Identification of Pin1 Chemical Probes for Studying Phosphorylation Signaling

用于研究磷酸化信号转导的 Pin1 化学探针的鉴定

• 批准号: 8213459
• 负责人: Kun Ping Lu
• 金额: $ 4.35万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-01-15 至 2013-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8213459
• 关键词: Adopted; Affect; Aging; Alzheimer' s Disease; Animal Model; Basic Science; Binding; Biological; Biological Assay; Breast Cancer Cell; Cell physiology; Cells; Chemicals; Clinic; Coupled; Cyclophilin A; Cyclosporine; Development; Disease; ERBB2 gene; Enzymes; FK506; Family; Fluorescence; Fluorescence Polarization; Fluorogenic Substrate; Future; Genes; Genomics; Grant; Growth; HER2 inhibition; Human; Image Analysis; In Vitro; Isomerism; Laboratories; Malignant Neoplasms; Methods; Microscope; Molecular; Molecular Conformation; Nature; Peptides; Peptidylprolyl Isomerase; Phosphorylation; Physiological; Play; Procedures; Process; Proline; Proteins; Regulation; Role; Screening procedure; Series; Serine; Signal Pathway; Signal Transduction; Sirolimus; Specificity; Structure; Structure-Activity Relationship; Tacrolimus Binding Protein 1A; Technology; Testing; Threonine; Time; Titrations; Trastuzumab; United States National Institutes of Health; alanylalanine; alanylproline; analog; base; chymotrypsin; follow-up; high throughput screening; human disease; in vivo; inhibitor/antagonist; interest; malignant breast neoplasm; mouse model; novel; novel therapeutics; overexpression; protein degradation; protein function; protein structure; public health relevance; response; small molecule; stereochemistry; tool

DESCRIPTION (provided by applicant): A common and central regulatory mechanism in the cell is proline-directed phosphorylation on certain Ser/Thr- Pro (phos.Ser/Thr-Pro) motifs. The unique stereochemistry of Pro means that it can adopt two distinct cis or trans conformations, but the biological significance of these conformational switches was unappreciated for a long time. We have recently identified a unique prolyl isomerase, Pin1 (Gene ID, NM_006221; Protein ID, NP_006212) that catalyzes the conformational switches of specific phos.Ser/Thr-Pro motifs in a subset of proteins to regulate cell signaling. Such Pin1-catalyzed conformational regulation can have a profound impact on many key proteins in diverse cellular processes. Importantly, Pin1 deregulation plays a pivotal role in the development of an increasing number of diseases and provides a potential new therapeutic option. However, chemical probes to inhibit Pin1 function identified so far either lack the critically needed specificity or simply cannot enter cells. We have now developed a series of robust and sensitive procedures to identify and evaluate Pin1 probes in vitro and in vivo and also identified interesting hits in our pilot screen. Therefore, in this proposal, we will collaborate with Drs. Douglas Auld and Craig Thomas at NIH Chemical Genomics Center to identify Pin1 chemical probes, with the following specific aims: (1) To identify inhibitors of human Pin1 using a quantitative high-throughput screening approach against the MLSMR containing 300,000 small molecules. (2) To validate the potency and specificity of the hits in secondary assays to identify those compounds that specifically inhibit Pin1, but not other non-phosphorylation-specific prolyl isomerases. (3) To characterize and optimize Pin1 chemical probes by tertiary cell-based assays and structure-activity relationship analysis, structure-based methods, analogue synthesis and medicinal chemical principles. (4) Beyond the granting period for this proposal, to test the most promising compounds for their ability to inhibit Pin1 function in several Pin1-relevant mouse models of cancer or Alzheimer<s disease that we have established in our laboratory. This proposal would allow us to identify urgently needed chemical probes to study Pin1-regulated Pro-directed phosphorylation signaling under physiological and pathological conditions. PUBLIC HEALTH RELEVANCE: We have recently identified a new enzyme called Pin1 that is a pivotal regulator of numerous cellular processes. Importantly, Pin1 deregulation plays a critical role in the development of an increasing number of human diseases, including aging, cancer and Alzheimer<s disease. However, currently there is no Pin1-specific chemical probe available. In this application, we propose to discover and optimize Pin1 chemical probe to study Pin1-regulated processes under physiological and pathological conditions.

