Design of Pin1 Inhibitors

项目总结/摘要 分子伴侣和脯氨酰异构酶Pin 1是一个有前途的癌症治疗靶点。敲除PIN 1 在正常小鼠中仅产生轻度退行性表型，但在致癌小鼠中极大地损害肿瘤发生。 在小鼠模型中，并降低前列腺癌和乳腺癌异种移植模型中的增殖。的依赖 癌细胞对Pin 1活性的影响似乎是通过其稳定多个癌基因并破坏其稳定性的能力而发生的。 多种肿瘤抑制剂。然而，尽管Pin 1作为治疗靶点具有巨大的潜力， 已经报道了既特异又具有生物活性抑制剂。挑战在于Pin 1具有 典型的“不可药物化”蛋白质的特征;它有一个浅的极性结合口袋，到目前为止， 阻止了活性抑制剂的发现。我们提出了一个以前从未探索过的选择， 现有的Pin 1的有效体外抑制剂，以及使用二硫键片段拴系扩展该SAR。一 本研究的主要创新点在于设计并初步测试了第一个高效的细胞 使用高亲和性含磷酸盐化合物并赋予渗透性的Pin 1的渗透性抑制剂 通过氨基磷酸酯笼化。这些分子渗透脂质双层， 释放成含磷酸盐的活性代谢物。在令人兴奋的初步结果中，我们的顶级化合物（DS-2（R）） 对于细胞周期蛋白D1在体内的不稳定性具有单位数纳摩尔的EC 50-一种已知的Pin 1抑制生物标志物。 在此，我提出（SA 1）对第一个化合物的生物活性进行严格的物理和功能表征。 有效且具有细胞活性的Pin 1抑制剂。由于我们还不确定这些分子的特异性， 补充这种方法，我们提出（SA 2）的表征和改进的特异性，我们的 初步先导化合物DS-2（R）。这种方法源于观察到Pin 1能够容忍大体积的 取代苯并噻吩，这可能导致对脱靶的亲和力降低。因此在 与加州大学旧金山分校的先驱专家（吉姆威尔斯，米歇尔阿金）合作，我们将筛选一个含二硫化物的 用于结合本体耐受疏水架以制备类似物的药效团的片段文库 其将保持与Pin 1的有效相互作用，同时有望降低与off- 目标的我们将使用在二硫键连接筛选中获得的信息来合成和评估 定向系列DS-2（R）类似物，用于改善Pin 1特异性。总的来说，本项目的目标是 利用现代设计原理来表征和改进第一种具有生物活性的Pin 1化学品 probes.这些分子将促进Pin 1在肿瘤学中的一致性和质量研究，并可能导致 对它是否适合作为目标有了新的认识。重要的是，拟议的研究将大大扩展我的培训， 一个化学生物学家通过结合基于结构的设计，现代筛选方法，化合物和 目标验证方法

Project Summary/Abstract The molecular chaperone and prolyl isomerase Pin1 is a promising target for cancer therapy. Knockout of PIN1 yields only mild degenerative phenotypes in normal mice, but greatly impairs tumorigenesis in oncogenic mouse models and decreases proliferation in prostate and breast cancer xenograft models. The reliance of cancer cells on Pin1 activity seems to occur through its ability to stabilize multiple oncogenes and destabilize multiple tumor suppressors. However, despite Pin1's great potential as a therapeutic target, no chemical inhibitor which is both specific and biologically-active has been reported. The challenge is that Pin1 has the features of a classically “undruggable” protein; it has a shallow, polar binding pocket which has, thus far, prevented discovery of viable inhibitors. We propose a previously-unexplored option for imparting permeability on existing potent in vitro inhibitors of Pin1 as well as expand this SAR using disulfide fragment tethering. A key innovation of this study is that we have designed and preliminarily tested the first potent and cell permeable inhibitors of Pin1 using high affinity phosphate-baring compounds and imparting permeability through phosphoramidate caging. These permeate the lipid bilayer and are subsequently enzymatically liberated to a phosphate-baring active metabolite. In exciting preliminary results, our top compound (DS-2(R)) has single-digit nanomolar EC50 for cyclin D1 destabilization in vivo – a known Pin1 inhibition biomarker. Herein, I propose (SA 1) rigorous physical and functional characterization of the biological activity of the first potent and cell-active Pin1 inhibitors. Since we are yet unsure of the specificity of these molecules, to supplement this approach, we propose (SA 2) the characterization and improvement of the specificity of our preliminary lead compound DS-2(R). This approach arose from observations that Pin1 is able to tolerate bulky substitutions in place of the benzothiophene which would likely lead to reduced affinity for off-targets. Thus, in collaboration with pioneering experts at UCSF (Jim Wells, Michelle Arkin), we will screen a disulfide-containing fragment library for pharmacophores which engage the bulk-tolerant hydrophobic shelf to prepare analogues which will maintain potent interaction with Pin1 while hopefully reducing ligand complementarity with off- targets. We will use the information gained in the disulfide tethering screen to synthesize and evaluate a directed series of DS-2(R) analogues for improved Pin1 specificity. Overall, the goal of this project is to leverage modern design principles to characterize and improve the first biologically active Pin1 chemical probes. These molecules will facilitate consistent and quality research of Pin1 in oncology and potentially lead to new insights into its suitability as a target. Critically, the proposed studies will greatly expand my training as a chemical biologist by combining structure-based design, modern screening methods, and compound and target validation methods.