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Functional genetics of tissue factor in bleeding and thrombotic risk

出血和血栓风险中组织因子的功能遗传学

基本信息

批准号：
10249178
负责人：
Sol Schulman
金额：
$ 43.75万
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：
2019
资助国家：
美国
起止时间：
2019-09-16 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10249178
关键词：
Address Advisory Committees Anticoagulant therapy Anticoagulants Area Award Biochemistry Biological Assay Blood Coagulation Disorders Blood Coagulation Factor Blood Vessels Blood coagulation Budgets Cell surface Cells Cellular biology Choristoma Clinic Clinical Clustered Regularly Interspaced Short Palindromic Repeats Coagulation Process Collaborations Conflict (Psychology)Core Facility Custom Data Diagnostic Disease Disease susceptibility Disseminated Intravascular Coagulation Electron Transport Endothelium Ensure Equipment Etiology Explosion Faculty Fellowship Foundations Future Genes Genetic Genetic Models Genetic Variation Genomic approach Goals Grant Hematology Hemorrhage Hemostatic Disorders Hemostatic function Hereditary Disease Heritability Human Human Genetics Human Resources Individual Inherited Injury Institutional Review Boards International Investigation Israel Laboratories Lead Leadership Medical center Medicine Mentors Mentorship Metabolism Morbidity - disease rate Mus Myocardial Infarction Ownership Oxidation-Reduction Pathogenesis Pathologic Pathway interactions Patient Care Physicians Population Positioning Attribute Principal Investigator Prize Protein Biochemistry Proteins Protocols documentation Pulmonary Embolism Quantitative Trait Loci Readiness Reporting Research Research Activity Risk Rodent Model Scientist Secure Seminal Sickle Cell Anemia Smooth Muscle Myocytes Stroke Structure Students Testing Therapeutic Thromboplastin Thrombosis Training Variant Vascular Smooth Muscle Vision Vitamin K Vocational Guidance Wages Warfarin career development cell type design disorder risk functional genomics genetic approach genetic association genome-wide human population study improved in vivo induced pluripotent stem cell innovation insight intravital microscopy kindred loss of function medical schools mortality personalized medicine professor recruit senior faculty square foot stem cell differentiation thrombotic ubiquitin-protein ligase vascular injury venous thromboembolism

项目摘要

Project Summary/Abstract Tissue factor (TF) is the primary initiator of blood coagulation in vivo and is required to ensure hemostasis following injury. However, inappropriate TF procoagulant activity underlies substantial human suffering, including that due to myocardial infarction (MI), venous thromboembolism (VTE), stroke, and sickle cell disease. Because the cellular contribution of TF is not captured in any routine clinical assay of blood coagulation, the machinery influencing TF expression and procoagulant activity remains largely unknown. Our central hypothesis is that cellular TF and its modifiers contribute to the substantial unexplained heritability of hemostatic and thrombotic disorders, including VTE, MI, and stroke. To achieve this goal, we will intersect functional genetic and human genetic approaches to identify modifiers that contribute to TF-dependent heritability of bleeding and clotting risk. TF (“factor III”) is the only coagulation factor for which a hereditary deficiency has not been described, but in Aim 1 we will study a kindred with a bleeding diathesis in association with heterozygous TF deficiency using protein biochemistry, CRISPR-edited induced pluripotent stem cells (iPSCs) differentiated into endothelial and vascular smooth muscle cells not accessible in humans, and genetically modified mice via intravital microscopy following vascular injury; importantly, these efforts provide proof of concept that modifiers reducing TF expression by 50% can influence bleeding risk in vivo. In Aim 2, we will use an arrayed loss-of-function screen at genome-scale to identify modifiers of TF cell surface expression and procoagulant activity and intersect these findings with expression quantitative trait loci and existing population data capturing common and rare human genetic variation to identify areas of intersection, offering a generalizable strategy to use a functional screen to move beyond genetic association towards causation and mechanism. Modifiers likely contributing to TF-dependent heritability of bleeding and clotting risk will be mechanistically evaluated at scale using a custom CRISPR array and then evaluated in iPSCs, mice, and, where possible, humans. In Aim 3, we will study as a representative example one such discovery, an E3 ubiquitin ligase that strongly negatively regulates TF protein stability and procoagulant activity, employing cell biology, biochemistry, rodent models, and human genetics to mechanistically demonstrate how human genetic variation may contribute to VTE risk. Our findings will highlight the critical but clinically unmeasured contribution of cellular TF to the pathogenesis and inheritance of broadly defined hemorrhagic and thrombotic diseases. The findings will have immediate translational potential for diagnostics able to capture this risk to guide personalized treatment and suggest promise for new anticoagulant therapies with reduced bleeding risk that selectively target pathways leading to inappropriate TF procoagulant activity.

项目总结/摘要组织因子（TF）是体内血液凝固的主要引发剂，是确保止血所必需的受伤后。然而，不适当的TF促凝血活性导致了人类的巨大痛苦，包括心肌梗死（MI）、静脉血栓栓塞（VTE）、中风和镰状细胞病疾病因为TF的细胞贡献在任何常规的临床血液分析中都没有被捕获凝血，影响TF表达和促凝血活性的机制在很大程度上仍然未知。我们中心假设是细胞TF及其修饰物有助于实质性的无法解释的遗传性，止血和血栓性疾病，包括VTE、MI和卒中。为了实现这一目标，我们将功能遗传学和人类遗传学方法来鉴定有助于TF依赖性出血和凝血风险的遗传性。TF（“因子III”）是唯一的凝血因子，缺乏症尚未描述，但在目标1中，我们将研究一个与出血素质相关的家系。使用蛋白质生物化学，CRISPR编辑的诱导多能干细胞，诱导多能干细胞（iPSC）分化成人类无法获得的内皮细胞和血管平滑肌细胞，通过血管损伤后的活体显微镜检查转基因小鼠;重要的是，这些努力提供了概念证明，将TF表达降低50%的修饰剂可以影响体内出血风险。在目标2中，我们将在基因组水平上使用阵列功能缺失筛选来鉴定TF细胞表面修饰剂表达和促凝血活性，并将这些发现与表达数量性状基因座交叉，现有的人口数据捕获常见和罕见的人类遗传变异，以确定交叉区域，提供了一种可推广的策略，使用功能筛选超越遗传关联，原因和机制。可能导致TF依赖性出血和凝血遗传性的修饰因子风险将使用定制的CRISPR阵列进行大规模的机械评估，然后在iPSC中进行评估，老鼠，如果可能的话，还有人类。在目标3中，我们将作为一个代表性的例子来研究这样的发现，一种强烈负调节TF蛋白稳定性和促凝血活性的E3泛素连接酶，利用细胞生物学、生物化学、啮齿动物模型和人类遗传学，人类遗传变异可能导致静脉血栓栓塞风险。我们的研究结果将突出关键的，但临床上细胞TF对广泛定义的出血性胰腺炎的发病机制和遗传的不可测量的贡献和血栓性疾病。这些发现将具有立即转化的诊断潜力，捕捉这一风险，以指导个性化治疗，并提出新的抗凝治疗的承诺，降低出血风险，选择性靶向导致TF促凝血活性不适当的途径。