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）：细胞中的一种常见和中央调节机制是在某些Ser/thr-pro（phos.ser/thr-Pro）基序上定向的磷酸化。 Pro的独特立体化学意味着它可以采用两个不同的顺式或反式构象，但是这些构象开关的生物学意义很长一段时间都没有得到批准。我们最近确定了独特的蛋白酶异构酶，PIN1（基因ID，NM_006221;蛋白质ID，NP_006212），该蛋白质在蛋白质子集中催化特定的Phos/Thr-Pro基序的构象开关以调节细胞信号。这种PIN1催化的构象调节可以对各种细胞过程中的许多关键蛋白产生深远的影响。重要的是，PIN1放松管制在越来越多的疾病的发展中起关键作用，并提供了潜在的新治疗选择。但是，到目前为止，抑制PIN1功能的化学探针要么缺乏迫切需要的特异性，要么根本无法进入细胞。现在，我们已经开发了一系列强大而敏感的程序，以在体外和体内识别和评估PIN1探针，并在我们的飞行员屏幕中识别出有趣的命中。因此，在此提案中，我们将与Drs合作。 NIH化学基因组学中心的道格拉斯·奥尔德（Douglas Auld）和克雷格·托马斯（Craig Thomas）识别PIN1化学探针，以下具体目的：（1）使用定量的高通量筛选方法来鉴定人类PIN1的抑制剂，以针对含有300,000个小分子的MLSMR。 （2）验证次级测定中命中的效力和特异性，以识别那些专门抑制PIN1但没有其他非磷酸化特异性丙酸异构体的化合物。 （3）通过基于三级细胞的测定和结构活性关系分析，基于结构的方法，模拟合成和药用化学原理来表征和优化PIN1化学探针。 （4）除了该提案的授予期之外，还可以在我们在实验室中确定的几种癌症或阿尔茨海默氏病的疾病中抑制PIN1功能的最有希望的化合物。该提案将使我们能够识别出急需的化学探针，以研究生理和病理条件下的PIN1调节的磷酸化信号传导。 公共卫生相关性：我们最近确定了一种称为PIN1的新酶，该酶是众多细胞过程的关键调节剂。重要的是，PIN1放松管制在越来越多的人类疾病（包括衰老，癌症和阿尔茨海默氏病）的发展中起着至关重要的作用。但是，目前尚无PIN1特异性化学探针。在此应用中，我们建议在生理和病理条件下发现并优化PIN1化学探针以研究PIN1调节的过程。

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