Autoantigen delivery to induce tolerance in diabetes

自身抗原递送诱导糖尿病耐受

• 批准号: 6399954
• 负责人: WILLIAM R OSBORNE
• 金额: $ 14.43万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-30 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6399954
• 关键词: autoantigens; biotechnology; cell transplantation; disease /disorder prevention /control; gene therapy; genetic strain; glutamate decarboxylase; immune tolerance /unresponsiveness; immunotherapy; insulin dependent diabetes mellitus; isozymes; laboratory rat; prediabetic state; proinsulin; technology /technique development; transfection /expression vector

DESCRIPTION (provided by applicant) Diabetes mellitus is a common disorder with a prevalence of 4-5 percent and is classified into three major forms: type 1 diabetes often referred to as immune mediated diabetes, type 2 diabetes or non-insulin dependent diabetes, and diabetes due to mutations in genes controlling beta cell function or metabolism. The effect of age is marked and type 1 diabetes dominates among children and teenagers. The etiology and pathogenesis of type 1 diabetes is strongly associated with autoreactivity to glutamic acid decarboxylase 65 (GAD65), insulin or both. This research will test the hypothesis that constitutive expression of secretable GAD65 or proinsulin will tolerize diabetes prone rats and prevent or significantly delay the onset of diabetes. As a contingency plan, should single administration of autoantigen not be effective in inducing tolerance, we will co-express both GAD65 and proinsulin from bicistronic vectors. BB rats with the lyp/lyp genotype predictably develop diabetes between 60-90 days of age permitting the delivery of islet autoantigens before disease onset to study tolerance induction and protection from diabetes. Diabetes is prevented or delayed by injecting young diabetes-prone NOD mice with either one of these two autoantigens and we have shown T cell cytokine deviation in response to GAD65 injections in the BB rat. We believe that utilizing gene therapy for the constitutive expression of autoantigens presents a significant improvement over serial administration of autoantigen by injection. We propose to investigate 2 Specific Aims. In aim 1 we will construct novel retroviral vectors encoding secretable GAD65 and proinsulin under the control of a fibronectin promoter to transduce skin fibroblasts for long-term expression from cells introduced as skin equivalent grafts to control and BB diabetic rats. Regulation of autoantigen delivery will be achieved by controlling the number of fibroblasts implanted. When tolerance has been achieved skin grafts will be removed to monitor preservation of this altered immune state in the absence of autoantigen. In aim 2 we will generate HIV-1 based lentiviral vectors expressing proinsulin or secretable rat GAD65 elements for direct intramuscular injection into prediabetic BB rats. The levels of autoantigen delivered will be controlled by variation in number of lentivirus particles administered. The overall goal of this application is the development of a gene therapy method of tolerance induction that can be applied to patients with type 1 diabetes and their high risk relatives, and may be applied to other autoimmune diseases.

描述（由申请人提供） 糖尿病是一种常见的疾病，患病率为4- 5%， 糖尿病分为三种主要形式：1型糖尿病通常称为免疫性糖尿病， 介导的糖尿病、2型糖尿病或非胰岛素依赖型糖尿病，和 由于控制β细胞功能的基因突变导致的糖尿病，或 新陈代谢.年龄的影响是显着的，1型糖尿病占主导地位， 儿童和青少年。1型糖尿病的病因和发病机制是 与对谷氨酸脱羧酶65的自身反应性密切相关 （GAD 65）、胰岛素或两者。这项研究将检验以下假设： 分泌型GAD 65或胰岛素原的组成型表达将耐受 糖尿病易感大鼠和预防或显着延迟糖尿病的发病。 作为一种应急计划，单次给予自身抗原是否不应 为了有效诱导耐受，我们将共表达GAD 65和胰岛素原两者， 双顺反子载体。具有lyp/lyp基因型的BB大鼠可预测地发育为 年龄在60-90天之间的糖尿病，允许提供胰岛 在疾病发作前检测自身抗原，以研究耐受诱导和保护 糖尿病糖尿病是预防或延迟注射年轻 糖尿病易感NOD小鼠与这两个自身抗原之一，我们有 显示了BB大鼠中响应于GAD 65注射的T细胞细胞因子偏差。 我们认为，利用基因治疗的组成型表达， 自身抗原相对于连续施用的 注射自体抗原。我们提出了两个具体目标。 目的1：构建新型的分泌型GAD 65逆转录病毒载体 和在纤连蛋白启动子控制下的胰岛素原， 用于从作为皮肤等同物引入的细胞长期表达的成纤维细胞 移植到对照和BB糖尿病大鼠。自身抗原递送的调节将 通过控制植入的成纤维细胞的数量来实现。当宽容 已经实现的皮肤移植将被删除，以监测保存这一 在自身抗原不存在的情况下改变免疫状态。 在目标2中，我们将产生表达胰岛素原的基于HIV-1的慢病毒载体 或可分泌的大鼠GAD 65元件，用于直接肌内注射到 糖尿病前期BB大鼠。递送的自身抗原水平将通过以下方式控制： 施用的慢病毒颗粒数量的变化。 这项申请的总体目标是开发一种基因疗法， 可应用于1型糖尿病患者的耐受诱导方法 糖尿病及其高危亲属，并可能适用于其他自身免疫性疾病。 疾病