Embryonic Stem Cell-Derived Platelets as Cellular Therapeutics

胚胎干细胞衍生的血小板作为细胞治疗药物

• 批准号: 7939697
• 负责人: Mortimer Poncz
• 金额: $ 120.91万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7939697
• 关键词: Affect; Animal Model; Animals; Apoptosis; Beds; Biological Models; Blood Circulation; Blood Platelets; Blood Vessels; Bone Marrow; Canis familiaris; Cell Line; Cells; Clinical; Coagulation Process; Collaborations; Cytoplasmic Granules; Data; Dependency; Development; Drug usage; Ectopic Expression; Engineering; Environment; Erythroid; Erythroid Progenitor Cells; Exhibits; Fibrinolysis; Funding; Future; GATA1 gene; Goals; Half-Life; Hematopoiesis; Hematopoietic; Hemostatic function; Human; In Vitro; Individual; Infusion procedures; Injury; Knock-out; Lung; Marrow; Megakaryocytes; Megakaryocytopoieses; Mesoderm; Modeling; Mus; Patients; Platelet Activation; Platelet Count measurement; Principal Investigator; Process; Production; Proteins; Regenerative Medicine; Reporting; Resources; Signal Transduction; Simulate; Site; Source; Staging; Stem cells; System; Testing; Therapeutic; Therapeutic Agents; Thrombus; Transfusion; Translating; Urokinase; Venous; angiogenesis; base; clinical application; clinically relevant; cytokine; embryonic stem cell; experience; genetic manipulation; human GATA1 protein; human embryonic stem cell; immunodeficient mouse model; improved; in vivo; induced pluripotent stem cell; inhibitor/antagonist; interest; notch protein; novel strategies; preclinical study; progenitor; response; targeted delivery; therapeutic target; thrombolysis; venule

DESCRIPTION (provided by applicant): Present day delivery of platelets for various clinically relevant settings has a number of significant limitations, from issues of storage, contamination, inhibitors in multiply transfused individuals, variable quality and dependency on human donors. We propose a novel strategy for production of clinically relevant numbers of platelets from human embryonic stem cells (hESCs), supported by preliminary data. Such platelets may allow us to avoid many of the present limitations and allow us to develop platelets as a mechanism for the targeted delivery of therapeutic agents to sites of vascular injury. The approach will involve the following steps which will be pursued simultaneously building on our preliminary data: 1) Enhance directed differentiation of hESCs into megakaryocytes (Megs) by improving the efficiency of hESC to hematopoietic mesoderm development and directing hematopoietic progenitors into the Meg lineage. 2) Arrest cells at the Meg-erythroid progenitor (MEP) stage to form self-replicating cells by knockdown of the transcription factor GATA1. Upon re-induction of GATA1, these MEP cells will complete differentiation into Megs. Strategies to enhance MEP to mature Megs will also be pursued. 3) Infuse either ex vivo-generated platelets or mature Megs to generate a vigorous wave of new, functional platelets. 4) Either hESCs or the self-replicating MEP cells will also be modified so that the Megs express an ectopic protein of interest stored in their a-granules for release upon platelet activation. Based on preliminary data, we chose as proof-of-principle to express urinary plasminogen activator in the developing Megs. We will demonstrate that the resulting human platelets allow targeted delivery of this agent to growing thrombi without causing systemic fibrinolysis. These studies will be done in close collaboration with the Seattle Cluster and will involve 3 species: mice, dogs and humans. Mice studies will allow rapid advances that will then be tested in dogs as a large animal model as well as apply our advances to human studies. We believe that over the seven years of support that we will have achieved significant advances in the production of clinically relevant numbers of functional platelets beginning with hESCs and be ready for large-scale confirmatory studies and clinical application.

描述（由申请人提供）： 目前，用于各种临床相关环境的血小板输送存在许多显着的局限性，包括储存、污染、多次输血个体中的抑制剂、质量参差不齐以及对人类捐赠者的依赖等问题。我们提出了一种从人胚胎干细胞（hESC）生产临床相关数量的血小板的新策略，并得到了初步数据的支持。这种血小板可以让我们避免目前的许多限制，并让我们开发血小板作为将治疗剂靶向递送到血管损伤部位的机制。该方法将涉及以下步骤，这些步骤将在我们的初步数据的基础上同时进行：1）通过提高 hESC 造血中胚层发育的效率并引导造血祖细胞进入 Meg 谱系，增强 hESC 向巨核细胞（Meg）的定向分化。 2) 通过敲低转录因子GATA1，将细胞阻滞在巨红系祖细胞（MEP）阶段，形成自我复制细胞。重新诱导 GATA1 后，这些 MEP 细胞将完全分化为 Megs。还将采取增强 MEP 使其成熟的 Megs 的策略。 3) 输注离体产生的血小板或成熟的巨噬细胞，以产生一波充满活力的新的功能性血小板。 4) hESC 或自我复制的 MEP 细胞也将被修改，以便 Meg 表达储存在其 a 颗粒中的感兴趣的异位蛋白，以便在血小板激活时释放。根据初步数据，我们选择在发育中的巨噬细胞中表达尿纤溶酶原激活剂作为原理验证。我们将证明，由此产生的人血小板可以将这种药物定向输送到正在生长的血栓，而不会引起全身性纤维蛋白溶解。 这些研究将与西雅图集群密切合作进行，并将涉及 3 个物种：小鼠、狗和人类。小鼠研究将取得快速进展，然后将在狗作为大型动物模型中进行测试，并将我们的进展应用于人类研究。我们相信，经过七年的支持，我们将在从 hESC 开始生产临床相关数量的功能性血小板方面取得重大进展，并为大规模验证性研究和临床应用做好准备。