAK3 Kinase Inhibitors in Murine Models of Asthma

    Science.gov (United States)

    Wagh, Akshaya D.; Sharma, Manoranjan; Mahapatra, Jogeshwar; Chatterjee, Abhijeet; Jain, Mukul; Addepalli, Veeranjaneyulu

    2017-01-01

    Asthma is a clinical disorder commonly characterized by chronic eosinophilic inflammation, remodeling and hyper responsiveness of the airways. However, the kinases like Phosphoinositide 3 kinase (PI3K) and Janus kinase 3 (JAK3) are involved in mast cell proliferation, activation, recruitment, migration, and prolonged survival of inflammatory cells. The present study was designed to evaluate the in-vivo comparative effects of two kinase inhibitors on airway inflammation and airway remodeling in acute and chronic models of asthma. Mice were sensitized twice intra-peritoneally and then challenged by inhalation with ovalbumin (OVA). They developed an extensive inflammatory response, goblet cell hyperplasia, collagen deposition, airway smooth muscle thickening similar to pathologies observed in human asthma. The effects of PI3K inhibitor (30 mg/kg, p.o), JAK3 inhibitor (30 mg/kg, p.o) and Dexamethasone (0.3 mg/kg) on airway inflammation and remodeling in OVA sensitized/challenged BALB/c mice were evaluated. Twenty-four hours after the final antigen challenge, bronchoalveolar lavage (BAL) and histological examinations were carried out. It was observed that kinase inhibitors significantly reduced airway inflammation as evidenced by the decrease in pro inflammatory cytokines in BALF and lung homogenate and inflammatory cell count in sensitized mice after allergen challenge. Lung histological analysis showed increased infiltration of inflammatory cells, hyperplasia of goblet cells and the collagen deposition, which were significantly reduced with kinase inhibitor. In conclusion, our data suggest that PI3K and JAK3 inhibitors showed promising alternative therapeutic activity in asthma, which might significantly counteract the airway inflammation in patients with allergic asthma. PMID:28293189

  4. A high-content EMT screen identifies multiple receptor tyrosine kinase inhibitors with activity on TGFβ receptor

    Science.gov (United States)

    Ackerknecht, Sabine; Lehembre, François; Fink, Tobias; Stritt, Manuel; Wirth, Matthias; Pavan, Simona; Bill, Ruben; Regenass, Urs; Christofori, Gerhard; Meyer-Schaller, Nathalie

    2016-01-01

    An epithelial to mesenchymal transition (EMT) enables epithelial tumor cells to break out of the primary tumor mass and to metastasize. Understanding the molecular mechanisms driving EMT in more detail will provide important tools to interfere with the metastatic process. To identify pharmacological modulators and druggable targets of EMT, we have established a novel multi-parameter, high-content, microscopy-based assay and screened chemical compounds with activities against known targets. Out of 3423 compounds, we have identified 19 drugs that block transforming growth factor beta (TGFβ)-induced EMT in normal murine mammary gland epithelial cells (NMuMG). The active compounds include inhibitors against TGFβ receptors (TGFBR), Rho-associated protein kinases (ROCK), myosin II, SRC kinase and uridine analogues. Among the EMT-repressing compounds, we identified a group of inhibitors targeting multiple receptor tyrosine kinases, and biochemical profiling of these multi-kinase inhibitors reveals TGFBR as a thus far unknown target of their inhibitory spectrum. These findings demonstrate the feasibility of a multi-parameter, high-content microscopy screen to identify modulators and druggable targets of EMT. Moreover, the newly discovered “off-target” effects of several receptor tyrosine kinase inhibitors have important consequences for in vitro and in vivo studies and might beneficially contribute to the therapeutic effects observed in vivo. PMID:27036020

  5. A high-content EMT screen identifies multiple receptor tyrosine kinase inhibitors with activity on TGFβ receptor.

    Science.gov (United States)

    Lotz-Jenne, Carina; Lüthi, Urs; Ackerknecht, Sabine; Lehembre, François; Fink, Tobias; Stritt, Manuel; Wirth, Matthias; Pavan, Simona; Bill, Ruben; Regenass, Urs; Christofori, Gerhard; Meyer-Schaller, Nathalie

    2016-05-03

    An epithelial to mesenchymal transition (EMT) enables epithelial tumor cells to break out of the primary tumor mass and to metastasize. Understanding the molecular mechanisms driving EMT in more detail will provide important tools to interfere with the metastatic process. To identify pharmacological modulators and druggable targets of EMT, we have established a novel multi-parameter, high-content, microscopy-based assay and screened chemical compounds with activities against known targets. Out of 3423 compounds, we have identified 19 drugs that block transforming growth factor beta (TGFβ)-induced EMT in normal murine mammary gland epithelial cells (NMuMG). The active compounds include inhibitors against TGFβ receptors (TGFBR), Rho-associated protein kinases (ROCK), myosin II, SRC kinase and uridine analogues. Among the EMT-repressing compounds, we identified a group of inhibitors targeting multiple receptor tyrosine kinases, and biochemical profiling of these multi-kinase inhibitors reveals TGFBR as a thus far unknown target of their inhibitory spectrum. These findings demonstrate the feasibility of a multi-parameter, high-content microscopy screen to identify modulators and druggable targets of EMT. Moreover, the newly discovered "off-target" effects of several receptor tyrosine kinase inhibitors have important consequences for in vitro and in vivo studies and might beneficially contribute to the therapeutic effects observed in vivo.

  6. Potent quinoxaline-based inhibitors of PDGF receptor tyrosine kinase activity. Part 2: the synthesis and biological activities of RPR127963 an orally bioavailable inhibitor.

    Science.gov (United States)

    He, Wei; Myers, Michael R; Hanney, Barbara; Spada, Alfred P; Bilder, Glenda; Galzcinski, Helen; Amin, Dilip; Needle, Saul; Page, Ken; Jayyosi, Zaid; Perrone, Mark H

    2003-09-15

    RPR127963 demonstrates an excellent pharmacokinetic profile in several species and was found to be efficacious in the prevention of restenosis in a Yucatan mini-pig model upon oral administration of 1-5 mg/kg. The in vitro selectivity profile and SAR of the highly optimized PDGF-R tyrosine kinase inhibitor are highlighted.

  7. Selectivity analysis of protein kinase CK2 inhibitors DMAT, TBB and resorufin in cisplatin-induced stress responses

    DEFF Research Database (Denmark)

    Fritz, Gerhard; Issinger, Olaf-Georg; Olsen, Birgitte Brinkmann

    2009-01-01

    Targeting protein kinases as a therapeutic approach to treat various diseases, especially cancer is currently a fast growing business. Although many inhibitors are available, exhibiting remarkable potency, the major challenge is their selectivity. Here we show that the protein kinase CK2 inhibitors...... viability and inhibition of endogenous CK2 activity in the absence and presence of the anti-cancer drug cisplatin. The strongest effect on viability was observed with resorufin. In contrast to resorufin, TBB protected cells from cisplatin-induced cell killing. Furthermore, the inhibition of endogenous CK2...

  8. Inhibition of Lassa virus and Ebola virus infection in host cells treated with the kinase inhibitors genistein and tyrphostin.

    Science.gov (United States)

    Kolokoltsov, Andrey A; Adhikary, Shramika; Garver, Jennifer; Johnson, Lela; Davey, Robert A; Vela, Eric M

    2012-01-01

    Arenaviruses and filoviruses are capable of causing hemorrhagic fever syndrome in humans. Limited therapeutic and/or prophylactic options are available for humans suffering from viral hemorrhagic fever. In this report, we demonstrate that pre-treatment of host cells with the kinase inhibitors genistein and tyrphostin AG1478 leads to inhibition of infection or transduction in cells infected with Ebola virus, Marburg virus, and Lassa virus. In all, the results demonstrate that a kinase inhibitor cocktail consisting of genistein and tyrphostin AG1478 is a broad-spectrum antiviral that may be used as a therapeutic or prophylactic against arenavirus and filovirus hemorrhagic fever.

  9. Non-hinge-binding pyrazolo[1,5-a]pyrimidines as potent B-Raf kinase inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Berger, Dan M.; Torres, Nancy; Dutia, Minu; Powell, Dennis; Ciszewski, Greg; Gopalsamy, Ariamala; Levin, Jeremy I.; Kim, Kyung-Hee; Xu, Weixin; Wilhelm, James; Hu, YongBo; Collins, Karen; Feldberg, Larry; Kim, Steven; Frommer, Eileen; Wojciechowicz, Donald; Mallon, Robert; (Wyeth)

    2010-11-19

    As part of our research effort to discover B-Raf kinase inhibitors, we prepared a series of C-3 substituted N-(3-(pyrazolo[1,5-a]pyrimidin-7-yl)phenyl)-3-(trifluoromethyl)benzamides. X-ray crystallography studies revealed that one of the more potent inhibitors (10n) bound to B-Raf kinase without forming a hinge-binding hydrogen bond. With basic amine residues appended to C-3 aryl residues, cellular activity and solubility were enhanced over previously described compounds of this class.

  10. Neurite outgrowth resistance to rho kinase inhibitors in PC12 Adh cell.

    Science.gov (United States)

    Yin, Hua; Hou, Xiaolin; Tao, Tingrui; Lv, Xiaoman; Zhang, Luyong; Duan, Weigang

    2015-05-01

    Rho kinase (ROCK) inhibitor is a promising agent for neural injury disorders, which mechanism is associated with neurite outgrowth. However, neurite outgrowth resistance occurred when PC12 Adh cell was treated with ROCK inhibitors for a longer time. PC12 Adh cells were treated with ROCK inhibitor Y27632 or NGF for different durations. Neurite outgrowth resistance occurred when PC12 Adh cell exposed to Y27632 (33 µM) for 3 or more days, but not happen when exposed to nerve growth factor (NGF, 100 ng/mL). The gene expression in the PC12 Adh cells treated with Y27632 (33 µM) or NGF (100 ng/mL) for 2 or 4 days was assayed by gene microarray, and the reliability of the results were confirmed by real-time RT-PCR. Cluster analysis proved that the gene expression profile of PC12 Adh cell treated with Y27632 for 4 days was different from that treated with Y27632 for 2 days and those treated with NGF for 2 and 4 days, respectively. Pathway analysis hinted that the neurite outgrowth resistance could be associated with up-regulation of inflammatory pathways, especially rno04610 (complement and coagulation cascades), and down-regulation of cell cycle pathways, especially rno04110.

  11. Cyclin-Dependent Kinase Inhibitors in Maize Endosperm and Their Potential Role in Endoreduplication1

    Science.gov (United States)

    Coelho, Cintia M.; Dante, Ricardo A.; Sabelli, Paolo A.; Sun, Yuejin; Dilkes, Brian P.; Gordon-Kamm, William J.; Larkins, Brian A.

    2005-01-01

    Two maize (Zea mays) cyclin-dependent kinase (CDK) inhibitors, Zeama;KRP;1 and Zeama;KRP;2, were characterized and shown to be expressed in developing endosperm. Similar to the CDK inhibitors in Arabidopsis (Arabidopsis thaliana) and tobacco (Nicotiana tabacum), the maize proteins contain a carboxy-terminal region related to the inhibitory domain of the mammalian Cip/Kip inhibitors. Zeama;KRP;1 is present in the endosperm between 7 and 21 d after pollination, a period that encompasses the onset of endoreduplication, while the Zeama;KRP;2 protein declines during this time. Nevertheless, Zeama;KRP;1 accounts for only part of the CDK inhibitory activity that peaks coincident with the endoreduplication phase of endosperm development. In vitro assays showed that Zeama;KRP;1 and Zeama;KRP;2 are able to inhibit endosperm Cdc2-related CKD activity that associates with p13Suc1. They were also shown to specifically inhibit cyclin A1;3- and cyclin D5;1-associated CDK activities, but not cyclin B1;3/CDK. Overexpression of Zeama;KRP;1 in maize embryonic calli that ectopically expressed the wheat dwarf virus RepA protein, which counteracts retinoblastoma-related protein function, led to an additional round of DNA replication without nuclear division. PMID:16055680

  12. Third generation tyrosine kinase inhibitors and their development in advanced renal cell carcinoma.

    Science.gov (United States)

    Bukowski, Ronald M

    2012-01-01

    Angiogenesis in general and the vascular endothelial growth factor (VEGF) signaling axis in particular is a validated target in renal cell carcinoma (RCC). Clear-cell carcinoma of the kidney is now recognized as a malignancy that is sensitive to inhibitors of the VEGF pathway. Treatment options for patients with metastatic renal cell carcinoma have evolved in dramatic fashion over the past 6 years, and a new paradigm has developed. The cytokines interferon-α and interleukin-2 were previously utilized for therapy, but since December 2005, six new agents have been approved in the United States for the treatment of advanced RCC. Two are tyrosine kinase inhibitors (TKI's) including sunitinib and recently pazopanib, and the multikinase inhibitor sorafenib. The current review examines the evolving data with the next generation of TKI's, axitinib and tivozanib being developed for the treatment of advanced RCC. These agents were synthesized to provide increased target specificity and enhanced target inhibition. The preclinical and clinical data are examined, an overview of the development of these TKI's is provided, and discussion plus speculation concerning their potential roles as RCC therapy is provided.

  13. A new trend in pathogenetic treatment of rheumatoid arthritis: tofacitinib, the first inhibitor of Janus kinase

    Directory of Open Access Journals (Sweden)

    Dmitry Evgenyevich Karateev

    2014-01-01

    Full Text Available The Problem of effective treatment of rheumatoid arthritis (RA has not been solved yet. In patients with an inadequate response to the synthetic disease-modifying antirheumatic drugs (DMARDs, TNF inhibitors and other genetically engineered (GE drugs may give an incomplete effect or be totally ineffective in 20–40% of cases. The search for new therapeutic options is very active in many directions, including modulation of intracellular signaling pathways. The most impressive clinical results in RA patients have been achieved for Janus kinase (JAK inhibitor, tofacitinib, – Tofa (CP-690,550, a low molecular weight drug for oral administration. Unlike the conventional synthetic DMARD, Tofa belongs to targeted therapy drugs; its pharmacological effects largely resemble those of GE drugs (blockade of the biological effects of cytokines. In particular, some features of its mechanism of action (e.g., interleukin 6 inhibition make it similar to tocilizumab. In all trials, Tofa proved to exhibit high clinical efficacy and satisfactory safety. Tofa was shown to be significantly more effective than methotrexate and placebo (in monotherapy, as well as in combination with DMARDs in RA patients with an inadequate response to standard DMARDs and TNFa inhibitors (regarding response to treatment, individual parameters of disease activity, achievement of low disease activity and remission, functional impairment. Furthermore, Tofa can inhibit joint damage progression.

  14. Structure-Based Design of Potent and Selective 3-Phosphoinositide-Dependent Kinase-1 (PDK1) Inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Medina, Jesus R.; Becker, Christopher J.; Blackledge, Charles W.; Duquenne, Celine; Feng, Yanhong; Grant, Seth W.; Heerding, Dirk; Li, William H.; Miller, William H.; Romeril, Stuart P.; Scherzer, Daryl; Shu, Arthur; Bobko, Mark A.; Chadderton, Antony R.; Dumble, Melissa; Gardiner, Christine M.; Gilbert, Seth; Liu, Qi; Rabindran, Sridhar K.; Sudakin, Valery; Xiang, Hong; Brady, Pat G.; Campobasso, Nino; Ward, Paris; Axten, Jeffrey M. (GSKPA)

    2014-10-02

    Phosphoinositide-dependent protein kinase-1(PDK1) is a master regulator of the AGC family of kinases and an integral component of the PI3K/AKT/mTOR pathway. As this pathway is among the most commonly deregulated across all cancers, a selective inhibitor of PDK1 might have utility as an anticancer agent. Herein we describe our lead optimization of compound 1 toward highly potent and selective PDK1 inhibitors via a structure-based design strategy. The most potent and selective inhibitors demonstrated submicromolar activity as measured by inhibition of phosphorylation of PDK1 substrates as well as antiproliferative activity against a subset of AML cell lines. In addition, reduction of phosphorylation of PDK1 substrates was demonstrated in vivo in mice bearing OCl-AML2 xenografts. These observations demonstrate the utility of these molecules as tools to further delineate the biology of PDK1 and the potential pharmacological uses of a PDK1 inhibitor.

  15. Look out, aurorae about!

    Science.gov (United States)

    Wild, Jim

    2006-02-01

    Myths and legends abound concerning the aurora borealis; observations, at least in the UK, are considerably more rare, yet people continue to be interested in observing them on the few times a year they are visible. AuroraWatch hopes to boost this interest by providing an email alert service that, since 2000, has used real-time information from a network of magnetometers to assess when aurorae might be visible. Alerts can be sent to mobile phones as well as by conventional email, and have already enabled many people to see this beautiful natural phenomenon.

  16. Design, synthesis and evaluation of 3-quinoline carboxylic acids as new inhibitors of protein kinase CK2.

    Science.gov (United States)

    Syniugin, Anatolii R; Ostrynska, Olga V; Chekanov, Maksym O; Volynets, Galyna P; Starosyla, Sergiy A; Bdzhola, Volodymyr G; Yarmoluk, Sergiy M

    2016-01-01

    In this article, the derivatives of 3-quinoline carboxylic acid were studied as inhibitors of protein kinase CK2. Forty-three new compounds were synthesized. Among them 22 compounds inhibiting CK2 with IC50 in the range from 0.65 to 18.2 μM were identified. The most active inhibitors were found among tetrazolo-quinoline-4-carboxylic acid and 2-aminoquinoline-3-carboxylic acid derivatives.

  17. Discovery and Characterization of the N-phenyl-N 0 -Naphthylurea Class of p38 Kinase Inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Cirillo, P.; Hickey, E; Moss, N; Breitfelder, S; Betageri, R; Fadra, T; Gaenzler, F; Gilmore, T; Xiong, Z; et al.

    2009-01-01

    An effort aimed at exploring structural diversity in the N-pyrazole-N{prime}-naphthylurea class of p38 kinase inhibitors led to the synthesis and characterization of N-phenyl-N{prime}-naphthylureas. Examples of these compounds displayed excellent inhibition of TNF-{alpha} production in vitro, as well as efficacy in a mouse model of lipopolysaccharide induced endotoxemia. In addition, perspective is provided on the role of a sulfonamide functionality in defining inhibitor potency.

  18. Enzyme kinetics and distinct modulation of the protein kinase N family of kinases by lipid activators and small molecule inhibitors

    OpenAIRE

    Falk, Matthew D.; Liu, Wei; Bolaños, Ben; Unsal-Kacmaz, Keziban; Klippel, Anke; Grant, Stephan; Brooun, Alexei; Timofeevski, Sergei

    2014-01-01

    The PKN (protein kinase N) family of Ser/Thr protein kinases regulates a diverse set of cellular functions, such as cell migration and cytoskeletal organization. Inhibition of tumour PKN activity has been explored as an oncology therapeutic approach, with a PKN3-targeted RNAi (RNA interference)-derived therapeutic agent in Phase I clinical trials. To better understand this important family of kinases, we performed detailed enzymatic characterization, determining the kinetic mechanism and lipi...

  19. Localized Aurora B activity spatially controls non-kinetochore microtubules during spindle assembly.

    Science.gov (United States)

    Tanenbaum, Marvin E; Medema, René H

    2011-12-01

    Efficient spindle assembly involves the generation of spatial cues around chromosomes that locally stabilize microtubule (MT) plus-ends. In addition to the small GTPase Ran, there is evidence that Aurora B kinase might also generate a spatial cue around chromosomes but direct proof for this is still lacking. Here, we find that the Aurora B substrate MCAK localizes to MT plus-ends throughout the mitotic spindle, but its accumulation is strongly reduced on MT plus-ends near chromatin, suggesting that a signal emanating from chromosomes negatively regulates MCAK plus-end binding. Indeed, we show that Aurora B is the kinase responsible for producing this chromosome-derived signal. These results are the first to visualize spatially restricted Aurora B kinase activity around chromosomes on an endogenous substrate and explain how Aurora B could spatially control the dynamics of non-kinetochore MTs during spindle assembly.

  20. Structure-based design, discovery and development of checkpoint kinase inhibitors as potential anti-cancer therapies

    Science.gov (United States)

    Matthews, Thomas P; Jones, Alan M; Collins, Ian

    2014-01-01

    Introduction Checkpoint kinase inhibitors offer the promise of enhancing the effectiveness of widely prescribed cancer chemotherapies and radiotherapy by inhibiting the DNA damage response, as well as the potential for single agent efficacy. Areas covered This article surveys structural insights into the checkpoint kinases CHK1 and CHK2 that have been exploited to enhance the selectivity and potency of small molecule inhibitors. The use of mechanistic cellular assays to guide the optimisation of inhibitors is reviewed. The status of the current clinical candidates and emerging new clinical contexts for CHK1 and CHK2 inhibitors are discussed, including the prospects for single agent efficacy. Expert opinion Protein bound water molecules play key roles in structural features that can be targeted to gain high selectivity for either enzyme. The results of early phase clinical trials of checkpoint inhibitors have been mixed, but significant progress has been made in testing the combination of CHK1 inhibitors with genotoxic chemotherapy. Second generation CHK1 inhibitors are likely to benefit from increased selectivity and oral bioavailability. While the optimum therapeutic context for CHK2 inhibition remains unclear, the emergence of single agent preclinical efficacy for CHK1 inhibitors in specific tumour types exhibiting constitutive replication stress represents exciting progress in exploring the therapeutic potential of these agents. PMID:23594139

  1. Using 2 nd generation tyrosine kinase inhibitors in frontline management of chronic phase chronic myeloid leukemia

    Directory of Open Access Journals (Sweden)

    Vishal Jayakar

    2014-01-01

    Full Text Available Choices in medicine come with responsibility. With several TKI′s (Tyrosine kinase inhibitors available for front-line management of CML (Chronic Myeloid Leukemia, an astute clinician has to personalise, rationalise and take a pragmatic approach towards selection of the best drug for the ′patient in question′. Though it is hotly debated as to which TKI will triumph, the truth of this debate lies in individualising treatment rather than a general ′all size fits all′ approach with imatinib. I personally believe that the second generation TKI′s will suit most patient clinical profiles rather than prescribing imatinib to all and I have strived to make a strong case for them in front line treatment of CML. Though Imatinib may remain the first line choice for some patients, my efforts in this debate are mainly geared towards breaking the myth that imatinib is the sole ′block buster′ on the CML landscape

  2. The effect of cyclin-dependent kinases inhibitor treatment on experimental herpes simplex encephalitis mice.

    Science.gov (United States)

    Zhou, Yu; Zeng, Yan-Ping; Zhou, Qin; Guan, Jing-Xia; Lu, Zu-Neng

    2016-08-01

    Herpes simplex encephalitis(HSE) is the most common and serious viral encephalitis in humans. There is a lack of effective medication to date for HSE. A better understanding of the mediators of tissue damage is essential for finding new targets for therapeutic intervention. In this project, we explored the effect of cyclin-dependent kinases inhibitor olomoucine treatment on experimental HSE mice. The following results were obtained: (1) olomoucine increased survival in HSE mice; (2) olomoucine inhibited microglial activation and reduced HSV-1-induced cytokines release; (3) olomoucine prevented neural cells apoptosis and attenuated brain tissue pathological changes following HSV-1 infection; (4) olomoucine reduced brain edema and improved neurological function in HSE. Overall, olomoucine can induce a blunted inflammatory response, maintain the blood vessel wall intact, improve neurological function and increase survival in HSE mice.

  3. Tyrosine kinase inhibitors in the treatment of systemic sclerosis: from animal models to clinical trials.

    Science.gov (United States)

    Iwamoto, Naoki; Distler, Jörg H W; Distler, Oliver

    2011-02-01

    Efficient antifibrotic therapies are not available for patients with systemic sclerosis (SSc). This review summarizes the current preclinical and clinical evidence for imatinib and related tyrosine kinase inhibitors as potential antifibrotic therapies for SSc and other fibrotic diseases. In experimental models of SSc, imatinib, nilotinib, and dasatinib demonstrated strong antifibrotic effects. Imatinib not only prevented fibrosis in the bleomycin-induced model of dermal fibrosis and the tight skin mouse-1 model but also reduced established fibrosis in a modified bleomycin model. Open-label, proof-of-concept trials in SSc showed moderate effects on skin fibrosis, biological measures of skin fibrosis, and lung fibrosis compared with baseline measures. However, whether this reflects the natural course of the disease or is a result of treatment effects is unclear and needs to be analyzed in larger, multicenter, randomized, placebo-controlled trials. Toxicity is expected from cancer trials with frequent mild to moderate adverse events.