描述（由申请人提供）：细胞中常见且重要的调节机制是某些 Ser/Thr-Pro (phos.Ser/Thr-Pro) 基序上的脯氨酸定向磷酸化。 Pro独特的立体化学意味着它可以采用两种不同的顺式或反式构象，但这些构象开关的生物学意义长期以来未被认识到。我们最近发现了一种独特的脯氨酰异构酶 Pin1（基因 ID，NM_006221；蛋白质 ID，NP_006212），它可以催化蛋白质子集中特定 phos.Ser/Thr-Pro 基序的构象转换，从而调节细胞信号传导。这种 Pin1 催化的构象调节可以对不同细胞过程中的许多关键蛋白质产生深远的影响。重要的是，Pin1 失调在越来越多疾病的发展中发挥着关键作用，并提供了潜在的新治疗选择。然而，迄今为止发现的抑制 Pin1 功能的化学探针要么缺乏急需的特异性，要么根本无法进入细胞。我们现在已经开发了一系列稳健且灵敏的程序来识别和评估体外和体内的 Pin1 探针，并在我们的试点筛选中识别出有趣的命中。因此，在这项提案中，我们将与博士合作。 NIH 化学基因组中心的 Douglas Auld 和 Craig Thomas 旨在鉴定 Pin1 化学探针，具体目标如下：（1）使用针对包含 300,000 个小分子的 MLSMR 的定量高通量筛选方法来鉴定人类 Pin1 的抑制剂。 (2) 验证二次测定中命中的效力和特异性，以鉴定那些特异性抑制 Pin1 但不抑制其他非磷酸化特异性脯氨酰异构酶的化合物。 (3) 通过三级细胞分析和构效关系分析、基于结构的方法、类似物合成和药物化学原理来表征和优化Pin1化学探针。 (4) 在本提案的授权期之外，在我们实验室建立的几种与 Pin1 相关的癌症或阿尔茨海默病小鼠模型中测试最有前途的化合物抑制 Pin1 功能的能力。该提案将使我们能够识别急需的化学探针，以研究生理和病理条件下 Pin1 调节的 Pro 定向磷酸化信号传导。 公共健康相关性：我们最近发现了一种名为 Pin1 的新酶，它是许多细胞过程的关键调节剂。重要的是，Pin1 失调在越来越多的人类疾病（包括衰老、癌症和阿尔茨海默病）的发展中发挥着关键作用。然而，目前还没有针对 Pin1 的化学探针。在此应用中，我们建议发现并优化 Pin1 化学探针来研究生理和病理条件下 Pin1 的调节过程。

项目成果

Prolyl isomerase Pin1 acts downstream of miR200c to promote cancer stem-like cell traits in breast cancer.

• DOI: 10.1158/0008-5472.can-13-2785
• 发表时间: 2014-07-01
• 期刊: Cancer research
• 影响因子: 11.2
• 作者: Luo ML;Gong C;Chen CH;Lee DY;Hu H;Huang P;Yao Y;Guo W;Reinhardt F;Wulf G;Lieberman J;Zhou XZ;Song E;Lu KP
• 通讯作者: Lu KP

The Rab2A GTPase promotes breast cancer stem cells and tumorigenesis via Erk signaling activation.

• DOI: 10.1016/j.celrep.2015.03.002
• 发表时间: 2015-04-07
• 期刊: Cell reports
• 影响因子: 8.8
• 作者: Luo ML;Gong C;Chen CH;Hu H;Huang P;Zheng M;Yao Y;Wei S;Wulf G;Lieberman J;Zhou XZ;Song E;Lu KP
• 通讯作者: Lu KP