Dissecting and Targeting Deregulated Mitochondrial Apoptosis in Human Cancer

 描述（由申请人提供）：凋亡蛋白BCL-2家族调节细胞生命和死亡之间的关键平衡。这种重要信号转导网络的失调驱动了广谱人类癌症的发展、维持和耐药性。我的实验室正在解决BCL-2家族蛋白质之间的相互作用如何在稳态和恶性转化过程中调节其结构变化和信号功能的问题。由于BCL-2蛋白的相互作用决定了细胞在应激反应中的生存或死亡，我们相信这些结构-功能研究将为基本的生物学机制和对它们进行间接调节的机会提供信息。30年的BCL-2家族研究已经产生了第一个在BCL-2依赖性癌症中反应性凋亡的小分子药物。尽管这一显着的科学进步和靶向BCL-2蛋白在癌症中的新的承诺，基本问题仍然是关于这个复杂的蛋白质家族如何调节线粒体凋亡。事实上，它们的主要膜定位使得BCL-2家族蛋白的研究非常具有挑战性。BAX和巴克的活化形式如何自组装成毒性寡聚孔-介导细胞凋亡的死亡通道-尚不清楚。抗凋亡蛋白如BCL-2如何改变其在膜内的结构以阻断BAX/巴克活化的各个步骤尚不清楚。事实上，BCL-2蛋白的许多保守结构域之间的接触表面的全谱是未知的。也许最令人困惑的问题是，这些结构相似的蛋白质如何具有完全相反的功能，作为细胞死亡的抑制剂或激活剂，仍然是一个基本未知的问题。我们用广泛的实验方法来解决这些机制问题，包括化学、结构生物学、蛋白质组学、生物化学、细胞生物学、小鼠模型和药理学。例如，我们已经开发了新的化学工具来解剖和靶向BCL-2家族蛋白质相互作用，先进的新方法使用光反应性结构肽和质谱快速准确地识别相互作用位点，并且目前正在应用氢-氘交换质谱首次实时研究BCL-2家族在膜中的构象变化。我们对R35癌症研究计划的目标包括定义BAX和巴克的构象激活和同源寡聚化机制，表征抗凋亡BCL-2蛋白的BH 4结构域抑制BAX和巴克的新机制，并研究控制BCL-2蛋白凋亡功能的新变构机制。在每种情况下，结构-功能的见解将被利用来开发针对癌症中凋亡抗性的新方法。作为一名化学生物学家和执业儿科肿瘤学家，我致力于我的研究实验室破译BCL-2家族介导的癌症机制，以便对其蛋白质相互作用生物学的新见解可以为下一代人类癌症的促凋亡疗法提供信息。

 DESCRIPTION (provided by applicant): The BCL-2 family of apoptotic proteins regulates the critical balance between cellular life and death. Deregulation of this essential signal transductio network drives the development, maintenance, and chemoresistance of a broad spectrum of human cancers. My laboratory is addressing the questions of just how interactions among BCL-2 family proteins regulate their structural changes and signaling functions during homeostasis and malignant transformation. Because BCL-2 protein interactions determine whether the cell will live or die in response to stress, we believe these structure-function studies will both infor basic biological mechanisms and opportunities to pharmacologically modulate them. Thirty years of BCL-2 family research has led to the first small molecule drug to reactive apoptosis in BCL-2 dependent cancer. Despite this remarkable scientific progress and the renewed promise of targeting BCL-2 proteins in cancer, fundamental questions remain about how this complex protein family regulates mitochondrial apoptosis. Indeed, their predominant membrane localization has made BCL-2 family proteins remarkably challenging to study. How the activated forms of BAX and BAK self-assemble into toxic oligomeric pores - the very death channels that mediate apoptosis - is unknown. How anti-apoptotic proteins such as BCL-2 change their structure within the membrane to block the various steps of BAX/BAK activation is unknown. Indeed, the full spectrum of contact surfaces between the many conserved domains of BCL-2 proteins is unknown. Perhaps the most perplexing question of how such structurally similar proteins can have diametrically opposite functions, as either inhibitors or activators of cell deat, remains essentially unknown. We address these mechanistic questions with broad experimental approaches, spanning chemistry, structural biology, proteomics, biochemistry, cell biology, mouse modeling, and pharmacology. For example, we have developed new chemical tools to dissect and target BCL-2 family protein interactions, advanced new methods to rapidly and accurately identify interaction sites using photoreactive structured peptides and mass spectrometry, and are currently applying hydrogen- deuterium exchange mass spectrometry to study BCL-2 family conformational changes in the membrane in real-time for the first time. Our goals for this R35 cancer research program include defining the conformational activation and homo-oligomerization mechanism(s) of BAX and BAK, characterizing a novel mechanism for BAX and BAK suppression by the BH4 domains of anti-apoptotic BCL-2 proteins, and investigating a new allosteric mechanism that controls the apoptotic functionalities of BCL-2 proteins. In each case, the structure- function insights will be harnessed to develop new approaches for targeting apoptotic resistance in cancer. As a chemical biologist and practicing pediatric oncologist, I have dedicated my research laboratory to deciphering BCL-2 family-mediated cancer mechanisms so that fresh insights into their protein interaction biology can inform the next generation of pro-apoptotic therapies for human cancer.