  4. Ibrutinib: a novel Bruton's tyrosine kinase inhibitor with outstanding responses in patients with chronic lymphocytic leukemia.

    Science.gov (United States)

    Barrientos, Jacqueline; Rai, Kanti

    2013-08-01

    New treatment options are urgently needed for patients with relapsed chronic lymphocytic leukemia (CLL) who fail to respond to currently available therapies or cannot achieve a sustained response. Moreover, targeted agents with less myelotoxicity are necessary to treat patients with multiple comorbidities who would otherwise be unable to tolerate standard regimens. Ibrutinib, a Bruton's tyrosine kinase inhibitor, has shown highly encouraging results in phase I/II trials in patients with treatment-naive, relapsed and refractory CLL even in the presence of high risk disease or poor prognostic markers. In phase I/II trials, ibrutinib 420 mg or 840 mg - given continuously as single agent or at a dose of 420 mg daily in combination with a monoclonal antibody or chemoimmunotherapy - has been associated with high response rates and durable clinical remissions. Phase II and III trials are currently under way for treatment-naive patients, relapsed/refractory patients, and for those patients harboring a 17p deletion.

  5. 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.

  6. Design and synthesis of orally bioavailable serum and glucocorticoid-regulated kinase 1 (SGK1) inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Hammond, Marlys; Washburn, David G.; Hoang, Tram H.; Manns, Sharada; Frazee, James S.; Nakamura, Hiroko; Patterson, Jaclyn R.; Trizna, Walter; Wu, Charlene; Azzarano, Leonard M.; Nagilla, Rakesh; Nord, Melanie; Trejo, Rebecca; Head, Martha S.; Zhao, Baoguang; Smallwood, Angela M.; Hightower, Kendra; Laping, Nicholas J.; Schnackenberg, Christine G.; Thompson, Scott K.; (GSKPA)

    2010-09-27

    The lead serum and glucocorticoid-related kinase 1 (SGK1) inhibitors 4-(5-phenyl-1H-pyrrolo[2,3-b]pyridin-3-yl)benzoic acid (1) and {l_brace}4-[5-(2-naphthalenyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]phenyl{r_brace}acetic acid (2) suffer from low DNAUC values in rat, due in part to formation and excretion of glucuronic acid conjugates. These PK/glucuronidation issues were addressed either by incorporating a substituent on the 3-phenyl ring ortho to the key carboxylate functionality of 1 or by substituting on the group in between the carboxylate and phenyl ring of 2. Three of these analogs have been identified as having good SGK1 inhibition potency and have DNAUC values suitable for in vivo testing.

  7. Design and synthesis of orally bioavailable serum and glucocorticoid-regulated kinase 1 (SGK1) inhibitors.

    Science.gov (United States)

    Hammond, Marlys; Washburn, David G; Hoang, H Tram; Manns, Sharada; Frazee, James S; Nakamura, Hiroko; Patterson, Jaclyn R; Trizna, Walter; Wu, Charlene; Azzarano, Leonard M; Nagilla, Rakesh; Nord, Melanie; Trejo, Rebecca; Head, Martha S; Zhao, Baoguang; Smallwood, Angela M; Hightower, Kendra; Laping, Nicholas J; Schnackenberg, Christine G; Thompson, Scott K

    2009-08-01

    The lead serum and glucocorticoid-related kinase 1 (SGK1) inhibitors 4-(5-phenyl-1H-pyrrolo[2,3-b]pyridin-3-yl)benzoic acid (1) and {4-[5-(2-naphthalenyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]phenyl}acetic acid (2) suffer from low DNAUC values in rat, due in part to formation and excretion of glucuronic acid conjugates. These PK/glucuronidation issues were addressed either by incorporating a substituent on the 3-phenyl ring ortho to the key carboxylate functionality of 1 or by substituting on the group in between the carboxylate and phenyl ring of 2. Three of these analogs have been identified as having good SGK1 inhibition potency and have DNAUC values suitable for in vivo testing.

  8. Targeting Mitogen-Activated Protein Kinase-Activated Protein Kinase 2 (MAPKAPK2, MK2): Medicinal Chemistry Efforts To Lead Small Molecule Inhibitors to Clinical Trials.

    Science.gov (United States)

    Fiore, Mario; Forli, Stefano; Manetti, Fabrizio

    2016-04-28

    The p38/MAPK-activated kinase 2 (MK2) pathway is involved in a series of pathological conditions (inflammation diseases and metastasis) and in the resistance mechanism to antitumor agents. None of the p38 inhibitors entered advanced clinical trials because of their unwanted systemic side effects. For this reason, MK2 was identified as an alternative target to block the pathway but avoiding the side effects of p38 inhibition. However, ATP-competitive MK2 inhibitors suffered from low solubility, poor cell permeability, and scarce kinase selectivity. Fortunately, non-ATP-competitive inhibitors of MK2 have been already discovered that allowed circumventing the selectivity issue. These compounds showed the additional advantage to be effective at lower concentrations in comparison to the ATP-competitive inhibitors. Therefore, although the significant difficulties encountered during the development of these inhibitors, MK2 is still considered as an attractive target to treat inflammation and related diseases to prevent tumor metastasis and to increase tumor sensitivity to chemotherapeutics.

  9. Scaffold oriented synthesis. Part 4: design, synthesis and biological evaluation of novel 5-substituted indazoles as potent and selective kinase inhibitors employing heterocycle forming and multicomponent reactions.

    Science.gov (United States)

    Akritopoulou-Zanze, Irini; Wakefield, Brian D; Gasiecki, Alan; Kalvin, Douglas; Johnson, Eric F; Kovar, Peter; Djuric, Stevan W

    2011-03-01

    We report the synthesis and biological evaluation of 5-substituted indazoles as kinase inhibitors. The compounds were synthesized in a parallel synthesis fashion from readily available starting materials employing heterocycle forming and multicomponent reactions and were evaluated against a panel of kinase assays. Potent inhibitors were identified for Gsk3β, Rock2, and Egfr.

  10. Aurora final report

    Energy Technology Data Exchange (ETDEWEB)

    Robert, Dross; Amedeo, Conti

    2013-12-06

    Final Technical report detailing the work done by Nuvera and its partners to fulfill the goals of the program "Transport Studies Enabling Efficiency Optimization of Cost-Competitive Fuel Cell Stacks" (a.k.a. AURORA)

  11. Identification of a Glycogen Synthase Kinase-3[beta] Inhibitor that Attenuates Hyperactivity in CLOCK Mutant Mice

    Energy Technology Data Exchange (ETDEWEB)

    Kozikowski, Alan P.; Gunosewoyo, Hendra; Guo, Songpo; Gaisina, Irina N.; Walter, Richard L.; Ketcherside, Ariel; McClung, Colleen A.; Mesecar, Andrew D.; Caldarone, Barbara (Psychogenics); (Purdue); (UIC); (UTSMC)

    2012-05-02

    Bipolar disorder is characterized by a cycle of mania and depression, which affects approximately 5 million people in the United States. Current treatment regimes include the so-called 'mood-stabilizing drugs', such as lithium and valproate that are relatively dated drugs with various known side effects. Glycogen synthase kinase-3{beta} (GSK-3{beta}) plays a central role in regulating circadian rhythms, and lithium is known to be a direct inhibitor of GSK-3{beta}. We designed a series of second generation benzofuran-3-yl-(indol-3-yl)maleimides containing a piperidine ring that possess IC{sub 50} values in the range of 4 to 680 nM against human GSK-3{beta}. One of these compounds exhibits reasonable kinase selectivity and promising preliminary absorption, distribution, metabolism, and excretion (ADME) data. The administration of this compound at doses of 10 to 25 mg kg{sup -1} resulted in the attenuation of hyperactivity in amphetamine/chlordiazepoxide-induced manic-like mice together with enhancement of prepulse inhibition, similar to the effects found for valproate (400 mg kg{sup -1}) and the antipsychotic haloperidol (1 mg kg{sup -1}). We also tested this compound in mice carrying a mutation in the central transcriptional activator of molecular rhythms, the CLOCK gene, and found that the same compound attenuates locomotor hyperactivity in response to novelty. This study further demonstrates the use of inhibitors of GSK-3{beta} in the treatment of manic episodes of bipolar/mood disorders, thus further validating GSK-3{beta} as a relevant therapeutic target in the identification of new therapies for bipolar patients.

  12. Optimization of a homogeneous assay for kinase inhibitors in plant extracts.

    Science.gov (United States)

    Dufau, Isabelle; Lazzari, Anne; Samson, Arnaud; Pouny, Isabelle; Ausseil, Frederic

    2008-10-01

    To identify natural and original kinase inhibitors from plant extracts, we have developed and compared a heterogeneous enzyme-linked immunosorbent assay (ELISA) and a homogeneous time-resolved fluorescence (HTRF, Cisbio International, Bagnols/Cèze, France) assay. Kinase affinity for the ATP substrate was determined in both assays, and the same [ATP]/ATP Km ratio was used in each case to enable the identification of ATP competitive and noncompetitive inhibitors. Assays were then used to screen the same collection of chemical compounds and plant extracts. The intra-assay correlation analysis of each technology showed a very good screening precision in HTRF and an acceptable one in ELISA. When the two methods were compared, a poor correlation was obtained with a higher hit rate in the ELISA. We then performed a detailed study of the ELISA hits and showed that they also presented a strong antioxidant activity, associated with high adsorption into microplate wells, which interfered with the horseradish peroxidase-based detection system. These hits were then flagged as false-positives. We also showed that many plant extracts presented this kind of activity and that this interference could explain the lack of correlation between the assays. These findings suggest that assay design should be carefully adapted to the substances to be screened and that interferences should be extensively considered before any assay development process and comparison studies. In spite of a few interferences, our results showed that a homogeneous-phase assay like the HTRF assay could be more efficiently used for plant extract screening than a heterogeneous-phase assay like ELISA.

  13. Hepatitis B Virus Reactivation during Treatment with Multi-Tyrosine Kinase Inhibitor for Hepatocellular Carcinoma

    Directory of Open Access Journals (Sweden)

    Satoshi Shiba

    2012-09-01

    Full Text Available Hepatitis B virus (HBV reactivation is well documented in individuals with cancer who receive certain cytotoxic or immunosuppressive therapies including rituximab treatment. As a general rule, the risk is greatest upon withdrawal of chemotherapy. The risk ranges from approximately 20 to 50% among HBsAg-positive carriers. A 67-year-old man was diagnosed with inoperable multiple hepatocellular carcinoma accompanied by an increase in alpha-fetoprotein and protein induced by vitamin K absence or antagonist II level. Eighteen weeks after starting on the oral multi-tyrosine kinase inhibitor TSU-68, laboratory investigations showed a substantial increase in serum transaminase levels (AST: 302 IU/l; ALT: 324 IU/l and an elevation of the HBV-DNA level (6.9 log copies/ml. The diagnosis was that the cause of the acute hepatitis was HBV reactivation and we immediately administered entecavir. Two months after the initiation of daily entecavir treatment, laboratory findings showed that the serum levels of transaminases and ALP had improved (AST: 18 IU/l; ALT: 10 IU/l; ALP: 197 U/l. When the HBV markers were examined 4 months later, they were altered: HBeAg was negative and HBeAb was positive. Entecavir treatment was discontinued after 6 months. Although reactivation with rituximab has been reported, reactivation with a tyrosine kinase inhibitor is extremely unusual in a patient who is HBsAg negative but anti-HBc positive. This is the first report describing HBV reactivation with an increasing HBV-DNA level in a HBsAg-negative/HBcAb-positive/HBsAb-positive patient who was treated with TSU-68 for hepatocellular carcinoma.

  14. Tamoxifen and Src kinase inhibitors as neuroprotective/neuroregenerative drugs after spinal cord injury

    Institute of Scientific and Technical Information of China (English)

    Iris K Salgado; Aranza I Torrado; Jose M Santiago; Jorge D Miranda

    2015-01-01

    Spinal cord injury (SCI) is a devastating condition that produces signiifcant changes in the life-style of patients. Many molecular and cellular events are triggered after the initial physical impact to the cord. Two major phases have been described in the ifeld of SCI: an acute phase and late phase. Most of the therapeutic strategies are focused on the late phase because this provides an opportunity to target cellular events like apoptosis, demyelination, scar formation and axonal outgrowth. In this mini-review, we will focus on two agents (tamoxifen and a Src kinase family inhibitor known as PP2) that have been shown in our laboratory to produce neuroprotective (increase cell survival) and/or regenerative (axonal outgrowth) actions. The animal model used in our laboratory is adult female rat (~250 g) with a moderate contusion (12.5 mm) to the spinal cord at the T10 level, using the MASCIS impactor device. Tamoxifen or PP2 was administered by implantation of a 15 mg pellet (Innovative Research of America, Sarasota, FL, USA) or by intraperitoneal injections (1.5 mg/kg, every 3 days), respectively, to produce a long-term effect (28 days). Tamoxifen and the Src kinase inhibitor, PP2, are drugs that in rats with a moderate spinal cord injury promote functional locomotor recovery, increase spared white matter tissue, and stimulate axonal outgrowth. Moreover, tamoxifen reduces the formation of reactive oxygen species. Therefore, these drugs are possible therapeutic agents that have a neuroprotective/regen-erative activity in vertebrates with SCI.

  15. The dynamic aurora

    Energy Technology Data Exchange (ETDEWEB)

    Akasofu, Syunichi (Univ. of Alaska, Fairbanks (USA))

    1989-05-01

    Auroral physicists today know that auroras are lights emitted when atoms and molecules in the ionosphere are struck by electrons blowing in from the sun. The apparent motion of the auroral curtain is caused not by atmospheric turbulence but by changes in the electromagnetic conditions that propel the electrons, just as motion on a television screen is an illusion created by changes in the magnetic field that directs electrons from a cathode tube onto the screen. In the case of the aurora, what serves as the cathode tube Where is its power supply Why does that power seem to fluctuate from time to time, causing the aurora to ebb and flow across the polar sky More than 20 years ago it was determined that auroral emissions occur because the ionosphere is bombarded by electron beams generated by a complex interaction between the solar wind and the earth's magnetic envelope. The geomagnetic nature of the aurora can be seen clearly from outer space. Centered on each of the geomagnetic poles is a great luminous oval that is a permanent feature of the planet. The one around the North Pole is the aurora borealis. Its counterpart in the south is the aurora australis. The authors and his colleagues have now found a numerical relation between the solar wind and the generation of the power that drives the aurora, as well as other disturbances of the geomagnetic field. They are also gaining a better idea of how the sun's activity affects the solar wind. This progress raises the interesting possibility of developing a computer method to predict the intensity of auroral activity. Because auroras can interfere significantly with radio and satellite communications, as well as with power lines and some defense systems, the ability to predict auroral intensity has become increasingly important as human activity expands into polar regions and into space.

  16. Molecular mechanism of hepatitis B virus (HBV) on suppression of raf kinase inhibitor protein (RKIP) expression

    Science.gov (United States)

    Cheng, Xiao-Ke; Yu, Guo-Zheng; Li, Xiao-Dong; Ren, Xue-Qun

    2017-01-01

    Raf kinase inhibitor protein (RKIP) has been shown to be a suppressor of the mitogen-activated protein kinase pathway and is reported to be involved in human malignancy. However, the molecular mechanism of hepatitis B virus (HBV) in regulating RKIP expression is not yet clarified. In this study, we compared RKIP expression in 107 pairs of matched liver cancer and adjacent non-cancerous liver tissues. Among seven HBV-encoded proteins, we found HBV X (HBX) protein could significantly inhibit the expression level of RKIP, indicating that HBV could suppress RKIP expression through regulating HBX. To further elucidate the mechanism, analyses on transcriptional regulation and promoter methylation inhibition were conducted in Huh7 cells. Our results showed that HBX can interact with AP1 protein to inhibit the RKIP transcription. Moreover, we observed that the promoter methylation level of RKIP could be enhanced by HBV. In conclusion, our study revealed that RKIP could act as a molecular marker for HBV-infected liver cancer, but had no tumor-suppressing effect. PMID:27902472

  17. Rho kinase inhibitor Y-27632 and Accutase dramatically increase mouse embryonic stem cell derivation.

    Science.gov (United States)

    Zhang, Peng; Wu, Xinglong; Hu, Chunchao; Wang, Pengbo; Li, Xiangyun

    2012-01-01

    Although it has been 30 yr since the development of derivation methods for mouse embryonic stem (ES) cells, the biology of derivation of ES cells is poorly understood and the efficiency varies dramatically between cell lines. Recently, the Rho kinase inhibitor Y-27632 and the cell dissociation reagent Accutase were reported to significantly inhibit apoptosis of human ES cells during passaging. Therefore, in the current study, C57BL/6×129/Sv mouse blastocysts were used to evaluate the effect of the combination of the two reagents instead of using the conventional 129 line in mouse ES cell derivation. The data presented in this study suggests that the combination of Y-27632 and Accutase significantly increases the efficiency of mouse ES cell derivation; furthermore, no negative side effects were observed with Y-27632 and Accutase treatment. The newly established ES cell lines retain stable karyotype, surface markers expression, formed teratomas, and contributed to viable chimeras and germline transmission by tetraploid complementation assay. In addition, Y-27632 improved embryoid body formation of ES cells. During ES cell microinjection, Y-27632 prevented the formation of dissociation-induced cell blebs and facilitates the selection and the capture of intact cells. The methods presented in this study clearly demonstrate that inhibition of Rho kinase with Y-27632 and Accutase dissociation improve the derivation efficiently and reproducibility of mouse ES cell generation which is essential for reducing variability in the results obtained from different cell lines.

  18. Fasting potentiates the anticancer activity of tyrosine kinase inhibitors by strengthening MAPK signaling inhibition.

    Science.gov (United States)

    Caffa, Irene; D'Agostino, Vito; Damonte, Patrizia; Soncini, Debora; Cea, Michele; Monacelli, Fiammetta; Odetti, Patrizio; Ballestrero, Alberto; Provenzani, Alessandro; Longo, Valter D; Nencioni, Alessio

    2015-05-20

    Tyrosine kinase inhibitors (TKIs) are now the mainstay of treatment in many types of cancer. However, their benefit is frequently short-lived, mandating the search for safe potentiation strategies. Cycles of fasting enhance the activity of chemo-radiotherapy in preclinical cancer models and dietary approaches based on fasting are currently explored in clinical trials. Whether combining fasting with TKIs is going to be potentially beneficial remains unknown. Here we report that starvation conditions increase the ability of commonly administered TKIs, including erlotinib, gefitinib, lapatinib, crizotinib and regorafenib, to block cancer cell growth, to inhibit the mitogen-activated protein kinase (MAPK) signaling pathway and to strengthen E2F-dependent transcription inhibition. In cancer xenografts models, both TKIs and cycles of fasting slowed tumor growth, but, when combined, these interventions were significantly more effective than either type of treatment alone. In conclusion, cycles of fasting or of specifically designed fasting-mimicking diets should be evaluated in clinical studies as a means to potentiate the activity of TKIs in clinical use.

  19. Kinase-dead ATM protein is highly oncogenic and can be preferentially targeted by Topo-isomerase I inhibitors.

    Science.gov (United States)

    Yamamoto, Kenta; Wang, Jiguang; Sprinzen, Lisa; Xu, Jun; Haddock, Christopher J; Li, Chen; Lee, Brian J; Loredan, Denis G; Jiang, Wenxia; Vindigni, Alessandro; Wang, Dong; Rabadan, Raul; Zha, Shan

    2016-06-15

    Missense mutations in ATM kinase, a master regulator of DNA damage responses, are found in many cancers, but their impact on ATM function and implications for cancer therapy are largely unknown. Here we report that 72% of cancer-associated ATM mutations are missense mutations that are enriched around the kinase domain. Expression of kinase-dead ATM (Atm(KD/-)) is more oncogenic than loss of ATM (Atm(-/-)) in mouse models, leading to earlier and more frequent lymphomas with Pten deletions. Kinase-dead ATM protein (Atm-KD), but not loss of ATM (Atm-null), prevents replication-dependent removal of Topo-isomerase I-DNA adducts at the step of strand cleavage, leading to severe genomic instability and hypersensitivity to Topo-isomerase I inhibitors. Correspondingly, Topo-isomerase I inhibitors effectively and preferentially eliminate Atm(KD/-), but not Atm-proficientor Atm(-/-) leukemia in animal models. These findings identify ATM kinase-domain missense mutations as a potent oncogenic event and a biomarker for Topo-isomerase I inhibitor based therapy.

  20. Targeting Vascular Endothelial Growth Factor Receptor 2 and Protein Kinase D1 Related Pathways by a Multiple Kinase Inhibitor in Angiogenesis and Inflammation Related Processes In Vitro

    OpenAIRE

    Attila Varga; Pál Gyulavári; Zoltán Greff; Krisztina Futosi; Tamás Németh; Laura Simon-Szabó; Krisztina Kerekes; Csaba Szántai-Kis; Diána Brauswetter; Márton Kokas; Gábor Borbély; Anna Erdei; Attila Mócsai; György Kéri; Tibor Vántus

    2015-01-01

    Emerging evidence suggests that the vascular endothelial growth factor receptor 2 (VEGFR2) and protein kinase D1 (PKD1) signaling axis plays a critical role in normal and pathological angiogenesis and inflammation related processes. Despite all efforts, the currently available therapeutic interventions are limited. Prior studies have also proved that a multiple target inhibitor can be more efficient compared to a single target one. Therefore, development of novel inflammatory pathway-specific...

  1. The discovery of potent ribosomal S6 kinase inhibitors by high-throughput screening and structure-guided drug design.

    Science.gov (United States)

    Couty, Sylvain; Westwood, Isaac M; Kalusa, Andrew; Cano, Celine; Travers, Jon; Boxall, Kathy; Chow, Chiau Ling; Burns, Sam; Schmitt, Jessica; Pickard, Lisa; Barillari, Caterina; McAndrew, P Craig; Clarke, Paul A; Linardopoulos, Spiros; Griffin, Roger J; Aherne, G Wynne; Raynaud, Florence I; Workman, Paul; Jones, Keith; van Montfort, Rob L M

    2013-10-01

    The ribosomal P70 S6 kinases play a crucial role in PI3K/mTOR regulated signalling pathways and are therefore potential targets for the treatment of a variety of diseases including diabetes and cancer. In this study we describe the identification of three series of chemically distinct S6K1 inhibitors. In addition, we report a novel PKA-S6K1 chimeric protein with five mutations in or near its ATP-binding site, which was used to determine the binding mode of two of the three inhibitor series, and provided a robust system to aid the optimisation of the oxadiazole-substituted benzimidazole inhibitor series. We show that the resulting oxadiazole-substituted aza-benzimidazole is a potent and ligand efficient S6 kinase inhibitor, which blocks the phosphorylation of RPS6 at Ser235/236 in TSC negative HCV29 human bladder cancer cells by inhibiting S6 kinase activity and thus provides a useful tool compound to investigate the function of S6 kinases.

  2. Proteomic analysis of human metaphase chromosomes reveals Topoisomerase II alpha as an Aurora B substrate

    DEFF Research Database (Denmark)

    Morrison, Ciaran; Henzing, Alexander J; Jensen, Ole Nørregaard;

    2002-01-01

    The essential Aurora B kinase is a chromosomal passenger protein that is required for mitotic chromosome alignment and segregation. Aurora B function is dependent on the chromosome passenger, INCENP. INCENP, in turn, requires sister chromatid cohesion for its appropriate behaviour. Relatively few...

  3. Discovery and Optimization of Macrocyclic Quinoxaline-pyrrolo-dihydropiperidinones as Potent Pim-1/2 Kinase Inhibitors.

