Interrogating the ubiquitin pathway to understand and treat cytokine-induced beta-cell death in type 1 diabetes

探究泛素通路以了解和治疗 1 型糖尿病中细胞因子诱导的 β 细胞死亡

• 批准号: 10477373
• 负责人: ROHIT N. KULKARNI
• 金额: $ 51.69万
• 依托单位: BROAD INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-30 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10477373
• 关键词: Alberta province; Apoptosis; Area; Autoimmune Diabetes; Autoimmune Diseases; Beta Cell; Binding; Biology; Cell Death; Cell Line; Cell Survival; Cell physiology; Cellular biology; Chemicals; Clinical; Complex; Complications of Diabetes Mellitus; Cytokine Signaling; Development; Diabetes Mellitus; Diabetic mouse; Disease; Equilibrium; Event; Gene Expression Profiling; Genetic; Goals; Heart; Human; Hypoglycemia; Immune; Immune Targeting; In Vitro; Individual; Inflammatory; Insulin; Insulin-Dependent Diabetes Mellitus; Interferon Type II; Interleukin-1 beta; JAK2 gene; Kidney; Label; Lead; Lysine; Measurable; Measurement; Mediating; Methods; Modeling; Morbidity - disease rate; Mus; Mutate; Non obese; Outcome; Pancreas; Pathway interactions; Patients; Peripheral Nerves; Phenotype; Phosphorylation; Phosphotransferases; Post-Translational Protein Processing; Process; Proteins; Proteomics; Retina; Risk; Role; STAT1 gene; Signal Transduction; Structure of beta Cell of islet; TNF gene; Therapeutic; Tyrosine; Ubiquitin; Ubiquitination; Universities; activity-based protein profiling; analog; cellular targeting; chemoproteomics; cytokine; early detection biomarkers; improved; in vivo; innovation; islet; kinase inhibitor; knock-down; mortality risk; mouse model; novel therapeutic intervention; preservation; prevent; response; screening; small molecule; tool

PROJECT SUMMARY The loss of insulin-producing beta cells in the pancreas results in an absolute requirement for injected insulin, causing significant risks of mortality from hypoglycemia and morbidity from diabetic complications in peripheral nerves, the retina, the heart, and the kidney. A key goal of efforts to treat T1D is to stop this cellular attack, either by halting the immune mis-recognition of beta cells or by protecting beta cells from cell death. However, a critical barrier to progress in the field is a lack of complete understanding of the cellular events in the islet that contribute to the loss of beta-cell mass. Using a phenotypic screening approach, we discovered BRD0476, a compound that is selectively active against cytokine-mediated apoptosis. Further study of this compound revealed that it binds the deubiquitinase USP9X to halt JAK2 and STAT1 signaling in response to IFNγ. We determined that JAK2 can be rendered signaling incompetent by ubiquitination, and that by modulating USP9X, we can tip the balance toward reduced JAK2 kinase activity, even in the presence of IFNγ. These results point to an emerging role for ubiquitination in regulating beta-cell apoptosis in T1D, and suggest that a greater understanding of this process (and its potential dysregulation) in the early stages of T1D development could lead to 1) the ability to identify at-risk individuals, and 2) novel therapeutic strategies to preserve beta-cell mass in early-stage T1D. Using our probe BRD0476 and chemical biology tools not previously applied to islet biology, we will improve our understanding of the role of USP9X in beta-cell survival in vitro and in vivo through the following aims: In Aim 1, we will characterize the mode of JAK2 inhibition by USP9X in human islets. In Aim 2, we will assess effects of inhibiting USP9X-JAK2 (with BRD0476) on development and progression of autoimmune diabetes in a mouse model of type 1 diabetes. In Aim 3, we will profile deubiquitinase (DUB) expression and activity in human islets during early T1D development, using activity-based protein profiling (ABPP) and global ubiquitome measurements. The successful outcomes of this proposal are 1) a greater understanding of mechanisms to promote beta-cell survival in early T1D, and 2) a chemical probe to provide translational proof-of-concept. This project will set the stage for developing a biomarker of early-stage T1D development, as well as advanced therapeutic strategies for preventing beta-cell apoptosis in early-stage T1D, representing a potentially curative approach.

项目摘要 胰腺中产生胰岛素β细胞的丧失导致对注射胰岛素的绝对需求， 降低血糖和周围糖尿病并发症的发病率引起死亡的重大风险 神经，视网膜，心脏和肾脏。治疗T1D努力的关键目标是停止这种细胞攻击 通过停止对β细胞的免疫识别或保护β细胞免受细胞死亡的影响。但是，这是一个批判 该领域进步的障碍是缺乏对胰岛中蜂窝事件的完全理解 损失β细胞质量。使用表型筛选方法，我们发现了BRD0476，一种化合物 对于细胞因子介导的细胞凋亡，这是有选择性的。对这种化合物的进一步研究表明 响应于IFNγ，将去泛素酶USP9X与HALT JAK2和STAT1信号结合。我们确定了这一点 JAK2可以通过泛素化使信号传导无能，并且通过调节USP9X，我们可以向 即使在IFNγ存在下，平衡降低了JAK2激酶活性。这些结果指向新兴 泛素化在调节T1D中β细胞凋亡中的作用，并提出对此有更大的了解 在T1D开发的早期阶段，过程（及其潜在失调）可能导致1） 确定危险的个体，以及2）在早期T1D中保留β细胞质量的新型治疗策略。 使用我们的探针BRD0476和化学生物学工具以前不应用于胰岛生物学，我们将改善我们的 通过以下目的了解USP9X在体外和体内生存中的作用：在AIM 1中， 我们将表征USP9X在人类胰岛中抑制JAK2的模式。在AIM 2中，我们将评估 抑制USP9X-JAK2（具有BRD0476）对小鼠自身免疫性糖尿病的发育和进展 1型糖尿病的模型。在AIM 3中，我们将在人类胰岛中介绍去泛素酶（DUB）的表达和活性 在T1D早期开发期间，使用基于活动的蛋白质分析（ABPP）和全局泛素 测量。该提议的成功结果是1）对机制的更深入了解 在T1D早期促进β细胞存活，以及2）一种化学探针，以提供转化概念验证。 项目将为开发早期T1D开发的生物标志物奠定舞台，并高级 预防早期T1D中β细胞凋亡的治疗策略，代表了潜在的治疗方法 方法。