Mechanisms of mutant p53 reactivation

项目总结 抑癌蛋白P53是人类癌症中最常见的突变蛋白。约600,000 美国每年都有新的癌症患者被诊断出患有表达突变的p53的肿瘤。多数 其中的突变是错义突变，影响P53DNA结合域的六个热点位置之一。 这些癌症表达全长P53，它失去了肿瘤抑制活性，但获得了功能增强。 为癌细胞提供选择性优势的致癌特性。 大量受影响的癌症使p53成为癌症治疗的精致靶点。然而，治疗性的 治疗方法需要重新激活突变的P53。研发“再激活或矫正药物”在中国具有挑战性。 但由于在制药、生物技术和这一领域的学术界经验有限，这一问题变得更加复杂。 这些通过开发p53校正药物来探索新的治疗方法的挑战导致了 建议的P53复活剂化合物的临床试验非常缓慢，而且成功有限。它最近才浮出水面 已报道的几种化合物很可能在体内不作用于突变的p53，而是利用氧化还原- 表达P53突变体的细胞的敏感性。与P53结合的真p53突变校正药物的研究进展 并在p53癌突变体中恢复类似野生型的构象/活性，因此仍然是 潜在的非常大的影响。为了实现这一目标，从机制上理解p53癌突变体 重新激活过程是必不可少的，但目前主要缺乏，因为缺乏真正的p53校正器 除了专为相对罕见的P53-Y220C等位基因开发的化合物以外的其他分子。 我们已经广泛地研究了遗传和药理学上的P53重新激活。我们发现基因内营救 我们正在开发的突变和小分子会引起类似的构象变化，并稳定 P53热点突变体的活性构象。尽管重新激活的突变没有直接的治疗作用 潜在的，它们有助于我们理解p53突变的再激活机制，并可以指导纠正药物 发展。利用从重新激活第二位点突变获得的信息，我们开发了一种工具 与突变型p53结合从而恢复突变型p53的DNA结合活性的化合物 体外纯化系统。当含有p53热点突变体的细胞暴露时，p53靶基因被诱导 这些化合物。此外，在p53突变体中，细胞增殖被阻止，并被诱导凋亡。 依赖的态度。重要的是，携带p53突变体的肿瘤的生长被该化合物系列所阻断 动物模型。缺乏P53或表达野生型P53的肿瘤不受这种治疗的影响。这些 化合物为开发作为真正的p53突变体的类药物分子的可行性提供了强有力的支持 校正员。我们现在建议使用这些工具化合物以及特征良好的救援突变来 发展对P53热点突变体重新激活过程的详细分子理解。调查结果来自 这些研究对于启动化学成分多样的P53校正药物的开发至关重要。

PROJECT SUMMARY The tumor suppressor protein p53 is the most frequently mutated protein in human cancers. About 600,000 new cancer patients in the United States are diagnosed each year with tumors expressing mutated p53. Most of the mutations are missense mutations that affect one of six hotspot sites in the p53 DNA binding domain. These cancers express full length p53 that has lost tumor suppressor activity, but has acquired gain-of-function oncomorphic properties that provide selective advantage to cancer cells. The large number of affected cancers make p53 an exquisite target for cancer therapy. However, therapeutic approaches require reactivation of mutated p53. Developing “reactivation or corrector drugs” is challenging in itself, but further complicated by very limited experience in pharma, biotech, and academia in this domain. These challenges in exploring novel therapeutic approaches by developing p53 corrector drugs have led to very slow, and limited success in clinical trials with proposed p53 reactivator compounds. It recently emerged that several of the reported compounds are likely not acting on mutant p53 in vivo, but rather exploit redox- sensitivity of cells expressing p53 mutants. Development of bona fide p53 mutant corrector drugs that bind p53 and restore a wild-type like conformation/activity in p53 cancer mutants, thus remains a central goal with potentially very high impact. To achieve this goal mechanistic understanding of the p53 cancer mutant reactivation process is essential, but currently mostly lacking due to the lack of genuine p53 corrector molecules with the exception of compounds developed specifically for the relatively rare p53-Y220C allele. We have extensively studied genetic and pharmacological p53 reactivation. We found that Intragenic rescue mutations and small molecules we are developing induce a similar conformational change and stabilize an active conformation of p53 hotspot mutants. Although reactivation mutations have no direct therapeutic potential, they help in our understanding of p53 mutant reactivation mechanisms and can guide corrector drug development. Using information obtained from reactivating second-site mutations, we have developed tool compounds that bind mutant p53 and thereby restore DNA binding activity of mutant p53 in a reconstituted purified in vitro system. p53 target genes are induced when cells harboring p53 hotspot mutants are exposed to these compounds. Furthermore, cell proliferation is halted and apoptosis is induced in a p53 mutant dependent manner. Importantly, growth of tumors carrying p53 mutants is blocked by this compound series in animal models. Tumors lacking p53 or expressing wild-type p53 are not affected by such treatment. These compounds provide strong support for feasibility to develop drug-like molecules that act as genuine p53 mutant correctors. We now propose to use these tool compounds as well as well-characterized rescue mutations to develop detailed molecular understanding of the reactivation process for p53 hotspot mutants. Findings from these studies will be essential to jump start the development of chemically diverse p53 corrector drugs.