    Science.gov (United States)

    Cee, Victor J; Chavez, Frank; Herberich, Bradley; Lanman, Brian A; Pettus, Liping H; Reed, Anthony B; Wu, Bin; Wurz, Ryan P; Andrews, Kristin L; Chen, Jie; Hickman, Dean; Laszlo, Jimmy; Lee, Matthew R; Guerrero, Nadia; Mattson, Bethany K; Nguyen, Yen; Mohr, Christopher; Rex, Karen; Sastri, Christine E; Wang, Paul; Wu, Qiong; Wu, Tian; Xu, Yang; Zhou, Yihong; Winston, Jeffrey T; Lipford, J Russell; Tasker, Andrew S; Wang, Hui-Ling

    2016-04-14

    The identification of Pim-1/2 kinase overexpression in B-cell malignancies suggests that Pim kinase inhibitors will have utility in the treatment of lymphoma, leukemia, and multiple myeloma. Starting from a moderately potent quinoxaline-dihydropyrrolopiperidinone lead, we recognized the potential for macrocyclization and developed a series of 13-membered macrocycles. The structure-activity relationships of the macrocyclic linker were systematically explored, leading to the identification of 9c as a potent, subnanomolar inhibitor of Pim-1 and -2. This molecule also potently inhibited Pim kinase activity in KMS-12-BM, a multiple myeloma cell line with relatively high endogenous levels of Pim-1/2, both in vitro (pBAD IC50 = 25 nM) and in vivo (pBAD EC50 = 30 nM, unbound), and a 100 mg/kg daily dose was found to completely arrest the growth of KMS-12-BM xenografts in mice.

  4. Review : Third Generation Tyrosine Kinase Inhibitors and Their Development in Advanced Renal Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Ronald M Bukowski

    2012-02-01

    Full Text Available Angiogenesis in general and the VEGF signaling axis in particular is a validated target in renal cell carcinoma. Clear cell carcinoma of the kidney is now recognized as a malignancy that is sensitive to inhibitors of the vascular endothelial growth factor pathway. Treatment options for patients with metastatic renal cell carcinoma have evolved in dramatic fashion over the past six years, and a new paradigm has developed. The cytokines interferon-α and interleukin-2 were previously utilized for therapy, but since December 2005, six new agents have been approved in the United States for the treatment of advanced RCC. Three are tyrosine kinase inhibitors (TKI’s including sunitinib, sorafenib, and recently pazopanib. The current review examines the evolving data with the next generation of TKI’s, axitinib and tivozanib being developed for the treatment of advanced RCC. These agents were synthesized to provide increased target specificity and enhanced target inhibition. The preclinical and clinical data are examined, an overview of the development of these TKI’s is provided, and discussion plus speculation concerning their potential roles as RCC therapy is provided.

  5. Cyclin-dependent kinase inhibitor p20 controls circadian cell-cycle timing.

    Science.gov (United States)

    Laranjeiro, Ricardo; Tamai, T Katherine; Peyric, Elodie; Krusche, Peter; Ott, Sascha; Whitmore, David

    2013-04-23

    Specific stages of the cell cycle are often restricted to particular times of day because of regulation by the circadian clock. In zebrafish, both mitosis (M phase) and DNA synthesis (S phase) are clock-controlled in cell lines and during embryo development. Despite the ubiquitousness of this phenomenon, relatively little is known about the underlying mechanism linking the clock to the cell cycle. In this study, we describe an evolutionarily conserved cell-cycle regulator, cyclin-dependent kinase inhibitor 1d (20 kDa protein, p20), which along with p21, is a strongly rhythmic gene and directly clock-controlled. Both p20 and p21 regulate the G1/S transition of the cell cycle. However, their expression patterns differ, with p20 predominant in developing brain and peak expression occurring 6 h earlier than p21. p20 expression is also p53-independent in contrast to p21 regulation. Such differences provide a unique mechanism whereby S phase is set to different times of day in a tissue-specific manner, depending on the balance of these two inhibitors.

  6. Anilides of (R)-trifluoro-2-hydroxy-2-methylpropionic acid as inhibitors of pyruvate dehydrogenase kinase.

    Science.gov (United States)

    Bebernitz, G R; Aicher, T D; Stanton, J L; Gao, J; Shetty, S S; Knorr, D C; Strohschein, R J; Tan, J; Brand, L J; Liu, C; Wang, W H; Vinluan, C C; Kaplan, E L; Dragland, C J; DelGrande, D; Islam, A; Lozito, R J; Liu, X; Maniara, W M; Mann, W R

    2000-06-01

    The optimization of a series of anilide derivatives of (R)-3,3, 3-trifluoro-2-hydroxy-2-methylpropionic acid as inhibitors of pyruvate dehydrogenase kinase (PDHK) is described that started from N-phenyl-3,3,3-trifluoro-2-hydroxy-2-methylpropanamide 1 (IC(50) = 35 +/- 1.4 microM). It was found that small electron-withdrawing groups on the ortho position of the anilide, i.e., chloro, acetyl, or bromo, increased potency 20-40-fold. The oral bioavailability of the compounds in this series is optimal (as measured by AUC) when the anilide is substituted at the 4-position with an electron-withdrawing group (i.e., carboxyl, carboxyamide, and sulfoxyamide). N-(2-Chloro-4-isobutylsulfamoylphenyl)-(R)-3,3, 3-trifluoro-2-hydroxy-2-methylpropionamide (10a) inhibits PDHK in the primary enzymatic assay with an IC(50) of 13 +/- 1.5 nM, enhances the oxidation of [(14)C]lactate into (14)CO(2) in human fibroblasts, lowers blood lactate levels significantly 2.5 and 5 h after oral doses as low as 30 micromol/kg, and increases the ex vivo activity of PDH in muscle, kidney, liver, and heart tissues. However, in contrast to sodium dichloroacetate (DCA), these PDHK inhibitors did not lower blood glucose levels. Nevertheless, they are effective at increasing the utilization and disposal of lactate and could be of utility to ameliorate conditions of inappropriate blood lactate elevation.

  7. Tirosin Kinase Inhibitors in Chronic Graft versus Host Disease: From Bench to Bedside

    Directory of Open Access Journals (Sweden)

    Jacopo Olivieri

    2011-01-01

    Full Text Available Chronic Graft Versus Host Disease (cGVHD is a major complication of allogeneic stem-cell transplantation (SCT. In many inflammatory fibrotic diseases, such as Systemic Scleroderma (SSc and cGVHD with fibrotic features, an abnormal activation of transforming growth factor (TGFβ and platelet-derived growth factor receptor (PDGF-R pathways have been observed. Tyrosin Kinase Inhibitors (TKIs, which are currently used for treatment of patients with Chronic Myeloid Leukemia (CML, share potent antifibrotic and antiinflammatory properties, being powerful dual inhibitors of both PDGF-R and TGFβ pathways. Moreover accumulating in vitro data confirm that TKIs, interacting with the TCR and other signalling molecules, carry potent immunomodulatory effects, being involved in both T-cell and B-cell response. Translation to the clinical setting revealed that treatment with Imatinib can achieve encouraging responses in patients with autoimmune diseases and steroid-refractory cGVHD, showing a favourable toxicity profile. While the exact mechanisms leading to such efficacy are still under investigation, use of TKIs in the context of clinical trials should be promoted, aiming to evaluate the biological changes induced in vivo by TKIs and to assess the long term outcome of these patients. Second-generation TKIs, with more favourable toxicity profile are under evaluation in the same setting.

  8. Phosphatidylinositol 3-kinase inhibitor, LY294002, induced senescence-like changes in human diploid fibroblasts

    Institute of Scientific and Technical Information of China (English)

    李淑萍; 张宗玉; 童坦君

    2003-01-01

    Objective To reveal the role of Phosphatidylinositol 3-kinases (PI3Ks) in regulating human diploid fibroblast (2BS cell) senescence as well as the possible mechanisms involved.Methods Using a PI3Ks specific inhibitor, LY294002, cell cycle, apoptosis, proliferation, senescence association β-galactosidase staining as well as senescence association CKIs, p16 INK4 and p21 Cip1 protein expressions were all measured in the low passages of 2BS cells.Results Both 25 μmol/L and 50 μmol/L concentrations of LY294002 could cause a significant decrease in cells entering into S phase, and this cell cycle of G 1 phase arrest was dose-dependent. Meanwhile, LY294002 contributed to apoptosis, caused 2BS cell growth arrest, and activated senescence association β-galactosidase (P<0.05). In addition, LY294002 could induce time-course expressions of p16 INK4 and p21 Cip1 in 2BS cell lines.Conclusions PI3Ks inhibitor LY294002 could induce senescence-like changes in 2BS cell lines. Two enescence associated CKIs,p16 INK4 and p21 Cip1, might be involved in this senescence phenotype proceeding in 2BS cell lines.

  9. 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.

  10. Genetic and Pharmacological Inhibition of PDK1 in Cancer Cells: Characterization of a Selective Allosteric Kinase Inhibitor

    Energy Technology Data Exchange (ETDEWEB)

    Nagashima, Kumiko; Shumway, Stuart D.; Sathyanarayanan, Sriram; Chen, Albert H.; Dolinski, Brian; Xu, Youyuan; Keilhack, Heike; Nguyen, Thi; Wiznerowicz, Maciej; Li, Lixia; Lutterbach, Bart A.; Chi, An; Paweletz, Cloud; Allison, Timothy; Yan, Youwei; Munshi, Sanjeev K.; Klippel, Anke; Kraus, Manfred; Bobkova, Ekaterina V.; Deshmukh, Sujal; Xu, Zangwei; Mueller, Uwe; Szewczak, Alexander A.; Pan, Bo-Sheng; Richon, Victoria; Pollock, Roy; Blume-Jensen, Peter; Northrup, Alan; Andersen, Jannik N. (Merck)

    2013-11-20

    Phosphoinositide-dependent kinase 1 (PDK1) is a critical activator of multiple prosurvival and oncogenic protein kinases and has garnered considerable interest as an oncology drug target. Despite progress characterizing PDK1 as a therapeutic target, pharmacological support is lacking due to the prevalence of nonspecific inhibitors. Here, we benchmark literature and newly developed inhibitors and conduct parallel genetic and pharmacological queries into PDK1 function in cancer cells. Through kinase selectivity profiling and x-ray crystallographic studies, we identify an exquisitely selective PDK1 inhibitor (compound 7) that uniquely binds to the inactive kinase conformation (DFG-out). In contrast to compounds 1-5, which are classical ATP-competitive kinase inhibitors (DFG-in), compound 7 specifically inhibits cellular PDK1 T-loop phosphorylation (Ser-241), supporting its unique binding mode. Interfering with PDK1 activity has minimal antiproliferative effect on cells growing as plastic-attached monolayer cultures (i.e. standard tissue culture conditions) despite reduced phosphorylation of AKT, RSK, and S6RP. However, selective PDK1 inhibition impairs anchorage-independent growth, invasion, and cancer cell migration. Compound 7 inhibits colony formation in a subset of cancer cell lines (four of 10) and primary xenograft tumor lines (nine of 57). RNAi-mediated knockdown corroborates the PDK1 dependence in cell lines and identifies candidate biomarkers of drug response. In summary, our profiling studies define a uniquely selective and cell-potent PDK1 inhibitor, and the convergence of genetic and pharmacological phenotypes supports a role of PDK1 in tumorigenesis in the context of three-dimensional in vitro culture systems.

  11. AuroraMAX!

    Science.gov (United States)

    Donovan, E.; Spanswick, E. L.; Chicoine, R.; Pugsley, J.; Langlois, P.

    2011-12-01

    AuroraMAX is a public outreach and education initiative that brings auroral images to the public in real time. AuroraMAX utilizes an observing station located just outside Yellowknife, Canada. The station houses a digital All-Sky Imager (ASI) that collects full-colour images of the night sky every six seconds. These images are then transmitted via satellite internet to our web server, where they are made instantly available to the public. Over the last two years this program has rapidly become one of the most successful outreach programs in the history of Space Science in Canada, with hundreds of thousands of distinct visitors to the CSA AuroraMAX website, thousands of followers on social media, and hundreds of newspaper, magazine, radio, and television spots. Over the next few years, the project will expand to include a high-resolution SLR delivering real-time auroral images (also from Yellowknife), as well as a program where astronauts on the ISS will take pictures of the aurora with a handheld SLR. The objectives of AuroraMAX are public outreach and education. The ASI design, operation, and software were based on infrastructure that was developed for the highly successful ASI component of the NASA THEMIS mission as well as the Canadian Space Agency (CSA) Canadian GeoSpace Monitoring (CGSM) program. So from an education and public outreach perspective, AuroraMAX is a single camera operating in the Canadian north. On the other hand, AuroraMAX is one of nearly 40 All-Sky Imagers that are operating across North America. The AuroraMAX camera produces data that is seamlessly integrated with the CGSM ASI data, and made widely available to the Space Science community through open-access web and FTP sites. One of our objectives in the next few years is to incorporate some of the data from the THEMIS and CGSM imagers into the AuroraMAX system, to maximize viewing opportunities and generate more real-time data for public outreach. This is an exemplar of a program that

  12. Converting potent indeno[1,2-b]indole inhibitors of protein kinase CK2 into selective inhibitors of the breast cancer resistance protein ABCG2.

    Science.gov (United States)

    Jabor Gozzi, Gustavo; Bouaziz, Zouhair; Winter, Evelyn; Daflon-Yunes, Nathalia; Aichele, Dagmar; Nacereddine, Abdelhamid; Marminon, Christelle; Valdameri, Glaucio; Zeinyeh, Waël; Bollacke, Andre; Guillon, Jean; Lacoudre, Aline; Pinaud, Noël; Cadena, Silvia M; Jose, Joachim; Le Borgne, Marc; Di Pietro, Attilio

    2015-01-08

    A series of indeno[1,2-b]indole-9,10-dione derivatives were synthesized as human casein kinase II (CK2) inhibitors. The most potent inhibitors contained a N(5)-isopropyl substituent on the C-ring. The same series of compounds was found to also inhibit the breast cancer resistance protein ABCG2 but with totally different structure-activity relationships: a N(5)-phenethyl substituent was critical, and additional hydrophobic substituents at position 7 or 8 of the D-ring or a methoxy at phenethyl position ortho or meta also contributed to inhibition. The best ABCG2 inhibitors, such as 4c, 4h, 4i, 4j, and 4k, behaved as very weak inhibitors of CK2, whereas the most potent CK2 inhibitors, such as 4a, 4p, and 4e, displayed limited interaction with ABCG2. It was therefore possible to convert, through suitable substitutions of the indeno[1,2-b]indole-9,10-dione scaffold, potent CK2 inhibitors into selective ABCG2 inhibitors and vice versa. In addition, some of the best ABCG2 inhibitors, which displayed a very low cytotoxicity, thus giving a high therapeutic ratio, and appeared not to be transported, constitute promising candidates for further investigations.

  13. A platinum-based hybrid drug design approach to circumvent acquired resistance to molecular targeted tyrosine kinase inhibitors

    Science.gov (United States)

    Wei, Yuming; Poon, Daniel C.; Fei, Rong; Lam, Amy S. M.; Au-Yeung, Steve C. F.; To, Kenneth K. W.

    2016-05-01

    Three molecular targeted tyrosine kinase inhibitors (TKI) were conjugated to classical platinum-based drugs with an aim to circumvent TKI resistance, predominately mediated by the emergence of secondary mutations on oncogenic kinases. The hybrids were found to maintain specificity towards the same oncogenic kinases as the original TKI. Importantly, they are remarkably less affected by TKI resistance, presumably due to their unique structure and the observed dual mechanism of anticancer activity (kinase inhibition and DNA damage). The study is also the first to report the application of a hybrid drug approach to switch TKIs from being efflux transporter substrates into non-substrates. TKIs cannot penetrate into the brain for treating metastases because of efflux transporters at the blood brain barrier. The hybrids were found to escape drug efflux and they accumulate more than the original TKI in the brain in BALB/c mice. Further development of the hybrid compounds is warranted.

  14. A platinum-based hybrid drug design approach to circumvent acquired resistance to molecular targeted tyrosine kinase inhibitors.

    Science.gov (United States)

    Wei, Yuming; Poon, Daniel C; Fei, Rong; Lam, Amy S M; Au-Yeung, Steve C F; To, Kenneth K W

    2016-05-06

    Three molecular targeted tyrosine kinase inhibitors (TKI) were conjugated to classical platinum-based drugs with an aim to circumvent TKI resistance, predominately mediated by the emergence of secondary mutations on oncogenic kinases. The hybrids were found to maintain specificity towards the same oncogenic kinases as the original TKI. Importantly, they are remarkably less affected by TKI resistance, presumably due to their unique structure and the observed dual mechanism of anticancer activity (kinase inhibition and DNA damage). The study is also the first to report the application of a hybrid drug approach to switch TKIs from being efflux transporter substrates into non-substrates. TKIs cannot penetrate into the brain for treating metastases because of efflux transporters at the blood brain barrier. The hybrids were found to escape drug efflux and they accumulate more than the original TKI in the brain in BALB/c mice. Further development of the hybrid compounds is warranted.

  15. Combination of bortezomib and mitotic inhibitors down-modulate Bcr-Abl and efficiently eliminates tyrosine-kinase inhibitor sensitive and resistant Bcr-Abl-positive leukemic cells.

    Science.gov (United States)

    Bucur, Octavian; Stancu, Andreea Lucia; Goganau, Ioana; Petrescu, Stefana Maria; Pennarun, Bodvael; Bertomeu, Thierry; Dewar, Rajan; Khosravi-Far, Roya

    2013-01-01

    Emergence of resistance to Tyrosine-Kinase Inhibitors (TKIs), such as imatinib, dasatinib and nilotinib, in Chronic Myelogenous Leukemia (CML) demands new therapeutic strategies. We and others have previously established bortezomib, a selective proteasome inhibitor, as an important potential treatment in CML. Here we show that the combined regimens of bortezomib with mitotic inhibitors, such as the microtubule-stabilizing agent Paclitaxel and the PLK1 inhibitor BI2536, efficiently kill TKIs-resistant and -sensitive Bcr-Abl-positive leukemic cells. Combined treatment activates caspases 8, 9 and 3, which correlate with caspase-induced PARP cleavage. These effects are associated with a marked increase in activation of the stress-related MAP kinases p38MAPK and JNK. Interestingly, combined treatment induces a marked decrease in the total and phosphorylated Bcr-Abl protein levels, and inhibits signaling pathways downstream of Bcr-Abl: downregulation of STAT3 and STAT5 phosphorylation and/or total levels and a decrease in phosphorylation of the Bcr-Abl-associated proteins CrkL and Lyn. Moreover, we found that other mitotic inhibitors (Vincristine and Docetaxel), in combination with bortezomib, also suppress the Bcr-Abl-induced pro-survival signals and result in caspase 3 activation. These results open novel possibilities for the treatment of Bcr-Abl-positive leukemias, especially in the imatinib, dasatinib and nilotinib-resistant CML cases.

  16. The phosphatidylinositol 3-kinases (PI3K) inhibitor GS-1101 synergistically potentiates histone deacetylase inhibitor-induced proliferation inhibition and apoptosis through the inactivation of PI3K and extracellular signal-regulated kinase pathways.

    Science.gov (United States)

    Bodo, Juraj; Zhao, Xiaoxian; Sharma, Arishya; Hill, Brian T; Portell, Craig A; Lannutti, Brian J; Almasan, Alexandru; Hsi, Eric D

    2013-10-01

    Previously, we showed that inhibition of the protein kinase C β (PKCβ)/AKT pathway augments engagement of the histone deacetylase inhibitor (HDI)-induced apoptosis in lymphoma cells. In the present study, we investigated the cytotoxicity and mechanisms of cell death induced by the delta isoform-specific phosphatidylinositide 3-kinase (PI3K) inhibitor, GS-1101, in combination with the HDI, panobinostat (LBH589) and suberoylanilide hydroxamic acid (SAHA). Lymphoma cell lines, primary non-Hodgkin Lymphoma (NHL) and chronic lymphocytic leukaemia (CLL) cells were simultaneously treated with the HDI, LBH589 and GS-1101. An interaction of the LBH589/GS-1101 combination was formally examined by using various concentrations of LBH589 and GS-1101. Combined treatment resulted in a synergistic inhibition of proliferation and showed synergistic effect on apoptotic induction in all tested cell lines and primary NHL and CLL cells. This study indicates that interference with PI3K signalling dramatically increases HDI-mediated apoptosis in malignant haematopoietic cells, possibly through both AKT-dependent or AKT- independent mechanisms. Moreover, the increase in HDI-related apoptosis observed in PI3K inhibitor-treated cells appears to be related to the disruption of the extracellular signal-regulated kinase (ERK) signalling pathway. This study provides a strong rational for testing the combination of PI3K inhibitors and HDI in the clinic.

  17. Inhibition of the signal transduction through the AtoSC system by histidine kinase inhibitors in Escherichia coli.

    Science.gov (United States)

    Theodorou, Evaggelos C; Theodorou, Marina C; Kyriakidis, Dimitrios A

    2011-08-01

    AtoSC two-component system participates in many indispensable processes of Escherichia coli. We report here that the AtoSC signal transduction is inhibited by established histidine kinase inhibitors. Closantel, RWJ-49815 and TNP-ATP belonging to different chemical classes of inhibitors, abrogated the in vitro AtoS kinase autophosphorylation. However, when AtoS was embedded in the membrane fractions, higher inhibitor concentrations were required for total inhibition. When AtoS interacted with AtoC forming complex, the intrinsic histidine kinase was protected by the response regulator, requiring increased inhibitors concentrations for partially AtoS autophosphorylation reduction. The inhibitors exerted an additional function on AtoSC, blocking the phosphotransfer from AtoS to AtoC, without however, affecting AtoC~P dephosphorylation. Their in vivo consequences through the AtoSC inhibition were elucidated on atoDAEB operon expression, which was inhibited only in AtoSC-expressing bacteria where AtoSC was induced by acetoacetate or spermidine. The inhibitor effects were extended on the AtoSC regulatory role on cPHB [complexed poly-(R)-3-hydroxybutyrate] biosynthesis. cPHB was decreased upon the blockers only in acetoacetate-induced AtoSC-expressing cells. Increased ATP amounts during bacterial growth reversed the inhibitory TNP-ATP-mediated effect on cPHB. The alteration of pivotal E. coli processes as an outcome of AtoSC inhibition, establish this system as a target of two-component systems inhibitors.

  18. Receptor tyrosine kinase (c-Kit inhibitors: a potential therapeutic target in cancer cells

    Directory of Open Access Journals (Sweden)

    Abbaspour Babaei M

    2016-08-01

    Full Text Available Maryam Abbaspour Babaei,1 Behnam Kamalidehghan,2,3 Mohammad Saleem,4–6 Hasniza Zaman Huri,1,7 Fatemeh Ahmadipour1 1Department of Pharmacy, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia; 2Department of Medical Genetics, National Institute of Genetic Engineering and Biotechnology (NIGEB, Shahrak-e Pajoohesh, 3Medical Genetics Department, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran; 4Department of Urology, 5Department of Laboratory Medicine and Pathology, Masonic Cancer Center, University of Minnesota, 6Section of Molecular Therapeutics & Cancer Health Disparity, The Hormel Institute, Austin, MN, USA; 7Clinical Investigation Centre, University Malaya Medical Centre, Lembah Pantai, Kuala Lumpur, Malaysia Abstract: c-Kit, a receptor tyrosine kinase, is involved in intracellular signaling, and the mutated form of c-Kit plays a crucial role in occurrence of some cancers. The function of c-Kit has led to the concept that inhibiting c-Kit kinase activity can be a target for cancer therapy. The promising results of inhibition of c-Kit for treatment of cancers have been observed in some cancers such as gastrointestinal stromal tumor, acute myeloid leukemia, melanoma, and other tumors, and these results have encouraged attempts toward improvement of using c-Kit as a capable target for cancer therapy. This paper presents the findings of previous studies regarding c-Kit as a receptor tyrosine kinase and an oncogene, as well as its gene targets and signaling pathways in normal and cancer cells. The c-Kit gene location, protein structure, and the role of c-Kit in normal cell have been discussed. Comprehending the molecular mechanism underlying c-Kit-mediated tumorogenesis is consequently essential and may lead to the identification of future novel drug targets. The potential mechanisms by which c-Kit induces cellular transformation have been described. This study aims to elucidate the function of c

  19. Discovery of a novel allosteric inhibitor-binding site in ERK5: comparison with the canonical kinase hinge ATP-binding site.

