Mediator Kinase Inhibitor Selectivity and Activity in Colorectal Cancer.

IF 3.5 2区 生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY
Maria J Ortiz-Ruiz, Olajumoke Popoola, Konstantinos Mitsopoulos, Robert Te-Poele, Rahul S Samant, Gary Box, Will Court, Alexis De Haven Brandon, Sharon Gowan, Aurelie Mallinger, Toby Roe, Kate Swabey, Melanie Valenti, Bissan Al-Lazikani, Julian Blagg, Christina Esdar, Kai Schiemann, Dirk Wienke, Suzanne A Eccles, Paul Workman, Paul A Clarke
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引用次数: 0

Abstract

The Mediator complex is a regulator of gene expression, influencing chromatin structure and RNA polymerase II-mediated transcription. Its activity is controlled by a protein kinase module, which includes cyclin-dependent kinases 8 and 19, that phosphorylates RNA polymerase II and transcription factors to regulate gene expression. Using orthogonal approaches combining chemical and genetic tools, we demonstrated the selectivity of our small-molecule inhibitors derived from 3,4,5-trisubstituted pyridine and 3-methyl-1H-pyrazolo[3,4-b]pyridine chemical series in human colorectal cell culture and tumor xenograft models. The lack of activity of our inhibitors in CDK8/19 double knockout models, with respect to molecular, proliferative, and antitumor end points, revealed their specificity and dependence on these kinases. Using our chemical probes and knockout models, we explored Mediator kinase function in human colorectal cancer cells. Phospho-proteome profiling revealed substrates enriched with transcription and chromatin regulators, while promoter reporter experiments identified transcription factor binding sites, including TCF/LEF and AP1, regulated by Mediator kinases. Additionally, altered phosphorylation of several Mediator subunits suggests a mechanism for the rapid regulation of the Mediator complex. Overall, our results demonstrate that CDK8 and CDK19 play pivotal roles in regulating gene expression associated with oncogene activation and signaling pathways. Further studies are warranted to elucidate their broader cellular roles and regulatory mechanisms. The selective inhibitors validated in this study will provide valuable tools for such mechanistic investigations into Mediator kinase functions and their potential therapeutic exploitation.

结直肠癌中介体激酶抑制剂的选择性和活性。
中介复合物是基因表达的调节因子,影响染色质结构和RNA聚合酶ii介导的转录。其活性受蛋白激酶模块控制,其中包括细胞周期蛋白依赖性激酶8和19,磷酸化RNA聚合酶II和转录因子以调节基因表达。利用化学和遗传工具相结合的正交方法,我们证明了从3,4,5-三取代吡啶和3-甲基- 1h -吡唑啉[3,4-b]吡啶化学系列衍生的小分子抑制剂在人类结直肠癌细胞培养和肿瘤异种移植模型中的选择性。我们的抑制剂在CDK8/19双敲除模型中缺乏活性,在分子、增殖和抗肿瘤终点方面,揭示了它们对这些激酶的特异性和依赖性。利用我们的化学探针和敲除模型,我们探索了介导激酶在人类结直肠癌细胞中的功能。磷酸化蛋白质组分析显示底物富含转录和染色质调节因子,而启动子报告实验发现转录因子结合位点,包括TCF/LEF和AP1,由中介激酶调节。此外,几个中介亚基磷酸化的改变提示了中介复合物快速调节的机制。总之,我们的研究结果表明,CDK8和CDK19在调节与癌基因激活和信号通路相关的基因表达中起着关键作用。需要进一步的研究来阐明它们更广泛的细胞作用和调控机制。本研究中验证的选择性抑制剂将为中介激酶功能的机制研究及其潜在的治疗开发提供有价值的工具。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
ACS Chemical Biology
ACS Chemical Biology 生物-生化与分子生物学
CiteScore
7.50
自引率
5.00%
发文量
353
审稿时长
3.3 months
期刊介绍: ACS Chemical Biology provides an international forum for the rapid communication of research that broadly embraces the interface between chemistry and biology. The journal also serves as a forum to facilitate the communication between biologists and chemists that will translate into new research opportunities and discoveries. Results will be published in which molecular reasoning has been used to probe questions through in vitro investigations, cell biological methods, or organismic studies. We welcome mechanistic studies on proteins, nucleic acids, sugars, lipids, and nonbiological polymers. The journal serves a large scientific community, exploring cellular function from both chemical and biological perspectives. It is understood that submitted work is based upon original results and has not been published previously.
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