Gene-Editing of Human Pluripotent Stem Cell-Derived Cardiovascular Therapy Grafts to Improve Transplant Outcomes

对人类多能干细胞衍生的心血管治疗移植物进行基因编辑以改善移植结果

• 批准号: 10683804
• 负责人: Matthew E Brown
• 金额: $ 38.21万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-13 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10683804
• 关键词: Ablation; Adherence; Allogenic; American; Anatomy; Animal Experiments; Anti-Inflammatory Agents; Antiinflammatory Effect; Apoptosis; Binding; Biological Assay; Biology; Blood Vessels; CCL2 gene; CCL22 gene; CRISPR/Cas technology; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Cardiovascular system; Cell Adhesion; Cell Adhesion Molecules; Cell Communication; Cell Therapy; Cell-Mediated Cytolysis; Cells; Cellular immunotherapy; Clinical; Clinical Trials; Cues; Cytolysis; Data; Discipline; Disease; Elements; Endothelial Cells; Extravasation; Female; Fibroblasts; Free Radical Scavenging; Functional disorder; Future; Gene Expression; Genes; Goals; HLA Antigens; Health; Histocompatibility Antigens Class I; Human; Immune; Immune Targeting; Immune Tolerance; Immune response; Immunocompetent; Immunology; Immunosuppression; Immunotherapy; In Vitro; Individual; Infarction; Infiltration; Inflammatory; Inflammatory Response; Intercellular adhesion molecule 1; Interleukin-10; Intervention; Knock-out; Laboratories; Leukocytes; Malignant Neoplasms; Mediating; Mission; Modeling; Myocardial Infarction; Natural Killer Cells; Outcome; Oxidative Stress; Pathology; Pathway interactions; Patients; Pharmacologic Substance; Phenotype; Pluripotent Stem Cells; Public Health; Quality of life; Research; Research Project Grants; Resistance; Risk; Source; T-Lymphocyte; TNF gene; Testing; Therapeutic Intervention; Thick; Transplantation; Transplantation Tolerance; United States National Institutes of Health; Validation; Vascular Graft; Ventricular; Work; cell type; chemokine; clinical translation; cytokine; cytotoxic; design; disability; donor-specific antibody; heart function; human pluripotent stem cell; immunogenicity; immunological synapse formation; improved; improved functioning; in vivo; in vivo Model; innovation; insight; male; mouse model; next generation; nonhuman primate; novel; novel strategies; post-transplant; preclinical study; prevent; repair function; reparative capacity; stem cell therapy; therapeutic genome editing; translational study; transplantation therapy

ABSTRACT After a first myocardial infarction (MI), 36% of male and 47% of female patients die within 5 years. This illustrates the inadequacy of current therapeutic interventions. The long-term goal of our laboratory is to develop reparative pluripotent stem cell (PSC)-based therapies that are immune-tolerated and meaningfully improve patient health and quality of life. The overall objectives of this R01 application are to: 1) use CRISPR/Cas9 gene-editing approaches to target adhesion molecules (AMs) on human PSC-derived cardiovascular therapies (PSC-CVTs) to disrupt the adherence, infiltration, and destruction of vascularized grafts by allogeneic immune cells; and 2) define optimal cellular composition and immunogenicity profiles of next-generation hypoimmune PSC-CVT grafts to maximize their reparative capacity in the inflammatory setting of MI. Our central hypothesis is that targeted deletion of AM genes will facilitate immune tolerance of PSC- CVTs via two mechanisms: 1) diminished immune cell contact-mediated destruction; and 2) anti-inflammatory effects (e.g., secreted factor and gene expression changes) directly associated with genetically disrupting AM function. The rationale for this project is that hypoimmune PSCs will be key clinical platforms in the coming years and improved gene-editing approaches are needed to achieve effective immune tolerance of PSC grafts. Additionally, successful validation of AM gene-editing in this project will provide a new avenue for advancing future transplantation therapies for other diseases. To attain our objectives, we will pursue the following specific aims (SAs): SA1) Define the effects of AM ablation on immune cell contact-mediated PSC-CVT graft destruction; SA2) Define the inflammatory responses initiated by immune cell:PSC-CVT graft interactions; and SA3) Determine the in vivo reparative capacity and immune-tolerance potential of AM knockout PSC-CVT grafts in the inflammatory MI setting. This research is significant because it validates a new graft strategy and testing platform for hypoimmune PSC therapies, with great potential to save lives and improve quality of life for many patients with MI and other pathologies characterized by cellular dysfunction in immune-competent anatomical sites. It is innovative because: 1) of the new approach of targeting immune cell adhesion in a manner anticipated to impede both adaptive and innate immune cell-mediated graft destruction; 2) it uses a tri- cellular PSC-CVT graft optimized for superior reparative function and hypoimmunogenicity; and 3) we rigorously interrogate the human immune response using advanced assays and models developed in our lab. Ultimately, this work will develop a breakthrough cardiac therapy well-suited for clinical trials, with the potential to save lives and improve the quality-of-life for millions of patients.

抽象的 首次心肌梗塞 (MI) 后，36% 的男性患者和 47% 的女性患者会在 5 年内死亡。这 说明了当前治疗干预措施的不足。我们实验室的长期目标是 开发基于修复性多能干细胞 (PSC) 的免疫耐受且有意义的疗法 改善患者的健康和生活质量。该 R01 应用程序的总体目标是： 1) 使用 CRISPR/Cas9 基因编辑方法靶向人类 PSC 来源的粘附分子 (AM) 心血管疗法（PSC-CVT）破坏血管化的粘附、浸润和破坏 同种异体免疫细胞移植； 2) 确定最佳细胞组成和免疫原性特征 下一代低免疫 PSC-CVT 移植物可最大限度地提高其在炎症环境中的修复能力 心肌梗死。我们的中心假设是 AM 基因的定向删除将促进 PSC-的免疫耐受 CVT 通过两种机制：1）减少免疫细胞接触介导的破坏； 2) 抗炎 与基因破坏 AM 直接相关的影响（例如，分泌因子和基因表达变化） 功能。该项目的基本原理是低免疫 PSC 将成为未来的关键临床平台 需要多年的时间和改进的基因编辑方法才能实现 PSC 移植物的有效免疫耐受。 此外，该项目中 AM 基因编辑的成功验证将为推进技术进步提供新途径。 未来其他疾病的移植疗法。为了实现我们的目标，我们将追求以下目标 特定目标 (SA)：SA1) 定义 AM 消融对免疫细胞接触介导的 PSC-CVT 移植物的影响 破坏； SA2) 定义免疫细胞引发的炎症反应：PSC-CVT移植物相互作用；和 SA3) 确定AM敲除PSC-CVT的体内修复能力和免疫耐受潜力 炎症性心肌梗死环境中的移植物。这项研究意义重大，因为它验证了一种新的移植策略 低免疫 PSC 疗法的测试平台，具有拯救生命和提高生活质量的巨大潜力 许多患有心肌梗死和其他以免疫功能正常的细胞功能障碍为特征的疾病的患者 解剖部位。它具有创新性，因为：1）针对免疫细胞粘附的新方法 预期阻止适应性和先天免疫细胞介导的移植物破坏的方式； 2）它使用三 细胞 PSC-CVT 移植物经过优化，具有卓越的修复功能和低免疫原性； 3）我们 使用我们实验室开发的先进检测方法和模型严格询问人体免疫反应。 最终，这项工作将开发出一种非常适合临床试验的突破性心脏疗法，具有潜在的潜力 拯救生命并提高数百万患者的生活质量。