    Science.gov (United States)

    Chen, Hongming; Tucker, Julie; Wang, Xiaotao; Gavine, Paul R; Phillips, Chris; Augustin, Martin A; Schreiner, Patrick; Steinbacher, Stefan; Preston, Marian; Ogg, Derek

    2016-05-01

    MAP kinases act as an integration point for multiple biochemical signals and are involved in a wide variety of cellular processes such as proliferation, differentiation, regulation of transcription and development. As a member of the MAP kinase family, ERK5 (MAPK7) is involved in the downstream signalling pathways of various cell-surface receptors, including receptor tyrosine kinases and G protein-coupled receptors. In the current study, five structures of the ERK5 kinase domain co-crystallized with ERK5 inhibitors are reported. Interestingly, three of the compounds bind at a novel allosteric binding site in ERK5, while the other two bind at the typical ATP-binding site. Binding of inhibitors at the allosteric site is accompanied by displacement of the P-loop into the ATP-binding site and is shown to be ATP-competitive in an enzymatic assay of ERK5 kinase activity. Kinase selectivity data show that the most potent allosteric inhibitor exhibits superior kinase selectivity compared with the two inhibitors that bind at the canonical ATP-binding site. An analysis of these structures and comparison with both a previously published ERK5-inhibitor complex structure (PDB entry 4b99) and the structures of three other kinases (CDK2, ITK and MEK) in complex with allosteric inhibitors are presented.

  20. The CUL3-KLHL18 ligase regulates mitotic entry and ubiquitylates Aurora-A.

    Science.gov (United States)

    Moghe, Saili; Jiang, Fei; Miura, Yoshie; Cerny, Ronald L; Tsai, Ming-Ying; Furukawa, Manabu

    2012-02-15

    The cullin-RING family of ubiquitin ligases regulates diverse cellular functions, such as cell cycle control, via ubiquitylation of specific substrates. CUL3 targets its substrates through BTB proteins. Here we show that depletion of CUL3 and the BTB protein KLHL18 causes a delay in mitotic entry. Centrosomal activation of Aurora-A, a kinase whose activity is required for entry into mitosis, is also delayed in depleted cells. Moreover, we identify Aurora-A as a KLHL18-interacting partner. Overexpression of KLHL18 and CUL3 promotes Aurora-A ubiquitylation in vivo, and the CUL3-KLHL18-ROC1 ligase ubiquitylates Aurora-A in vitro. Our study reveals that the CUL3-KLHL18 ligase is required for timely entry into mitosis, as well as for the activation of Aurora-A at centrosomes. We propose that the CUL3-KLHL18 ligase regulates mitotic entry through an Aurora-A-dependent pathway.

  1. The CUL3-KLHL18 ligase regulates mitotic entry and ubiquitylates Aurora-A

    Directory of Open Access Journals (Sweden)

    Saili Moghe

    2012-02-01

    The cullin-RING family of ubiquitin ligases regulates diverse cellular functions, such as cell cycle control, via ubiquitylation of specific substrates. CUL3 targets its substrates through BTB proteins. Here we show that depletion of CUL3 and the BTB protein KLHL18 causes a delay in mitotic entry. Centrosomal activation of Aurora-A, a kinase whose activity is required for entry into mitosis, is also delayed in depleted cells. Moreover, we identify Aurora-A as a KLHL18-interacting partner. Overexpression of KLHL18 and CUL3 promotes Aurora-A ubiquitylation in vivo, and the CUL3-KLHL18-ROC1 ligase ubiquitylates Aurora-A in vitro. Our study reveals that the CUL3-KLHL18 ligase is required for timely entry into mitosis, as well as for the activation of Aurora-A at centrosomes. We propose that the CUL3-KLHL18 ligase regulates mitotic entry through an Aurora-A-dependent pathway.

  2. Identification of green tea catechins as potent inhibitors of the polo-box domain of polo-like kinase 1.

    Science.gov (United States)

    Shan, Hong-Mei; Shi, Yanxia; Quan, Junmin

    2015-01-01

    Polo-like kinase 1 (PLK1) plays crucial functions in multiple stages of mitosis and is considered to be a potential drug target for cancer therapy. The functions of PLK1 are mediated by its N-terminal kinase domain and C-terminal polo-box domain (PBD). Most inhibitors targeting the kinase domain of PLK1 have a selectivity issue because of a high degree of structural conservation within kinase domains of all protein kinases. Here, we combined virtual and experimental screenings to identify green tea catechins as potent inhibitors of the PLK1 PBD. Initially, (-)-epigallocatechin, one of the main components of green tea polyphenols, was found to significantly block the binding of fluorescein-labeled phosphopeptide to the PBD at a concentration of 10 μm. Next, additional catechins were evaluated for their dose-dependent inhibition of the PBD and preliminary structure-activity relationships were derived. Cellular analysis further showed that catechins interfere with the proper subcellular localization of PLK1, lead to cell-cycle arrest in the S and G2M phases, and induce growth inhibition of several human cancer cell types, such as breast adenocarcinoma (MCF7), lung adenocarcinoma (A549), and cervical adenocarcinoma (HeLa). Our data provides new insight into understanding the anticancer activities of green tea catechins.

  3. Urolithin as a converging scaffold linking ellagic acid and coumarin analogues: design of potent protein kinase CK2 inhibitors.

    Science.gov (United States)

    Cozza, Giorgio; Gianoncelli, Alessandra; Bonvini, Paolo; Zorzi, Elisa; Pasquale, Riccardo; Rosolen, Angelo; Pinna, Lorenzo A; Meggio, Flavio; Zagotto, Giuseppe; Moro, Stefano

    2011-12-09

    Casein kinase 2 (CK2) is a ubiquitous, essential, and highly pleiotropic protein kinase; its abnormally high constitutive activity is suspected to underlie its pathogenic potential in neoplasia and other relevant diseases. Previously, using different in silico screening approaches, two potent and selective CK2 inhibitors were identified by our group: ellagic acid, a naturally occurring tannic acid derivative (K(i)=20 nM) and 3,8-dibromo-7-hydroxy-4-methylchromen-2-one (DBC, K(i)=60 nM). Comparing the crystallographic binding modes of both ellagic acid and DBC, an X-ray structure-driven merging approach was taken to design novel CK2 inhibitors with improved target affinity. A urolithin moiety is proposed as a possible bridging scaffold between the two known CK2 inhibitors, ellagic acid and DBC. Optimization of urolithin A as the bridging moiety led to the identification of 4-bromo-3,8-dihydroxy-benzo[c]chromen-6-one as a novel, potent and selective CK2 inhibitor, which shows a K(i) value of 7 nM against the protein kinase, representing a significant improvement in affinity for the target compared with the two parent fragments.

  4. Bora and Aurora-A continue to activate Plk1 in mitosis.

    Science.gov (United States)

    Bruinsma, Wytse; Macurek, Libor; Freire, Raimundo; Lindqvist, Arne; Medema, René H

    2014-02-15

    Polo-like kinase-1 (Plk1) is required for proper cell division. Activation of Plk1 requires phosphorylation on a conserved threonine in the T-loop of the kinase domain (T210). Plk1 is first phosphorylated on T210 in G2 phase by the kinase Aurora-A, in concert with its cofactor Bora. However, Bora was shown to be degraded prior to entry into mitosis, and it is currently unclear how Plk1 activity is sustained in mitosis. Here we show that the Bora-Aurora-A complex remains the major activator of Plk1 in mitosis. We show that a small amount of Aurora-A activity is sufficient to phosphorylate and activate Plk1 in mitosis. In addition, a fraction of Bora is retained in mitosis, which is essential for continued Aurora-A-dependent T210 phosphorylation of Plk1. We find that once Plk1 is activated, minimal amounts of the Bora-Aurora-A complex are sufficient to sustain Plk1 activity. Thus, the activation of Plk1 by Aurora-A may function as a bistable switch; highly sensitive to inhibition of Aurora-A in its initial activation, but refractory to fluctuations in Aurora-A activity once Plk1 is fully activated. This provides a cell with robust Plk1 activity once it has committed to mitosis.

  5. Elevated Aurora B expression contributes to chemoresistance and poor prognosis in breast cancer.

    Science.gov (United States)

    Zhang, Yiqian; Jiang, Chunling; Li, Huilan; Lv, Feng; Li, Xiaoyan; Qian, Xiaolong; Fu, Li; Xu, Bo; Guo, Xiaojing

    2015-01-01

    Aurora-B is a major kinase responsible for appropriate mitotic progression. Elevated expression of Aurora-B has been frequently associated with several types of cancer, including breast cancer. However, it is not clear whether the alteration contributes to tumor responses to therapies and prognosis. In this study, we conducted immunohistochemistry using antibodies against Aurora-B, S1981p-ATM, Ki67, and p53 in paraffin-embedded tumor tissues from 312 invasive breast cancer patients. The correlation between disease-free-survival (DFS) and Aurora-B expression was analyzed using the Kaplan-Meier method and log-rank test. A Cox proportional hazards regression analysis was used to determine whether Aurora-B was an independent prognostic factor for breast cancer. We found that Aurora-B expression was correlated with the proliferation index (P breast cancer tissues. Further we found that Aurora-B expression was associated with lymph node metastasis (P = 0.002) and histological grade (P = 0.001). Multivariate analyses indicated that elevated Aurora-B expression predicted a poor survival. In a subgroup of patients that received neoadjuvant chemotherapy, we found that elevated Aurora-B contributed to chemoresistance (P = 0.011). In conclusion, elevated Aurora-B expression in breast cancer patients contributes to chemoresistance and predicts poor prognosis.

  6. Ca2+/calmodulin-dependent kinase II contributes to inhibitor of nuclear factor-kappa B kinase complex activation in Helicobacter pylori infection.

    Science.gov (United States)

    Maubach, Gunter; Sokolova, Olga; Wolfien, Markus; Rothkötter, Hermann-Josef; Naumann, Michael

    2013-09-15

    Helicobacter pylori, a class I carcinogen, induces a proinflammatory response by activating the transcription factor nuclear factor-kappa B (NF-κB) in gastric epithelial cells. This inflammatory condition could lead to chronic gastritis, which is epidemiologically and biologically linked to the development of gastric cancer. So far, there exists no clear knowledge on how H. pylori induces the NF-κB-mediated inflammatory response. In our study, we investigated the role of Ca(2+) /calmodulin-dependent kinase II (CAMKII), calmodulin, protein kinases C (PKCs) and the CARMA3-Bcl10-MALT1 (CBM) complex in conjunction with H. pylori-induced activation of NF-κB via the inhibitor of nuclear factor-kappa B kinase (IKK) complex. We use specific inhibitors and/or RNA interference to assess the contribution of these components. Our results show that CAMKII and calmodulin contribute to IKK complex activation and thus to the induction of NF-κB in response to H. pylori infection, but not in response to TNF-α. Thus, our findings are specific for H. pylori infected cells. Neither the PKCs α, δ, θ, nor the CBM complex itself is involved in the activation of NF-κB by H. pylori. The contribution of CAMKII and calmodulin, but not PKCs/CBM to the induction of an inflammatory response by H. pylori infection augment the understanding of the molecular mechanism involved and provide potential new disease markers for the diagnosis of gastric inflammatory diseases including gastric cancer.

  7. Regeneration of human epidermis on acellular dermis is impeded by small-molecule inhibitors of EGF receptor tyrosine kinase.

    Science.gov (United States)

    Forsberg, Sofi; Ostman, Arne; Rollman, Ola

    2008-10-01

    The family of human epidermal growth factor receptors (EGFR, HER2-4) exerts key functions in normal and malignant epithelial cells. Both EGFR and HER2 are valuable targets for anti-cancer drugs by interfering with ligand binding, receptor dimerization, or tyrosine kinase activity. A similar therapeutic strategy has been advocated for chronic psoriasis since plaque lesions overexpress EGFR and its ligands. Our aim was to characterize EGFR/HER2 protein expression in skin cultures and to evaluate the effects of tyrosine kinase inhibitors on epidermal outgrowth, morphology, and EGFR activation. Human skin explants were established on cell-free dermis and cultured at the air-liquid interface. The impact of small-molecule HER inhibitors on outgrowth was assayed by fluorescence-based image analysis and histometry. Effects of a dual EGFR/HER2 kinase inhibitor, PKI166, on neoepidermis were studied by immunohistochemistry and Western blot. Receptor immunostaining showed in vivo-like distributions with highest EGFR intensity in the proliferative layers whereas HER2 was mainly expressed by suprabasal keratinocytes. Reepithelialization was associated with EGFR autophosphorylation irrespective of exogenous ligand stimulation. PKI166 inhibited neoepidermal EGFR activation, keratinocyte proliferation, and outgrowth from normal and psoriatic skin explants. The rate of epidermalization in presence of other HER inhibitors varied suggesting that drug specificity, potency, and reversibility determine the dynamic outcome. Overall, agents predominantly targeting EGFR kinase were more efficient inhibitors of epidermal regeneration than an HER2-selective drug. The study illustrates the usefulness of a dynamic skin model and emphasizes the potential of HER-directed approaches to control epidermal growth in hyperproliferative skin disorders.

  8. Next-Generation Covalent Irreversible Kinase Inhibitors in NSCLC: Focus on Afatinib.

    Science.gov (United States)

    Hirsh, Vera

    2015-06-01

    First-generation, reversible epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs), erlotinib and gefitinib, represented an important addition to the treatment armamentarium for non-small-cell lung cancer (NSCLC) patients with activating EGFR mutations. However, all patients inevitably develop acquired resistance to these agents, primarily due to secondary EGFR mutations, molecular aberrations affecting other signaling pathways, or transformation to small-cell histology. It was hypothesized that development of second-generation TKIs with broader inhibitory profiles could confer longer-lasting clinical activity and overcome acquired resistance to first-generation inhibitors. Here, we review the development of afatinib, an irreversible ErbB family blocker that potently inhibits signaling of all homodimers and heterodimers formed by the EGFR, human epidermal growth factor receptor (HER)-2, HER3, and HER4 receptors. In two phase III trials in patients with EGFR mutation-positive NSCLC, first-line afatinib significantly improved progression-free survival (PFS) and health-related quality of life versus standard-of-care chemotherapy. Moreover, in preplanned sub-analyses, afatinib significantly improved overall survival in patients harboring EGFR Del19 mutations. Afatinib has also demonstrated clinical activity in NSCLC patients who had progressed on erlotinib/gefitinib, particularly when combined with cetuximab, and offers 'treatment beyond progression' benefit when combined with paclitaxel versus chemotherapy alone. Furthermore, a recent phase III study demonstrated that PFS was significantly improved with afatinib versus erlotinib for the second-line treatment of patients with squamous cell carcinoma of the lung. The activity of afatinib in both first-line and relapsed/refractory settings may reflect its ability to irreversibly inhibit all ErbB family members. Afatinib has a well-defined safety profile with characteristic gastrointestinal (diarrhea

  9. Novel protein kinase D inhibitors cause potent arrest in prostate cancer cell growth and motility

    Directory of Open Access Journals (Sweden)

    Lazo John S

    2010-05-01

    Full Text Available Abstract Background Protein kinase D (PKD has been implicated in a wide range of cellular processes and pathological conditions including cancer. However, targeting PKD therapeutically and dissecting PKD-mediated cellular responses remains difficult due to lack of a potent and selective inhibitor. Previously, we identified a novel pan-PKD inhibitor, CID755673, with potency in the upper nanomolar range and high selectivity for PKD. In an effort to further enhance its selectivity and potency for potential in vivo application, small molecule analogs of CID755673 were generated by modifying both the core structure and side-chains. Results After initial activity screening, five analogs with equal or greater potencies as CID755673 were chosen for further analysis: kb-NB142-70, kb-NB165-09, kb-NB165-31, kb-NB165-92, and kb-NB184-02. Our data showed that modifications to the aromatic core structure in particular significantly increased potency while retaining high specificity for PKD. When tested in prostate cancer cells, all compounds inhibited PMA-induced autophosphorylation of PKD1, with kb-NB142-70 being most active. Importantly, these analogs caused a dramatic arrest in cell proliferation accompanying elevated cytotoxicity when applied to prostate cancer cells. Cell migration and invasion were also inhibited by these analogs with varying potencies that correlated to their cellular activity. Conclusions Throughout the battery of experiments, the compounds kb-NB142-70 and kb-NB165-09 emerged as the most potent and specific analogs in vitro and in cells. These compounds are undergoing further testing for their effectiveness as pharmacological tools for dissecting PKD function and as potential anti-cancer agents in the treatment of prostate cancer.

  10. Differential regulation of cyclin-dependent kinase inhibitors in neuroblastoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Qiao, Lan [Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Department of Pharmaceutical Sciences, Jilin University, Changchun 130021 (China); Paul, Pritha; Lee, Sora [Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Qiao, Jingbo [Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Wang, Yongsheng [Department of Pharmaceutical Sciences, Jilin University, Changchun 130021 (China); Chung, Dai H., E-mail: dai.chung@vanderbilt.edu [Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, TN 37232 (United States)

    2013-05-31

    Highlights: •GRP-R signaling differentially regulated the expression of p21 and p27. •Silencing GRP/GRP-R downregulated p21, while p27 expression was upregulated. •Inhibition of GRP/GRP-R signaling enhanced PTEN expression, correlative to the increased expression of p27. •PTEN and p27 co-localized in cytoplasm and silencing PTEN decreased p27 expression. -- Abstract: Gastrin-releasing peptide (GRP) and its receptor (GRP-R) are highly expressed in undifferentiated neuroblastoma, and they play critical roles in oncogenesis. We previously reported that GRP activates the PI3K/AKT signaling pathway to promote DNA synthesis and cell cycle progression in neuroblastoma cells. Conversely, GRP-R silencing induces cell cycle arrest. Here, we speculated that GRP/GRP-R signaling induces neuroblastoma cell proliferation via regulation of cyclin-dependent kinase (CDK) inhibitors. Surprisingly, we found that GRP/GRP-R differentially induced expressions of p21 and p27. Silencing GRP/GRP-R decreased p21, but it increased p27 expressions in neuroblastoma cells. Furthermore, we found that the intracellular localization of p21 and p27 in the nuclear and cytoplasmic compartments, respectively. In addition, we found that GRP/GRP-R silencing increased the expression and accumulation of PTEN in the cytoplasm of neuroblastoma cells where it co-localized with p27, thus suggesting that p27 promotes the function of PTEN as a tumor suppressor by stabilizing PTEN in the cytoplasm. GRP/GRP-R regulation of CDK inhibitors and tumor suppressor PTEN may be critical for tumoriogenesis of neuroblastoma.

  11. Thermodynamics parameters for binding of halogenated benzotriazole inhibitors of human protein kinase CK2α.

    Science.gov (United States)

    Winiewska, Maria; Kucińska, Katarzyna; Makowska, Małgorzata; Poznański, Jarosław; Shugar, David

    2015-10-01

    The interaction of human CK2α (hCK2α) with nine halogenated benzotriazoles, TBBt and its analogues representing all possible patterns of halogenation on the benzene ring of benzotriazole, was studied by biophysical methods. Thermal stability of protein-ligand complexes, monitored by calorimetric (DSC) and optical (DSF) methods, showed that the increase in the mid-point temperature for unfolding of protein-ligand complexes (i.e. potency of ligand binding to hCK2α) follow the inhibitory activities determined by biochemical assays. The dissociation constant for the ATP-hCK2α complex was estimated with the aid of microscale thermophoresis (MST) as 4.3±1.8 μM, and MST-derived dissociation constants determined for halogenated benzotriazoles, when converted according to known ATP concentrations, perfectly reconstruct IC50 values determined by the biochemical assays. Ligand-dependent quenching of tyrosine fluorescence, together with molecular modeling and DSC-derived heats of unfolding, support the hypothesis that halogenated benzotriazoles bind in at least two alternative orientations, and those that are efficient hCK2α inhibitors bind in the orientation which TBBt adopts in its complex with maize CK2α. DSC-derived apparent heat for ligand binding (ΔΔHbind) is driven by intermolecular electrostatic interactions between Lys68 and the triazole ring of the ligand, as indicated by a good correlation between ΔΔHbind and ligand pKa. Overall results, additionally supported by molecular modeling, confirm that a balance of hydrophobic and electrostatic interactions contribute predominantly (~40 kJ/mol), relative to possible intermolecular halogen/hydrogen bonding (less than 10 kJ/mol), in binding of halogenated benzotriazoles to the ATP-binding site of hCK2α. This article is part of a Special Issue entitled: Inhibitors of Protein Kinases.

  12. Type II kinase inhibitors show an unexpected inhibition mode against Parkinson's disease-linked LRRK2 mutant G2019S.

    Science.gov (United States)

    Liu, Min; Bender, Samantha A; Cuny, Gregory D; Sherman, Woody; Glicksman, Marcie; Ray, Soumya S

    2013-03-12

    A number of well-known type II inhibitors (ATP-noncompetitive) that bind kinases in their DFG-out conformation were tested against wild-type LRRK2 and the most common Parkinson's disease-linked mutation, G2019S. We found that traditional type II inhibitors exhibit surprising variability in their inhibition mechanism between the wild type (WT) and the G2019S mutant of LRRK2. The type II kinase inhibitors were found to work in an ATP-competitive fashion against the G2019S mutant, whereas they appear to follow the expected noncompetitive mechanism against WT. Because the G2019S mutation lies in the DXG motif (DYG in LRRK2 but DFG in most other kinases) of the activation loop, we explored the structural consequence of the mutation on loop dynamics using an enhanced sampling method called metadynamics. The simulations suggest that the G2019S mutation stabilizes the DYG-in state of LRRK2 through a series of hydrogen bonds, leading to an increase in the conformational barrier between the active and inactive forms of the enzyme and a relative stabilization of the active form. The conformational bias toward the active form of LRRK2 mutants has two primary consequences. (1) The mutant enzyme becomes hyperactive, a known contributor to the Parkinsonian phenotype, as a consequence of being "locked" into the activated state, and (2) the mutation creates an unusual allosteric pocket that can bind type II inhibitors but in an ATP-competitive fashion. Our results suggest that developing type II inhibitors, which are generally considered superior to type I inhibitors because of desirable selectivity profiles, might be especially challenging for the G2019S LRRK2 mutant.

  13. Long-term culture of human odontoma-derived cells with a Rho kinase inhibitor.

    Science.gov (United States)

    Uzawa, Katsuhiro; Kasamatsu, Atsushi; Saito, Tomoaki; Takahara, Toshikazu; Minakawa, Yasuyuki; Koike, Kazuyuki; Yamatoji, Masanobu; Nakashima, Dai; Higo, Morihiro; Sakamoto, Yosuke; Shiiba, Masashi; Tanzawa, Hideki

    2016-09-10

    Because of cellular senescence/apoptosis, no effective culture systems are available to maintain replication of cells from odontogenic tumors especially for odontoma, and, thus, the ability to isolate human odontoma-derived cells (hODCs) for functional studies is needed. The current study was undertaken to develop an approach to isolate hODCs and fully characterize the cells in vitro. The hODCs were cultured successfully with a Rho-associated protein kinase inhibitor (Y-27632) for an extended period with stabilized lengths of the telomeres to sustain a similar phenotype/property as the primary tumoral cells. While the hODCs showed stable long-term expansion with expression of major dental epithelial markers including dentin sialophosphoprotein (DSPP) even in the three-dimensional microenvironment, they lack the specific markers for the characteristics of stem cells. Moreover, cells from dental pulp showed significant up-regulation of DSPP when co-cultured with the hODCs, while control fibroblasts with the hODCs did not. Taken together, we propose that the hODCs can be isolated and expanded over the long term with Y-27632 to investigate not only the development of the hODCs but also other types of benign human tumors.

