RIPK-dependent necrosis in development and cancer

发育和癌症中的 RIPK 依赖性坏死

• 批准号: 8345283
• 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 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8345283
• 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. PUBLIC HEALTH RELEVANCE: Cell death is crucial for normal homeostasis, and defects in this process underlie many human diseases. This project explores how cell death is controlled at the level of precise molecular interactions, amenable to pharmacologic manipulation, testing a new model of this process in development and cancer.

描述(由申请人提供)：虽然caspase在细胞凋亡中的作用已得到很好的证实，但对这些蛋白水解酶在程序性坏死过程中的作用知之甚少。这一应用是基于令人震惊的发现，这些发现表明，由于切除caspase-8或其适配蛋白FADD而导致的胚胎死亡，可以通过缺失RIPK3完全挽救，RIPK3是程序性坏死所需的一种激酶。我们的研究表明，FADD、caspase-8和flip(一种缺乏催化半胱氨酸的类似caspase的分子)的复合体可以预防RIPK依赖的坏死。我们发现FADD-Flip-RIPK3 TKO小鼠发育正常，进一步支持了这一点。我们建议进行以下研究来描述这些蛋白在发育和癌症中的功能：目的1.FADD-caspase-8-FLIPL复合体保护的发育靶点是什么？Caspase-8、FADD和Flipl基因敲除的表型都显示出胚胎在e10.5附近的致死性，与卵黄囊血管形成缺陷有关。在这个目标中，我们将测试早期的血管内皮细胞和造血细胞前体细胞是这种发育缺陷的最早和最重要的靶点的想法。在这样做的过程中，我们将确定RIPK3-坏死的其他靶点，并研究参与这种胚胎死亡的信号通路。目的2.RIPK依赖的坏死在肿瘤发生中是如何调节的？Caspase-8在人类中存在于2q33，在人类神经母细胞瘤、小细胞肺癌和其他癌症中经常被沉默或缺失。然而，这是一个悖论，因为许多细胞类型中的这种丢失使细胞对RIPK依赖的坏死敏感。在这里，我们探讨了caspase-8的缺失如何不能使肿瘤细胞对RIPK依赖的坏死敏感。我们的研究包括RIPK3在原发组织和转化组织中的转录是如何控制的，以及RIPK1和肿瘤抑制因子CyLD在控制RIPK依赖的坏死中的作用。目的3.依赖RIPK的坏死是否代表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，这可能会也可能不会促进辅助的抗肿瘤免疫。通过将信号转移到RIPK-坏死，我们可以a)防止抵抗这种形式的死亡的组织(例如肝脏)中的医源性损伤，同时b)促进肿瘤细胞死亡的炎症模式。我们将模拟“纯”RIK3诱导的坏死与凋亡，以检查抗肿瘤的后果，并将探索一种反直觉的方法，通过体内死亡受体结扎在自体和移植肿瘤中触发RIPK依赖的坏死。还将探讨肿瘤新生血管作为靶点的可能性(基于目标1的考虑)。这些研究为我们提出的新模型提供了一些测试和探索，并有可能极大地提高对这一控制生命细胞和死亡的基本过程的理解，无论是在正常发育还是在癌症中。 与公共卫生相关：细胞死亡对正常的动态平衡至关重要，而这一过程中的缺陷是许多人类疾病的基础。这个项目探索了如何在精确的分子相互作用水平上控制细胞死亡，服从于药物操作，在发育和癌症中测试这一过程的新模型。