Discovering and Exploiting Selectivity within Tandem Bromodomains

发现和利用串联布罗莫结构域内的选择性

• 批准号: 10241303
• 负责人: Brian Christopher Smith
• 金额: $ 23.1万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10241303
• 关键词: Acetylation; Achievement; Acyl Coenzyme A; Acylation; Address; Binding; Binding Proteins; Biology; Biophysics; Bromodomain; Cell model; Chemicals; Chromatin; Coronary Arteriosclerosis; Cysteine; DNA Sequence; Data; Diabetes Mellitus; Disease model; Distal; Elements; Enhancers; Epigenetic Process; Family; Genetic; Genetic Transcription; Genomic Segment; Goals; Histones; Individual; Knowledge; Ligands; Link; Location; Lymphoid; Lysine; Malignant Neoplasms; Memory Loss; Metabolic; Metabolism; Modification; Molecular Conformation; Non-Insulin-Dependent Diabetes Mellitus; Nucleosomes; Oncogenic; Pharmaceutical Preparations; Phase I/II Clinical Trial; Post-Translational Protein Processing; Proteins; Proteomics; Publications; Research; Rodent Model; Role; Specificity; Structure; Techniques; Tertiary Protein Structure; Toxic effect; Writing; base; biophysical techniques; combinatorial; human disease; inhibitor/antagonist; innovation; member; nanomolar; novel; phase III trial; promoter; scaffold; side effect; small molecule; structural biology; therapeutic target; tool; transcription factor

PROJECT SUMMARY Members of the bromodomain and extra-terminal domain (BET) family (Brd2, Brd3, Brd4, Brdt) each contain two bromodomains that bind acetyl-lysines on histones and transcription factors. The importance of BET- regulated transcription in human disease is well appreciated with pan-BET bromodomain inhibitors in phase I/II clinical trials for multiple cancers and phase III trials for type 2 diabetes subjects with coronary artery disease. Despite these achievements, several critical questions remain. For example, BET proteins are localized disproportionately at super-enhancers, genomic regions with large clusters of elements that enhance gene transcription. The basis of this localization is unknown but important given that super-enhancers are enriched at loci with oncogenic potential. Our unpublished data support the hypothesis that tandem bromodomains act as a scaffold for acetylation-dependent reorganization of chromatin; for instance, joining promotors with their corresponding distal enhancers to drive transcription (Focus 1). However, the ability of tandem bromodomains to scaffold nucleosomes and transcription factors in an acetylation-dependent manner has not been shown. We take an innovative structural and biophysical approach to investigate the role of Brd4 in maintaining chromatin conformations that facilitate enhancer-driven oncogenic gene transcription. This mechanism of chromatin reorganization, if true, is paradigm shifting and would have broad impact on studies of tandem histone-binding domains. We also hypothesize that metabolic changes induce distinct post-translational modifications on histones that are “read” by bromodomains. Yet, the broader acylation and protein binding specificity of bromodomains is poorly understood. We have begun to address this knowledge gap in our recent publication that highlights how metabolically-derived acylations and neighboring modifications tune BET bromodomain binding to histones. To continue to address this broad metabolic question, we are using biophysical, structural biology, and proteomic techniques to investigate BET bromodomain acylation and protein selectivity in linking acyl-CoA metabolism with transcription (Focus 2). To aid our mechanistic inquiries, we are removing a critical barrier in the study of BET bromodomain biology: the lack of inhibitors and chemical probes that selectively target individual BET proteins. Currently, all existing BET inhibitors target Brd2, Brd3, Brd4, and Brdt with equal nanomolar potency. This lack of selectivity may be responsible for the side effects of memory loss and lymphoid toxicity recently associated with existing pan-BET inhibitors. We are overcoming these barriers with a novel fragment-based ligand discovery and chemical biology strategy to discover selective Brd4 inhibitors by covalently targeting a unique cysteine within Brd4 (Focus 3). These chemical tools will be necessary to distinguish the differential activities of BET proteins in cell and rodent models of disease and may also be useful in developing therapeutics targeting the Brd4 axis in cancer and diabetes.

项目摘要 溴d和末端域（BET）家族（BRD2，BRD3，BRD4，BRDT）的成员都包含 两个结合乙酰赖氨酸的溴化植物对组蛋白和转录因子的结合。赌注的重要性 在I/II期间，泛面条溴ab抑制剂对人类疾病的调节转录得到了很好的认识 多种癌症和III期试验的2型糖尿病患者的临床试验。 尽管取得了这些成就，但仍然存在一些关键问题。例如，BET蛋白是局部的 在超级增强剂的基因组区域中，具有大量元素的基因组区域增强基因 转录。这种本地化的基础尚不清楚，但考虑到超级增强剂很丰富 在基因座具有致癌潜力。我们未发表的数据支持串联溴构成法的假设 作为乙酰化依赖性染色质重组的支架；例如，加入发起人与他们的 相应的远端增强子驱动转录（焦点1）。但是，串联溴化组的能力 尚未显示以乙酰化依赖性方式进行脚手架核体和转录因子。 我们采用一种创新的结构和生物物理方法来研究BRD4在维持的作用 染色质构象，促进增强子驱动的致癌基因转录。这种机制 染色质的重组（如果为true）是范式转移，将对串联的研究产生广泛的影响 Hisstone结合域。我们还假设代谢变化会影响不同的翻译后不同 对溴结构域“读取”组蛋白的修改。然而，较宽的酰化和蛋白质结合 溴结构域的特异性知之甚少。我们已经开始在我们的最新知识中解决这个知识差距 出版物，强调了代谢衍生的酰基化和相邻修改如何调节 溴结构域与组蛋白结合。为了继续解决这个广泛的代谢问题，我们正在使用 生物物理，结构生物学和蛋白质组学技术，以研究β溴构域酰化和 将酰基-COA代谢与转录联系起来的蛋白质选择性（焦点2）。为了帮助我们的机械查询， 我们正在删除BET BET BOTOMODOMAIN生物学研究中的关键障碍：缺乏抑制剂和化学物质 选择性针对单个BET蛋白的问题。目前，所有现有的BET抑制剂目标BRD2，BRD3， BRD4和BRDT具有相等的纳摩尔效力。缺乏选择性可能是导致 最近与现有的泛膏抑制剂相关的记忆丧失和淋巴毒性。我们正在克服 这些障碍具有新颖的基于碎片的配体发现和化学生物学策略，以发现 选择性BRD4抑制剂通过共价靶向BRD4内的独特半胱氨酸（焦点3）。这些化学工具 有必要区分细胞和啮齿动物模型中BET蛋白的差异活性 并且在开发靶向癌症和糖尿病的BRD4轴的治疗中也可能很有用。