  14. The Tyrosine Kinase Inhibitor Sunitinib Affects Ovulation but Not Ovarian Reserve in Mouse: A Preclinical Study

    Science.gov (United States)

    Bernard, Valérie; Bouilly, Justine; Kramer, Piet; Carré, Nadège; Schlumberger, Martin; Visser, Jenny A.; Young, Jacques; Binart, Nadine

    2016-01-01

    The aim of the study was to evaluate ovarian toxicity of tyrosine kinase inhibitor (TKI) sunitinib, since only scarce data are available on gonadal function after this treatment. Six-week-old female mice received orally, once daily, vehicle or sunitinib (50 mg/kg/d) during 5 weeks. Fertility parameters were analyzed from ovulation to litter assessment. Sunitinib exposure significantly reduced (i) corpora lutea number per ovary (1.1 ± 0.38 in sunitinib group versus 4 ± 0.79 in control group, p<0.01) and (ii) serum Anti Müllerian hormone (AMH) levels in sunitinib treated mice (12.01 ± 1.16) compared to control mice (14.33 ± 0.87 ng/ml, p< 0.05). However, primordial and growing follicles numbers per ovary were not different in both groups. After treatment withdrawal, female mice in both groups were able to obtain litters. These data could be helpful to counsel clinicians and patients, when fertility preservation methods are discussed, before TKI treatment in girls and young women. PMID:27035144

  15. Tau kinase inhibitors protect hippocampal synapses despite of insoluble tau accumulation.

    Science.gov (United States)

    Hinners, Ina; Hill, Anika; Otto, Ulrike; Michalsky, Anke; Mack, Till G A; Striggow, Frank

    2008-03-01

    A better understanding of the cellular and molecular pathomechanisms of Alzheimer's disease (AD) is a prerequisite for the development of efficient treatments. We have used a novel assay system based on virus-transduced organotypic hippocampal slice cultures that mimics important aspects of tau-driven AD pathology in a short time frame. Human tau P301L, when expressed in pyramidal neurons of hippocampal slice cultures, was increasingly phosphorylated at several disease-relevant epitopes, leading to progressive neuronal dystrophy and formation of RIPA-insoluble tau. AD-like tau hyperphosphorylation was reduced by the tau kinase inhibitors lithium and SRN-003-556, but RIPA-insoluble tau remained unaffected after treatment with any of these substances. Only SRN-003-556 was able to protect hippocampal neurons from synaptic damage that was presumably caused by a toxic soluble tau fraction. These data provide first mechanistic insights towards the functional benefits of SRN-003-556 that have been observed in vivo.

  16. Lenalidomide in chronic lymphocytic leukemia: the present and future in the era of tyrosine kinase inhibitors.

    Science.gov (United States)

    Maffei, Rossana; Colaci, Elisabetta; Fiorcari, Stefania; Martinelli, Silvia; Potenza, Leonardo; Luppi, Mario; Marasca, Roberto

    2016-01-01

    Lenalidomide is an immunomodulatory agent (IMiD) clinically active in chronic lymphocytic leukemia (CLL), both in heavily pre-treated patients and upfront. Lenalidomide has a unique mechanism of action in CLL. Its efficacy relies on a multifactorial mode-of-action (MOA), comprising a plethora of immunomodulatory actions, the disruption of mutualistic interactions inside CLL microenvironment and direct effects against leukemic cells. In the last few years, a number of new and highly effective drugs appeared in the scenario of CLL therapeutic options, i.e. tyrosine kinase inhibitors (TKIs), showing a good safety profile and impressive clinical response, also in high-risk patients. In this review, we describe the data from clinical studies about lenalidomide efficacy in CLL and we critically dissect the different mechanisms of action of this drug. We point the attention on open issues, including drug dosage and administration schedule, prediction of clinical response to lenalidomide, and combination therapeutic strategies. This overview would be useful to envision a possible role of lenalidomide in the treatment flow-chart of CLL, exploiting its peculiar MOA and also exploring the possible synergetic effect with new drugs.

  17. Nanosuspension for the delivery of a poorly soluble anti-cancer kinase inhibitor.

    Science.gov (United States)

    Danhier, Fabienne; Ucakar, Bernard; Vanderhaegen, Marie-Lyse; Brewster, Marcus E; Arien, Tina; Préat, Véronique

    2014-09-01

    We hypothesized that nanosuspensions could be promising for the delivery of the poorly water soluble anti-cancer multi-targeted kinase inhibitor, MTKi-327. Hence, the aims of this work were (i) to evaluate the MTKi-327 nanosuspension for parenteral and oral administrations and (ii) to compare this nanosuspension with other nanocarriers in terms of anti-cancer efficacy and pharmacokinetics. Therefore, four formulations of MTKi-327 were studied: (i) PEGylated PLGA-based nanoparticles, (ii) self-assembling PEG₇₅₀-p-(CL-co-TMC) polymeric micelles, (iii) nanosuspensions of MTKi-327; and (iv) Captisol solution (pH=3.5). All the nano-formulations presented a size below 200 nm. Injections of the highest possible dose of the three nano-formulations did not induce any side effects in mice. In contrast, the maximum tolerated dose of the control Captisol solution was 20-fold lower than its highest possible dose. The highest regrowth delay of A-431-tumor-bearing nude mice was obtained with MTKi-327 nanosuspension, administered intravenously, at a dose of 650 mg/kg. After intravenous and oral administration, the AUC₀₋∞ of MTKi-327 nanosuspension was 2.4-fold greater than that of the Captisol solution. Nanosuspension may be considered as an effective anti-cancer MTKi-327 delivery method due to (i) the higher MTKi-327 maximum tolerated dose, (ii) the possible intravenous injection of MTKi-327, (iii) its ability to enhance the administered dose and (iv) its higher efficacy.

  18. High-throughput virtual screening using quantum mechanical probes: discovery of selective kinase inhibitors.

    Science.gov (United States)

    Zhou, Ting; Caflisch, Amedeo

    2010-07-05

    A procedure based on semi-empirical quantum mechanical (QM) calculations of interaction energy is proposed for the rapid screening of compound poses generated by high-throughput docking. Small molecules (consisting of 2-10 atoms and termed "probes") are overlapped with polar groups in the binding site of the protein target. The interaction energy values between each compound pose and the probes, calculated by a semi-empirical Hamiltonian, are used as filters. The QM probe method does not require fixed partial charges and takes into account polarization and charge-transfer effects which are not captured by conventional force fields. The procedure is applied to screen approximately 100 million poses (of 2.7 million commercially available compounds) obtained by high-throughput docking in the ATP binding site of the tyrosine kinase erythropoietin-producing human hepatocellular carcinoma receptor B4 (EphB4). Three QM probes on the hinge region and one at the entrance pocket are employed to select for binding affinity, while a QM probe on the side chain of the so-called gatekeeper residue (a hypervariable residue in the kinome) is used to enforce selectivity. The poses with favorable interactions with the five QM probes are filtered further for hydrophobic matching and low ligand strain. In this way, a single-digit micromolar inhibitor of EphB4 with a relatively good selectivity profile is identified in a multimillion-compound library upon experimental tests of only 23 molecules.

  19. Myocardial steatosis and necrosis in atria and ventricles of rats given pyruvate dehydrogenase kinase inhibitors.

    Science.gov (United States)

    Jones, Huw Bowen; Reens, Jaimini; Johnson, Elizabeth; Brocklehurst, Simon; Slater, Ian

    2014-12-01

    Pharmaceutical therapies for non-insulin-dependent diabetes mellitus (NIDDM) include plasma glucose lowering by enhancing glucose utilization. The mitochondrial pyruvate dehydrogenase (PDH) complex is important in controlling the balance between glucose and fatty acid substrate oxidation. Administration of pyruvate dehydrogenase kinase inhibitors (PDHKIs) to rats effectively lowers plasma glucose but results in myocardial steatosis that in some instances is associated primarily with atrial and to a lesser degree with ventricular pathology. Induction of myocardial steatosis is not dose-dependent, varies from minimal to moderate severity, and is either of multifocal or diffuse distribution. Ventricular histopathology was restricted to few myocardial degenerative fibers, while that in the atrium/atria was of either acute or chronic appearance with the former showing myocardial degeneration/necrosis, acute myocarditis, edema, endothelial activation (rounding up), endocarditis, and thrombosis associated with moderate myocardial steatosis and the latter with myocardial loss, replacement fibrosis, and no apparent or minimal association with steatosis. The evidence from these evaluations indicate that excessive intramyocardial accumulation of lipid may be either primarily adverse or represents an indicator of other adversely affected cellular processes.

  20. Recent developments in the chromatographic bioanalysis of approved kinase inhibitor drugs in oncology.

    Science.gov (United States)

    Rood, Johannes J M; Schellens, Jan H M; Beijnen, Jos H; Sparidans, Rolf W

    2016-10-25

    In recent years (2010-present) there has been an increase in the number of publications reporting the development, validation and use of bioanalytical methods in the rapidly expanding drug class of small molecule protein kinase inhibitors. Most reports describe the technological set-up of the methods that have allowed for drug concentration measurements from various sample types. This includes plasma, dried blood-spot, and tissue-analysis. Also method development, exploration of various techniques, as well as measurement and identification of metabolites were addressed. For the bioanalysis, a variety of sample-pretreatment methods like protein-precipitation, liquid-liquid extraction, and solid-phase extraction have been employed, all varying in complexity, cleanliness and time-consumption. Chromatographic separation, nowadays, is more focused on separating components from ion-suppressive effects, since for MS/MS detection, various components do not have to be baseline separated. For detection multiple types of detectors were used, ranging from state-of-the-art high resolution, and tandem mass spectrometry with low picogram per milliliter detection limits to the classical UV-detector with several nanograms per milliliter limits. As new bioanalytical methods have arisen that do rely on chromatographic separation, for example for high-throughput analysis, these are addressed in this review as well.

  1. Human metabolism of lapatinib, a dual kinase inhibitor: implications for hepatotoxicity.

    Science.gov (United States)

    Castellino, Stephen; O'Mara, Michael; Koch, Kevin; Borts, David J; Bowers, Gary D; MacLauchlin, Christopher

    2012-01-01

    Lapatinib (Tykerb, Tyverb) is an important orally active dual tyrosine kinase inhibitor efficacious in combination therapy for patients with progressive human epidermal receptor 2-overexpressing metastatic breast cancer. However, clinically significant liver injury, which may be associated with lapatinib metabolic activation, has been reported. We describe the metabolism and excretion of [(14)C]lapatinib in six healthy human volunteers after a single oral dose of 250 mg and the potential relationships between metabolism and clinical hepatotoxicity. Overall, elimination showed high intersubject variability, with fecal elimination being the predominant pathway, representing a median of 92% of the dose with lapatinib as the largest component (approximate median 27% of the dose). In plasma, approximately 50% of the observed radioactivity was attributed to metabolites. Analysis of a 4-h pooled plasma extract identified seven metabolites related by an N- and α-carbon oxidation cascade. Fecal metabolites derived from three prominent pathways: N- and α-carbon oxidation, fluorobenzyl oxidative cleavage, and hydroxypyridine formation. Several of the lapatinib metabolites can undoubtedly be linked to reactive species such as aldehydes or quinone imines. In addition to the contribution of these potentially reactive metabolites as suspects in clinical liver injury, the role of other disposition factors, including interaction with drug transporters, pharmacogenetics, or magnitude of the therapeutic dose, should not be discounted.

  2. The Tyrosine Kinase Inhibitor Sunitinib Affects Ovulation but Not Ovarian Reserve in Mouse: A Preclinical Study.

    Directory of Open Access Journals (Sweden)

    Valérie Bernard

    Full Text Available The aim of the study was to evaluate ovarian toxicity of tyrosine kinase inhibitor (TKI sunitinib, since only scarce data are available on gonadal function after this treatment. Six-week-old female mice received orally, once daily, vehicle or sunitinib (50 mg/kg/d during 5 weeks. Fertility parameters were analyzed from ovulation to litter assessment. Sunitinib exposure significantly reduced (i corpora lutea number per ovary (1.1 ± 0.38 in sunitinib group versus 4 ± 0.79 in control group, p<0.01 and (ii serum Anti Müllerian hormone (AMH levels in sunitinib treated mice (12.01 ± 1.16 compared to control mice (14.33 ± 0.87 ng/ml, p< 0.05. However, primordial and growing follicles numbers per ovary were not different in both groups. After treatment withdrawal, female mice in both groups were able to obtain litters. These data could be helpful to counsel clinicians and patients, when fertility preservation methods are discussed, before TKI treatment in girls and young women.

  3. Discovery of Inhibitors of Trypanosoma brucei by Phenotypic Screening of a Focused Protein Kinase Library.

    Science.gov (United States)

    Woodland, Andrew; Thompson, Stephen; Cleghorn, Laura A T; Norcross, Neil; De Rycker, Manu; Grimaldi, Raffaella; Hallyburton, Irene; Rao, Bhavya; Norval, Suzanne; Stojanovski, Laste; Brun, Reto; Kaiser, Marcel; Frearson, Julie A; Gray, David W; Wyatt, Paul G; Read, Kevin D; Gilbert, Ian H

    2015-11-01

    A screen of a focused kinase inhibitor library against Trypanosoma brucei rhodesiense led to the identification of seven series, totaling 121 compounds, which showed >50 % inhibition at 5 μm. Screening of these hits in a T. b. brucei proliferation assay highlighted three compounds with a 1H-imidazo[4,5-b]pyrazin-2(3H)-one scaffold that showed sub-micromolar activity and excellent selectivity against the MRC5 cell line. Subsequent rounds of optimisation led to the identification of compounds that exhibited good in vitro drug metabolism and pharmacokinetics (DMPK) properties, although in general this series suffered from poor solubility. A scaffold-hopping exercise led to the identification of a 1H-pyrazolo[3,4-b]pyridine scaffold, which retained potency. A number of examples were assessed in a T. b. brucei growth assay, which could differentiate static and cidal action. Compounds from the 1H-imidazo[4,5-b]pyrazin-2(3H)-one series were found to be either static or growth-slowing and not cidal. Compounds with the 1H-pyrazolo[3,4-b]pyridine scaffold were found to be cidal and showed an unusual biphasic nature in this assay, suggesting they act by at least two mechanisms.

  4. Cip/Kip cyclin-dependent protein kinase inhibitors and the road to polyploidy

    Directory of Open Access Journals (Sweden)

    DePamphilis Melvin L

    2009-06-01

    Full Text Available Abstract Cyclin-dependent kinases (CDKs play a central role in the orderly transition from one phase of the eukaryotic mitotic cell division cycle to the next. In this context, p27Kip1 (one of the CIP/KIP family of CDK specific inhibitors in mammals or its functional analogue in other eukarya prevents a premature transition from G1 to S-phase. Recent studies have revealed that expression of a second member of this family, p57Kip2, is induced as trophoblast stem (TS cells differentiate into trophoblast giant (TG cells. p57 then inhibits CDK1 activity, an enzyme essential for initiating mitosis, thereby triggering genome endoreduplication (multiple S-phases without an intervening mitosis. Expression of p21Cip1, the third member of this family, is also induced in during differentiation of TS cells into TG cells where it appears to play a role in suppressing the DNA damage response pathway. Given the fact that p21 and p57 are unique to mammals, the question arises as to whether one or both of these proteins are responsible for the induction and maintenance of polyploidy during mammalian development.

  5. Small molecule kinase inhibitors alleviate different molecular features of myotonic dystrophy type 1.

    Science.gov (United States)

    Wojciechowska, Marzena; Taylor, Katarzyna; Sobczak, Krzysztof; Napierala, Marek; Krzyzosiak, Wlodzimierz J

    2014-01-01

    Expandable (CTG)n repeats in the 3' UTR of the DMPK gene are a cause of myotonic dystrophy type 1 (DM1), which leads to a toxic RNA gain-of-function disease. Mutant RNAs with expanded CUG repeats are retained in the nucleus and aggregate in discrete inclusions. These foci sequester splicing factors of the MBNL family and trigger upregulation of the CUGBP family of proteins resulting in the mis-splicing of their target transcripts. To date, many efforts to develop novel therapeutic strategies have been focused on disrupting the toxic nuclear foci and correcting aberrant alternative splicing via targeting mutant CUG repeats RNA; however, no effective treatment for DM1 is currently available. Herein, we present results of culturing of human DM1 myoblasts and fibroblasts with two small-molecule ATP-binding site-specific kinase inhibitors, C16 and C51, which resulted in the alleviation of the dominant-negative effects of CUG repeat expansion. Reversal of the DM1 molecular phenotype includes a reduction of the size and number of foci containing expanded CUG repeat transcripts, decreased steady-state levels of CUGBP1 protein, and consequent improvement of the aberrant alternative splicing of several pre-mRNAs misregulated in DM1.

  6. UV-Vis spectroscopy and solvatochromism of the tyrosine kinase inhibitor AG-1478.

    Science.gov (United States)

    Khattab, Muhammad; Wang, Feng; Clayton, Andrew H A

    2016-07-05

    The effect of twenty-one solvents on the UV-Vis spectrum of the tyrosine kinase inhibitor AG-1478 was investigated. The absorption spectrum in the range 300-360nm consisted of two partially overlapping bands at approximately 340nm and 330nm. The higher energy absorption band was more sensitive to solvent and exhibited a peak position that varied from 327nm to 336nm, while the lower energy absorption band demonstrated a change in peak position from 340nm to 346nm in non-chlorinated solvents. The fluorescence spectrum of AG-1478 was particularly sensitive to solvent. The wavelength of peak intensity varied from 409nm to 495nm with the corresponding Stokes shift in the range of 64nm to 155nm (4536cm(-1) to 9210cm(-1)). We used a number of methods to assess the relationship between spectroscopic properties and solvent properties. The detailed analysis revealed that for aprotic solvents, the peak position of the emission spectrum in wavenumber scale correlated with the polarity (dielectric constant or ET(30)) of the solvent. In protic solvents, a better correlation was observed between the hydrogen bonding power of the solvent and the position of the emission spectrum. Moreover, the fluorescence quantum yields were larger in aprotic solvents as compared to protic solvents. This analysis underscores the importance of polarity and hydrogen-bonding environment on the spectroscopic properties of AG-1478. These studies will assume relevance in understanding the interaction of AG-1478 in vitro and in vivo.

  7. Efficacy of cyclin dependent kinase 4 inhibitors as potent neuroprotective agents against insults relevant to Alzheimer's disease.

    Directory of Open Access Journals (Sweden)

    Priyankar Sanphui

    Full Text Available Alzheimer's disease (AD is a progressive neurodegenerative disease with no cure till today. Aberrant activation of cell cycle regulatory proteins is implicated in neurodegenerative diseases including AD. We and others have shown that Cyclin dependent kinase 4 (Cdk4 is activated in AD brain and is required for neuron death. In this study, we tested the efficiency of commercially available Cdk4 specific inhibitors as well as a small library of synthetic molecule inhibitors targeting Cdk4 as neuroprotective agents in cellular models of neuron death. We found that several of these inhibitors significantly protected neuronal cells against death induced by nerve growth factor (NGF deprivation and oligomeric beta amyloid (Aβ that are implicated in AD. These neuroprotective agents inhibit specifically Cdk4 kinase activity, loss of mitochondrial integrity, induction of pro-apoptotic protein Bim and caspase3 activation in response to NGF deprivation. The efficacies of commercial and synthesized inhibitors are comparable. The synthesized molecules are either phenanthrene based or naphthalene based and they are synthesized by using Pschorr reaction and Buchwald coupling respectively as one of the key steps. A number of molecules of both kinds block neurodegeneration effectively. Therefore, we propose that Cdk4 inhibition would be a therapeutic choice for ameliorating neurodegeneration in AD and these synthetic Cdk4 inhibitors could lead to development of effective drugs for AD.

  8. Different Persistence of the Cellular Effects Promoted by Protein Kinase CK2 Inhibitors CX-4945 and TDB

    Directory of Open Access Journals (Sweden)

    Cristina Girardi

    2015-01-01

    Full Text Available We compare the cellular efficacy of two selective and cell permeable inhibitors of the antiapoptotic kinase CK2. One inhibitor, CX-4945, is already in clinical trials as antitumor drug, while the other, TDB, has been recently successfully employed to demonstrate the implication of CK2 in cellular (disregulation. We found that, upon treatment of cancer cells with these compounds, the extent of inhibition of endocellular CK2 is initially comparable but becomes significantly different after the inhibitors are removed from the cellular medium: while in CX-4945 treated cells CK2 activity is restored to control level after 24 h, in the case of TDB it is still strongly reduced after 4 days from removal. The biological effects of the two inhibitors have been analyzed by performing clonogenic, spheroid formation, and wound-healing assays: we observed a permanent inhibition of cell survival and migration in TDB-treated cells even after the inhibitor removal, while in the case of CX-4945 only its maintenance for the whole duration of the assay insured a persisting effect. We suggest that the superiority of TDB in maintaining kinase activity inhibited and perpetuating the consequent effects is an added value to be considered when planning new therapies based on CK2 targeting.

  9. Different Persistence of the Cellular Effects Promoted by Protein Kinase CK2 Inhibitors CX-4945 and TDB

    Science.gov (United States)

    Girardi, Cristina; Ottaviani, Daniele; Pinna, Lorenzo A.; Ruzzene, Maria

    2015-01-01

    We compare the cellular efficacy of two selective and cell permeable inhibitors of the antiapoptotic kinase CK2. One inhibitor, CX-4945, is already in clinical trials as antitumor drug, while the other, TDB, has been recently successfully employed to demonstrate the implication of CK2 in cellular (dis)regulation. We found that, upon treatment of cancer cells with these compounds, the extent of inhibition of endocellular CK2 is initially comparable but becomes significantly different after the inhibitors are removed from the cellular medium: while in CX-4945 treated cells CK2 activity is restored to control level after 24 h, in the case of TDB it is still strongly reduced after 4 days from removal. The biological effects of the two inhibitors have been analyzed by performing clonogenic, spheroid formation, and wound-healing assays: we observed a permanent inhibition of cell survival and migration in TDB-treated cells even after the inhibitor removal, while in the case of CX-4945 only its maintenance for the whole duration of the assay insured a persisting effect. We suggest that the superiority of TDB in maintaining kinase activity inhibited and perpetuating the consequent effects is an added value to be considered when planning new therapies based on CK2 targeting. PMID:26558259

  10. Discovery of 7-aminofuro[2,3-c]pyridine inhibitors of TAK1: optimization of kinase selectivity and pharmacokinetics.

    Science.gov (United States)

    Hornberger, Keith R; Chen, Xin; Crew, Andrew P; Kleinberg, Andrew; Ma, Lifu; Mulvihill, Mark J; Wang, Jing; Wilde, Victoria L; Albertella, Mark; Bittner, Mark; Cooke, Andrew; Kadhim, Salam; Kahler, Jennifer; Maresca, Paul; May, Earl; Meyn, Peter; Romashko, Darlene; Tokar, Brianna; Turton, Roy

    2013-08-15

    The kinase selectivity and pharmacokinetic optimization of a series of 7-aminofuro[2,3-c]pyridine inhibitors of TAK1 is described. The intersection of insights from molecular modeling, computational prediction of metabolic sites, and in vitro metabolite identification studies resulted in a simple and unique solution to both of these problems. These efforts culminated in the discovery of compound 13a, a potent, relatively selective inhibitor of TAK1 with good pharmacokinetic properties in mice, which was active in an in vivo model of ovarian cancer.

  11. 8-THP-DHI analogs as potent Type I dual TIE-2/VEGF-R2 receptor tyrosine kinase inhibitors.

    Science.gov (United States)

    Hudkins, Robert L; Zulli, Allison L; Underiner, Ted L; Angeles, Thelma S; Aimone, Lisa D; Meyer, Sheryl L; Pauletti, Daniel; Chang, Hong; Fedorov, Elena V; Almo, Steven C; Fedorov, Alexander A; Ruggeri, Bruce A

    2010-06-01

    A novel series of 8-(2-tetrahydropyranyl)-12,13-dihydroindazolo[5,4-a]pyrrolo[3,4-c]carbazoles (THP-DHI) was synthesized and evaluated as dual TIE-2 and VEGF-R2 receptor tyrosine kinase inhibitors. Development of the structure-activity relationships (SAR) with the support of X-ray crystallography led to identification of 7f and 7g as potent, selective dual TIE-2/VEGF-R2 inhibitors with excellent cellular potency and acceptable pharmacokinetic properties. Compounds 7f and 7g were orally active in tumor models with no observed toxicity.

