Survival Mechanisms for Apoptotic Caspase

描述（由申请人提供）：自从大约20年前发现半胱天冬酶以来，已有关于凋亡半胱天冬酶的非凋亡作用的报道。也许最清楚的范例是半胱天冬酶-8（Casp 8），虽然它对触发外源性凋亡途径至关重要，但也是某些细胞群体在发育过程中生存所必需的。例如，在小鼠中的缺失研究表明，半胱氨酸天冬氨酸蛋白8，其适配器FADD，及其调节剂FLIPL可以驱动淋巴细胞，心肌细胞和一些上皮细胞类型的克隆扩增/增殖或凋亡。该提案旨在为生存和死亡之间的选择提供生物化学证据，同时也有可能揭示生存途径的下游目标。因此，我们提供了概念验证工具和知识，可以导致化合物的开发，以治疗性地干扰增殖性疾病如癌症的存活/死亡轴。 总体假设“进入增殖程序的细胞的存活取决于Casp 8活性，其关键依赖于Casp 8酶的构象和有限数量的下游靶点的切割”将在本研究中进行剖析。我们提出了三个相互关联的目标来检验这一假设 通过定义驱动存活事件的Casp 8酶的生物化学性质，定义这种酶的靶点，并理解在特定的实验细胞环境中，Casp 8如何促进存活而不是凋亡。每个目标都可以独立完成，每个目标的结果将大大加深对Casp 8介导的细胞存活机制的了解。然而，当来自目标1（Casp 8特异性的基本机制）的数据与目标2（内源性促存活Casp 8靶点）结合时，该项目的全部力量得以实现，并且目标1和2的结果用于通知用于定义细胞在存活和凋亡之间的决定的适当实验范例（目标3）。这一提议意义重大，因为它试图揭示Casp 8用于描绘从死亡中存活的分子机制，并且成功的结果可能直接影响正在临床开发作为癌症治疗剂的蛋白质和小分子药理学工具的选择。

DESCRIPTION (provided by applicant): Ever since the discovery of the caspases almost 20 years ago there have been reports of non-apoptotic roles for apoptotic caspases. Perhaps the clearest paradigm is caspase-8 (casp8), which although essential for triggering the extrinsic apoptotic pathway, is also required for the survival of certain cell populations during development. For example, deletion studies in mice reveal that casp8, its adapter FADD, and its regulator FLIPL may drive either clonal expansion/proliferation or apoptosis of lymphocytes, cardiomyocytes, and some epithelial cell types. This proposal seeks to provide biochemical evidence for the choice between survival and death, while also having the potential to reveal downstream targets of the survival pathway. Thus we provide proof-of-concept tools and knowledge that can lead to the development of compounds to interfere therapeutically in the survival/death axis of proliferative diseases such as cancer. The overarching hypothesis "The survival of cells entering the proliferation program that depends on casp8 activity relies criticaly on the conformation of the casp8 enzyme and cleavage of a limited number of downstream targets" will be dissected in this study. We propose three interlinked aims to test this hypothesis by defining the biochemical nature of the casp8 enzyme that drives the survival events, defining the targets of this enzyme, and understanding how casp8 promotes survival, and not apoptosis, in specific experimental cellular contexts. Each aim can be accomplished independently, and the outcome of each aim will substantially further knowledge of the mechanisms of cell survival mediated by casp8. However, the full power of this project is realized when the data from Aim 1 (fundamental mechanisms of casp8 specificity) is coupled to Aim 2 (endogenous pro-survival casp8 targets), and the outcomes of Aims 1 and 2 used to inform the appropriate experimental paradigms for defining the cell's decision between survival and apoptosis (Aim 3). This proposal is significant because it seeks to reveal the molecular mechanism used by casp8 to delineate survival from death, and a successful outcome may impact directly on the selection of protein and small molecule pharmacologic tools that are under clinical development as cancer therapeutics.