Small molecule screen reveals synergy of cell cycle checkpoint kinase inhibitors with DNA-damaging chemotherapies in medulloblastoma (RNAseq dataset)
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PROVIDER: EGAS00001004748 | EGA |
REPOSITORIES: EGA
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Project description:Medulloblastoma (MB) consists of four core molecular subgroups with distinct clinical features and prognoses. Treatment consists of surgery, followed by radiotherapy and cytotoxic chemotherapy. Despite this intensive approach, outcome remains dismal for patients with certain subtypes of MB, namely, MYC-amplified Group 3 and TP53-mutated SHH. Using high-throughput assays, six human MB cell lines were screened against a library of 3208 unique compounds. We identified 45 effective compounds from the screen and found that cell cycle checkpoint kinase (CHK1/2) inhibition synergistically enhanced the cytotoxic activity of clinically used chemotherapeutics cyclophosphamide, cisplatin, and gemcitabine. To identify the best-in-class inhibitor, multiple CHK1/2 inhibitors were assessed in mice bearing intracranial MB. When combined with DNA-damaging chemotherapeutics, CHK1/2 inhibition reduced tumor burden and increased survival of animals with high-risk MB, across multiple different models. In total, we tested 14 different models, representing distinct MB subgroups, and data were validated in three independent laboratories. Pharmacodynamics studies confirmed central nervous system penetration. In mice, combination treatment significantly increased DNA damage and apoptosis compared to chemotherapy alone, and studies with cultured cells showed that CHK inhibition disrupted chemotherapy-induced cell cycle arrest. Our findings indicated CHK1/2 inhibition, specifically with LY2606368 (prexasertib), has strong chemosensitizing activity in MB that warrants further clinical investigation. Moreover, these data demonstrated that we developed a robust and collaborative preclinical assessment platform that can be used to identify potentially effective new therapies for clinical evaluation for pediatric MB.
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Project description:The members of the Aurora kinase family play critical roles in the regulation of the cell cycle and mitotic spindle assembly and have been intensively investigated as potential targets for a new class of anticancer drugs. We describe a new highly potent and selective class of Aurora kinase inhibitors discovered using a phenotypic cellular screen. Optimized inhibitors display many of the hallmarks of Aurora inhibition including endoreduplication, polyploidy, and loss of cell viability in cancer cells. Structure-activity relationships with respect to kinome-wide selectivity and guided by an Aurora B co-crystal structure resulted in the identification of key selectivity determinants and discovery of a subseries with selectivity toward Aurora A. A direct comparison of biochemical and cellular profiles with respect to published Aurora inhibitors including VX-680, AZD1152, MLN8054, and a pyrimidine-based compound from Genentech demonstrates that compounds 1 and 3 will become valuable additional pharmacological probes of Aurora-dependent functions.
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Project description:Mps1, a dual-specificity kinase, is required for the proper functioning of the spindle assembly checkpoint and for the maintenance of chromosomal stability. As Mps1 function has been implicated in numerous phases of the cell cycle, the development of a potent, selective small-molecule inhibitor of Mps1 should facilitate dissection of Mps1-related biology. We describe the cellular effects and Mps1 cocrystal structures of new, selective small-molecule inhibitors of Mps1. Consistent with RNAi studies, chemical inhibition of Mps1 leads to defects in Mad1 and Mad2 establishment at unattached kinetochores, decreased Aurora B kinase activity, premature mitotic exit and gross aneuploidy, without any evidence of centrosome duplication defects. However, in U2OS cells having extra centrosomes (an abnormality found in some cancers), Mps1 inhibition increases the frequency of multipolar mitoses. Lastly, Mps1 inhibitor treatment resulted in a decrease in cancer cell viability.
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Project description:Cell cycle checkpoint kinases serve as important therapeutic targets for various cancers. When they are inhibited by small molecules, checkpoint abrogation can induce cell death or further sensitize cancer cells to other genotoxic therapies. Particularly aberrant Cdk1 activation at the G2/M checkpoint by kinase inhibitors causing unscheduled mitotic entry and mitotic arrest was found to lead to DNA damage and cell death selectively in cancer cells. Promising drugs inhibiting kinases like Wee1 (Adavosertib), Wee1+Myt1 (PD166285), ATR (AZD6738) and Chk1 (UCN-01) have been developed, but clinical data has shown variable efficacy for them with poorly understood mechanisms of resistance. Our lab recently identified Myt1 as a predictive biomarker of acquired resistance to the Wee1 kinase inhibitor, Adavosertib. Here, we investigate the role of Myt1 overexpression in promoting resistance to inhibitors (PD166285, UCN-01 and AZD6738) of other kinases regulating cell cycle progression. We demonstrate that Myt1 confers resistance by compensating Cdk1 inhibition in the presence of these different kinase inhibitors. Myt1 overexpression leads to reduced premature mitotic entry and decreased length of mitosis eventually leading to increased survival rates in Adavosertib treated cells. Elevated Myt1 levels also conferred resistance to inhibitors of ATR or Chk1 inhibitor. Our data supports that Myt1 overexpression is a common mechanism by which cancer cells can acquire resistance to a variety of drugs entering the clinic that aim to induce mitotic catastrophe by abrogating the G2/M checkpoint.
