Genetic and therapeutic studies of hemostatic and thrombotic disorders using zebrafish

使用斑马鱼进行止血和血栓性疾病的遗传和治疗研究

• 批准号: 10375366
• 负责人: JORDAN A SHAVIT
• 金额: $ 78万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2027-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10375366
• 关键词: Affect; Anticoagulants; Anticoagulation; Biological Assay; Blood Coagulation Disorders; Blood Coagulation Factor; Chemicals; Clustered Regularly Interspaced Short Palindromic Repeats; Coagulation Process; Diagnostic; Disease; Family; Fishes; General Population; Genes; Genetic; Genetic study; Genome; Health; Hemorrhage; Hemorrhagic Disorders; Hemostatic Disorders; Hemostatic function; Human; Individual; Intravenous infusion procedures; Lead; Life; Light; Mutagenesis; Oral; Pathologic; Pathology; Patients; Penetrance; Protein Engineering; Recombinant Proteins; Recombinants; Regulator Genes; Severities; Severity of illness; System; Therapeutic; Therapeutic Studies; Thrombopoiesis; Thrombosis; Work; Zebrafish; blood product; experimental study; genetic technology; genome editing; high risk; human morbidity; human mortality; individualized medicine; innovation; mutant; new therapeutic target; next generation sequencing; novel; novel diagnostics; novel therapeutics; nuclease; prospective; public health relevance; therapeutic target; thrombotic; tool; transcription activator-like effector nucleases

PROJECT SUMMARY/ABSTRACT Pathologic dysregulation of the coagulation system is a major contributor to human morbidity and mortality, resulting in either excessive bleeding or clotting. Significant progress has been made in the identification of genetic regulators of the coagulation cascade, but many unknown modifier genes contribute to the variable disease severity and penetrance observed among patients and families with hemostatic and thrombotic disorders. Understanding such modifiers could help classify patients at higher risk for pathology as well as identify novel therapeutic targets. Although major improvements to treatment of hemorrhagic and bleeding disorders have been made with blood products and human derived or recombinant coagulation factors, these have limited shelf life and storage conditions, and require intravenous infusion. Anticoagulation has seen a surge in recent years with many new direct acting oral anticoagulants, but their mechanisms of action are limited to the coagulation cascade. Building on our previous work, this project will take advantage of powerful genetic tools, including genome editing nucleases, next generation sequencing, and the zebrafish. We will conduct a large scale interrogation of the genome to discover hemostasis regulatory genes with the potential to modify the severity of human coagulation disorders. We have developed a panel of clotting factor mutant zebrafish using robust genome editing nucleases (TALENS and CRISPR/Cas) and conducted chemical mutagenesis experiments that have identified potential suppressor mutant lines harboring prospective thrombosis and hemostasis modifier genes. This panel of mutant fish will also be used for unbiased assays to identify novel lead molecules that suppress hemorrhage or thrombosis. The approaches described in this proposal will lead to the identification of the key non-canonical factors regulating hemostasis and thrombopoiesis, some of which will likely prove to be important genetic modifying factors in humans, or will suggest novel species-specific but biologically insightful regulatory mechanisms. This will shed light on the regulatory mechanisms of hemostasis, and the modifiers will also be candidate diagnostic and therapeutic targets for human thrombotic and hemorrhagic diseases. These targets will be utilized to develop potential innovative agents and new therapeutic classes for treatment of hemorrhage and thrombosis that could benefit the general population, as well as patients with bleeding and thrombotic disorders.

项目概要/摘要 凝血系统的病理性失调是人类发病和死亡的主要原因 死亡率，导致过度出血或凝血。方面已取得重大进展 凝血级联的遗传调节因子的鉴定，但许多未知的修饰基因有助于 在使用止血剂和止血剂的患者和家庭中观察到的不同疾病严重程度和外显率 血栓性疾病。了解这些修饰因素可以帮助将病理风险较高的患者分类为 以及确定新的治疗靶点。尽管出血性和出血性疾病的治疗取得了重大进展 出血性疾病是由血液制品和人源性或重组凝血引起的 因素，这些药物的保质期和储存条件有限，并且需要静脉输注。抗凝 近年来，许多新型直接作用口服抗凝剂的数量激增，但它们的作用机制 作用仅限于凝血级联。在我们之前的工作基础上，该项目将利用 强大的遗传工具，包括基因组编辑核酸酶、下一代测序和斑马鱼。 我们将对基因组进行大规模询问，以发现止血调控基因 改变人类凝血障碍严重程度的潜力。我们开发了一组凝血因子 使用强大的基因组编辑核酸酶（TALENS 和 CRISPR/Cas）并进行化学分析突变斑马鱼 诱变实验已鉴定出潜在的抑制突变系 血栓形成和止血调节基因。这组突变鱼也将用于无偏分析 识别抑制出血或血栓形成的新型先导分子。本文中描述的方法 该提案将导致确定调节止血的关键非典型因素和 血小板生成，其中一些可能被证明是人类重要的基因修饰因素，或者将 提出了新颖的物种特异性但具有生物学洞察力的调节机制。这将揭示 止血的调节机制，修饰剂也将成为诊断和治疗的候选药物 人类血栓和出血性疾病的目标。这些目标将被用来开发潜力 用于治疗出血和血栓形成的创新药物和新治疗类别可能会受益 一般人群以及患有出血和血栓性疾病的患者。