Mechanisms of Non-Canonical NF-kB Activation in Transplant Arteriosclerosis

移植动脉硬化中非典型 NF-kB 激活机制

• 批准号: 8805540
• 负责人: DANIEL JANE-WIT
• 金额: $ 13.19万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8805540
• 关键词: Accounting; Affect; Alleles; Allografting; Anaphylatoxins; Antibodies; Arteriosclerosis; Attenuated; Binding; Biopsy; Biopsy Specimen; Blood specimen; CD4 Positive T Lymphocytes; Cardiac; Cell Death; Cell membrane; Cell surface; Cells; Chronic; Clinical; Complement; Complement 3a; Complement 5a; Complement Activation; Complement Membrane Attack Complex; Complication; Coronary Vessels; Coronary artery; Deposition; Development; Diagnosis; Diagnostic tests; Diffuse; Elements; Endocytosis; Endothelial Cells; FDA approved; Generations; Genes; Goals; Heart Transplantation; High Prevalence; Human; Immune system; Immunodeficient Mouse; Implant; In Vitro; Indium; Inflammation; Inflammatory; Interferon Type II; Intervention; Kidney Transplantation; Label; Lasers; Lesion; Mediating; Mediator of activation protein; Medical; Methods; Modeling; Morbidity - disease rate; NF-kappa B; Operative Surgical Procedures; Organelles; Outcome; Pathogenesis; Pathway interactions; Patients; Pharmaceutical Preparations; Phosphorylation Site; Phosphotransferases; Predictive Value; Process; Protein Isoforms; Protein-Serine-Threonine Kinases; Proteins; RNA; Recruitment Activity; Risk Factors; Role; Sampling; Serum; Signal Transduction; Site; Small Interfering RNA; Specimen; Stenosis; Structure; T-Cell Activation; T-Lymphocyte; Testing; Therapeutic Agents; Transcript; Transplant Recipients; Transplantation; Work; base; clinically relevant; cytokine; genome-wide; graft failure; improved; in vivo; mortality; mouse model; novel; novel diagnostics; novel marker; novel therapeutics; patient oriented; peripheral blood; protein expression; renal artery; response; tool; trafficking

DESCRIPTION (provided by applicant): PROJECT SUMMARY Transplant arteriosclerosis (TA) is a highly prevalent complication of heart transplantation and is the leading cause of late graft failure. TA lesions cause diffuse, irreversible narrowing of graft coronary arteries and account for an annualized rate of graft loss of ~5% per year. There are no effective medical therapies for TA, reflecting deficits in the understanding of its underlying causes. Donor specific antibody (DSA) binding MHC on graft endothelial cells (EC) is a strong risk factor for TA, but the mechanism(s) of how DSA causes TA are not well understood. To study this problem, DSA was modeled on EC using high panel reactive antibody (PRA) sera taken from sensitized transplant candidates. PRA sera deposited DSA on EC and activated complement, a system of immune-related proteins that when activated forms pore-like structures called membrane attack complexes (MAC) on cell surfaces. PRA caused MAC assembly on EC which induced inflammatory genes and increased the ability of EC to recruit and activate CD4+ T cells producing IFN-γ, a vasculopathic cytokine. These processes had an overall effect of exacerbating TA lesions in human coronary arteries in a humanized mouse model. MAC mediated these effects through a novel effector pathway involving NF-κB-inducing kinase (NIK), a critical mediator of non-canonical NF-κB signaling. This proposal defines mechanisms of how MAC activates NIK in EC, explores how these mechanisms operate in a humanized mouse model for TA, and uses NIK in patient EC as a platform to develop novel diagnostic tests for TA. In Aim 1, endocytosis of MAC will be explored as a mechanism for NIK stabilization. MAC+ subcellular compartments will be isolated and characterized to identify novel non-canonical NF-κB signaling elements, Akt activation in MAC+ compartments will be explored as a mechanism for stabilizing NIK, and off-label use of FDA-approved drugs inhibiting endocytosis and Akt will be tested in vivo in a humanized mouse model of TA. Aim 2 explores clinical correlations between NIK and TA. The positive predictive value of NIK expression in two microvessel EC compartments will be assessed in patient samples, and novel markers correlating with TA will be identified using laser capture and RNA profiling of NIK+ EC in patient biopsy specimens. The long-term goal of this work is to improve outcomes for patients with TA. To this end the present application uses patient-oriented approaches to exploit novel mechanisms of TA to identify clinically relevant therapies and to develop diagnostic tests for this condition.

描述（由申请人提供）：项目总结移植动脉硬化（TA）是心脏移植的一种非常普遍的并发症，是晚期移植失败的主要原因。TA病变导致移植冠状动脉弥漫性、不可逆性狭窄，每年约5%的移植物丢失率。TA没有有效的医学治疗方法，反映了对其根本原因的理解不足。供体特异性 结合移植物内皮细胞（EC）MHC的抗体（DSA）是TA的重要危险因素，但DSA引起TA的机制尚不清楚。为了研究这个问题，DSA是模仿EC使用高群体反应性抗体（PRA）血清从致敏的移植候选人。PRA血清将DSA沉积在EC上并激活补体，补体是一种免疫相关蛋白质系统，当激活时在细胞表面形成称为膜攻击复合物（MAC）的孔样结构。PRA引起EC上的MAC组装，其诱导炎性基因并增加EC募集和激活产生IFN-γ（一种血管病理性细胞因子）的CD 4 + T细胞的能力。这些过程在人源化小鼠模型中具有加剧人冠状动脉中TA病变的总体效应。MAC通过一种涉及NF-κ B诱导激酶（NIK）的新型效应子途径介导这些效应，NIK是非经典NF-κB信号传导的关键介质。 该提案定义了MAC如何激活EC中的NIK的机制，探讨了这些机制如何在TA的人源化小鼠模型中起作用，并使用患者EC中的NIK作为开发TA的新诊断测试的平台。在目标1中，MAC的内吞作用将被探索为NIK稳定的机制。将分离MAC+亚细胞区室并表征以鉴定新型非经典NF-κB信号传导元件，将探索MAC+区室中的Akt活化作为稳定NIK的机制，并将在TA的人源化小鼠模型中体内测试FDA批准的抑制内吞作用和Akt的药物的标签外使用。目的2探讨NIK与TA的临床相关性。将在患者样本中评估两个微血管EC隔室中NIK表达的阳性预测值，并将使用患者活检标本中NIK+ EC的激光捕获和RNA分析来鉴定与TA相关的新标志物。这项工作的长期目标是改善TA患者的预后。为此，本申请使用面向患者的方法来开发TA的新机制，以鉴定临床相关疗法并开发用于该病症的诊断测试。