Photoaffinity labeling probes for development of novel isoform selective HDAC inh

用于开发新型亚型选择性 HDAC inh 的光亲和标记探针

• 批准号: 7648175
• 负责人: Pavel A Petukhov
• 金额: $ 31.78万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7648175
• 关键词: Abbreviations; Address; Affect; Azides; Binding; Biological Testing; Biology; Biotin; Cell Death; Chemicals; Clinical Trials; Complex; Computer Assisted; Development; Disease; Docking; Drug Design; Enzymes; FDA approved; Fingerprint; Gene Expression Regulation; Goals; HDAC7 histone deacetylase; Hela Cells; Histone Deacetylase; Histone Deacetylase Inhibitor; Homology Modeling; Image; Individual; Label; Life; Ligand Binding; Ligands; Malignant Neoplasms; Maps; Mass Spectrum Analysis; Methods; Modification; Nature; Pharmaceutical Chemistry; Photoaffinity Labels; Problem Solving; Protein Isoforms; Proteins; Proteomics; Recombinants; Research; Resolution; Series; Site; Structure; Study models; Surface; Testing; Therapeutic; Toxic effect; Transcription factor genes; Treatment Efficacy; Zinc; anti-cancer therapeutic; base; cancer therapy; cell growth; cytotoxicity; design; drug discovery; histone acetyltransferase; human HDAC1 protein; human HDAC11 protein; improved; inhibitor/antagonist; innovation; knowledge of results; molecular dynamics; multidisciplinary; neoplastic cell; novel; novel strategies; public health relevance; research study; small molecule; success; three dimensional structure

DESCRIPTION (provided by applicant): This proposal is intended to charter new directions for design of more selective and more active histone deacetylase (HDAC) inhibitors by mapping the binding "fingerprints" of the HDAC ligands in the different HDAC isoforms using small molecule photoaffinity probes. Since many cancers are associated with aberrant transcriptional activity, and the HDACs can affect transcription factors and gene regulation, these enzymes have been identified as attractive targets for cancer therapy. Indeed, chemical inhibitors of HDACs have been shown to inhibit tumor cell growth and induce differentiation and cell death. At least one non-selective HDAC inhibitor was approved by FDA for cancer and several non-selective HDAC inhibitors are in clinical trials. Multiple studies have shown that therapeutics selective for one or a limited number of specific HDAC isoforms may improve overall efficacy and lower toxicity of these compounds. While some rather limited degree of isoform selectivity has been shown by a few compounds, the problem of identifying selective inhibitors is far from solved. Recent success in the elucidation of the crystal structure of HDAC7, 8 and homologous HDAC proteins has opened a possibility for structure-based drug design. Despite the progress, both ligand-based and structure-based approaches are limited in their applicability to design of isoform selective HDAC inhibitors either because only few isoform selective inhibitors are available as a starting point for ligand based drug design or because high resolution three-dimensional structures are available only for 2 out of 11 class I and II HDAC isoforms. It is also poorly understood how the binding modes of those portions of the HDAC ligands that bind on the surface of the HDAC protein affect the activity of HDAC inhibitors. Clearly, a new approach to this problem is needed. We hypothesize that it will be possible to map the binding modes available to the ligands in different HDAC isoforms by small molecule photoaffinity probes (PAP) and use the resulting knowledge to design isoform selective HDAC inhibitors as potential therapeutics for cancer. Our specific aims in this proposal are as follow: Aim 1: Design and synthesize at least three series of photoaffinity probes (PAPs) consisting of (1) an HDAC ligand, (2) an aromatic azide group for photoaffinity labeling (PAL) of the protein residues directly involved in the binding of the ligand, and (3) an aliphatic azide (a imaging tag attachment group - imTAG) for attaching a fluorescent or a biotin tag. Aim 2: Evaluate the ligands containing either PAL or PAL+imTAG or both groups in photoaffinity labeling experiments with available recombinant purified class I and class II HDACs and determine the modification sites on the proteins using mass-spectrometry proteomics methods. Match the proteomics results with the modeling studies and refine later to better reproduce the experimental findings. Aim 3. Evaluate the cytotoxicity, selectivity, and binding "fingerprints" of the most potent photoaffinity probes in live Hela cells. Aim 4. Incorporate the findings of Aims 1-3 in the structure-based drug design and perform several rounds of CADD, medicinal chemistry, and biological tests to improve the activity and selectivity of the HDAC ligands. PUBLIC HEALTH RELEVANCE: This proposal is intended to charter new directions for design of more selective and more active histone deacetylase (HDAC) inhibitors by mapping the binding "fingerprints" of the HDAC ligands in the different HDAC isoforms using small molecule photoaffinity probes. The ultimate goal of this proposal is to design HDAC inhibitors suitable as HDAC based therapeutics for cancer and other HDAC relevant diseases and conditions.