  12. Imaging Sunlit Aurora from Balloon

    Science.gov (United States)

    Lummerzheim, D.; Zhou, X.

    2012-12-01

    Since 1892 when aurora was first imaged by Martin Brendel, useful auroral images have been obtained only when the aurora is in darkness. While UV imagers onboard satellite provide global auroral dynamics, the sunlit aurora in the UV band is overwhelmed by the UV airglow that has the same wavelengths with the UV auroral emissions. The visible band imaging is feasible only when the aurora is in darkness to avoid the sunlight contamination. However, sunlit aurora (such as dayside aurora) is closely related to the solar wind - magnetosphere - ionosphere coupling. In addition, limited land area has badly restricted the capability of imaging dayside aurora from the northern hemisphere and nightside aurora from the southern hemisphere. We have confirmed that sunlit aurora can be imaged using a near-infrared (NIR) camera on board the balloon platform flying in Antarctica or Arctic. This method provides a unique capability for dayside and conjugate auroral investigations. Scientific questions that can be addressed by such observations include how does the dayside aurora respond to solar wind transient variations? Are auroras hemispherically symmetric? Are auroral forms and their variations under sunlight the same as those in darkness? etc. The new method is also cost effective comparing to space-borne imagers, and offers capabilities not obtainable from space and the ground. With the accomplishment of identifying auroral dynamics in sunlight and darkness, in the south and north simultaneously, our knowledge and understanding of auroral phenomenon and its causes will be expanded.

  13. Expression of the Apoptosis Inhibitor Survivin and its correlation with Thymidine Kinase and Axillary Lymph Node Metastasis in Breast Cancer

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    1 Introduction Many molecular factors have been demonstrated to interfere with cellular proliferation and programmed cell death.One of these factors is a recently discovered member of the “inhibitor of apoptosis protein(IAP)” called survivin. Survivin is abundantly expressed in most solid and hematologic malignancies, but undetectable in normal adult tissues. Interference with survivin function induces pleiotropic cell-division defects and apoptosis. Cytosolic thymidine kinase (TK) is a marker for prolifera...

  14. [Efficacy of levocarnitine for tyrosine kinase inhibitor-induced painful muscle cramps in patients with chronic myelogenous leukemia].

    Science.gov (United States)

    Yamada, Michiko; Kuroda, Hiroyuki; Shimoyama, Saori; Ito, Ryo; Sugama, Yusuke; Sato, Ken; Yamauchi, Natsumi; Horiguchi, Hiroto; Nakamura, Hajime; Hamaguchi, Kota; Abe, Tomoyuki; Fujii, Shigeyuki; Maeda, Masahiro; Kato, Junji

    2016-04-01

    Muscle cramps are side effects commonly associated with tyrosine kinase inhibitor (TKI) treatment. Patients suffering from muscle cramps are treated with various medications such as calcium, magnesium and vitamin supplements, but these therapies are often ineffective. We report two patients with chronic myelogenous leukemia who developed muscle cramps caused by TKI. These patients were treated successfully with levocarnitine. Both of our cases revealed the beneficial effects of levocarnitine treatment on TKI-induced muscle cramps.

  15. Inhibitors of LRRK2 kinase attenuate neurodegeneration and Parkinson-like phenotypes in Caenorhabditis elegans and Drosophila Parkinson's disease models

    Science.gov (United States)

    Liu, Zhaohui; Hamamichi, Shusei; Dae Lee, Byoung; Yang, Dejun; Ray, Arpita; Caldwell, Guy A.; Caldwell, Kim A.; Dawson, Ted M.; Smith, Wanli W.; Dawson, Valina L.

    2011-01-01

    Mutations in leucine-rich repeat kinase 2 (LRRK2) have been identified as a genetic cause of familial Parkinson's disease (PD) and have also been found in the more common sporadic form of PD, thus positioning LRRK2 as important in the pathogenesis of PD. Biochemical studies of the disease-causing mutants of LRRK2 implicates an enhancement of kinase activity as the basis of neuronal toxicity and thus possibly the pathogenesis of PD due to LRRK2 mutations. Previously, a chemical library screen identified inhibitors of LRRK2 kinase activity. Here, two of these inhibitors, GW5074 and sorafenib, are shown to protect against G2019S LRRK2-induced neurodegeneration in vivo in Caenorhabditis elegans and in Drosophila. These findings indicate that increased kinase activity of LRRK2 is neurotoxic and that inhibition of LRRK2 activity can have a disease-modifying effect. This suggests that inhibition of LRRK2 holds promise as a treatment for PD. PMID:21768216

  16. Tabletop Aurora Demonstrator.

    Science.gov (United States)

    Zaffo, Peter Alfred

    1991-01-01

    A simple apparatus for studying various forms of the aurora borealis on a miniature scale can be constructed from a neon-storage flask, aluminum tape, a cardboard box, and a small piezoelectric ion gun of the type used to neutralize static charges on phonograph records. (PR)

  17. Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening.

    Science.gov (United States)

    Nienberg, Christian; Garmann, Claudia; Gratz, Andreas; Bollacke, Andre; Götz, Claudia; Jose, Joachim

    2017-01-05

    Human protein kinase CK2 has emerged as promising target for the treatment of neoplastic diseases. The vast majority of kinase inhibitors known today target the ATP binding site, which is highly conserved among kinases and hence leads to limited selectivity. In order to identify non-ATP competitive inhibitors, a 12-mer peptide library of 6 × 10⁵ variants was displayed on the surface of E. coli by autodisplay. Screening of this peptide library on variants with affinity to CK2 was performed by fluorophore-conjugated CK2 and subsequent flow cytometry. Single cell sorting of CK2-bound E. coli yielded new peptide variants, which were tested on inhibition of CK2 by a CE-based assay. Peptide B2 (DCRGLIVMIKLH) was the most potent inhibitor of both, CK2 holoenzyme and the catalytic CK2α subunit (IC50 = 0.8 µM). Using different ATP concentrations and different substrate concentrations for IC50 determination, B2 was shown to be neither ATP- nor substrate competitive. By microscale thermophoresis (MST) the KD value of B2 with CK2α was determined to be 2.16 µM, whereas no binding of B2 to CK2β-subunit was detectable. To our surprise, besides inhibition of enzymatic activity, B2 also disturbed the interaction of CK2α with CK2β at higher concentrations (≥25 µM).

  18. Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening

    Directory of Open Access Journals (Sweden)

    Christian Nienberg

    2017-01-01

    Full Text Available Human protein kinase CK2 has emerged as promising target for the treatment of neoplastic diseases. The vast majority of kinase inhibitors known today target the ATP binding site, which is highly conserved among kinases and hence leads to limited selectivity. In order to identify non-ATP competitive inhibitors, a 12-mer peptide library of 6 × 105 variants was displayed on the surface of E. coli by autodisplay. Screening of this peptide library on variants with affinity to CK2 was performed by fluorophore-conjugated CK2 and subsequent flow cytometry. Single cell sorting of CK2-bound E. coli yielded new peptide variants, which were tested on inhibition of CK2 by a CE-based assay. Peptide B2 (DCRGLIVMIKLH was the most potent inhibitor of both, CK2 holoenzyme and the catalytic CK2α subunit (IC50 = 0.8 µM. Using different ATP concentrations and different substrate concentrations for IC50 determination, B2 was shown to be neither ATP- nor substrate competitive. By microscale thermophoresis (MST the KD value of B2 with CK2α was determined to be 2.16 µM, whereas no binding of B2 to CK2β-subunit was detectable. To our surprise, besides inhibition of enzymatic activity, B2 also disturbed the interaction of CK2α with CK2β at higher concentrations (≥25 µM.

  19. Fragment-Based Discovery of a Dual pan-RET/VEGFR2 Kinase Inhibitor Optimized for Single-Agent Polypharmacology.

    Science.gov (United States)

    Frett, Brendan; Carlomagno, Francesca; Moccia, Maria Luisa; Brescia, Annalisa; Federico, Giorgia; De Falco, Valentina; Admire, Brittany; Chen, Zhongzhu; Qi, Wenqing; Santoro, Massimo; Li, Hong-yu

    2015-07-20

    Oncogenic conversion of the RET (rearranged during transfection) tyrosine kinase is associated with several cancers. A fragment-based chemical screen led to the identification of a novel RET inhibitor, Pz-1. Modeling and kinetic analysis identified Pz-1 as a type II tyrosine kinase inhibitor that is able to bind the "DFG-out" conformation of the kinase. Importantly, from a single-agent polypharmacology standpoint, Pz-1 was shown to be active on VEGFR2, which can block the blood supply required for RET-stimulated growth. In cell-based assays, 1.0 nM of Pz-1 strongly inhibited phosphorylation of all tested RET oncoproteins. At 1.0 mg kg(-1)  day(-1) per os, Pz-1 abrogated the formation of tumors induced by RET-mutant fibroblasts and blocked the phosphorylation of both RET and VEGFR2 in tumor tissue. Pz-1 featured no detectable toxicity at concentrations of up to 100.0 mg kg(-1), which indicates a large therapeutic window. This study validates the effectiveness and usefulness of a medicinal chemistry/polypharmacology approach to obtain an inhibitor capable of targeting multiple oncogenic pathways.

  20. Interruption of the blood-stage cycle of the malaria parasite, Plasmodium chabaudi, by protein tyrosine kinase inhibitors

    Directory of Open Access Journals (Sweden)

    M.L. Gazarini

    2003-11-01

    Full Text Available Malaria is a devastating disease caused by a unicellular protozoan, Plasmodium, which affects 3.7 million people every year. Resistance of the parasite to classical treatments such as chloroquine requires the development of new drugs. To gain insight into the mechanisms that control Plasmodium cell cycle, we have examined the effects of kinase inhibitors on the blood-stage cycle of the rodent malaria parasite, Plasmodium chabaudi. In vitro incubation of red blood cells for 17 h at 37ºC with the inhibitors led to a decrease in the percent of infected cells, compared to control treatment, as follows: genistein (200 µM - 75%, staurosporine (1 µM - 58%, R03 (1 µM - 75%, and tyrphostins B44 (100 µM - 66% and B46 (100 µM - 68%. All these treatments were shown to retard or prevent maturation of the intraerythrocytic parasites. The diverse concentration ranges at which these inhibitors exert their effects give a clue as to the types of signals that initiate the transitions between the different developmental stages of the parasite. The present data support our hypothesis that the maturation of the intraerythrocytic cycle of malaria parasites requires phosphorylation. In this respect, we have recently reported a high Ca2+ microenvironment surrounding the parasite within red blood cells. Several kinase activities are modulated by Ca2+. The molecular identification of the targets of these kinases could provide new strategies against malaria.

  1. Reduced intensity conditioning is superior to nonmyeloablative conditioning for older chronic myelogenous leukemia patients undergoing hematopoietic cell transplant during the tyrosine kinase inhibitor era

    DEFF Research Database (Denmark)

    Warlick, Erica; Ahn, Kwang Woo; Pedersen, Tanya L;

    2012-01-01

    Tyrosine kinase inhibitors (TKIs) and reduced intensity conditioning (RIC)/nonmyeloablative (NMA) conditioning hematopoietic cell transplants (HCTs) have changed the therapeutic strategy for chronic myelogenous leukemia (CML) patients. We analyzed post-HCT outcomes of 306 CML patients reported to...

  2. IkB kinase-beta inhibitor attenuates hepatic fibrosis in mice

    Institute of Scientific and Technical Information of China (English)

    Jue Wei; Min Shi; Wei Qi Wu; Ting Wang; Na Wang; Jia-Li Ma; Yu-Gang Wang

    2011-01-01

    AIM: To investigate the anti-fibrosis effect of IκkB kinase-beta inhibitor (IKK2 inhibitor IMD0354) in liver fibrosis. METHODS: Twenty male C57BL6 mice were divided into four groups. Five high-fat fed mice were injected with lipopolysaccharide (LPS, 10 mg/kg) intraperitoneally and five high-fat fed mice were without LPS injection to build models of liver injury, and the intervention group (five mice) was injected intraperitoneally with IKK2 inhibitor (IMD 30 mg/kg for 14 d), while the remaining five mice received a normal diet as controls. Hepatic function, pathological evaluation and liver interleukin- 6 (IL-6) expression were examined. Western blotting and real-time polymerase chain reaction were used to detect the expressions of nuclear factor-κkB (NF-κkB), alpha-smooth muscle actin (α-SMA), tumor growth factor-beta1 (TGF-β1), tumor necrosis factor-alpha (TNF-α), typeⅠand type Ⅲ collagen proteins and mRNA. RESULTS: A mouse model of liver injury was successfully established, and IMD decreased nuclear translocation translocation of NF-κkB p65 in liver cells. In the IMD-treated group, the levels of alanine aminotransferase (103 ± 9.77 μ/L vs 62.4 ± 7.90 μ/L, P < 0.05) and aminotransferase (295.8 ± 38.56 μ/L vs 212 ± 25.10 μ/L, P < 0.05) were significantly decreased when compared with the model groups. The histological changes were significantly ameliorated. After treatment, the expressions of IL-6 (681 ± 45.96 vs 77 ± 7.79, P < 0.05), TGF-β1 (Western blotting 5.65% ± 0.017% vs 2.73% ± 0.005%, P < 0.05), TNF-α (11.58% ± 0.0063% vs 8.86% ± 0.0050%, P < 0.05), typeⅠcollagen (4.49% ± 0.014% vs 1.90% ± 0.0006%, P < 0.05) and type Ⅲ collagen (3.46% ± 0.008% vs 2.29% ± 0.0035%, P < 0.05) as well as α-SMA (6.19 ± 0.0036 μ/L vs 2.16 ± 0.0023 μ/L, P < 0.05) protein and mRNA were downregulated in the IMD group compared to the fibrosis control groups (P < 0.05).CONCLUSION: IKK2 inhibitor IMD markedly improved non-alcoholic fatty liver

  3. High throughput screens yield small molecule inhibitors of Leishmania CRK3:CYC6 cyclin-dependent kinase.

    Directory of Open Access Journals (Sweden)

    Roderick G Walker

    Full Text Available BACKGROUND: Leishmania species are parasitic protozoa that have a tightly controlled cell cycle, regulated by cyclin-dependent kinases (CDKs. Cdc2-related kinase 3 (CRK3, an essential CDK in Leishmania and functional orthologue of human CDK1, can form an active protein kinase complex with Leishmania cyclins CYCA and CYC6. Here we describe the identification and synthesis of specific small molecule inhibitors of bacterially expressed Leishmania CRK3:CYC6 using a high throughput screening assay and iterative chemistry. We also describe the biological activity of the molecules against Leishmania parasites. METHODOLOGY/PRINCIPAL FINDINGS: In order to obtain an active Leishmania CRK3:CYC6 protein kinase complex, we developed a co-expression and co-purification system for Leishmania CRK3 and CYC6 proteins. This active enzyme was used in a high throughput screening (HTS platform, utilising an IMAP fluorescence polarisation assay. We carried out two chemical library screens and identified specific inhibitors of CRK3:CYC6 that were inactive against the human cyclin-dependent kinase CDK2:CycA. Subsequently, the best inhibitors were tested against 11 other mammalian protein kinases. Twelve of the most potent hits had an azapurine core with structure activity relationship (SAR analysis identifying the functional groups on the 2 and 9 positions as essential for CRK3:CYC6 inhibition and specificity against CDK2:CycA. Iterative chemistry allowed synthesis of a number of azapurine derivatives with one, compound 17, demonstrating anti-parasitic activity against both promastigote and amastigote forms of L. major. Following the second HTS, 11 compounds with a thiazole core (active towards CRK3:CYC6 and inactive against CDK2:CycA were tested. Ten of these hits demonstrated anti-parasitic activity against promastigote L. major. CONCLUSIONS/SIGNIFICANCE: The pharmacophores identified from the high throughput screens, and the derivatives synthesised, selectively

  4. Glycogen Synthase Kinase-3 Inhibitor Protects Against Microvascular Hyperpermeability Following Hemorrhagic Shock

    Science.gov (United States)

    Sawant, Devendra A.; Tharakan, Binu; Hunter, Felicia A.; Childs, Ed W.

    2015-01-01

    Background Hemorrhagic shock (HS)-induce microvascular hyperpermeability involves disruption of endothelial cell adherens junctions leading to increase in paracellular permeability. β-Catenin, an integral component of the adherens junctional complex and Wnt pathway, and caspase-3 via its apoptotic signaling regulate endothelial cell barrier integrity. We have hypothesized that inhibiting phosphorylation of β-catenin and caspase-3 activity using glycogen synthase kinase-3 (GSK-3) specific inhibitor SB216763, would attenuate microvascular hyperpermeability following HS. Methods In Sprague-Dawley rats, HS was induced by withdrawing blood to reduce mean arterial pressure to 40 mmHg for 60 minutes followed by resuscitation. Rats were given SB216763 (600 μg/kg) intravenously 10 minutes prior to shock. To study microvascular permeability, the rats were injected intravenously with FITC-albumin (50 mg/kg) and its flux across the mesenteric post-capillary venules was determined using intravital microscopy. In cell-culture studies, rat lung microvascular endothelial cell (RLMEC) monolayers grown on Transwell plates were pre-treated with SB216763 (5 μM) followed by BAK (5 μg/mL) and caspase-3 (5 μg/mL) protein transfection. FITC-albumin (5 mg/mL) flux across cell monolayers indicates change in monolayer permeability. Activity of canonical Wnt pathway was determined by luciferase assay. Caspase-3 enzyme activity was assayed fluorometrically. Results The HS group showed significant increase in FITC-albumin extravasation (p<0.05) compared with sham. SB216763 significantly decrease HS-induced FITC-albumin extravasation (p<0.05). Pre-treatment with SB216763, protected against a BAK-induced increase in RLMEC monolayer permeability and caspase-3 activity, but failed to show similar results with a caspase-3-induced increase in monolayer permeability. Wnt3a treatment showed an increase in β-catenin dependent TCF-mediated transcription. Conclusion Inhibiting phosphorylation of

  5. The Crystal Structure of BRAF in Complex with an Organoruthenium Inhibitor Reveals a Mechanism for Inhibition of an Active Form of BRAF Kinase

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Peng; Streu, Craig; Qin, Jie; Bregman, Howard; Pagano, Nicholas; Meggers, Eric; Marmorstein, Ronen (Wistar); (UPENN)

    2012-06-19

    Substitution mutations in the BRAF serine/threonine kinase are found in a variety of human cancers. Such mutations occur in 70% of human malignant melanomas, and a single hyperactivating V600E mutation is found in the activation segment of the kinase domain and accounts for more than 90% of these mutations. Given this correlation, the molecular mechanism for BRAF regulation as well as oncogenic activation has attracted considerable interest, and activated forms of BRAF, such as BRAF{sup V600E}, have become attractive targets for small molecule inhibition. Here we report on the identification and subsequent optimization of a potent BRAF inhibitor, CS292, based on an organometallic kinase inhibitor scaffold. A cocrystal structure of CS292 in complex with the BRAF kinase domain reveals that CS292 binds to the ATP binding pocket of the kinase and is an ATP competitive inhibitor. The structure of the kinase-inhibitor complex also demonstrates that CS292 binds to BRAF in an active conformation and suggests a mechanism for regulation of BRAF by phosphorylation and BRAF{sup V600E} oncogene-induced activation. The structure of CS292 bound to the active form of the BRAF kinase also provides a novel scaffold for the design of BRAF{sup V600E} oncogene selective BRAF inhibitors for therapeutic application.

  6. Growth-Factor-Driven Rescue to Receptor Tyrosine Kinase (RTK) Inhibitors through Akt and Erk Phosphorylation in Pediatric Low Grade Astrocytoma and Ependymoma

    NARCIS (Netherlands)

    Sie, Mariska; den Dunnen, Wilfred F. A.; Lourens, Harm Jan; Meeuwsen-de Boer, Tiny G. J.; Scherpen, Frank J. G.; Zomerman, Walderik W.; Kampen, Kim R.; Hoving, Eelco W.; de Bont, Eveline S. J. M.

    2015-01-01

    Up to now, several clinical studies have been started investigating the relevance of receptor tyrosine kinase (RTK) inhibitors upon progression free survival in various pediatric brain tumors. However, single targeted kinase inhibition failed, possibly due to tumor resistance mechanisms. The present

  7. A phase II study of enzastaurin, a protein kinase C beta inhibitor, in patients with relapsed or refractory mantle cell lymphoma

    NARCIS (Netherlands)

    F. Morschhauser; J.F. Seymour; H.C. Kluin-Nelemans (Hanneke); A. Grigg; M. Wolf; M. Pfreundschuh (Michael); H. Tilly (Herve); J. Raemaekers; M.B. van 't Veer (Mars); N. Milpied; G. Cartron; A. Pezzutto; A. Spencer; F. Reyes; M. Dreyling (Martin)

    2008-01-01

    textabstractBackground: Protein kinase C beta (PKCβ), a pivotal enzyme in B-cell signaling and survival, is overexpressed in most cases of mantle cell lymphoma (MCL). Activation of PI3K/AKT pathway is involved in pathogenesis of MCL. Enzastaurin, an oral serine/threonine kinase inhibitor, suppresses

  8. A phase II study of enzastaurin, a protein kinase C beta inhibitor, in patients with relapsed or refractory mantle cell lymphoma

    NARCIS (Netherlands)

    Morschhauser, F.; Seymour, J. F.; Kluin-Nelemans, H. C.; Grigg, A.; Wolf, M.; Pfreundschuh, M.; Tilly, H.; Raemaekers, J.; van 't Veer, M. B.; Milpied, N.; Cartron, G.; Pezzutto, A.; Spencer, A.; Reyes, F.; Dreyling, M.

    2008-01-01

    Background: Protein kinase C beta (PKC beta), a pivotal enzyme in B-cell signaling and survival, is overexpressed in most cases of mantle cell lymphoma (MCL). Activation of PI3K/AKT pathway is involved in pathogenesis of MCL. Enzastaurin, an oral serine/threonine kinase inhibitor, suppresses signali

  9. Effects of inhibitor of κB kinase activity in the nucleus accumbens on emotional behavior.

    Science.gov (United States)

    Christoffel, Daniel J; Golden, Sam A; Heshmati, Mitra; Graham, Ami; Birnbaum, Shari; Neve, Rachael L; Hodes, Georgia E; Russo, Scott J

    2012-11-01

    Inhibitor of κB kinase (IκK) has historically been studied in the context of immune response and inflammation, but recent evidence demonstrates that IκK activity is necessary and sufficient for regulation of neuronal function. Chronic social defeat stress of mice increases IκK activity in the nucleus accumbens (NAc) and this increase is strongly correlated to depression-like behaviors. Inhibition of IκK signaling results in a reversal of chronic social defeat stress-induced social avoidance behavior. Here, we more completely define the role of IκK in anxiety and depressive-like behaviors. Mice underwent stereotaxic microinjection of a herpes simplex virus expressing either green fluorescent protein, a constitutively active form of IκK (IκKca), or a dominant negative form of IκK into the NAc. Of all three experimental groups, only mice expressing IκKca show a behavioral phenotype. Expression of IκKca results in a decrease in the time spent in the non-periphery zones of an open field arena and increased time spent immobile during a forced swim test. No baseline differences in sucrose preference were observed, but following the acute swim stress we noted a marked reduction in sucrose preference. To determine whether IκK activity alters responses to other acute stressors, we examined behavior and spine morphology in mice undergoing an acute social defeat stress. We found that IκKca enhanced social avoidance behavior and promoted thin spine formation. These data show that IκK in NAc is a critical regulator of both depressive- and anxiety-like states and may do so by promoting the formation of immature excitatory synapses.

