LUPUS ANTICOAGULANTS AND THE PROTEIN C PATHWAY

狼疮抗凝剂和蛋白质 C 途径

• 批准号: 6302361
• 负责人: NAOMI L ESMON
• 金额: $ 22.97万
• 依托单位: UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-15 至 2001-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6302361
• 关键词: antiantibody; anticoagulants; chemical binding; chimeric proteins; clinical research; coagulation factor V; disease /disorder proneness /risk; enzyme linked immunosorbent assay; human tissue; liposomes; molecular pathology; monoclonal antibody; phosphatidylethanolamines; platelets; polymerase chain reaction; protein C; prothrombin; systemic lupus erythematosus; thrombomodulin; venous thrombosis

Lupus anticoagulants and antiphospholipid antibodies (APAs) are associated with an increased incidence of thrombosis. Although a causal relationship has not been established, the correlation suggests that these antibodies contribute directly to the thrombotic process. The protein C pathway has been proposed by many, including ourselves, as a possible target for involvement of these APAs. Using a newly developed ELISA assay, we have found that a relatively high percentage of APA patients (20/61) and patients with unexplained thrombosis (20/200) have an antithrombomodulin (TM) antibody at least some of which inhibit protein C activation. Furthermore, our recent finding that optimal activated protein C (APC) anticoagulant activity, but not procoagulant activity, requires the presence of phosphatidylethanolamine (PE) in the membranes provides a potential mechanism to explain selective reduction in APC activity by antibodies that bind to membrane surfaces or protein- membrane complexes. Patients with APAs have been identified whose plasma inhibits APC activity to a greater extent than prothrombin activation and do so in a PE dependent fashion. Preliminary screening suggests that high levels of anti APC activity in APA patients is associated with a history of thrombosis. The goals of the present application are to determine the prevalence of anti-APC and anti-TM activity in APA patients with and without histories of thrombosis and to obtain a molecular understanding of how PE facilitates APC function and participates in APA inhibition of APC activity. This will aid in the design of better assays for detecting APAs with anti-APC activity. The molecular basis of PE function will be approached by determining the nature of PE involvement in facilitating APC binding interactions, by mapping the sites in APC responsible for PE dependent membrane interaction and by determining the mechanisms of inhibition (i.e., direct binding to membranes and the influence of membrane composition vs. binding to protein C or protein S-membrane complexes and whether the antibodies selectively displace APC or protein S), and by using chimeric proteins to map the antigenic determinants. The ability of APAs to inhibit platelet mediated factor Va inactivation vs. prothrombin activation will be examined. Monoclonal antibodies from APA patient B cells will be prepared to study their mechanism of action and potential thrombogenicity. These studies should provide information on the mechanisms by which APAs function, may allow initial insights into which specificities contribute directly to thrombogenicity, and may provide improved assays to monitor the effectiveness of therapeutic approaches.

狼疮抗凝剂和抗磷脂抗体（APA）为 与血栓形成的发生率增加有关。虽然是因果关系 关系尚未建立，相关性表明 这些抗体直接有助于血栓形成过程。 这 包括我们自己在内的许多人都提出了蛋白质C途径 这些APA参与的可能目标。使用新开发 ELISA分析，我们发现APA比例较高 患者（20/61）和无法解释的血栓形成患者（20/200）患有 一种抗鲜活蛋白（TM）抗体至少一些抑制 蛋白C激活。此外，我们最近的发现 活化的蛋白C（APC）抗凝剂活性，但没有凝聚剂 活性需要在磷脂酰乙醇胺（PE）中 膜提供了解释选择性减少的潜在机制 在APC活性中通过与膜表面或蛋白质结合的抗体 膜络合物。已经鉴定出患有APA的患者血浆 抑制APC活性的程度大于凝血酶原激活和 以体育依赖的方式进行。初步筛查表明高 APA患者的抗APC活性水平与病史有关 血栓形成。本应用的目标是确定 APA患者的抗APC和抗TM活性的患病率 没有血栓形成的历史并获得分子理解 PE如何促进APC功能并参与APA抑制 APC活动。这将有助于设计更好的测定 具有抗APC活性的APA。 PE功能的分子基础将是 通过确定PE参与的性质来促进 APC绑定相互作用，通过映射负责PE的APC中的位点 依赖性膜相互作用并通过确定的机制 抑制（即直接结合与膜的结合和 膜组成与与蛋白C或蛋白S-膜结合的结合 复合物以及抗体是否有选择地置换APC或蛋白质 S），并使用嵌合蛋白来绘制抗原决定因素。这 APA抑制血小板介导的因子VA失活VS的能力。 将检查凝血酶原激活。 APA的单克隆抗体 患者B细胞将准备研究其作用机理，并 潜在的血栓形成。这些研究应提供有关 APA功能的机制可能允许初始见解 哪些特异性直接有助于血栓形成，并且可能 提供改进的测定法以监测治疗的有效性 方法。