Development of Zinc Metallochaperones as Mutant p53 Targeted Anti-cancerDrugs

 描述(申请人提供)：几十年来，研究人员已经承认，在肿瘤中恢复野生型p53功能的小分子是抗癌药物开发的最终目标之一，但却难以实现。TP53是肿瘤中最常见的突变基因，其功能缺失是肿瘤发生的核心驱动因素之一。大多数突变是错义的，会产生一种在癌细胞中高水平存在的缺陷蛋白质。P53-R175H是最常见的错义突变体，错误折叠是因为替代削弱了蛋白质与锌结合的能力。我们发现了一种锌螯合小分子(ZMC1)，它可以通过恢复锌结合和促进正确的折叠(所谓的锌金属配位蛋白(ZMC)机制)来恢复P53-R175H的野生型结构/功能。在体内和体外，ZMC1通过P53介导的凋亡程序重新激活突变型P53并选择性地杀死癌细胞。通过金属离子传递对金属蛋白折叠缺陷的药物修复在药物开发中是前所未有的。我们发现，其他对锌有类似亲和力(Kd)的锌络合剂也可以起到ZMC的作用，ZMC可以重新激活其他锌结合受损的突变体。因此，我们推测ZMCs可以作为有效的突变型P53靶向抗癌药物。这将通过以下具体目标进行研究：1)确定锌稳态机制对ZMC1药效学的影响。细胞对锌水平的ZMC1激增作出反应，通过体内平衡机制使锌正常化。我们假设这些机制调节ZMC1的活性。我们将通过测量操纵肿瘤细胞中锌调控基因对ZMC1活性的影响的机制研究来探索这一点。2)确定易受ZMC再激活影响的p53错义突变谱。ZMC1被建议用来重新激活锌结合受损的突变类；然而，这一类的全部光谱尚不清楚。我们推测，最接近锌结合口袋的突变体最有可能具有受损的锌结合。我们将研究锌结合口袋10？范围内最常见的15个突变。这将定义ZMC潜在的患者粪便。3)设计一种药效、毒性和体内疗效优化的ZMC。我们设计了具有ZMC活性的新型化学类型，在体外比ZMC1更有效。我们将在体内验证这些结果。我们的团队由领导这一研究领域的三个实验室组成。他们的专长如下：1)P53生物学(达伦·卡皮索)、锌/P53折叠生物物理学(Stewart Loh)和锌螯合物药物设计(David Augeri)。与公众健康相关：该提案中进行的研究将为开发一类针对人类癌症最常见突变基因的新型抗癌药物提供基础。这些药物将对所有类型的癌症具有广泛的活性。

 DESCRIPTION (provided by applicant): Researchers have acknowledged for decades that the small-molecule restoration of wild type p53 function in tumors is one of the ultimate, yet elusive goals in cancer drug development. TP53 is the most commonly mutated gene in cancer and loss of its function is one of the central drivers of tumorigenesis. The majority of mutations are missense and generate a defective protein found at high levels in cancer cells. The p53-R175H is the most common missense mutant and is misfolded because the substitution impairs the protein's ability to bind zinc. We discovered a zinc chelating small-molecule (ZMC1) that can restore wild type structure/function to the p53-R175H by restoring zinc-binding and facilitating proper folding (so-called zinc metallochaperone (ZMC) mechanism). ZMC1 "reactivates" mutant p53 and selectively kills cancer cells by a p53 mediated apoptotic program both in vitro and in vivo. The pharmacologic restoration of a folding defect in a metalloprotein by metal ion delivery is unprecedented in drug development. We found that other zinc chelators with a similar affinity (Kd) for zinc can function as ZMCs and ZMCs can reactivate other mutants with impaired zinc-binding. Therefore, we hypothesize that ZMCs can be developed as effective mutant p53 targeted anti-cancer drugs. This will be investigated through the following specific aims: 1) Determine the impact of zinc homeostatic mechanisms on ZMC1 pharmacodynamics. Cells respond to the ZMC1 surge in zinc levels by normalizing zinc through homeostatic mechanisms. We hypothesize that these mechanisms regulate ZMC1 activity. We will explore this through mechanistic studies that measure the effect on ZMC1 activity of manipulating zinc regulatory genes in tumors cells. 2) Define the p53 mis-sense mutational spectrum that is amenable to ZMC reactivation. ZMC1 is proposed to reactivate the mutant class with impaired zinc binding; however the full spectrum of this class is unknown. We hypothesize that the mutants closest to the zinc-binding pocket are most likely to have impaired zinc binding. We will study the 15 most frequent mutants within 10Å of the zinc-binding pocket. This will define the potential patient poo for ZMCs. 3) Design a ZMC optimized for potency, toxicity, and efficacy in vivo. We have designed novel chemotypes that have ZMC activity and are more potent than ZMC1 in vitro. We will validate these results in vivo. Our team is composed of three laboratories that are leading this area of research. Their expertise is the following: 1) p53 biology (Darren Carpizo), Zinc/p53 folding biophysics (Stewart Loh), and zinc chelator drug design (David Augeri). Relevance to public health: The research performed in this proposal will provide the foundation for the development of a new class of anti-cancer drugs that target the most commonly mutated gene in human cancer. These drugs will have broad activity against all types of cancers.