  10. Transcriptional regulation of human polo-like kinases and early mitotic inhibitors

    Institute of Scientific and Technical Information of China (English)

    Moe Tategu; Hiroki Nakagawa; Kaori Sasaki; Rieko Yamauchi; Sota Sekimachi; Yuka Suita; Naoko Watanabe; Kenichi Yoshida

    2008-01-01

    Human polo-like kinases (PLK1-PLK4) have been implicated in mitotic regulation and carcinogenesis.PLK1 phosphorylates early mitotic inhibitor 1 (Emil) to ensure mitosis entry,whereas Emi2 plays a key role during the meiotic cell cycle.Transcription factor E2F is primarily considered to regulate the G1/S transition of the cell cycle but its involvement in the regulation of mitosis has also been recently suggested.A gap still exists between the molecular basis of E2F and mitotic regulation.The present study was designed to characterize the transcriptional regulation of human PLK and Emi genes.Adenoviral overexpression of E2F1 increased PLK1 and PLK3 mRNA levels in A549 cells.A reporter gene assay revealed that the putative promoter regions of PLK1,PLK3,and PLK4 genes were responsive to ac-tivators E2F,E2F1-E2F3.We further characterized the putative promoter regions of Emil and Emi2 genes,and these could be regulated by activators E2F and E2F1-E2F4,respectively.Finally,PLK1-PLK4,Emil,and Emi2 mRNA expression levels in human adult,fetal tis-sues,and several cell lines indicated that each gene has a unique expression pattern but is uniquely expressed in common tissues and cells such as the testes and thymus.Collectively,these results indicate that E2F can integrate G1/S and G2/Mto oscillate the cell cycle by regu-lating mitotic genes PLK and Emi,leading to determination of the cell fate.

  11. Research progress of protein kinase B/Akt inhibitors%蛋白激酶B/Akt抑制剂

    Institute of Scientific and Technical Information of China (English)

    郝茜; 钟嫄; 王朋; 赵桂森

    2011-01-01

    PI3K (phosphatidylinositol 3-kinase)/Akt (protein kinase B, PKB) signaling pathway plays a key role in cell growth and survival. Excessive activation of PI3K/Akt pathway has been found in many tumors. In more than 50% of human tumors, Akt or/and its upstream regulatory molecules (such as PTEN and PI3K) changes abnormally. So Akt has become a hot target for cancer prevention and therapy. Recently, many effective small-molecule Akt inhibitors with different mechanisms have been found. According to the binding site and/or the chemical structures of various Akt inhibitors, they are divided into ATP competitive inhibitor, Akt allosteric inhibitor and phosphatidylinositol analog inhibitor. This paper reviews the relationship between PI3K/Akt pathway and tumors, and the research progress of Akt inhibitor, which will contribute to the design of new anti-tumor drugs.%磷脂酰肌醇-3-激酶(phosphatidylinositol 3-kinase,PI3K)/蛋白激酶B(protein kinase B,PKB/Akt)信号通路在细胞生长与存活中起着关键作用,PI3K/Akt通路的过度激活在多种肿瘤中常见.Akt激酶本身以及Akt激酶上游调节分子,例如PTEN和PI3K,在超过50%的人类肿瘤中均有异常变化.因此Akt成为肿瘤预防和肿瘤靶向治疗的热点之一.许多小分子化合物通过不同机制抑制Akt活性,根据小分子抑制剂与激酶的结合部位和化学结构不同,主要分为ATP竞争性抑制剂、Akt变构抑制剂和磷脂酰肌醇类似物抑制剂.本文综述了PI3K/Akt通路与肿瘤的关系和Akt抑制剂的研究现状,为新型抗癌药物的设计研究提供参考.

  12. Efficacy of the polo-like kinase inhibitor rigosertib, alone or in combination with Abelson tyrosine kinase inhibitors, against break point cluster region-c-Abelson-positive leukemia cells.

    Science.gov (United States)

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

    2015-08-21

    The potency of Abelson (ABL) tyrosine kinase inhibitors (TKIs) against chronic myeloid leukemia (CML) has been demonstrated. However, ABL TKI resistance can develop. In this study, we investigated the efficacy of a combination therapy including rigosertib (ON 01910.Na), a polo-like kinase (PLK) and phosphoinositide 3-kinase (PI3K) inhibitor, and ABL TKIs. A 72-h rigosertib treatment was found to inhibit cell growth, induce apoptosis, reduce phosphorylation of the breakpoint cluster region-c (BCR)-ABL and its substrate Crk-L, and increase the activities of caspase 3 and poly (ADP-ribose) polymerase (PARP). This combination therapy also exerted a synergistic inhibitory effect on Philadelphia chromosome (Ph)-positive cell proliferation and reduced the phosphorylation of BCR-ABL and Crk-L while increasing that of cleaved PARP and the H2A.X histone. Rigosertib also potently inhibited the growth of ABL TKI-resistant cells, and cotreatment with ABL TKIs and rigosertib induced higher cytotoxicity. These results indicate that rigosertib treatment may be a powerful strategy against ABL TKI-resistant cells and could enhance the cytotoxic effects of ABL TKIs.

  13. Identification of AKN-032, a novel 2-aminopyrazine tyrosine kinase inhibitor, with significant preclinical activity in acute myeloid leukemia.

    Science.gov (United States)

    Eriksson, Anna; Höglund, Martin; Lindhagen, Elin; Aleskog, Anna; Hassan, Sadia Bashir; Ekholm, Carina; Fhölenhag, Karin; Jensen, Annika Jenmalm; Löthgren, Agneta; Scobie, Martin; Larsson, Rolf; Parrow, Vendela

    2010-11-15

    Aberrant signal transduction by mutant or overexpressed protein kinases has emerged as a promising target for treatment of acute myeloid leukemia (AML). We here present a novel low molecular weight kinase inhibitor, AKN-032, targeting the FMS-like tyrosine kinase 3 (FLT3) and discovered in a new type of screening funnel combining the target therapy approach with sequential cellular screens. AKN-032 was identified among 150 selected hits from three different high throughput kinase screens. Further characterization showed inhibitory activity on FLT3 enzyme with an IC(50) of 70 nM. Western blot analysis revealed reduced autophosphorylation of the FLT3-receptor in AML cell line MV4-11 cells after exposure to AKN-032. Flow cytometry disclosed cytotoxic activity against MV4-11, but not against non-malignant 3T3-L1 fibroblast cells. Using a fluorometric microculture cytotoxicity assay, AKN-032 was tested against 15 cell lines and displayed a potent cytotoxic activity in AML cell lines MV4-11 (IC(50)=0.4 μM) and Kasumi-1 (IC(50)=2.3 μM). AKN-032 was also highly cytotoxic in tumor cells from AML patients in vitro. Furthermore, AKN-032 demonstrated significant antileukemic effect in a relatively resistant in vivo hollow fiber mouse model. No major toxicity was observed in the animals. In conclusion, AKN-032 is a promising new kinase inhibitor with significant in vivo and in vitro activity in AML. Results from the hollow fiber mouse assay suggest a favorable toxicity profile. Future studies will focus on pharmacokinetic properties, toxicity as well as further clarifying the mechanisms of action of AKN-032 in AML.

  14. 3-Amido pyrrolopyrazine JAK kinase inhibitors: development of a JAK3 vs JAK1 selective inhibitor and evaluation in cellular and in vivo models.

    Science.gov (United States)

    Soth, Michael; Hermann, Johannes C; Yee, Calvin; Alam, Muzaffar; Barnett, Jim W; Berry, Pamela; Browner, Michelle F; Frank, Karl; Frauchiger, Sandra; Harris, Seth; He, Yang; Hekmat-Nejad, Mohammad; Hendricks, Than; Henningsen, Robert; Hilgenkamp, Ramona; Ho, Hoangdung; Hoffman, Ann; Hsu, Pei-Yuan; Hu, Dong-Qing; Itano, Andrea; Jaime-Figueroa, Saul; Jahangir, Alam; Jin, Sue; Kuglstatter, Andreas; Kutach, Alan K; Liao, Cheng; Lynch, Stephen; Menke, John; Niu, Linghao; Patel, Vaishali; Railkar, Aruna; Roy, Douglas; Shao, Ada; Shaw, David; Steiner, Sandra; Sun, Yongliang; Tan, Seng-Lai; Wang, Sandra; Vu, Minh Diem

    2013-01-10

    The Janus kinases (JAKs) are involved in multiple signaling networks relevant to inflammatory diseases, and inhibition of one or more members of this class may modulate disease activity or progression. We optimized a new inhibitor scaffold, 3-amido-5-cyclopropylpyrrolopyrazines, to a potent example with reasonable kinome selectivity, including selectivity for JAK3 versus JAK1, and good biopharmaceutical properties. Evaluation of this analogue in cellular and in vivo models confirmed functional selectivity for modulation of a JAK3/JAK1-dependent IL-2 stimulated pathway over a JAK1/JAK2/Tyk2-dependent IL-6 stimulated pathway.

  15. Selectivity Profiling and Biological Activity of Novel β-Carbolines as Potent and Selective DYRK1 Kinase Inhibitors.

    Directory of Open Access Journals (Sweden)

    Katharina Rüben

    Full Text Available DYRK1A is a pleiotropic protein kinase with diverse functions in cellular regulation, including cell cycle control, neuronal differentiation, and synaptic transmission. Enhanced activity and overexpression of DYRK1A have been linked to altered brain development and function in Down syndrome and neurodegenerative diseases such as Alzheimer's disease. The β-carboline alkaloid harmine is a high affinity inhibitor of DYRK1A but suffers from the drawback of inhibiting monoamine oxidase A (MAO-A with even higher potency. Here we characterized a series of novel harmine analogs with minimal or absent MAO-A inhibitory activity. We identified several inhibitors with submicromolar potencies for DYRK1A and selectivity for DYRK1A and DYRK1B over the related kinases DYRK2 and HIPK2. An optimized inhibitor, AnnH75, inhibited CLK1, CLK4, and haspin/GSG2 as the only off-targets in a panel of 300 protein kinases. In cellular assays, AnnH75 dose-dependently reduced the phosphorylation of three known DYRK1A substrates (SF3B1, SEPT4, and tau without negative effects on cell viability. AnnH75 inhibited the cotranslational tyrosine autophosphorylation of DYRK1A and threonine phosphorylation of an exogenous substrate protein with similar potency. In conclusion, we have characterized an optimized β-carboline inhibitor as a highly selective chemical probe that complies with desirable properties of drug-like molecules and is suitable to interrogate the function of DYRK1A in biological studies.

  16. Differential sensitivities of trastuzumab (Herceptin)-resistant human breast cancer cells to phosphoinositide-3 kinase (PI-3K) and epidermal growth factor receptor (EGFR) kinase inhibitors.

    Science.gov (United States)

    Chan, Carmel T; Metz, Marianne Z; Kane, Susan E

    2005-05-01

    Her2 (erbB2/neu) is overexpressed in 25-30% of human breast cancers. Herceptin is a recombinant humanized Her2 antibody used to treat breast cancer patients with Her2 overexpression. Over a 5-month selection process, we isolated clones of BT474 (BT) human breast carcinoma cells (BT/Her(R)) that were resistant to Herceptin in vitro. In BT/Her(R) subclones, cell-surface, phosphorylated and total cellular Her2 protein remained high in the continuous presence of Herceptin. Likewise, the levels of cell-surface, phosphorylated, and total cellular Her3 and EGFR were either unchanged or only slightly elevated in BT/Her(R) subclones relative to BT cells. One BT/Her(R) subclone had substantially upregulated cell-surface EGFR, but this did not correlate with a higher relative resistance to Herceptin. In looking at the downstream PI-3K/Akt signaling pathway, phosphorylated and total Akt levels and Akt kinase activities were all sustained in BT/Her(R) subclones in the presence of Herceptin, but significantly downregulated in BT cells exposed to Herceptin. Whereas BT cells lost sensitivity to the PI-3K inhibitor LY294002 in the presence of Herceptin, BT/Her(R) subclones were equally sensitive to this agent in the presence and absence of Herceptin. This suggests that BT/Her(R) subclones acquired a Herceptin-resistant mechanism of PI-3K signaling. BT/Her(R) subclones were also sensitive to the EGFR kinase inhibitor AG1478 in the presence of Herceptin, to the same extent as BT cells. The BT/Her(R) subclones provide new insights into mechanisms of Herceptin resistance and suggest new treatment strategies in combination with other inhibitors targeted to signal transduction pathways.

  17. Steady-state concentrations of mRNA encoding two inhibitors of protein kinase C in ovine luteal tissue.

    Science.gov (United States)

    Juengel, J L; Melner, M H; Clapper, J A; Turzillo, A M; Moss, G E; Nett, T M; Niswender, G D

    1998-07-01

    Prostaglandin F2 alpha (PGF2 alpha) decreases secretion of progesterone from the corpus luteum in domestic ruminants. However, it is less effective during the early part of the oestrous cycle (Louis et al., 1973) and at the time of maternal recognition of pregnancy (Silvia and Niswender, 1984; Lacroix and Kann, 1986). Decreased luteal responsiveness may be due to failure of PGF2 alpha to activate fully its normal second messenger system, protein kinase C (PKC). Alternatively, increased resistance of the corpus luteum to PGF2 alpha might be attributable to greater concentrations of recently identified biological inhibitors of PKC. These possibilities were addressed by measuring steady-state concentrations of mRNA encoding PGF2 alpha receptor and two inhibitors of PKC, protein kinase C inhibitor-1 (PKCI-1) and kinase C inhibitor protein-1 (KCIP-1, brain 14-3-3 protein), in corpora lutea collected from ewes on days 4, 10 and 15 of the oestrous cycle (n = 5 per day) and day 15 of pregnancy (n = 7). There were no differences in mean concentrations of mRNA encoding PGF2 alpha receptor among the groups. However, concentrations of mRNA encoding both inhibitors of PKC were higher (P day 4 of the oestrous cycle compared with the other groups. Treatment of ewes with a luteolytic dose of PGF2 alpha, which activates PKC, did not change concentrations of mRNA encoding either PKCI-1 or KCIP-I up to 24 h later. Luteal expression of mRNA encoding the PKC inhibitors and PGF2 alpha receptor was also examined in ewes treated with oestradiol in vivo for 16 h in the midluteal phase. High concentrations of oestradiol in serum (20 and 70 pg ml-1) did not influence quantities of any of the mRNAs examined. Therefore, an increase in PKC inhibitors may be involved in resistance of the corpus luteum to PGF2 alpha during the early part of the oestrous cycle but does not appear to mediate the increased resistance of the corpus luteum to PGF2 alpha during maternal recognition of pregnancy

  18. Fragment-based discovery of a dual pan-RET/VEGFR2 kinase inhibitor optimized for single-agent polypharmacology1

    OpenAIRE

    Frett, Brendan; Carlomagno, Francesca; Moccia, Maria Luisa; Brescia, Annalisa; Federico, Giorgia; De Falco, Valentina; Admire, Brittany; Chen, Zhongzhu; Qi, Wenqing; Santoro, Massimo; Li, Hong-yu

    2015-01-01

    Oncogenic conversion of the RET (rearranged during transfection) tyrosine kinase is associated with several cancers. A fragment-based chemical screen lead to the identification of a novel RET inhibitor, Pz-1. Modeling and kinetic analysis identified Pz-1 as a Type-II tyrosine kinase inhibitor, able to bind the DFG-out conformation of the kinase. Importantly, from a single-agent polypharmacology standpoint, Pz-1 was shown active on VEGFR2, which can block blood supply required for RET-stimulat...

  19. Expression of Raf kinase inhibitor protein is downregulated in response to Newcastle disease virus infection to promote viral replication.

    Science.gov (United States)

    Yin, Renfu; Liu, Xinxin; Bi, Yuhai; Xie, Guangyao; Zhang, Pingze; Meng, Xin; Ai, Lili; Xu, Rongyi; Sun, Yuzhang; Stoeger, Tobias; Ding, Zhuang

    2015-09-01

    Newcastle disease virus (NDV) causes a severe and economically significant disease affecting almost the entire poultry industry worldwide. However, factors that affect NDV replication in host cells are poorly understood. Raf kinase inhibitory protein (RKIP) is a physiological inhibitor of c-RAF kinase and NF-κB signalling, known for their functions in the control of immune response as well as tumour invasion and metastasis. In the present study, we investigated the consequences of overexpression of host RKIP during viral infection. We demonstrate that NDV infection represses RKIP expression thereby promoting virus replication. Experimental upregulation of RKIP in turn acts as a potential antiviral defence mechanism in host cells that restricts NDV replication by repressing the activation of Raf/MEK/ERK and IκBα/NF-κB signalling pathways. Our results not only extend the concept of linking NDV-host interactions, but also reveal RKIP as a new class of protein-kinase-inhibitor protein that affects NDV replication with therapeutic potential.

  20. Lipid raft localization of EGFR alters the response of cancer cells to the EGFR tyrosine kinase inhibitor gefitinib.

    Science.gov (United States)

    Irwin, Mary E; Mueller, Kelly L; Bohin, Natacha; Ge, Yubin; Boerner, Julie L

    2011-09-01

    Epidermal growth factor receptor (EGFR) is overexpressed in many cancer types including ~30% of breast cancers. Several small molecule tyrosine kinase inhibitors (TKIs) targeting EGFR have shown clinical efficacy in lung and colon cancers, but no benefit has been noted in breast cancer. Thirteen EGFR expressing breast cancer cell lines were analyzed for response to EGFR TKIs. Seven were found to be EGFR TKI resistant; while shRNA knockdown of EGFR determined that four of these cell lines retained the requirement of EGFR protein expression for growth. Interestingly, EGFR localized to plasma membrane lipid rafts in all four of these EGFR TKI-resistant cell lines, as determined by biochemical raft isolation and immunofluorescence. When lipid rafts were depleted of cholesterol using lovastatin, all four cell lines were sensitized to EGFR TKIs. In fact, the effects of the cholesterol biosynthesis inhibitors and gefitinib were synergistic. While gefitinib effectively abrogated phosphorylation of Akt- and mitogen-activated protein kinase in an EGFR TKI-sensitive cell line, phosphorylation of Akt persisted in two EGFR TKI-resistant cell lines, however, this phosphorylation was abrogated by lovastatin treatment. Thus, we have shown that lipid raft localization of EGFR correlates with resistance to EGFR TKI-induced growth inhibition and pharmacological depletion of cholesterol from lipid rafts decreases this resistance in breast cancer cell lines. Furthermore, we have presented evidence to suggest that when EGFR localizes to lipid rafts, these rafts provide a platform to facilitate activation of Akt signaling in the absence of EGFR kinase activity.

  1. Bisubstrate analog probes for the insulin receptor protein tyrosine kinase: molecular yardsticks for analyzing catalytic mechanism and inhibitor design.

    Science.gov (United States)

    Hines, Aliya C; Parang, Keykavous; Kohanski, Ronald A; Hubbard, Stevan R; Cole, Philip A

    2005-08-01

    Bisubstrate analogs have the potential to provide enhanced specificity for protein kinase inhibition and tools to understand catalytic mechanism. Previous efforts led to the design of a peptide-ATP conjugate bisubstrate analog utilizing aminophenylalanine in place of tyrosine and a thioacetyl linker to the gamma-phosphate of ATP which was a potent inhibitor of the insulin receptor kinase (IRK). In this study, we have examined the contributions of various electrostatic and structural elements in the bisubstrate analog to IRK binding affinity. Three types of changes (seven specific analogs in all) were introduced: a Tyr isostere of the previous aminophenylalanine moiety, modifications of the spacer between the adenine and the peptide, and deletions and substitutions within the peptide moiety. These studies allowed a direct evaluation of the hydrogen bond strength between the anilino nitrogen of the bisubstrate analog and the enzyme catalytic base Asp and showed that it contributes 2.5 kcal/mol of binding energy, in good agreement with previous predictions. Modifications of the linker length resulted in weakened inhibitory affinity, consistent with the geometric requirements of an enzyme-catalyzed dissociative transition state. Alterations in the peptide motif generally led to diminished inhibitory potency, and only some of these effects could be rationalized based on prior kinetic and structural studies. Taken together, these results suggest that a combination of mechanism-based design and empirical synthetic manipulation will be necessary in producing optimized protein kinase bisubstrate analog inhibitors.

  2. Why Earth aurorae shine?

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ By using the data obtained from three satellites of the Cluster mission launched by the European Space Agency (ESA), CAO Jinbin from the CAS Center for Space Science and Applied Research (CSSAR) and his US and European co-workers have clarified why Earth's aurorae shine.Their work entitled Joint Observations by Cluster Satellites of Bursty Bulk Flows in the Magnetotail was published in a recent issue of Journal of Geophysical Research.

  3. Noninvasive magnetic resonance spectroscopic pharmacodynamic markers of the choline kinase inhibitor MN58b in human carcinoma models.

    Science.gov (United States)

    Al-Saffar, Nada M S; Troy, Helen; Ramírez de Molina, Ana; Jackson, Laura E; Madhu, Basetti; Griffiths, John R; Leach, Martin O; Workman, Paul; Lacal, Juan C; Judson, Ian R; Chung, Yuen-Li

    2006-01-01

    MN58b is a novel anticancer drug that inhibits choline kinase, resulting in inhibition of phosphocholine synthesis. The aim of this work was to develop a noninvasive and robust pharmacodynamic biomarker for target inhibition and, potentially, tumor response following MN58b treatment. Human HT29 (colon) and MDA-MB-231 (breast) carcinoma cells were examined by proton (1H) and phosphorus (31P) magnetic resonance spectroscopy (MRS) before and after treatment with MN58b both in culture and in xenografts. An in vitro time course study of MN58b treatment was also carried out in MDA-MB-231 cells. In addition, enzymatic assays of choline kinase activity in cells were done. A decrease in phosphocholine and total choline levels (P < 0.05) was observed in vitro in both cell lines after MN58b treatment, whereas the inactive analogue ACG20b had no effect. In MDA-MB-231 cells, phosphocholine fell significantly as early as 4 hours following MN58b treatment, whereas a drop in cell number was observed at 48 hours. Significant correlation was also found between phosphocholine levels (measured by MRS) and choline kinase activities (r2 = 0.95, P = 0.0008) following MN58b treatment. Phosphomonoesters also decreased significantly (P < 0.05) in both HT29 and MDA-MB-231 xenografts with no significant changes in controls. 31P-MRS and 1H-MRS of tumor extracts showed a significant decrease in phosphocholine (P < or = 0.05). Inhibition of choline kinase by MN58b resulted in altered phospholipid metabolism both in cultured tumor cells and in vivo. Phosphocholine levels were found to correlate with choline kinase activities. The decrease in phosphocholine, total choline, and phosphomonoesters may have potential as noninvasive pharmacodynamic biomarkers for determining tumor response following treatment with choline kinase inhibitors.

  4. Identification and characterization of a small-molecule inhibitor of death-associated protein kinase 1

    DEFF Research Database (Denmark)

    Wilbek, Theis S; Skovgaard, Tine; Sorrell, Fiona J

    2015-01-01

    A novel imidazo-pyramidazine inhibitor of DAPK1 that undergoes class-specific interactions and extends into the substrate recognition site has been identified. This inhibitor is a good starting point for the development of selective and potent inhibitors of DAPK1, with potential use against stroke...

  5. The Sensitivity of Memory Consolidation and Reconsolidation to Inhibitors of Protein Synthesis and Kinases: Computational Analysis

    Science.gov (United States)

    Zhang, Yili; Smolen, Paul; Baxter, Douglas A.; Byrne, John H.