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Project description:Here we performed a targeted small molecule screen on a stable, SHH-dependent murine MB cell line (SMB21). A subset of the HDAC inhibitors tested significantly inhibit tumor growth of SMB21 cells by preventing SHH pathway activation. Of note, class I HDAC inhibitors were also efficacious in suppressing growth of diverse SMO inhibitor-resistant clones of SMB21 cells. Finally, we show that the novel HDAC inhibitor Quisinostat (JNJ) targets multiple class I HDACs, is well tolerated in mouse models and robustly inhibits growth of SHH MB cells in vivo as well as in vitro. Our data provide strong evidence that Quisinostat (JNJ) or other class I HDAC inhibitors might be therapeutically useful for patients with SHH MB including those resistant to SMO inhibition.
2019-04-10 | GSE129512 | GEO
Project description:Mutations in chromatin modifying proteins are associated with a variety of cancers. These mutations may result in characteristic patterns of accessible chromatin indicative of altered nucleosome positioning and depletion at regulatory elements. We describe the adaptation of Formaldehyde-Assisted Isolation of Regulatory Elements (FAIRE), a chemical method to enrich for nucleosome-depleted genomic regions, as a high-throughput, automated assay. We applied this method in a screen of epigenetically targeted small molecules by evaluating regions of aberrant nucleosome depletion mediated by EWS-FLI1, the chimeric transcription factor critical for the bone and soft tissue tumor Ewing Sarcoma. We identified a novel compound, UNC0621, which exhibited a dose-dependent reduction in chromatin accessibility, decreased Ewing Sarcoma cell viability, and inhibited anchorage-independent growth. We demonstrate that chromatin accessibility can be exploited for drug discovery efforts offering the advantage that it does not depend on the a priori selection of a single molecular target. Identified compounds can then be used for biological target discovery and potential therapeutic validation. HT-FAIRE was performed on Human Umbilical Vein Endothelial Cells (HUVEC) in duplicate. No reference/control experiment is necessary.
2015-09-24 | E-GEOD-61735 | biostudies-arrayexpress
Project description:BackgroundThe cell-cycle checkpoint kinase Chk1 is essential in mammalian cells due to its roles in controlling processes such as DNA replication, mitosis and DNA-damage responses. Despite its paramount importance, how Chk1 controls these functions remains unclear, mainly because very few Chk1 substrates have hitherto been identified.ResultsHere, we combine a chemical genetics approach with high-resolution mass spectrometry to identify novel Chk1 substrates and their phosphorylation sites. The list of targets produced reveals the potential impact of Chk1 function not only on processes where Chk1 was already known to be involved, but also on other key cellular events such as transcription, RNA splicing and cell fate determination. In addition, we validate and explore the phosphorylation of transcriptional co-repressor KAP1 Ser473 as a novel DNA-damage-induced Chk1 site.ConclusionsBy providing a substantial set of potential Chk1 substrates, we present opportunities for studying unanticipated functions for Chk1 in controlling a wide range of cellular processes. We also refine the Chk1 consensus sequence, facilitating the future prediction of Chk1 target sites. In addition, our identification of KAP1 Ser473 phosphorylation as a robust readout for Chk1 activity could be used to explore the in vivo effects of Chk1 inhibitors that are being developed for clinical evaluation.
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Project description:Small molecule kinase inhibitors (SMKIs) are being approved at a fast pace under expedited programs for anticancer treatment. In this study, we construct a multi-domain dataset from a total of 4638 patients in the registrational trials of 16 FDA-approved SMKIs and employ a machine-learning model to examine the relationships between kinase targets and adverse events (AEs). Internal and external (datasets from two independent SMKIs) validations have been conducted to verify the usefulness of the established model. We systematically evaluate the potential associations between 442 kinases with 2145 AEs and made publicly accessible an interactive web application "Identification of Kinase-Specific Signal" ( https://gongj.shinyapps.io/ml4ki ). The developed model (1) provides a platform for experimentalists to identify and verify undiscovered KI-AE pairs, (2) serves as a precision-medicine tool to mitigate individual patient safety risks by forecasting clinical safety signals and (3) can function as a modern drug development tool to screen and compare SMKI target therapies from the safety perspective.
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Project description:The inhibition of the DNA damage response (DDR) pathway in the treatment of cancers has recently reached an exciting stage with several cell cycle checkpoint inhibitors that are now being tested in several clinical trials in cancer patients. Although the great amount of pre-clinical and clinical data are from the solid tumor experience, only few studies have been done on leukemias using specific cell cycle checkpoint inhibitors. This review aims to summarize the most recent data found on the biological mechanisms of the response to DNA damages highlighting the role of the different elements of the DDR pathway in normal and cancer cells and focusing on the main genetic alteration or aberrant gene expression that has been found on acute and chronic leukemias. This review, for the first time, outlines the most important pre-clinical and clinical data available on the efficacy of cell cycle checkpoint inhibitors in single agent and in combination with different agents normally used for the treatment of acute and chronic leukemias.
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