Regulation of TRAIL Induced Apoptosis in Cancer Cells

描述（由申请人提供）：TNF超家族死亡配体TRAIL基于其选择性杀死癌细胞和抑制肿瘤转移的能力，正在成为治疗许多癌症的潜在工具。不幸的是，许多肿瘤细胞系对TRAIL具有耐药性，这表明TRAIL作用的分子基础以及如何恢复TRAIL敏感性对于最大限度地发挥这种有前景的癌症治疗的潜力至关重要。长期目标是精确地了解癌细胞如何对TRAIL治疗产生耐药性，以及这些知识如何反过来导致新的治疗干预措施，恢复TRAIL敏感性。这项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转换为凋亡效应物来杀死癌细胞，以及PACS-2的失调如何使癌症抵抗TRAIL诱导的细胞死亡。具体来说，akt磷酸化的PACS-2通过协调抗凋亡钙通道定位到内质网（ER）和内质网-线粒体的通讯来调节健康细胞的内稳态。在TRAIL的响应下，PACS-2发生去磷酸化，通过协调凋亡钙信号和介导Bid激活的溶酶体-线粒体通讯，促进线粒体通透性和细胞死亡。核心假设是，PACS-2的失调使癌细胞能够以两种不同的方式抵抗TRAIL的杀伤：在Akt升高的癌症中，PACS-2的凋亡活性受到Akt持续磷酸化的抑制，而在PACS-2位点缺失的癌症中，TRAIL诱导的凋亡受到抑制，与Akt状态无关。这项研究的基本原理是，成功完成将建立PACS-2失调与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，协调线粒体通透性和癌细胞死亡所需的复杂的细胞器间通讯。在强有力的初步数据的指导下，这一假设将通过三个具体目标进行验证：1)确定PACS-2的缺失或抑制在多大程度上加速了肿瘤进展；2)确定TRAIL诱导PACS-2凋亡激活介导er线粒体钙信号传导的机制；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切割。该方法是创新的，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。这项研究具有重要意义，因为它提出了一个新的、可测试的模型，说明PACS-2如何作为一个分子开关，将细胞稳态与TRAIL诱导的细胞凋亡结合起来，因此，PACS-2失调如何加速肿瘤进展并导致对TRAIL治疗的耐药性。

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.