Dissecting and targetting HDMX-medicated chemoresistance in human cancer

项目摘要/摘要 P53肿瘤抑制蛋白调节细胞内稳态的关键控制点，包括 保护细胞DNA免受易患癌症的损害。事实上，一个完全可运作的p53信号 网络是许多化疗药物具有促凋亡活性所必需的。而P53可以是 在癌症中发生突变或缺失，以避免细胞周期停滞或凋亡，这是P53的一种常见替代模式 抑制依赖于负调控基因hdmx和hdm2的过度表达，这两个基因中和了p53 通过蛋白质的相互作用。Hdmx和hdm2在P53抑制和抑制中的单独作用 人类癌症的化疗耐药性在很大程度上是未知的。有趣的是，我们观察到一个相对一致的水平 Hdm2在一大群不同的癌细胞中的表达，而HDMX表现出更多的变量 表达，亚型，如急性髓系白血病表现出最高的HDMX水平。什么 此外，HDMX在几种AML细胞株中的基因依赖程度最高，这些细胞株也保持着野生型p53 表情。这些发现表明，选择性靶向HDMX既可以告知机制的作用 肿瘤对HDMX的依赖，并为在HDMX驱动的肿瘤中恢复P53通路提供治疗策略 癌症。而小分子和装订的多肽抑制剂分别选择性地抑制Hdm2或 针对HDM2和HDMX，现在已进入临床测试，没有经过验证的HDMX专属药物 到目前为止已经开发或验证过。此外，HDMX和HdM2的配体结合的影响 对于蛋白质结构和它们的P53复合体的构象动力学基本上是未知的。在这里，我 目的应用化学的、结构的、细胞的和活体的方法来确定结合决定因素 选择性靶向HDMX，评估选择性HDMX的结构和功能后果 参与，并提出重新激活HDMX驱动的P53介导的细胞凋亡的治疗策略 化疗耐药癌症。为了实现这些目标，我提出了三个实验目标：(1)合成 根据P53反式激活结构域模拟结构增强的α-螺旋以表征其 与HDMX和Hdm2的相互作用，并确定选择性决定因素，(2)阐明构象 SAH-P53多肽结合对HDMX和Hdm2蛋白的单独及与P53的复合体的影响 将HDMX特异性多肽抑制剂引入细胞和体内测试以验证其特异性抑制作用 HDMX作为克服野生型p53癌化疗耐药的治疗策略 表情。我渴望开始一项严格的、多学科的培训计划，以完成 以上建议在哈佛医学院和Dana-Farber癌症研究所攻读研究生课程，请看 期待在化学、癌症生物学、实验等领域从事有影响力的医生-科学家职业生涯 治疗学和临床肿瘤学。

PROJECT SUMMARY/ABSTRACT The p53 tumor suppressor protein regulates critical control points of cellular homeostasis, including protecting cellular DNA from damage that could predispose to cancer. Indeed, a fully operational p53 signaling network is required for the pro-apoptotic activity of many chemotherapeutic agents. Whereas p53 can be mutated or deleted in cancer to avoid cell cycle arrest or apoptosis, a frequent alternative mode of p53 suppression relies on overexpression of the negative regulators HDMX and HDM2, which neutralize p53 through protein interaction. The individual contributions of HDMX and HDM2 to p53 suppression and chemoresistance in human cancers are largely unknown. Interestingly, we observe a relatively consistent level of HDM2 expression across a large and diverse panel of cancer cells, whereas HDMX manifests more variable expression, with subtypes such as acute myeloid leukemia exhibiting among the highest HDMX levels. What's more, HDMX scored in the top gene dependencies for several AML cell lines that also maintain wild-type p53 expression. These findings suggest that selective targeting of HDMX could both inform the mechanistic role of HDMX dependency in cancer and provide a therapeutic strategy for restoring the p53-pathway in HDMX-driven cancers. Whereas small molecule and stapled peptide inhibitors, which respectively inhibit HDM2 selectively or target both HDM2 and HDMX, have now been advanced to clinical testing, no validated HDMX-specific agent has been developed or validated to date. In addition, the influence of ligand engagement of HDMX and HDM2 on protein structure and the conformational dynamics of their p53 complexes is essentially unknown. Here, I aim to apply chemical, structural, cellular, and in vivo approaches to identify the binding determinants for selective targeting of HDMX, evaluate the structural and functional consequences of selective HDMX engagement, and advance a therapeutic strategy for reactivating p53-mediated apoptosis in HDMX-driven chemoresistant cancers. To achieve these goals, I propose three experimental aims: (1) synthesize libraries of structurally-reinforced alpha-helices modeled after the p53 transactivation domain to characterize their interactions with HDMX and HDM2, and identify selectivity determinants, (2) elucidate the conformational effects of SAH-p53 peptide engagement on HDMX and HDM2 proteins, alone and in complex with p53, and (3) advance lead HDMX-specific peptide inhibitors to cellular and in vivo testing to validate specific inhibition of HDMX as a therapeutic strategy for overcoming chemoresistance in cancers that retain wildtype p53 expression. I am eager to embark on a rigorous and multidisciplinary training program to accomplish the above-proposed graduate studies at Harvard Medical School and the Dana-Farber Cancer Institute, and look forward to an impactful physician-scientist career at the interface of chemistry, cancer biology, experimental therapeutics, and clinical oncology.