Specific Chemical Probes for Histone Demethylases

描述（由申请人提供）：共价组蛋白修饰被认为是最重要的表观遗传现象之一。组蛋白去甲基化酶是最近发现的一类组蛋白修饰酶。到目前为止，已经鉴定了十几种含有Jumonji C结构域的组蛋白去甲基化酶（JHDMs）。它们可以在所有甲基化状态下修饰组蛋白上许多不同位置的赖氨酸残基。它们通常表现出组织特异性表达，并在多种细胞过程中发挥关键作用，如 如基因表达、减数分裂和胚胎干细胞自我更新。此外，几种JHDMs如JMJD 2C与人类疾病有关，如精神发育迟滞和癌症。然而，JHDMs的细胞功能和底物范围仍不清楚。目前，研究JHDM及其细胞功能的研究工具非常有限。目前尚不存在针对JHDM的高活性和特异性化学探针。各种JHDM亚型的统一生化测定也不发达。 在这项提案中，我们计划开发一系列针对JHDMs的特异性化学探针，包括一种高活性和特异性的细胞探针，一种荧光小分子探针，以及一种特异性靶向H3 K9-JHDMs的高度功能化的肽探针。该细胞探针可用于广泛的生物学过程和疾病过程中JHDMs的细胞功能和组蛋白甲基化动力学的研究。荧光探针被应用于开发基于荧光偏振的测定，这不仅使我们能够验证我们的二价抑制剂设计，而且还提供了一个统一的JHDM生化测定来分析JHDM抑制剂的亚型特异性。肽探针不仅对研究JHDMs的底物范围很重要，而且对组蛋白串扰提供有价值的信息。此外，我们在这里开发的策略可以应用于发现修饰H3 K79 me 3和H4 K20 me 3的新型JHDM，最终导致组蛋白甲基化及其相应的JHDM的系统图。此外，这些探针还可用于验证JHDM作为疾病（如癌症）的潜在新型治疗靶点，并可用作进一步开发这些疾病的新型治疗剂的先导化合物。

DESCRIPTION (provided by applicant): Covalent histone modifications are considered one of the most important epigenetic phenomena. Histone demethylases are the most recently discovered class of histone-modifying enzymes. Over a dozen Jumonji C domain-containing histone demethylases (JHDMs) have been characterized so far. They can modify lysine residues at many different positions on histone proteins at all methylation states. They often display tissue-specific expression and play critical roles in a variety of cellular processes, such as gene expression, meiosis, and embryonic stem cell self-renewal. In addition, several JHDMs such as JMJD2C have been associated with human diseases, such as mental retardation and cancers. However, the cellular functions and the substrate scope of JHDMs are still unclear. Currently, research tools to study JHDMs and their cellular functions are very limited. Highly active and specific chemical probes for JHDMs currently do not exist. A uniform biochemical assay for various JHDM isoforms is also underdeveloped. In this proposal, we plan to develop a series of specific chemical probes for JHDMs, including a highly active and specific cellular probe, a fluorescent small-molecule probe, and a highly functionalized peptidic probe specifically targeting H3K9-JHDMs. The cellular probe can be used for the studies of the cellular functions of JHDMs and histone methylation dynamics in a wide range of biological processes and disease processes. The fluorescent probe is being applied to develop a fluorescence polarization-based assay, which will not only enable us to validate our bivalent inhibitor design, but also provide a uniform JHDM biochemical assay to analyze isoform specificity of JHDM inhibitors. The peptidic probe is important not only to study the substrate scope of JHDMs, but also to provide valuable information on the histone crosstalk. In addition, the strategy we develop here can be applied to the discovery of novel JHDMs that modify H3K79me3 and H4K20me3, eventually leading to a systematic map of histone methylations and their corresponding JHDMs. Furthermore, these probes can also be used to validate JHDMs as potential novel therapeutic targets for diseases such as cancers, and can serve as lead compounds for further development of novel therapeutics for these diseases.