Targeting MLL3 histone methyltransferase

 描述(由申请人提供)：越来越多的证据表明，表观遗传途径在癌症的顺序进展中起着不可或缺的作用。研究表明，许多表观遗传酶是癌症患者突变和去调控的靶点。包括DNA甲基转移酶(DNMT)和组蛋白脱乙酰基酶(HDAC)在内的几种关键表观遗传酶的抑制剂已被证明有助于基本理解这些酶的各自活性的功能和在癌症表观遗传治疗中的应用。然而，组蛋白甲基转移酶抑制剂的开发工作仍在进行中 它还处于初级阶段。该项目的总体目标是开发有效和选择性的组蛋白甲基转移酶MLI3抑制剂。此外，我们还将开发一系列简便、灵敏和可重复性的高通量筛选试验，为确定其他组蛋白甲基转移酶(HMT)活性的有效、选择性抑制剂做好准备。至少有六个MLL家族HMT。MLL蛋白利用SAM辅因子将组蛋白H3赖氨酸残基4甲基化，并与其他组蛋白修饰酶协同工作，在染色质改变和基因激活之间提供机制联系；并通过招募各种H3K4‘Reader’蛋白来调节不同的生物学过程。MLL基因突变和调控异常与多种人类疾病相关，包括各种癌症(白血病、淋巴瘤、结直肠癌、胃癌和肺癌)、肥胖和人类智能缺陷。由于MLLS介导的有趣的生物学特性及其与人类疾病的关系，开发新型的小分子抑制剂作为生物探针和药物开发的先导分子是人们感兴趣的。对于这一应用，我们将重点筛选ML3蛋白，尽管该检测方法应该普遍适用于其他MLL家族HMT。该提案的具体目的是(1)开发MLL3抑制剂的高通量筛选和计数器筛选，(2)开发二次测定以量化效力并对高通量筛选中确定的候选命中化合物进行生化表征，以及(3)在细胞分析和全基因组研究中评估化合物的靶标特异性。我们希望这些研究定义生化和基于细胞的分析，以筛选大的小分子文库，以确定有效和选择性的MLI3组蛋白甲基转移酶抑制剂，并获得一些针对全球H3K4甲基化的泛MLL抑制剂。我们预期这些研究将会导致 鉴定有效和选择性的第一代MLL3抑制剂，以供进一步开发作为化学探针。

 DESCRIPTION (provided by applicant): Accumulating evidence suggests that epigenetic pathways play an integral role in the sequential progression of cancer. It is shown that many epigenetic enzymes are targeted for mutation and deregulation in cancer patients. Inhibitors of several key epigenetic enzymes including DNA methyltransferases (DNMTs) and histone deacetylases (HDACs) have proven to be instrumental for both basic understanding of the function of the respective activity of the enzymes and applications in cancer epigenetic therapy. However, efforts in development of inhibitors for histone methyltransferase inhibitors are still at its infancy. The overall goal of this project is to develop potent and selective inhibitors for histone methyltransferase MLL3. Furthermore, we will also develop a series of facile, sensitive and reproducible high-throughput screening assays ready to identify potent, selective inhibitors for other histone methyltransferase (HMT) activity. There are at least six MLL family HMTs. MLL proteins use the SAM cofactor to methylate histone H3 lysine residue 4 and work in concert with other histone modification enzymes to provide a mechanistic link between chromatin alteration and gene activation; and regulate different biological processes through recruitment of various H3K4 'reader' proteins. MLL mutations and dys-regulation have been correlated with several human diseases including various cancers (leukemia, lymphoma, colorectal, gastric and lung), obesity and human intelligent deficiency. Because of the interesting biology that is mediated by MLLs and their involvement to human diseases, it is of interest to develop novel small molecule inhibitors that might serve as biological probes as well as lead molecules for drug development. For this application, we will focus our screening efforts on the MLL3 protein, although the assay should be generally applicable to other MLL family HMTs. The specific aims of the proposal are to (1) Develop a high-throughput screen and counter screen for MLL3 inhibitors, (2) Develop an secondary assay to quantify the potency and to biochemically characterize candidate hit compounds identified in high-throughput screens, and (3) Assess compound on-target specificity in cellular assays and genome-wide studies. We expect these studies to define biochemical and cell based assays to screen large libraries of small molecules to identify potent and selective inhibitors of MLL3 histone methyltransferase, and to obtain some pan-MLL inhibitors that target global H3K4 methylation. We anticipate that these studies will result in the identification of potent and selective first-generation MLL3 inhibitors for further development as chemical probes.