Molecular Mechanisms and Treatment Of Autoimmunity In Man And Animal Models

人和动物模型中自身免疫的分子机制和治疗

• 批准号: 8555808
• 负责人: michael j lenardo
• 金额: $ 72.89万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8555808
• 关键词: Age; Animal Model; Animals; Antibodies; Antigen Targeting; Antigens; Apoptosis; Area; Autoantibodies; Autoimmune Diabetes; Autoimmune Diseases; Autoimmune Process; Autoimmunity; Beta Cell; Biological Assay; Blocking Antibodies; Blood Coagulation Factor; Blood Glucose; Cells; Cessation of life; Clinic; Clinical; Clinical Trials; Cooperative Research and Development Agreement; Detection; Development; Diabetes Mellitus; Diagnosis; Diagnostic tests; Disease; Disease model; Early treatment; Epitopes; Factor VIII; Female; Future; Gerda brand of difluprednate; Goals; Gold; Health; Human; Immune; Immune response; Immunology; Immunotherapy; Inbred NOD Mice; Incidence; Individual; Insulin; Insulin Antibodies; Insulin-Dependent Diabetes Mellitus; Intervention; Investigation; Islet Cell; Laboratories; Lymphocyte; Maryland; Measures; Mediating; Medical; Molecular; Multiple Sclerosis; Mus; Myelin Proteins; Nature; Organ; Pathogenesis; Phase; Plasma; Prediabetes syndrome; Process; Proteins; Radiolabeled; Recombinant Proteins; Recombinants; Regulation; Reproducibility; Research; Risk; Role; Sampling; Screening procedure; Severities; Signal Transduction; Specificity; Staging; System; T-Cell Immunologic Specificity; T-Cell Receptor; T-Lymphocyte; Technology; Testing; Therapeutic; Therapy Clinical Trials; Time; Translations; United Kingdom; United States National Institutes of Health; Universities; Variant; Working Women; apoptosis in lymphocytes; base; cohort; diabetic; human subject; improved; insight; instrument; islet; man; mouse model; non-diabetic; prevent; programs; radiotracer; response; success; tool; type I diabetic

We are studying the cellular and molecular basis of autoimmune diseases with two purposes. First, we want to understand the pathogenic role and antigen-specificity of T cells that cause autoimmune diseases such as multiple sclerosis, clotting factor inhibition, insulin-dependent diabetes, among others. Second, we would like to test specific antigen-induced apoptosis as a means of treating such autoimmune diseases. To these ends, we have made progress in the following areas: 1) we have reinitiated studies of recombinant molecules containing antigens potentially involved in multiple sclerosis with the goal of establishing a Cooperative Research and Development Agreement to test such a form of therapy in a clinical trial. At present there is increasing evidence that myelin proteins antigens are the target of the autoimmune attack. By programmed the T cells that recognize such antigens to die, the effect of eliminating these cells on the disease can be demonstrated. 2) We are studying new highly sensitive diagnostic tests to detect end organ damage during autoimmune diseases to determine if these can provide an early warning system of autoimmune attack; and 3) we are initiating studies of antigen-specific therapy to prevent the formation of blocking antibodies following factor VIII administration to hemophiliacs. These studies will employ new recombinant proteins constructed to contain the principal epitopic regions of Factor VIII to which T cells react. We will also be initiating studies in experimental animals of other autoimmune conditions. In particular, we are focusing on Type I diabetes mellitus and have been studying immune responses against insulin as a harbinger of disease in prediabetic mice and humans. The prediabetic state, known as insulitis, involves both cellular and numeral responses against the islet cells with insulin as the primary antigen. The focused nature of the immune response, which precedes any evident epitope spreading, may allow the use of insulin or congeners thereof as a therapeutic entity. As part of these studies we are trying to understand the molecular regulation of antigen-induced death by T cell receptor stimulation. The deployment of a highly sensitive early warning system as a screening tool to identify individuals with early immune-mediated organ damage with early intervention using antigen-specific treatment approaches, we hope to provide targeted therapy to minimize end-organ damage and clinical disease. to this end, we have prepared extremely sensitive electrochemiluminescence assays that can sensitively and specifically detect FVIII antibodies and insulin autoantibodies. This could better dispose our efforts to intervene early successfully. We believe these investigations will provide important new insights into the pathogenesis of autoimmune diseases and hopefully stimulate the development of new forms of highly specific immune therapy. In 2012, we have made notable progress in developing the test for insulin autoantibodies (IAA). The detection of IAA aids in the prediction of autoimmune diabetes development. However, the long-standing, gold standard 125I-insulin radiobinding assay (RBA) has low reproducibility between laboratories, long sample processing times and requires the use of newly synthesized radiolabeled insulin for each set of assays. Therefore, a rapid, non-radioactive, and reproducible assay is of great medical important. We have developed electrochemiluminescence (ECL)-based assays that over come these deficiencies that can measure IAA and anti-insulin antibodies (IA) in non-obese diabetic (NOD) mice and in type 1 diabetic individuals, respectively. Using the murine IAA ECL assay, we correlated IAA, histopathological insulitis, and blood glucose in a cohort of female NOD mice from 4 up to 36 weeks of age. We found that our human IA ECL assay compared favorably to conventional RBA and validated using samples from 34 diabetic and 59 non-diabetic individuals in three independent laboratories. The ECL assay technology was rapid and sensitive with a broad dynamic range and low background. In the NOD mouse model, IAA ECl signal was positively correlated with insulitis severity, and positive ECL values measured at 8-10 weeks of age were predictive of diabetes onset at 20 weeks of age. Using human serum and plasma samples, our IA ECL assay yielded reproducible and accurate results with an average sensitivity of 84% at 95% specificity. We carried out the assay with identical samples using instruments at the Wellstat company headquarters in Gaithersburg, Maryland, at the NIH in Bethesda, Maryland, and in the diabetes immunology laboratory at the University of Cambridge in the United Kingdom and found no statistically significant variation between laboratories. We concluded that highly sensitive, non-radioactive ECL-based assays should facilitate reliable and fast detection of antibodies to insulin and its precursors sera and plasma in a standardized manner between laboratories in both research and clinical settings. Our next step is to evaluate the human IA assay in the detection of IAA in prediabetic human subjects or those who harbor features that confer risk of type 1 diabetes and to develop similar assays for other autoantibodies directed at other islet beta cell antigens that together are predictive for the diagnosis of this common disorder, in order to improve prediction and facilitate future therapeutic trials. Given that the incidence of type I diabetes has been increasing over the last several decades, these tests might permit new interventions to prevent or forestall the development of this disease.

