Mechanisms of Coagulation Factor XII Recruitment and Activation at the Platelet Surface

血小板表面凝血因子 XII 招募和激活的机制

基本信息

批准号：
10424806
负责人：
Pavan Bendapudi
金额：
$ 8.65万
依托单位：
BETH ISRAEL DEACONESS MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10424806
关键词：
Animal Model Binding Biological Assay Blood Platelets Cell Line Clinical Co-Immunoprecipitations Coagulation Process Competitive Binding Complication Development Disease susceptibility Drug Targeting Enzyme Precursors Event Exhibits Factor XII Factor XII Deficiency Factor XIIa Fibrinolytic Agents Flow Cytometry Generations Hemorrhage Hemostatic function Human ITGB3 gene Integrin Binding Integrins Kinetics Knockout Mice Length Maps Mass Spectrum Analysis Membrane Proteins Molecular Molecular Conformation Pathway interactions Patients Phase Physiological Plasma Play Pre-Clinical Model Process Property Proteomics RGD (sequence)Reagent Recombinants Research Personnel Risk Role Signal Transduction Site Solid Specificity Surface Surface Plasmon Resonance System Testing Therapeutic Thrombosis Treatment Efficacy Wild Type Mouse Work base experimental study factor A follow-up in vivo insight interest member new therapeutic target novel novel strategies novel therapeutics pre-clinical reaction rate receptor recruit treatment strategy vascular injury

项目摘要

PROJECT SUMMARY All currently available antithrombotic medications carry the risk of hemorrhage, an often devastating complication in many patients that can rapidly reverse the benefits of therapy. Given this reality, coagulation factor XII (FXII) has emerged as a promising new drug target that could potentially transform antithrombotic therapy. Blockade or deletion of FXII in preclinical animal models has consistently been shown to be protective against thrombosis, yet severe congenital FXII deficiency is not associated with a bleeding diathesis. Therefore, inhibiting FXII could represent the long-sought means to “decouple” hemostasis from thrombosis and achieve antithrombotic efficacy without bleeding complications. However, despite its potential clinical importance, little is known about the mechanisms underlying platelet-dependent FXII activation in vivo. Using a mass spectrometry proteomic screen, we have identified integrin αIIbβ3 as the putative platelet receptor for FXII. These results have been followed up with a number of additional studies reproducibly demonstrating the FXII-integrin αIIbβ3 interaction. Our central hypothesis is that FXII zymogen exhibits specific binding to integrin αIIbβ3 on the platelet surface, which enhances its proteolytic activity and is necessary and sufficient for FXII-dependent coagulation. In Aim 1 of this proposal, we will map the region(s) of FXII responsible for binding to integrin αIIbβ3 via competitive inhibition assays using recombinantly-generated fragments of FXII. We will also utilize surface plasmon resonance to evaluate the specificity of the FXII- integrin αIIbβ3 interaction and obtain binding kinetics. In Aim 2, we will explore the role of integrin αIIbβ3 binding in the activation of FXII using integrin αIIbβ3-coated beads and platelets from wild- type and integrin β3 (ITGB3)-null mice. The proposed work will provide important new insights into the molecular mechanism of FXII recruitment, activation, and propagation at the platelet surface and inform efforts to develop novel therapeutics based on FXII inhibition.

项目概要目前所有可用的抗血栓药物都有出血的风险，这是一种常见的情况许多患者出现了毁灭性的并发症，可以迅速逆转治疗的效果。鉴于这一现实，凝血因子 XII (FXII) 已成为一种有前途的新药物靶点，可能会改变临床前的抗血栓治疗。动物模型一直被证明可以预防血栓形成，但严重先天性 FXII 缺陷与出血素质无关，因此，抑制 FXII。可以代表人们长期寻求的将止血与血栓“脱钩”的方法，并实现然而，尽管其具有潜在的临床潜力，但其抗血栓功效却无出血并发症。重要性，但人们对血小板依赖性 FXII 激活的潜在机制知之甚少使用质谱蛋白质组学筛选，我们已将整合素 αIIbβ3 鉴定为整合素 αIIbβ3。 FXII 的推定血小板受体这些结果已得到许多后续研究。其他研究可重复证明 FXII-整合素 αIIbβ3 相互作用。假设 FXII 酶原表现出与血小板表面整合素 αIIbβ3 的特异性结合，这增强了其蛋白水解活性，对于 FXII 依赖性来说是必要和充分的在该提案的目标 1 中，我们将绘制 FXII 负责结合的区域。使用重组生成的片段通过竞争性抑制测定整合αIIbβ3 我们还将利用表面等离子共振来评估 FXII-的特异性。整合素 αIIbβ3 相互作用和结合获得的动力学在目标 2 中，我们将探讨整合素的作用。使用整合素 αIIbβ3 包被的珠子和野生血小板激活 FXII 时的 αIIbβ3 结合 β3 型和整合素 β3 (ITGB3) 缺失小鼠的研究工作将提供重要的新见解。研究血小板中 FXII 募集、激活和增殖的分子机制浮出水面并为开发基于 FXII 抑制的新型疗法的努力提供信息。