Screening for small molecule inhibitors of Stat5 (RMI)

Stat5 小分子抑制剂 (RMI) 的筛选

• 批准号: 6879801
• 负责人: Robert A. Kirken
• 金额: $ 7.43万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-30 至 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6879801
• 关键词: biotechnology; cell line; combinatorial chemistry; drug discovery /isolation; fluorescence polarization; fluorescent dye /probe; gene induction /repression; genetic transcription; high throughput technology; peptide library; pharmacokinetics; phosphopeptides; protein purification; small molecule; transcription factor

DESCRIPTION (provided by applicant): Signal transducers and activators of transcription (Stats) mediate a variety of extracellular signals. In response to ligand binding to its receptor, these proteins become phosphorylated on an invariant tyrosine residue located near the C-terminus, either by the kinase activity of the receptor, or by Janus kinases (Jaks) recruited to the receptor. Upon tyrosine phosphorylation, Stat proteins dimerize via opposing SH2 domain of one monomer with the pTyr of the other monomer. They also become serine phosphorylated on a residue proximal to the tyrosine phosphorylation residue that can regulate its activity. The phosphorylated bi-dentate ligated dimer then translocates to the nucleus, binds specific DNA promoter sequences and initiates transcription of designated response elements. In normal cells Stats mediate signaling from growth factor receptors and immune stimuli. However in several cancer types, Stats are constitutively activated, including two highly homologous proteins Stat5a and Stat5b, which are present in many different cancers of breast, prostate, leukemias and lymphomas. We and others have shown that disruption of Stat5a/b activity can induce cell death in some tumor cell lines. It would therefore be advantageous to have a high throughput screen (HTS) to identify small molecule inhibitors of Stat5 to block their aberrant activation of this tumor pro-survival pathway. The objective of this project is to develop a HTS to identify small molecule inhibitors that block oligomerization of Stat5 and gene transcriptional activity. There has been little effort reported in the literature on the development of small molecule inhibitors of Stat5 function. Our specific aims are as follows: 1. Discover and optimize small molecule inhibitors that will be derived from either a) Stat5 SH2 domain inhibitor combinatorial phosphopeptide library of known Stat5 docking sites on receptors and b) screening the NCI Diversity Set of small drug-like compounds. Both readouts will employ monitoring changes in fluorescence polarization. 2. Assay and confirm potential inhibitory compounds of Stat5 oligomerization and DNA binding/function by testing the most promising compounds in a confirmatory HTS cell based reporter assay to determine their effects on gene transcription. At the conclusion of this research we expect to have established a method to rapidly screen and identify small molecule inhibitors of Stat5. This screen should allow for the more rapid development of therapeutic agents to treat a variety of diseases.

描述（由申请人提供）： 信号转导子和转录激活子（Stats）介导多种细胞外信号。 响应于配体与其受体的结合，这些蛋白质通过受体的激酶活性或通过Janus激酶（Jaks）募集到受体而在位于C-末端附近的不变酪氨酸残基上磷酸化。 在酪氨酸磷酸化后，Stat蛋白通过一个单体的相对SH 2结构域与另一个单体的pTyr二聚化。 它们也在靠近酪氨酸磷酸化残基的残基上变成丝氨酸磷酸化，这可以调节其活性。 磷酸化的双齿连接二聚体然后易位到细胞核，结合特异性DNA启动子序列并启动指定应答元件的转录。 在正常细胞中，Stats介导来自生长因子受体和免疫刺激的信号传导。 然而，在几种癌症类型中，Stats是组成性激活的，包括两种高度同源的蛋白Stat 5a和Stat 5 b，它们存在于许多不同的乳腺癌、前列腺癌、白血病和淋巴瘤中。 我们和其他人已经表明，Stat 5a/B活性的破坏可以诱导某些肿瘤细胞系的细胞死亡。 因此，具有高通量筛选（HTS）来鉴定Stat 5的小分子抑制剂以阻断其对该肿瘤促存活途径的异常激活将是有利的。 本项目的目的是开发一种HTS，以鉴定阻断Stat 5寡聚化和基因转录活性的小分子抑制剂。 在文献中几乎没有关于开发Stat 5功能的小分子抑制剂的报道。 我们的具体目标如下： 1. 发现和优化小分子抑制剂，其将衍生自a）受体上已知的Stat 5对接位点的Stat 5 SH 2结构域抑制剂组合磷酸肽文库和B）筛选小药物样化合物的NCI多样性集。 两个读数都将采用监测荧光偏振的变化。 2.通过在确证性HTS细胞报告基因试验中测试最有希望的化合物，测定并确认Stat 5寡聚化和DNA结合/功能的潜在抑制化合物，以确定其对基因转录的影响。 在本研究的结论中，我们期望建立一种快速筛选和鉴定Stat 5小分子抑制剂的方法。 这种筛选应该允许更快速地开发治疗多种疾病的治疗剂。