    2010-01-01

    Memory consolidation and reconsolidation require kinase activation and protein synthesis. Blocking either process during or shortly after training or recall disrupts memory stabilization, which suggests the existence of a critical time window during which these processes are necessary. Using a computational model of kinase synthesis and…

  6. Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design

    Energy Technology Data Exchange (ETDEWEB)

    Han, Seungil; Mistry, Anil; Chang, Jeanne S.; Cunningham, David; Griffor, Matt; Bonnette, Peter C.; Wang, Hong; Chrunyk, Boris A.; Aspnes, Gary E.; Walker, Daniel P.; Brosius, Arthur D.; Buckbinder, Leonard; Pfizer

    2009-05-21

    Proline-rich tyrosine kinase 2 (PYK2) is a cytoplasmic, non-receptor tyrosine kinase implicated in multiple signaling pathways. It is a negative regulator of osteogenesis and considered a viable drug target for osteoporosis treatment. The high-resolution structures of the human PYK2 kinase domain with different inhibitor complexes establish the conventional bilobal kinase architecture and show the conformational variability of the DFG loop. The basis for the lack of selectivity for the classical kinase inhibitor, PF-431396, within the FAK family is explained by our structural analyses. Importantly, the novel DFG-out conformation with two diarylurea inhibitors (BIRB796, PF-4618433) reveals a distinct subclass of non-receptor tyrosine kinases identifiable by the gatekeeper Met-502 and the unique hinge loop conformation of Leu-504. This is the first example of a leucine residue in the hinge loop that blocks the ATP binding site in the DFG-out conformation. Our structural, biophysical, and pharmacological studies suggest that the unique features of the DFG motif, including Leu-504 hinge-loop variability, can be exploited for the development of selective protein kinase inhibitors.

  7. Inhibition of thyrotropin-stimulated DNA synthesis by microinjection of inhibitors of cellular Ras and cyclic AMP-dependent protein kinase.

    Science.gov (United States)

    Kupperman, E; Wen, W; Meinkoth, J L

    1993-08-01

    Microinjection of a dominant interfering mutant of Ras (N17 Ras) caused a significant reduction in thyrotropin (thyroid-stimulating hormone [TSH])-stimulated DNA synthesis in rat thyroid cells. A similar reduction was observed following injection of the heat-stable protein kinase inhibitor of the cyclic AMP-dependent protein kinase. Coinjection of both inhibitors almost completely abolished TSH-induced DNA synthesis. In contrast to TSH, overexpression of cellular Ras protein did not stimulate the expression of a cyclic AMP response element-regulated reporter gene. Similarly, injection of N17 Ras had no effect on TSH-stimulated reporter gene expression. Moreover, overexpression of cellular Ras protein stimulated similar levels of DNA synthesis in the presence or absence of the heat-stable protein kinase inhibitor. Together, these results suggest that in Wistar rat thyroid cells, a full mitogenic response to TSH requires both Ras and cyclic APK-dependent protein kinase.

  8. Several Human Cyclin-Dependent Kinase Inhibitors, Structurally Related to Roscovitine, As New Anti-Malarial Agents

    Directory of Open Access Journals (Sweden)

    Sandrine Houzé

    2014-09-01

    Full Text Available In Africa, malaria kills one child each minute. It is also responsible for about one million deaths worldwide each year. Plasmodium falciparum, is the protozoan responsible for the most lethal form of the disease, with resistance developing against the available anti-malarial drugs. Among newly proposed anti-malaria targets, are the P. falciparum cyclin-dependent kinases (PfCDKs. There are involved in different stages of the protozoan growth and development but share high sequence homology with human cyclin-dependent kinases (CDKs. We previously reported the synthesis of CDKs inhibitors that are structurally-related to (R-roscovitine, a 2,6,9-trisubstituted purine, and they showed activity against neuronal diseases and cancers. In this report, we describe the synthesis and the characterization of new CDK inhibitors, active in reducing the in vitro growth of P. falciparum (3D7 and 7G8 strains. Six compounds are more potent inhibitors than roscovitine, and three exhibited IC50 values close to 1 µM for both 3D7 and 7G8 strains. Although, such molecules do inhibit P. falciparum growth, they require further studies to improve their selectivity for PfCDKs.

  9. Paroxetine Is a Direct Inhibitor of G Protein-Coupled Receptor Kinase 2 and Increases Myocardial Contractility

    Energy Technology Data Exchange (ETDEWEB)

    Thal, David M. [Univ. of Michigan, Ann Arbor, MI (United States); Homan, Kristoff T. [Univ. of Michigan, Ann Arbor, MI (United States); Chen, Jun [Univ. of New Mexico Health Sciences Center, Albuquerque, NM (United States); Wu, Emily K. [Univ. of Michigan, Ann Arbor, MI (United States); Hinkle, Patricia M. [Univ. of Rochester Medical Center, Rochester, NY (United States); Huang, Z. Maggie [Temple Univ. School of Medicine, Philadelphia, Pennsylvania (United States); Chuprun, J. Kurt [Temple Univ. School of Medicine, Philadelphia, Pennsylvania (United States); Song, Jianliang [Temple Univ. School of Medicine, Philadelphia, Pennsylvania (United States); Gao, Erhe [Temple Univ. School of Medicine, Philadelphia, Pennsylvania (United States); Cheung, Joseph Y. [Temple Univ. School of Medicine, Philadelphia, Pennsylvania (United States); Sklar, Larry A. [Univ. of New Mexico Health Sciences Center, Albuquerque, NM (United States); Koch, Walter J. [Temple Univ. School of Medicine, Philadelphia, Pennsylvania (United States); Tesmer, John J.G. [Univ. of Michigan, Ann Arbor, MI (United States)

    2012-08-10

    G protein-coupled receptor kinase 2 (GRK2) is a well-established therapeutic target for the treatment of heart failure. In this paper we identify the selective serotonin reuptake inhibitor (SSRI) paroxetine as a selective inhibitor of GRK2 activity both in vitro and in living cells. In the crystal structure of the GRK2·paroxetine–Gβγ complex, paroxetine binds in the active site of GRK2 and stabilizes the kinase domain in a novel conformation in which a unique regulatory loop forms part of the ligand binding site. Isolated cardiomyocytes show increased isoproterenol-induced shortening and contraction amplitude in the presence of paroxetine, and pretreatment of mice with paroxetine before isoproterenol significantly increases left ventricular inotropic reserve in vivo with no significant effect on heart rate. Neither is observed in the presence of the SSRI fluoxetine. Our structural and functional results validate a widely available drug as a selective chemical probe for GRK2 and represent a starting point for the rational design of more potent and specific GRK2 inhibitors.

  10. Synthesis of new pyrrolo[1,2-a]quinoxaline derivatives as potential inhibitors of Akt kinase.

    Science.gov (United States)

    Desplat, Vanessa; Geneste, Ambre; Begorre, Marc-Antoine; Fabre, Solene Belisle; Brajot, Stephane; Massip, Stephane; Thiolat, Denis; Mossalayi, Djavad; Jarry, Christian; Guillon, Jean

    2008-10-01

    Akt kinases are attractive targets for small molecule drug discovery because of their key role in tumor cell survival/proliferation and their overexpression/activation in many human cancers. Recent efforts in the development and biological evaluation of small molecule inhibitors of Akt have led to the identification of novel Akt kinase inhibitors, based on a quinoxaline or pyrazinone scaffold. A series of new substituted pyrrolo[1,2-a]quinoxaline derivatives, structural analogues of these active quinoxaline or pyrazinone pharmacophores, was synthesized from various substituted 2-nitroanilines or 1,2-phenylenediamine via multistep heterocyclization process. These new compounds were tested for their in vitro ability to inhibit the proliferation of the human leukemic cell lines K562, U937 and HL60, and the breast cancer cell line MCF7. Three of these human cell lines (K562, U937 and MCF7) exhibited an active phosphorylated Akt form. The most promising active pyrroloquinoxalines were found to be 1a that inhibited K562 cell line proliferation with an IC(50) of 4.5 microM, and 1h that inhibited U937 and MCF7 cell lines with IC(50) of 5 and 8 microM, respectively. These two candidates exhibited more potent activities than the reference inhibitor A6730.

  11. Overcoming the drug resistance problem with second-generation tyrosine kinase inhibitors: from enzymology to structural models.

    Science.gov (United States)

    Crespan, E; Zucca, E; Maga, G

    2011-01-01

    Protein phosphorylation is one of the major pathways used by eukaryotic cells to propagate signals to the final effectors, regulating multiple aspects of the living cell, such as metabolism, growth, differentiation, adhesion, motility, genome stability and death. In this context, tyrosine kinases (TKs) play a central role in signal transduction and their overexpression or disregulated activity has been implicated in tumor onset and malignancy progression. To date, eight TKs inhibitors have been approved by FDA for the treatment of specific tumors. In spite of their efficacy, insurgence of resistance is a common feature after prolonged administration. The selective pressure by these drugs, in fact, induces clonal expansion of subsets of cancer cells harboring TKs mutations, leading to decreased inhibition potency. Alternatively, resistance to TK inhibitors can be acquired through the activation of others, often unrelated, TKs. For this reason, while stringent target selectivity of TKs inhibitors has been always considered a desirable feature in order to limit toxicity, molecules targeting different TKs have been recently shown to be promising anti-cancer agents as well. Understanding the molecular mechanisms that confer resistance to TK inhibitors, through a combination of enzymatic, structural and cellular studies, is essential in the development of second generation inhibitors active also towards drug resistant tumors.

  12. Biological and Molecular Effects of Small Molecule Kinase Inhibitors on Low-Passage Human Colorectal Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Falko Lange

    2014-01-01

    Full Text Available Low-passage cancer cell lines are versatile tools to study tumor cell biology. Here, we have employed four such cell lines, established from primary tumors of colorectal cancer (CRC patients, to evaluate effects of the small molecule kinase inhibitors (SMI vemurafenib, trametinib, perifosine, and regorafenib in an in vitro setting. The mutant BRAF (V600E/V600K inhibitor vemurafenib, but also the MEK1/2 inhibitor trametinib efficiently inhibited DNA synthesis, signaling through ERK1/2 and expression of genes downstream of ERK1/2 in BRAF mutant cells only. In case of the AKT inhibitor perifosine, three cell lines showed a high or intermediate responsiveness to the drug while one cell line was resistant. The multikinase inhibitor regorafenib inhibited proliferation of all CRC lines with similar efficiency and independent of the presence or absence of KRAS, BRAF, PIK3CA, and TP53 mutations. Regorafenib action was associated with broad-range inhibitory effects at the level of gene expression but not with a general inhibition of AKT or MEK/ERK signaling. In vemurafenib-sensitive cells, the antiproliferative effect of vemurafenib was enhanced by the other SMI. Together, our results provide insights into the determinants of SMI efficiencies in CRC cells and encourage the further use of low-passage CRC cell lines as preclinical models.

  13. New auroras on the roles of the Chromosomal Passenger Complex in cytokinesis: implications for cancer therapies

    Directory of Open Access Journals (Sweden)

    Pier Paolo eD'Avino

    2015-10-01

    Full Text Available The Chromosomal Passenger Complex (CPC, composed of a kinase component, Aurora B, the scaffolding subunit Inner Centromeric Protein (INCENP, Borealin, and Survivin, is a key regulator of cell division. It controls multiple events, from chromosome condensation in prophase to the final separation or abscission of the two daughter cells. The essential functions of the CPC during metaphase, however, have always hindered an accurate study of its role during cytokinesis. The recent development of small molecule inhibitors against Aurora B and the use of elegant technologies such as chemical genetics have offered new approaches to study the functions of the CPC at the end of cell division. Here we review the recent findings about the roles of the CPC in controlling the assembly of the cleavage furrow, central spindle and midbody. We will also discuss the crucial function of this complex in controlling abscission timing in order to prevent abscission when lagging chromatin is present at the cleavage site, thereby avoiding the formation of genetically abnormal daughter cells. Finally, we offer our perspective on how to exploit the potential therapeutic applications of inhibiting CPC activity during cytokinesis in cancer cells.

  14. Myeloproliferative Neoplasms Associated with Mutation in JAK2V617F and Tyrosine Kinase Inhibitors as Therapeutic Strategy

    Directory of Open Access Journals (Sweden)

    Kaveh Tari

    2015-04-01

    Full Text Available MPNs including a heterogeneous group of clonal or oligoclonal hamtopathies characterized by proliferation and accumulation of mature myeloid cells. JAK2 tyrosine kinase mutation is the most common molecular lesion identified in 90% of cases. JAK2 is involved in EPO signaling pathway, and mutations in it lead to EPO-independent spontaneous phosphorylation. Most tyrosine kinase inhibitors (TKI are small molecules that compete with ATP for binding the ATP-binding site in tyrosine kinase domains, since ATP is a source of phosphate groups used by Tks to phosphorylate the target protein.there are many TKI agent that are studing for treatment of the MPNs with JAK2 tyrosine kinase mutation.the most important TKI drugs including CEP701, CYT387, LY2784544, SB1518, TG101348, XL019, INCB18424. Most important mechanism of them are reduse the splenomegaly, improvement of constitutional symptoms(improvement of bone marrow fibrosis and anemia. Although this drugs are useful but they have some side effect that common of them including Gastrointestinal disease (GI, diarrhea, nausea and vomiting, anemia, thrombocytopenia, thrombosis, leukocytosis, thrombocytosis, peripheral neuropathy, transient loss of blood pressure and lightheadedness.

  15. A novel anticancer diarylurea derivative HL-40 as a multi-kinases inhibitor with good pharmacokinetics in Wistar rats.

    Science.gov (United States)

    Lu, Yu-Yin; Zhao, Cui-Rong; Wang, Rui-Qi; Li, Wen-Bao; Qu, Xian-Jun

    2015-02-01

    HL-40, N-(4-(1-(4-chlorine indazole)) phenyl)-N-(4-chloro-3-three fluorine methyl phenyl) urea, is a novel diarylurea derivative. In this study, we investigated the kinases activities and binding constants, pharmacokinetics of HL-40, and then evaluated its anticancer efficacy by both in vitro and in vivo methods. Enzyme activities assays in vitro were employed to identify eight candidate kinase targets. The competition binding assays against eight candidate kinases suggested that HL-40 showed strong affinity to c-Kit, PDGFRβ and FLT3. The pharmacokinetic studies in Wistar rats showed that HL-40 could maintain high compound concentration and long residence time in the blood circulation. HL-40 possessed strong inhibition activities against 12 human cancer cells. Meanwhile, HL-40 effectively delayed the growth of cancer xenografts without significant toxicity to mice. Based on these in vitro and in vivo results, we suggested that HL-40 might be developed as a potential multi-kinases inhibitor for cancer treatment.

  16. The pan-HER family tyrosine kinase inhibitor afatinib overcomes HER3 ligand heregulin-mediated resistance to EGFR inhibitors in non-small cell lung cancer.

    Science.gov (United States)

    Yonesaka, Kimio; Kudo, Keita; Nishida, Satomi; Takahama, Takayuki; Iwasa, Tsutomu; Yoshida, Takeshi; Tanaka, Kaoru; Takeda, Masayuki; Kaneda, Hiroyasu; Okamoto, Isamu; Nishio, Kazuto; Nakagawa, Kazuhiko

    2015-10-20

    Afatinib is a second generation epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) characterized as an irreversible pan-human EGFR (HER) family inhibitor. Afatinib remains effective for a subpopulation of patients with non-small cell lung cancer (NSCLC) with acquired resistance to first generation EGFF-TKIs such as erlotinib. Heregulin activates HER3 in an autocrine fashion and causes erlotinib resistance in NSCLC. Here we examine whether afatinib is effective against heregulin-overexpressing NSCLCs harboring EGFR activating mutations. Afatinib but not erlotinib decreased EGFR mutant NSCLC PC9HRG cell proliferation in vitro and in mouse xenografts. Afatinib inhibited phosphorylation of the cell signaling pathway proteins HER3, EGFR, HER2, and HER4, likely by prevention of trans-phosphorylation as HER3 kinase activity is inadequate for auto-phosphorylation. Afatinib, unlike erlotinib, inhibited AKT activation, resulting in elevated apoptosis in PC9HRG cells. Clinically, a subpopulation of 33 patients with EGFR mutations and NSCLC who had received first generation EGFR-TKIs exhibited elevated plasma heregulin levels compared to healthy volunteers; one of these achieved a response with afatinib therapy despite having previously developed erlotinib resistance. Afatinib can overcome heregulin-mediated resistance to erlotinib in EGFR mutant NSCLC. Further studies are necessary to determine whether heregulin can predict afatinib efficacy after development offirst generation EGFR-TKI resistance.

  17. Solvatochromism and linear solvation energy relationship of the kinase inhibitor SKF86002

    Science.gov (United States)

    Khattab, Muhammad; Van Dongen, Madeline; Wang, Feng; Clayton, Andrew H. A.

    2017-01-01

    We studied the spectroscopic characteristics of SKF86002, an anti-inflammatory and tyrosine kinase inhibitor drug candidate. Two conformers SKF86002A and SKF86002B are separated by energy barriers of 19.68 kJ·mol- 1 and 6.65 kJ·mol- 1 due to H-bonds, and produce the three major UV-Vis absorption bands at 325 nm, 260 nm and 210 nm in cyclohexane solutions. This environment-sensitive fluorophore exhibited emission in the 400-500 nm range with a marked response to changes in environment polarity. By using twenty-two solvents for the solvatochromism study, it was noticed that solvent polarity, represented by dielectric constant, was well correlated with the emission wavelength maxima of SKF86002. Thus, the SKF86002 fluorescence peak red shifted in aprotic solvents from 397.5 nm in cyclohexane to 436 nm in DMSO. While the emission maximum in hydrogen donating solvents ranged from 420 nm in t-butanol to 446 nm in N-methylformamide. Employing Lippert-Mataga, Bakhshiev and Kawski models, we found that one linear correlation provided a satisfactory description of polarity effect of 18 solvents on the spectral changes of SKF86002 with R2 values 0.78, 0.80 and 0.80, respectively. Additionally, the multicomponent linear regression analysis of Kamlet-Taft (R2 = 0.94) revealed that solvent acidity, basicity and polarity accounted for 31%, 24% and 45% of solvent effects on SKF86002 emission, respectively. While Catalán correlation (R2 = 0.92) revealed that solvatochromic change of SKF86002 emission was attributed to changes in solvent dipolarity (71%), solvent polarity (12%), solvent acidity (11%) and solvent basicity (6%). Plot of Reichardt transition energies and emission energies of SKF86002 in 18 solvents showed also a linear correlation with R2 = 0.90. The dipole moment difference between excited and ground state was calculated to be 3.4-3.5 debye.

  18. Up-regulation of Raf kinase inhibitor protein enhances chemosensitivity of cervical cancer cell

    Institute of Scientific and Technical Information of China (English)

    Xiao Chu; Xinqiang Ji; Mingcui Wang; Wenqing Zhang; Hui Ou; Chong Li

    2014-01-01

    Objective:The purpose of the study is to investigate the ef ects of up-regulation of Raf kinase inhibitor protein (RKlP) on the chemosensitivity of cervical cancer Hela cells. Methods:Eukaryotic expression plasmid pcDNA3.1(+)-ssRKIP containing human overal length RKIPcDNA was transfected into cervical cancer Hela cellby lipofectin assay, establishing a stable cellline containing a target gene by G418. Expression of RKIP in Hela cells was measured by Western blot analysis. After treatment with cisplatin of dif erent concentrations and intervals of time, the ef ect of RKIP on the proliferation of Hela cells was evaluated by MTT method. The flow cytometry was used to investigate whether the RKIP could inhibit apoptosis in Hela cells induced by cisplatin. Results:The expression of RKIP in Hela cells transfected with pcDNA3.1-ssRKIP was increased obviously. After dif erent concentrations of cisplatin treatment cells for 24, 48 and 72 h, the growth inhibition rate in Hela cells transfected with pcDNA3.1-ssRKIP was significantly higher than in control cells (P<0.05). With 5μg/mL cisplatin treatment for 24 h, pcDNA3.1-ssRKIP-transfected Hela cells had an obviously higher percentage of apoptosis (23.2 ± 0.24)%than non-transfected cells (12.4 ± 0.31)%and empty vector-transfected cells (13.4 ± 0.47)%. Without treatment of cisplatin, the percentage of apoptosis for Hela cells transfected with pcDNA3.1-ssRKIP was (5.7 ± 0.12)%, which was stil higher than those of the non-transfected cells (2.9 ± 0.21)%and empty vector-transfected cells (3 ± 0.08)%. Conclusion:Higher expres-sion of RKIP gene can improve chemosensitivitv of cervical cancer Hela cells to cisplatin.

  19. Mixed-lineage kinase inhibitors require the activation of Trk receptors to maintain long-term neuronal trophism and survival.

    Science.gov (United States)

    Wang, Leo H; Paden, Andrew J; Johnson, Eugene M

    2005-03-01

    Small-molecule mixed-lineage kinase (MLK) inhibitors, such as CEP-1347 [3,9-bis[(ethylthio)methyl]-(8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H, 11H-2,7b,11a-triazadibenzo(a,g)cycloocta(cde)trinden-1-one] and CEP-11004 [3,9-bis-[(isopropylthio)methyl]-(8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo(a,g)cycloocta(cde)trinden-1-one], prevent c-Jun NH(2)-terminal kinase (JNK) pathway activation as well as the consequent neuronal cell death in many cell culture and animal models. In the cell culture model of nerve growth factor (NGF)-deprived sympathetic neurons, we find that CEP-11004 induced a approximately 3-fold increase in the mRNA and protein levels of TrkA, the NGF receptor. This resulted in ligand-independent activation of the TrkA receptor and the downstream phosphatidylinositol 3-kinase (PI3-kinase) pathway. Addition of the Trk inhibitor K252a [(8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo(a,g)cycloocta(cde)-trinden-1-one] or the PI3-kinase inhibitor LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one] significantly decreased the protein synthesis rates, mitochondrial function, and neuronal survival maintained by CEP-11004. In contrast to sympathetic neurons, MLK inhibitors maintain only short-term survival of potassium- and serum-deprived rat cerebellar granule neurons (CGNs), despite continuous inhibition of the JNK pathway. We found that similar to sympathetic neurons, CEP-11004 increased the levels of the Trk receptor expressed in CGNs, TrkB. However, CGNs required the addition of the exogenous ligand brain-derived neurotrophic factor (BDNF) to activate the PI3-kinase pathway and to maintain long-term survival. BDNF activated TrkB, but caused rapid down-regulation of activated receptors and maintained only minimal survival. Therefore, increase in TrkB levels

  20. Synthesis, biological evaluation and molecular modeling studies of imidazo[1,2-a]pyridines derivatives as protein kinase inhibitors.

    Science.gov (United States)

    Lawson, Marie; Rodrigo, Jordi; Baratte, Blandine; Robert, Thomas; Delehouzé, Claire; Lozach, Olivier; Ruchaud, Sandrine; Bach, Stéphane; Brion, Jean-Daniel; Alami, Mouad; Hamze, Abdallah

    2016-11-10

    We report here the synthesis, the biological evaluation and the molecular modeling studies of new imidazo[1,2-a]pyridines derivatives designed as potent kinase inhibitors. This collection was obtained from 2-aminopyridines and 2-bromoacetophenone which afforded final compound in only one step. The bioactivity of this family of new compounds was tested using protein kinase and ATP competition assays. The structure-activity relationship (SAR) revealed that six compounds inhibit DYRK1A and CLK1 at a micromolar range. Docking studies provided possible explanations that correlate with the SAR data. The most active compound 4c inhibits CLK1 (IC50 of 0.7 μM) and DYRK1A (IC50 of 2.6 μM).

  1. Discovery and optimization of novel 4-phenoxy-6,7-disubstituted quinolines possessing semicarbazones as c-Met kinase inhibitors.

    Science.gov (United States)

    Qi, Baohui; Mi, Bin; Zhai, Xin; Xu, Ziyi; Zhang, Xiaolong; Tian, Zeru; Gong, Ping

    2013-09-01

    A novel series of N(1)-(3-fluoro-4-(6,7-disubstituted-quinolin-4-yloxy)phenyl)-N(4)-arylidenesemicarbazide derivatives were synthesized and evaluated for their c-Met kinase inhibition and cytotoxicity against A549, HT-29, MKN-45 and MDA-MB-231 cancer cell lines in vitro. Several potent compounds were further evaluated against three other cancer cell lines (U87MG, NCI-H460 and SMMC7721). Most of compounds tested exhibited moderate to excellent activity. The studies of SARs identified the most promising compound 28 (c-Met IC50=1.4nM) as a c-Met kinase inhibitor. In this study, a promising compound 28 was identified