Regulation of TRAIL Induced Apoptosis in Cancer Cells

描述(申请人提供)：基于其选择性杀死癌细胞和抑制肿瘤转移的能力，肿瘤坏死因子超家族死亡配体试验正在成为许多癌症治疗的潜在工具。不幸的是，许多肿瘤细胞株对TRAIL具有耐药性，这表明TRAIL作用的分子基础以及如何恢复TRAIL的敏感性对于最大限度地发挥这种有希望的癌症治疗的潜力至关重要。长期目标是准确地了解癌细胞如何对TRAIL治疗产生抗药性，以及这种知识如何反过来导致恢复TRAIL敏感性的新的治疗干预措施。这一特殊应用的目的是确定TRAIL如何通过将多功能分选蛋白PACS-2转换为凋亡效应器来杀死癌细胞，以及PACS-2的失调如何使癌症抵抗TRAIL诱导的细胞死亡。具体地说，Akt磷酸化的PACS-2通过协调内质网(ER)与线粒体通讯的抗凋亡钙通道的定位来调节健康细胞的动态平衡。作为对TRAIL的响应，PACS-2被去磷酸化，通过协调凋亡钙信号和介导BID激活的溶酶体-线粒体通讯来促进线粒体通透性和细胞死亡。中心假说是，PACS-2的失调使癌细胞能够以两种不同的方式抵抗TRAIL杀伤：在Akt升高的癌症中，PACS-2的凋亡活性被持续的Akt磷酸化抑制，而在PACS-2基因缺失的癌症中，TRAIL诱导的凋亡被抑制，而不依赖Akt状态。这项拟议的研究的基本原理是，成功完成这项研究将建立PACS-2失调和肿瘤进展之间的因果关系，并将确定TRAIL如何将PACS-2转换为一种凋亡介质，以协调线粒体通透和癌细胞死亡所需的复杂的细胞器间通讯。在强大的初步数据的指导下，这一假说将通过追求三个具体目标来检验：1)确定PACS-2的缺失或抑制在多大程度上加速肿瘤的进展；2)确定TRAIL诱导PACS-2的凋亡激活以介导ER-线粒体钙信号转导的机制；以及3)确定TRAIL如何诱导PACS-2协调溶酶体膜通透性与组织蛋白酶B介导的线粒体BID裂解。这种方法是创新的，因为它使用了一个全面的、多学科的设计来剖析TRAIL作用的分子基础。这项拟议的研究具有重要意义，因为它提供了一个新的和可测试的模型，说明PACS-2如何作为分子开关将细胞稳态与TRAIL诱导的细胞凋亡整合在一起，从而研究PACS-2失调如何加速肿瘤进展并导致对TRAIL治疗的抵抗。 公共卫生相关性：拟议的研究与公共健康相关，因为准确了解癌细胞如何对TRAIL疗法产生抗药性，以及这些知识如何反过来导致恢复TRAIL敏感性的新治疗干预措施，对于最大限度地发挥这种有前景的癌症疗法的潜力至关重要。因此，拟议的研究与NIH支持最终治愈疾病的基础研究的使命的一部分相关。

DESCRIPTION (provided by applicant): The TNF superfamily death ligand TRAIL is emerging as a potential tool in the treatment of many cancers based on its ability to selectively kill cancer cells and repress tumor metastasis. Unfortunately, many tumor cell lines are resistant to TRAIL, suggesting the molecular basis of TRAIL action and how TRAIL-sensitivity can be restored is crucial for maximizing the potential of this promising cancer therapeutic. The long-term goal is to understand precisely how cancer cells become resistant to TRAIL therapy and how this knowledge can, in turn, lead to novel therapeutic interventions that restore TRAIL-sensitivity. The objective in this particular application is to determine how TRAIL kills cancer cells by switching the multifunctional sorting protein PACS-2 to an apoptotic effector and how dysregulation of PACS-2 enables cancers to resist TRAIL-induced cell death. Specifically, Akt-phosphorylated PACS-2 mediates homeostasis in healthy cells by coordinating the localization of antiapoptotic calcium channels to the endoplasmic reticulum (ER) with ER-mitochondria communication. In response to TRAIL, PACS-2 becomes dephosphorylated, promoting mitochondria permeabilization and cell death by coordinating apoptotic calcium signaling with lysosome-mitochondria communication that mediates Bid activation. The central hypothesis is that dysregulation of PACS-2 enables cancer cells to resist TRAIL killing in two different ways: in cancers with elevated Akt, PACS-2 apoptotic activity is repressed by persistent Akt phosphorylation, whereas in cancers with loss of the PACS-2 locus, TRAIL-induced apoptosis is repressed independent of Akt status. The rationale for the proposed research is that successful completion will establish a causal relationship between PACS-2 dysregulation and tumor progression and will determine how TRAIL switches PACS-2 to an apoptotic mediator that coordinates the complex interorganellar communication required for mitochondria permeabilization and cancer cell death. Guided by strong preliminary data, this hypothesis will be test- ed by pursuing three specific aims: 1) Determine to what extent loss or repression of PACS-2 accelerates tumor progression, 2) Identify the mechanism used by TRAIL to induce apoptotic activation of PACS-2 to mediate ER-mitochondria calcium signaling, and 3) Determine how TRAIL induces PACS-2 to coordinate lysosome membrane permeabilization with cathepsin B-mediated Bid cleavage on mitochondria. The approach is innovative because it uses a comprehensive, multi-disciplinary design to dissect the molecular basis of TRAIL action. The proposed research is significant because it presents a novel and testable model of how PACS-2 acts as a molecular switch to integrate cell homeostasis with TRAIL-induced apoptosis and, therefore, how PACS-2 dysregulation may accelerate tumor progression and cause resistance to TRAIL therapy. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because understanding precisely how cancer cells become resistant to TRAIL therapy and how this knowledge can lead, in turn, to novel therapeutic interventions that restore TRAIL-sensitivity is crucial for maximizing the potential of this promising cancer therapeutic. Thus, the proposed research is relevant to the part of NIH's mission supporting fundamental research that will ultimately cure disease.