RIPK-dependent necrosis in development and cancer

发育和癌症中的 RIPK 依赖性坏死

• 批准号: 8856171
• 负责人: DOUGLAS R GREEN
• 金额: $ 36.31万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-06 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8856171
• 关键词: 2q33; APAF1 gene; Ablation; Achievement; Animals; Apoptosis; Apoptotic; Appearance; Binding; Caspase; Cell Death; Cells; Cessation of life; Complex; Cysteine; Defect; Development; Elements; Embryo; Employee Strikes; Failure; Genetic; Genetic Transcription; Hematopoietic; Histocompatibility Testing; Homeostasis; Human; Inflammatory; Knock-out; Lead; Life; Ligands; Ligation; Liver; Malignant Neoplasms; Mediating; Mitochondria; Modeling; Mus; Necrosis; Neuroblastoma; Pathway interactions; Peptide Hydrolases; Phenotype; Phosphotransferases; Process; RIPK1 gene; RIPK3 gene; Regulation; Resistance; Role; Signal Pathway; Signal Transduction; TNFRSF6 gene; Testing; Tissues; Tumor Immunity; Tumor Suppressor Proteins; Vascular Endothelium; Vascularization; Yolk Sac; adapter protein; base; cancer therapy; caspase-3; caspase-8; caspase-9; cell type; human disease; in vivo; lung small cell carcinoma; mature animal; neoplastic cell; prevent; progenitor; programs; protein function; receptor; tumor; tumorigenesis

DESCRIPTION (provided by applicant): While the role of caspases in apoptosis is well established, little is known about the role of these proteases in the process of programmed necrosis. This application is based on startling findings that show that embryonic lethality as a result of ablation of caspase-8 or its adapter protein, FADD, is fully rescued by deletion of RIPK3, a kinase required for programmed necrosis. Our studies indicate that a complex of FADD, caspase-8, and FLIP, a caspase-like molecule that lacks a catalytic cysteine, protects against RIPK-dependent necrosis. This is further supported by our finding that the FADD-FLIP-RIPK3 TKO mouse develops normally. We propose the following studies to delineate the functions of these proteins in development and cancer: Aim 1. What is the developmental target protected by the FADD-caspase-8-FLIPL complex? The phenotypes of the caspase- 8, FADD, and FLIPL knockouts all show embryonic lethality around e10.5, associated with a defect in yolk sac vascularization. In this aim, we will test the idea that early progenitors of vascular endothelium and hematopoietic cells serve as the earliest and most important targets of this developmental defect. In so doing, we will identify additional targets of RIPK3-necrosis and investigate the signaling pathways engaged in this embryonic lethality. Aim 2. How is RIPK-dependent necrosis regulated in oncogenesis? Caspase-8, which in humans is present on 2q33, is often silenced or deleted in human neuroblastoma, small cell lung carcinoma, and other cancers. This represents a paradox, however, as such loss in many cell types sensitizes cells to RIPK-dependent necrosis. Here we explore how loss of caspase-8 can fail to sensitize tumor lines to RIPK-dependent necrosis. Our studies include how RIPK3 transcription is controlled in primary and transformed tissues and the role of RIPK1 and the tumor suppressor, CYLD, in controlling RIPK-dependent necrosis. Aim 3. Does RIPK-dependent necrosis represent a potential avenue for therapy? Many approaches to cancer therapy seek to promote apoptosis, which may or may not promote ancillary anti-tumor immunity. By shifting signals to RIPK-necrosis we may a) prevent iatrogenic damage in tissues resistant to this form of death (e.g., liver) while b) promoting an inflammatory mode of tumor cell death. We will model "pure" RIK3-induced necrosis versus apoptosis to examine the anti-tumor consequences, and will explore a counter- intuitive approach to triggering RIPK-dependent necrosis in autochthonous and grafted tumors by death receptor ligation in vivo. The possibility that tumor neo-vasculature is targeted (based on considerations from Aim 1) will also be explored. These studies provide a number of tests and explorations of the new model we propose, and hold the potential to greatly increase of understanding of this fundamental process controlling life cell and death, both in normal development and in cancer.

描述（由申请人提供）：虽然半胱天冬酶在细胞凋亡中的作用已得到充分证实，但对这些蛋白酶在程序性坏死过程中的作用知之甚少。该应用基于令人吃惊的发现，该发现表明，由于半胱天冬酶-8或其衔接蛋白FADD的消融而导致的胚胎致死性通过RIPK 3（程序性坏死所需的激酶）的缺失而被完全挽救。我们的研究表明，FADD，caspase-8和FLIP（一种缺乏催化半胱氨酸的caspase样分子）的复合物可防止RIPK依赖性坏死。我们发现FADD-FLIP-RIPK 3 TKO小鼠发育正常，这进一步支持了这一点。我们提出以下研究来描述这些蛋白质在发育和癌症中的功能：目的1。FADD-caspase-8-FLIPL复合物保护的发育靶点是什么？caspase- 8、FADD和FLIPL基因敲除的表型均显示在e10.5左右的胚胎致死率，与卵黄囊血管化缺陷相关。在这个目标中，我们将测试的想法，血管内皮细胞和造血细胞的早期祖细胞作为最早和最重要的目标，这种发展缺陷。在此过程中，我们将确定RIPK 3坏死的其他靶点，并研究参与这种胚胎致死性的信号通路。目标二。RIPK依赖性坏死在肿瘤发生中是如何调节的？Caspase-8在人类中存在于2 q33上，在人类神经母细胞瘤、小细胞肺癌和其他癌症中经常沉默或缺失。然而，这代表了一个矛盾，因为许多细胞类型的这种损失使细胞对RIPK依赖性坏死敏感。在这里，我们探讨了caspase-8的丢失如何不能使肿瘤细胞对RIPK依赖性坏死敏感。我们的研究包括RIPK 3的转录是如何在原发性和转化组织中控制的，以及RIPK 1和肿瘤抑制因子CYLD在控制RIPK依赖性坏死中的作用。目标3。RIPK依赖性坏死是否是一种潜在的治疗途径？许多癌症治疗方法寻求促进细胞凋亡，这可能会或可能不会促进辅助抗肿瘤免疫。通过将信号转移到RIPK-坏死，我们可以a）防止抵抗这种形式死亡的组织中的医源性损伤（例如，肝），而B）促进肿瘤细胞死亡的炎性模式。我们将对“纯”RIK 3诱导的坏死与凋亡进行建模，以检查抗肿瘤结果，并将探索通过体内死亡受体连接在自体和移植肿瘤中触发RIK 3依赖性坏死的反直觉方法。还将探索靶向肿瘤新生血管的可能性（基于目标1的考虑）。这些研究为我们提出的新模型提供了许多测试和探索，并有可能大大增加对正常发育和癌症中控制生命细胞和死亡的基本过程的理解。