A new epigenetic toolbox for inflammation research and drug discovery

用于炎症研究和药物发现的新表观遗传学工具箱

• 批准号: 10401943
• 负责人: Michael-Christopher Keogh
• 金额: $ 101.23万
• 依托单位: EPICYPHER, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-05 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10401943
• 关键词: Acetylation; Acylation; Adoption; Animal Model; Antibodies; Antibody Specificity; Area; Arginine; Binding; Biochemical; Biological Assay; Biology; Bone Marrow; Cells; ChIP-seq; Chemicals; Chromatin; Collection; Data; Dendritic Cells; Development; Disease; Employment; Engineering; Enzymes; Epigenetic Process; Feedback; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Genetic study; Genomics; Goals; High Pressure Liquid Chromatography; Histone H3; Histones; Human; Immune; Immunologics; Immunology; Inflammation; Inflammatory; Inflammatory Response; Link; Lipopolysaccharides; Lysine; Mammals; Mass Spectrum Analysis; Methods; Methylation; Mus; Natural Killer Cells; Nucleosomes; Orphan; Pharmaceutical Preparations; Phase; Phosphorylation; Post-Translational Protein Processing; Property; Reagent; Recombinants; Research; Research Personnel; Rest; Role; Running; Serine; Side; Site-Directed Mutagenesis; Technology; Testing; Validation; Variant; Work; biomarker discovery; commercialization; cross reactivity; dalton; drug development; drug discovery; epigenomics; human disease; immune system function; inhibitor; innovation; macrophage; medical schools; novel; research and development; response; screening; therapeutic development; tool; tool development

PROJECT SUMMARY Gene regulation is controlled in part by histone post-translational modifications (PTMs) on nucleosomes. EpiCypher® is developing fully defined recombinant designer nucleosomes (dNucs) carrying specific histone PTMs to enable epigenetics research and drug development. The power of EpiCypher’s dNuc platform comes from its broad chemical diversity. EpiCypher has commercialized > 100 unique dNucs, covering the most widely studied PTM classes (e.g. lysine methylation / acylation / ubiquitylation, arginine methylation, serine phosphorylation, etc.) and is leveraging the emergent property of this diversity for a range of high value applications: 1) antibody specificity testing (NucleoPlex® antibody validation: e.g. chromatinantibodies.com); 2) ultra-sensitive genomic mapping (CUTANA® CUT&RUN / CUT&Tag assays); and 3) high-throughput biochemical approaches for drug discovery and inhibitor screening (dCypher® assays). To date, EpiCypher’s dNuc technology (and related assay platforms) have been focused on PTMs with known associations with chromatin states and gene regulation. Relying on the field’s largely descriptive histone PTM studies as a guide is an inefficient way for us to expand our discovery platforms and maximize the potential of our nucleosome generating capability to target the most functionally important PTMs. Progress on the discovery side has been hindered by intractability of the multi-copy histone genes for functional genetics studies in mammals (vs. simpler model organisms). Here, EpiCypher is partnering with Dr. Steven Josefowicz (Weill Cornell Medical School) to expand epigenetic tool development for immunology research and biomarker discovery. The innovation of this project is employment of a first-in-class mammalian histone variant H3.3 genetic replacement method to identify orphaned / underappreciated residues (and PTMs) with roles in macrophage stimulation. We will then develop new dNucs containing these PTMs and validate their role in macrophage function. For proof of concept, we developed the histone replacement assay to characterize the role of some highly studied (e.g. H3.3K4, H3.3K36) and underappreciated (H3.3S31) residues in the macrophage stimulation response, and showed the resulting data can immediately be used to guide the delivery of new epigenetic reagents and assays to support the study of immune system function and disease. In Phase II, we will leverage this development pipeline to identify novel resides vital for macrophage stimulation (Aim 1). Next, we will develop a collection of dNucs carrying PTMs on these resides, which will be used in NucleoPlex assays to identify best-in-class antibodies to each target (Aim 2). Finally, we will validate the function of these novel PTMs in immune cell stimulation using CUT&Tag assays as well as share our expanded reagents and capabilities with key opinion leaders for external validation (Aim 3) Together, this work will result in the commercialization of an expanded epigenetics toolbox that will open new avenues of immunological research and drug development.

项目摘要 基因调控部分由核小体上的组蛋白翻译后修饰（PTM）控制。 EpiCypher®正在开发携带特定组蛋白的完全定义的重组设计核小体（dNucs） PTM使表观遗传学研究和药物开发成为可能。EpiCypher dNuc平台的强大功能来自于 从其广泛的化学多样性。EpiCypher已经商业化了超过100种独特的dNucs，覆盖了最广泛的 研究的PTM类别（例如赖氨酸甲基化/酰化/泛素化、精氨酸甲基化、丝氨酸甲基化、丝氨酸甲基化、精氨酸甲基化、精氨酸甲基化、丝氨酸甲基化、精氨酸甲基化、丝氨酸甲基化、精氨酸甲基化、丝氨酸甲基化、精氨酸甲基化、丝氨酸甲基 磷酸化等）并利用这种多样性的新兴特性来实现一系列高价值 应用：1）抗体特异性测试（NucleoTM ®抗体验证：例如chromatinantibodies.com）; 2） 超灵敏的基因组作图（CUTANA® CUT&RUN / CUT&Tag测定）;以及3）高通量 用于药物发现和抑制剂筛选的生物化学方法（dCypher®测定）。到目前为止，EpiCypher dNuc技术（和相关的分析平台）一直专注于已知与以下相关的PTM： 染色质状态和基因调控。依靠该领域的主要描述性组蛋白PTM研究作为指导 是一种低效的方式来扩展我们的发现平台并最大限度地发挥我们核小体的潜力 生成针对功能上最重要的PTM的能力。发现方面的进展 阻碍了多拷贝组蛋白基因在哺乳动物功能遗传学研究中的棘手性（与简单的 模式生物）。在这里，EpiCypher与Steven Josefowicz博士（Weill Cornell Medical 学校）扩大免疫学研究和生物标志物发现的表观遗传工具开发。的 该项目的创新之处在于首次采用了哺乳动物组蛋白变体H3.3遗传替代 鉴定在巨噬细胞刺激中具有作用的孤儿/低估残基（和PTM）的方法。我们 然后将开发含有这些PTM的新dNucs，并验证它们在巨噬细胞功能中的作用。为了证明 概念，我们开发了组蛋白替代测定来表征一些高度研究的作用（例如， H3.3K4、H3.3K36）和低估（H3.3S31）残基，和 结果表明，所得到的数据可以立即用于指导新的表观遗传试剂和测定的交付 以支持免疫系统功能和疾病的研究。在第二阶段，我们将利用这一发展 管道，以确定新的驻留至关重要的巨噬细胞刺激（目标1）。接下来，我们将开发一个集合， 在这些残基上携带PTM的dNucs，其将用于Nucleotide测定，以鉴定同类最佳 每个目标的抗体（目标2）。最后，我们将验证这些新的PTM在免疫细胞中的功能 使用CUT&Tag检测刺激，并分享我们扩展的试剂和功能， 外部验证的领导者（目标3）总之，这项工作将导致商业化的 扩展的表观遗传学工具箱，将开辟免疫学研究和药物治疗的新途径 发展