描述(由申请人提供)：这项提案旨在通过使用小分子光亲和探针绘制不同组蛋白脱乙酰酶(HDAC)亚型中组蛋白脱乙酰酶(HDAC)配体的结合“指纹”，为设计更具选择性和更具活性的组蛋白脱乙酰酶(HDAC)抑制剂提供新的方向。由于许多癌症与异常的转录活性有关，而HDAC可以影响转录因子和基因调控，这些酶已被确定为癌症治疗的有吸引力的靶点。事实上，HDAC的化学抑制剂已经被证明可以抑制肿瘤细胞的生长，并诱导分化和细胞死亡。至少有一种非选择性HDAC抑制剂已被FDA批准用于癌症，几种非选择性HDAC抑制剂正在进行临床试验。多项研究表明，针对一种或有限数量的特定HDAC亚型的治疗药物可能会提高这些化合物的整体疗效和较低的毒性。虽然一些化合物表现出了相当有限的异构体选择性，但识别选择性抑制剂的问题远未解决。最近对HDAC7、8及其同源HDAC蛋白晶体结构的研究取得成功，为基于结构的药物设计提供了可能。尽管取得了这些进展，但基于配体和基于结构的方法在设计异构体选择性HDAC抑制剂方面的适用性都受到限制，要么是因为几乎没有异构体选择性抑制剂可作为基于配体的药物设计的起点，要么是因为高分辨率三维结构仅适用于11个I类和II类HDAC异构体中的2个。人们也很少了解那些结合在HDAC蛋白表面的HDAC配体部分的结合模式如何影响HDAC抑制剂的活性。显然，需要一种新的方法来解决这个问题。我们假设，有可能通过小分子光亲和探针(PAP)来映射不同HDAC亚型的配体的结合模式，并利用所获得的知识来设计作为癌症潜在疗法的异构体选择性HDAC抑制剂。我们的具体目标如下：目标1：设计和合成至少三个系列的光亲和探针(PAPs)，包括(1)HDAC配体，(2)用于直接与配体结合的蛋白质残基的光亲和标记(PAL)的芳香叠氮基，和(3)用于连接荧光或生物素标记的脂肪族叠氮(成像标记连接基团-imTAG)。目的：对含有PAL或PAL+imTAG或同时含有PAL和PAL+imTAG的配体进行光亲和标记实验，并用质谱学蛋白质组学方法确定蛋白质的修饰位点。将蛋白质组学结果与建模研究相匹配，并在以后进行改进，以更好地重现实验结果。目的3.评价活体Hela细胞中最有效的光亲和探针的细胞毒性、选择性和结合“指纹”。目的4.将AIMS 1-3的发现融入到基于结构的药物设计中，并进行几轮CADD、药物化学和生物学测试，以提高HDAC配体的活性和选择性。与公共卫生相关：这项提案旨在通过使用小分子光亲和探针绘制不同HDAC亚型中HDAC配体的结合“指纹”，为设计更具选择性和更具活性的组蛋白脱乙酰酶(HDAC)抑制剂开辟新的方向。这项建议的最终目标是设计适合于作为基于HDAC的癌症和其他HDAC相关疾病的治疗的HDAC抑制剂。