Identification of Inhibitors of the Tip60 Histone Acetyltransferase

描述（由申请方提供）：组蛋白乙酰转移酶对赖氨酸残基的乙酰化调节蛋白质功能。细胞含有多个不同的组蛋白乙酰转移酶家族，包括Tip 60。Tip 60与几种人类疾病过程的发病机制有关，包括癌症进展、神经退行性疾病和病毒复制。Tip 60在这些不同疾病过程中的参与反映了Tip 60调节转录、病毒复制和细胞对DNA损伤的反应的能力。Tip 60通过其乙酰化和激活ATM蛋白激酶的能力来调节细胞对遗传毒性应激的反应。ATM功能的丧失与癌症发病率的增加、对电离辐射的敏感性和小脑神经变性有关。因此，Tip 60是细胞修复和存活DNA损伤能力的关键调节因子。 放射治疗是人类癌症的主要治疗方式。然而，许多肿瘤类型对放射治疗的细胞毒性作用具有相对抗性。因此，Tip 60的选择性药理学抑制提供了用于增强放疗和化疗在治疗人肿瘤中的功效的潜在的强有力的治疗靶标。目前，没有选择性抑制剂的组蛋白乙酰转移酶一般，特别是Tip 60，已被确定。因此，在这个建议中，我们建议开发一种高通量的检测，以确定新的，特异性的组蛋白乙酰转移酶的Tip 60抑制剂。将实现两个具体目标。在具体目标1中，将开发一种新的高通量ELISA测定法，其利用重组Tip 60乙酰化固定化肽的能力。在具体目标2中，将测定格式化为384孔格式，并进行活性文库化合物的初步筛选。 Tip 60代表了一个靶点，对于该靶点，存在一系列不充分的选择性和有效的小分子抑制剂;筛选和鉴定此类抑制剂将对研究Tip 60和相关HAT在人类疾病进展中的作用产生重大影响。该提案将开发Tip 60 HAT的特异性抑制剂，其可用于破坏细胞蛋白乙酰化水平，目的是改善人类肿瘤的治疗。这些抑制剂可用于动物和临床研究，以测试Tip 60 激活使人类肿瘤对放射治疗和化疗敏感。此外，Tip 60抑制剂可用于探测Tip 60发挥其生物学效应的基本信号传导机制，并鉴定新的Tip 60底物。最后，Tip 60抑制剂可用于复杂的生物体，如小鼠，以检查Tip 60对疾病过程的贡献，如癌症进展，神经退行性疾病和遗传毒性应激。

DESCRIPTION (provided by applicant): The acetylation of lysine residues by histone acetyltransferases regulates protein function. Cells contain multiple, distinct families of histone acetyltransferases, including Tip60. Tip60 is implicated in the pathogenesis of several human disease processes, including cancer progression, neurodegenerative diseases and viral replication. The participation of Tip60 in these diverse disease processes reflects Tip60's ability to regulate transcription, viral replication and the cells response to DNA damage. Tip60 regulates the cells response to genotoxic stress through its ability to acetylate and activate the ATM protein kinase. Loss of ATM function is associated with increased incidence of cancer, sensitivity to ionizing radiation and cerebellar neurodegeneration. Tip60 is therefore a key regulator of the cells ability to repair and survive DNA damage. Radiation therapy is a major treatment modality for human cancer. However, many tumor types are relatively resistant to the cytotoxic effects of radiation therapy. Accordingly, selective pharmacological inhibition of Tip60 provides a potentially powerful therapeutic target for enhancing the efficacy of radiotherapy and chemotherapy in the treatment of human tumors. Currently, no selective inhibitors of histone acetyltransferases in general, and particularly of Tip60, have been identified. Thus, in this proposal we propose to develop a high throughput assay to identify novel, specific inhibitors of the Tip60 histone acetyltransferases. 2 specific aims will be undertaken. In specific aim 1, a novel high throughput ELISA assay will be developed which utilizes the ability of recombinant Tip60 to acetylate immobilized peptides. In specific aim 2, the assay will be formatted to a 384-well format, and preliminary screening of active library compounds will be carried out. Tip60 represents a target for which an inadequate array of selective and potent small molecule inhibitors exist; screening and identifying such inhibitors would have a significant impact in studying the role of Tip60 and related HATs in the progression of human disease. This proposal will develop specific inhibitors of the Tip60 HAT which can be employed to disrupt cellular protein acetylation levels with the aim of improving treatment of human tumors. These inhibitors can be used in animal and clinical studies to test if Tip60 In activation sensitizes human tumors to radiation therapy and chemotherapy. In addition, Tip60 inhibitors can be employed to probe the basic signaling mechanisms by which Tip60 exerts its biological effects, and to identify new Tip60 substrates. Finally, Tip60 inhibitors can be used in complex organisms, such as mice, to examine the contribution of Tip60 to disease processes such as cancer progression, neurodegenerative diseases and genotoxic stress.