LUPUS ANTICOAGULANTS AND THE PROTEIN C PATHWAY

狼疮抗凝剂和抗磷脂抗体(APA)是 与血栓形成的发病率增加有关。虽然这是一个因果关系 关系尚未建立，相关关系表明 这些抗体直接促进血栓形成过程。这个 蛋白C途径已经被许多人提出，包括我们自己，作为一种 可能是这些APA参与的目标。使用一种新开发的 酶联免疫吸附试验，我们发现有较高比例的APA 患者(20/61)和不明原因血栓形成患者(20/200) 抗血栓调节蛋白(TM)抗体，至少部分抗体抑制 蛋白C的激活。此外，我们最近的发现是最佳的 活化蛋白C(APC)的抗凝血活性，但不是促凝剂 活性，需要磷脂酰乙醇胺(PE)存在于 膜为解释选择性还原提供了一种潜在的机制 结合在膜表面或蛋白质上的抗体的APC活性- 膜复合体。APAS患者已被确认其血浆 对APC活性的抑制程度大于凝血酶原激活和 以一种依赖PE的方式做到这一点。初步筛查显示， APA患者抗APC活性水平与病史相关 血栓形成的可能性。本申请的目标是确定 伴AND的APA患者抗APC和抗TM活性的患病率 没有血栓病史，并获得分子上的了解 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检测方法 具有抗APC活性的APAs。PE功能的分子基础将是 通过确定体育参与促进的性质来探讨 通过映射APC中负责PE的位置来实现APC绑定交互 依赖的膜相互作用和通过确定的机制 抑制(即与膜的直接结合和 膜成分与结合蛋白C或蛋白S的比较-膜 以及抗体是否选择性地取代APC或蛋白质 S)，并通过使用嵌合蛋白来定位抗原决定簇。这个 APAs抑制血小板介导的因子Va失活的能力与 将检查凝血酶原激活情况。来自APA的单抗 患者B细胞将准备研究它们的作用机制和 潜在的血栓形成能力。这些研究应该提供关于 APA的运作机制，可能会让人们对 哪些特异性直接导致血栓形成，并可能 提供改进的分析方法以监测治疗的有效性 接近了。

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.