Vital Tamara, Wali Aminah, Butler Kyle V, Xiong Yan, Foster Joseph P, Marcel Shelsa S, McFadden Andrew W, Nguyen Valerie U, Bailey Benton M, Lamb Kelsey N, James Lindsey I, Frye Stephen V, Mosely Amber L, Jin Jian, Pattenden Samantha G, Davis Ian J
Curriculum in Genetics and Molecular Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC, United States.
Department of Genetics, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC, United States.
Front Oncol. 2023 Jan 30;13:1099550. doi: 10.3389/fonc.2023.1099550. eCollection 2023.
Ewing sarcoma is a cancer of children and young adults characterized by the critical translocation-associated fusion oncoprotein EWSR1::FLI1. EWSR1::FLI1 targets characteristic genetic loci where it mediates aberrant chromatin and the establishment of enhancers. Ewing sarcoma thus provides a model to interrogate mechanisms underlying chromatin dysregulation in tumorigenesis. Previously, we developed a high-throughput chromatin-based screening platform based on the enhancers and demonstrated its utility in identifying small molecules capable of altering chromatin accessibility. Here, we report the identification of MS0621, a molecule with previously uncharacterized mechanism of action, as a small molecule modulator of chromatin state at sites of aberrant chromatin accessibility at EWSR1::FLI1-bound loci. MS0621 suppresses cellular proliferation of Ewing sarcoma cell lines by cell cycle arrest. Proteomic studies demonstrate that MS0621 associates with EWSR1::FLI1, RNA binding and splicing proteins, as well as chromatin regulatory proteins. Surprisingly, interactions with chromatin and many RNA-binding proteins, including EWSR1::FLI1 and its known interactors, were RNA-independent. Our findings suggest that MS0621 affects EWSR1::FLI1-mediated chromatin activity by interacting with and altering the activity of RNA splicing machinery and chromatin modulating factors. Genetic modulation of these proteins similarly inhibits proliferation and alters chromatin in Ewing sarcoma cells. The use of an oncogene-associated chromatin signature as a target allows for a direct approach to screen for unrecognized modulators of epigenetic machinery and provides a framework for using chromatin-based assays for future therapeutic discovery efforts.
尤因肉瘤是一种发生于儿童和年轻人的癌症，其特征是存在关键的与易位相关的融合癌蛋白EWSR1::FLI1。EWSR1::FLI1靶向特定的基因位点，在这些位点介导异常染色质和增强子的形成。因此，尤因肉瘤为探究肿瘤发生过程中染色质失调的机制提供了一个模型。此前，我们基于增强子开发了一个基于染色质的高通量筛选平台，并证明了其在鉴定能够改变染色质可及性的小分子方面的效用。在此，我们报告了MS0621的鉴定结果，该分子具有此前未被表征的作用机制，是一种在EWSR1::FLI1结合位点的异常染色质可及性位点调节染色质状态的小分子。MS0621通过细胞周期阻滞抑制尤因肉瘤细胞系的细胞增殖。蛋白质组学研究表明，MS0621与EWSR1::FLI1、RNA结合和剪接蛋白以及染色质调节蛋白相关联。令人惊讶的是，与染色质和许多RNA结合蛋白（包括EWSR1::FLI1及其已知相互作用蛋白）的相互作用不依赖于RNA。我们的研究结果表明，MS0621通过与RNA剪接机制和染色质调节因子相互作用并改变其活性来影响EWSR1::FLI1介导的染色质活性。对这些蛋白质进行基因调控同样会抑制尤因肉瘤细胞的增殖并改变染色质。使用与癌基因相关的染色质特征作为靶点，能够直接筛选未被识别的表观遗传机制调节剂，并为未来利用基于染色质的检测方法进行治疗发现工作提供了一个框架。