我们正在研究具有两个目的的自身免疫性疾病的细胞和分子基础。首先，我们想了解引起自身免疫性疾病的T细胞的致病作用和抗原特异性，例如多发性硬化症，凝血因子抑制，胰岛素依赖性糖尿病等。其次，我们想测试特定的抗原诱导的凋亡，作为治疗这种自身免疫性疾病的一种手段。对于这些目的，我们在以下领域取得了进展：1）我们对可能参与多发性硬化症的抗原的重组分子进行了重组研究，目的是建立合作研究和发展协议，以在临床试验中测试这种治疗形式。目前，有越来越多的证据表明髓磷脂蛋白抗原是自身免疫攻击的靶标。 通过对识别这种抗原死亡的T细胞进行编程，可以证明消除这些细胞对疾病的影响。 2）我们正在研究新的高度敏感诊断测试，以检测自身免疫性疾病期间的最终器官损伤，以确定这些测试是否可以提供自身免疫性攻击的预警系统； 3）我们正在启动对抗原特异性疗法的研究，以防止在对血友病的VIII因子给药后形成阻塞抗体。这些研究将采用新的重组蛋白，这些蛋白质构建，以包含T细胞反应的VIII因子的主要表现区域。我们还将在其他自身免疫性条件的实验动物中进行研究。特别是，我们专注于I型糖尿病，并一直在研究针对胰岛素的免疫反应，作为糖尿病前小鼠和人类疾病的预兆。 糖尿病前期被称为胰岛素，涉及针对胰岛素作为主要抗原的胰岛细胞的细胞和数字反应。在任何明显的表位扩散之前，免疫反应的重点性质可能允许使用胰岛素或同类物作为治疗实体。 作为这些研究的一部分，我们试图了解T细胞受体刺激对抗原诱导死亡的分子调节。 通过使用抗原特异性治疗方法，通过早期干预识别具有早期免疫介导器官损害的个体的筛查工具，将高度敏感的预警系统部署为筛查工具，我们希望提供有针对性的治疗，以最大程度地减少最终器官损伤和临床疾病。为此，我们准备了极灵敏的电化学测定法，可以敏感，特异性地检测FVIII抗体和胰岛素自身抗体。这可以更好地消除我们成功进行介绍的努力。我们认为，这些研究将为自身免疫性疾病的发病机理提供重要的新见解，并希望刺激新形式的高度特定免疫治疗。 在2012年，我们在开发胰岛素自身抗体（IAA）测试方面取得了显着进展。 IAA的检测有助于预测自身免疫性糖尿病的发展。然而，长期的黄金标准125i胰岛素放射性固定测定法（RBA）在实验室之间具有较低的可重复性，长期样本处理时间，需要在每种测定方面使用新合成的放射性标记的胰岛素。因此，快速，非放射性和可再现的测定法至关重要。我们已经开发了基于电化学发光（ECL）的测定法，这些分别可以在非肥胖糖尿病（NOD）小鼠和1型糖尿病患者中测量IAA和抗胰岛素抗体（IA）的缺陷。使用鼠IAA ECL测定法，我们将IAA，组织病理学胰岛炎和血糖相关联在一组雌性小鼠的同类中，从4至36周龄。我们发现，我们的人类IA ECL分析与常规RBA进行了比较，并使用来自三个独立实验室中34个糖尿病和59个非糖尿病个体的样品进行了验证。 ECL测定技术迅速而敏感，动态范围较大，背景较低。在NOD小鼠模型中，IAA ECL信号与胰岛炎的严重程度呈正相关，并且在8-10周龄时测得的正ECL值可预测20周龄时的糖尿病发作。使用人血清和血浆样品，我们的IA ECL分析得出了可再现和准确的结果，平均灵敏度为84％，在95％的特异性下。我们在马里兰州盖瑟斯堡的Wellstat Company总部，马里兰州贝塞斯达的NIH以及英国剑桥大学的糖尿病免疫学实验室中，使用相同的样品进行了测定，并在实验室之间没有统计学意义。我们得出的结论是，高度敏感的，非放射性ECL的测定应促进对胰岛素及其前体的可靠，快速检测，以研究和临床环境之间的实验室之间的标准化方式进行血清和血浆。我们的下一步是在糖尿病前受试者中检测IAA或具有赋予1型糖尿病风险的人的IAA时评估人类IA测定法，并为其他针对其他小岛β细胞抗原的自身抗体开发类似的测定法，从而可以预测这种共同的无序，以诊断出这种常见的诊断，以提高预测和面对面的预测和面对面的试验。鉴于在过去的几十年中，I型糖尿病的发生率一直在增加，因此这些测试可能允许采取新的干预措施，以防止或阻止这种疾病的发展。