A molecular toolbox to accelerate drug development for histone lysine methylation regulators

加速组蛋白赖氨酸甲基化调节剂药物开发的分子工具箱

• 批准号: 10481092
• 负责人: JONATHAN MICHAEL BURG
• 金额: $ 102.51万
• 依托单位: EPICYPHER, INC.
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10481092
• 关键词: ASH1L gene; Address; Affinity; Antibodies; Area; Automobile Driving; Binding; Biological Assay; Biomedical Research; Caymans; Chemicals; Chromatin; Chromatin Structure; Custom; Development; Disease; Drug Design; Engineering; Enzymes; Epigenetic Process; G-Protein-Coupled Receptors; Gene Expression; High Pressure Liquid Chromatography; Histone-Lysine N-Methyltransferase; Histones; KDM5B gene; Length; Letters; Libraries; Lysine; MLL gene; Mass Spectrum Analysis; Methylation; Molecular; Noise; Nucleosomes; Parents; Peptides; Pharmaceutical Chemistry; Pharmacologic Substance; Phase; Phosphotransferases; Physiological; Play; Positioning Attribute; Post-Translational Protein Processing; Process; Protocols documentation; Reagent; Research; Resolution; Role; Series; Services; Signal Transduction; Specificity; Structure; System; Technology; Tertiary Protein Structure; Validation; Work; assay development; base; commercialization; cost; design; drug development; drug discovery; enzyme activity; high throughput screening; improved; inhibitor; innovation; liquid chromatography mass spectrometry; novel; open source; preference; screening; small molecule libraries; targeted cancer therapy; therapeutic development; therapeutic enzyme; therapeutic target; tool; user-friendly

PROJECT SUMMARY Nucleosomes (Nucs) are the repeating unit of chromatin structure and are decorated with diverse post- translational modifications (PTMs) to regulate gene expression. The enzymes that add and remove lysine methylation (KMTs & KDMs) on Nucs play driving roles in many diseases and are important targets for cancer therapy. However, the complexity of chromatin structure has greatly challenged the accurate characterization of these enzymes for drug development. Indeed, many KMTs & KDMs contain multiple domains that engage distinct Nuc features in a multivalent manner, underscoring the need to use full-length enzymes and Nucs to define their activity. Notably, these reagents are difficult / costly to produce, require extensive / customized assay optimization, and are not widely supported by existing high-throughput screening (HTS) platforms, driving the use of protein domains and non-physiological histone peptide-based assays. Further, the field has been stalled by the lack of target-focused chemical compound sets, which are crucial to KMT & KDM inhibitor development, mechanistic analysis, and drug design. Better tools are needed to support this key area of biomedical research. Here, EpiCypher® is directly addressing these problems with the development of EpiVance™, a comprehensive toolbox comprising dNuc substrates, full-length enzymes, validated assays, user-friendly protocols, and a KMT & KDM focused chemical library to advance chromatin research. This innovative, integrated system will enable reliable and sensitive characterization of diverse KMTs & KDMs, which will improve our understanding of these enzymes for therapeutic development. For Phase I proof-of-concept, we developed HTS-compatible enzyme assays for nine KMTs & KDMs using dNuc (or peptide) substrates. We then worked with expert medicinal chemists Drs. Jian Jin and H. Ümit Kaniskan to apply an innovative structure-based optimization strategy, expanding existing KMT & KDM inhibitors into a 200-compound set for HTS. We discovered several compounds with novel target selectivity and found that a defined G9a inhibitor also displayed potent activity towards KDM7A, illustrating the importance of rigorous counter-screening to related enzymes and demonstrating strong feasibility for our approach. In Phase II, we are partnering with Cayman Chemical to exclusively develop and commercialize the EpiVance toolbox and services for drug discovery research. In Aim 1, we will develop a series of HTS assays using full-length KMT & KDM enzymes, dNuc substrates, and highly specific antibodies. In Aim 2, we will work with Drs. Jin / Kaniskan and Cayman Chemical to expand open-source KMT & KDM inhibitors, generating an ~500 compound set. In Aim 3, we will complete end-to-end validation of this system, performing HTS using select enzyme assays from Aim 1 and the chemical set from Aim 2. This project will demonstrate the power of EpiVance for accurate enzyme characterization and inhibitor development, thereby fulfilling a major need in the chromatin field. Our team’s expertise in assay development and medicinal chemistry uniquely positions us to deliver this system, which is expected to have significant market impact.

项目摘要 核小体（NUC）是染色质结构的重复单位 转化修饰（PTM）调节基因表达。添加和删​​除歌词的酶 NUC上的甲基化（KMTS和KDMS）在许多疾病中起驱动作用，是癌症的重要靶标 治疗。但是，染色质结构的复杂性极大地挑战了准确的表征 这些用于药物开发的酶。实际上，许多KMT和KDM都包含多个参与的领域 以多价方式独特的NUC特征，强调需要使用全长酶和NUC到 定义他们的活动。值得注意的是，这些试剂很难产生，需要广泛 /定制的测定 优化，并未得到现有的高通量筛选（HTS）平台的广泛支持，驱动 使用蛋白质结构域和基于Hisstone肽的非生理学结构域的使用。此外，该领域已经停滞不前 由于缺乏针对目标的化合物集，这对于KMT和KDM抑制剂的发展至关重要 机械分析和药物设计。需要更好的工具来支持生物医学研究的关键领域。 在这里，EpicyPher®直接解决了这些问题，而Epivance™的发展， 全面的工具箱综合DNUC底物，全长酶，经过验证的测定，用户友好 方案和KMT＆KDM集中的化学文库以推进染色质研究。这个创新的， 集成系统将实现潜水KMT和KDM的可靠和敏感的特征，这将改善 我们对这些酶进行热发育的理解。对于第一阶段的概念证明，我们开发了 使用DNUC（或胡椒）底物对HTS兼容的酶测定为9 kmts＆kdms。然后我们工作 与专家药物学家Drs一起。 Jian Jin和H.ümitKaniskan应用基于创新的结构 优化策略，将现有的KMT＆KDM抑制剂扩展为HTS的200个集合集。我们 发现了具有新型目标选择性的几种化合物，发现定义的G9A抑制剂也显示 对KDM7A的潜在活动，说明了严格的反筛查对相关酶和 证明我们的方法可行性很强。在第二阶段，我们正在与开曼化学化学合作 专门开发和商业化药物发现研究的Epivance工具箱和服务。目标 1，我们将使用全长KMT＆KDM酶，DNUC底物和高度开发一系列HTS测定法 特定抗体。在AIM 2中，我们将与Drs一起工作。 Jin / Kaniskan和Cayman Chemical扩展开源 KMT＆KDM抑制剂，生成约500个化合物集。在AIM 3中，我们将完成端到端的验证 该系统，使用AIM 1中的精选酶测定和AIM 2的化学组进行HTS进行HTS。 项目将证明尤其是准确酶表征和抑制剂发展的力量， 从而满足染色质领域的主要需求。我们团队在测定开发和医疗方面的专业知识 化学唯一地定位了我们提供该系统，预计该系统将产生重大的市场影响。