The role of protease activated receptors on platelets

蛋白酶激活受体对血小板的作用

基本信息

批准号：
9241436
负责人：
Marvin Thomas Nieman
金额：
$ 31.7万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-08-15 至 2021-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9241436
关键词：
Address Adverse effects Affect Agonist Alleles Allosteric Site Amides Amino Acids Antiplatelet Drugs Aspirin Binding Binding Sites Blood Platelets Blood Vessels Blood coagulation Cells Deuterium Development Dominant-Negative Mutation Effectiveness F2R gene FDA approved Family Frequencies G-Protein-Coupled Receptors Genetic Polymorphism Genotype Goals Hemorrhage Hemostatic function Heritability Histidine Homo Human Hybrids Hydrogen Hydroxyl Radical Individual Intracranial Hemorrhages Lead Ligand Binding Ligands Mass Spectrum Analysis Membrane Molecular Molecular Conformation Molecular Structure Monitor Myocardial Infarction Output Pathologic Pathway interactions Patients Pharmacogenomics Pharmacology Study Physiological Platelet Activation Platelet aggregation Population Proteinase-Activated Receptors Proteins Receptor Activation Receptor Signaling Recording of previous events Reporting Research Resistance Resolution Risk Role Signal Pathway Signal Transduction Single Nucleotide Polymorphism Stroke Structure Surface Techniques Testing Therapeutic Thrombin Thrombin Receptor Thrombus Treatment Protocols Variant acute coronary syndrome clopidogrel experience insight inter-individual variation interest molecular rearrangement new therapeutic target prevent public health relevance receptor receptor function response standard of care structural biology therapeutic target therapy resistant

项目摘要

DESCRIPTION (provided by applicant): Platelets are defined by their essential roles in hemostasis and the formation of pathological thrombi. Therefore, the major focus of platelet research is focused on identifying and developing safer and more effective anti- thrombotic therapies. A second goal is to have the ability to predict the effectiveness of specific therapies across populations. Pharmacogenomics may allow us to predict who will respond to or be resistant to therapies. In order for this to be successful, there needs to be a clear relationship between specific polymorphisms and physiological output. Finally, the alleles need to be present at high enough frequency to warrant testing prior to the start of therapy. In the current project w will examine common polymorphisms of a platelet receptor (PAR4) to explain the differences in reactivities and response to an antagonist at the molecular level. We will determine the structural rearrangement of PAR4 upon activation by thrombin using structural mass spectrometry (amide hydrogen deuterium exchange (HDX), histidine HDX, and hydroxyl radical foot-printing) with purified PAR4 and PAR4 on cells and platelets. These studies will determine how the tethered ligand influences the overall conformation of the receptor. Pharmacological studies in human platelet show that the PAR4 sequence variants have dramatically different responses. For example, PAR4-120T is hyper-reactive and PAR4-296V is resistant to signaling. We will determine how the PAR4 variants affect the ligand binding site and the transition to the active conformation at the molecular level. Further, PAR4-296V suppresses the activity of other alleles suggesting that it forms dominant negative homodimers. We will examine the physical interaction between PAR4 variants, PAR1 and P2Y12 to determine how these receptors influence signaling pathways. Finally, we will determine how the PAR4 sequence variants influence platelet reactivity in acute coronary syndrome patients on the current standard of care (aspirin and a P2Y12 antagonist) and if the altered platelet reactivity due to PAR4 variants is exacerbated with the PAR1 antagonist vorapaxar. By analyzing the structural rearrangements of PAR4 following activation by thrombin, these studies will provide the first detailed description of the tethered ligand activation mechanism for the PAR family and have the potential to uncover allosteric sites that can be exploited therapeutically. More specifically, understanding how naturally occurring sequence variants influence PAR4's response may allow us to predict the most appropriate antiplatelet therapy for patients depending on their genotype.

描述（由适用提供）：血小板由它们在止血和病理性血栓形成中的基本作用定义。因此，血小板研究的主要重点是识别和开发更安全，更有效的抗血栓形成疗法。第二个目标是有能力预测跨人群的特定疗法的有效性。药物基因组学可能使我们能够预测谁将对疗法做出反应或抗药性。命令要成功，需要在特定的多态性和身体产出之间存在明确的关系。最后，等位基因需要以足够高的频率存在，以在开始治疗之前进行测试。在当前的项目中，W将检查血小板受体（PAR4）的常见多态性，以解释反应性和对拮抗剂在分子水平上的差异。我们将使用结构质谱法（酰胺氢氘交换（HDX），组氨酸HDX和羟基自由基脚印）在通过凝血酶激活时确定PAR4的结构重排，并在细胞和血静脉上纯化PAR4和PAR4。这些研究将确定束缚配体如何影响接收器的整体考虑。人血小板中的药理学研究表明，PAR4序列变体具有截然不同的反应。例如，PAR4-120T具有高反应性，PAR4-296V对信号传导具有抵抗力。我们将确定PAR4变体如何影响配体结合位点以及分子水平上的活动会议的过渡。此外，PAR4-296V抑制了其他等位基因的活性，表明它形成了主要的负同二聚体。我们将检查PAR4变体，PAR1和P2Y12之间的物理相互作用，以确定这些受体如何影响信号通路。最后，我们将确定PAR4序列变体如何影响急性冠状动脉综合征患者的血小板反应性（阿司匹林和P2Y12拮抗剂）以及PAR1拮抗剂Vorapaxar导致PAR4变体引起的血小板反应性改变。通过分析凝血酶激活后PAR4的结构重排，这些研究将提供第一个详细描述 PAR家族的束缚配体激活机制，有可能发现可以热探索的变构位点。更具体地说，了解自然发生的序列变体如何影响PAR4的反应可能使我们能够根据其基因型预测最合适的抗血小板疗法。