Engineered Stem Cells for Cardiac Repair

用于心脏修复的工程干细胞

• 批准号: 10293039
• 负责人: Charles E Murry
• 金额: $ 5.4万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-02-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10293039
• 关键词: ATP Synthesis Pathway; Actins; Acute; Award; Binding; Cardiac; Cardiac Myocytes; Cardiotonic Agents; Cell Line; Cell Survival; Cell Therapy; Cell Transplantation; Cells; Chronic; Depressed mood; Devices; Dilated Cardiomyopathy; Drug Delivery Systems; Engineering; Environment; Enzymes; Gap Junctions; Heart; Heart Contractilities; Heart failure; Hip region structure; Ischemia; Modeling; Muscle Contraction; Muscular Dystrophies; Mutation; Myocardial Infarction; Myocardial Ischemia; Myocardium; Myosin ATPase; Nucleotides; Nude Rats; Parents; Power stroke; Pump; Rattus; Research Project Grants; Rest; Ribonucleotide Reductase; Testing; Tissues; Transgenic Model; Translational Research; Transplantation; Variant; adeno-associated viral vector; base; cardiac repair; cell replacement therapy; engineered stem cells; experimental study; gene therapy; graft function; heart function; improved; improved functioning; improved outcome; in vivo; mouse model; novel; overexpression; parent project; small molecule; ATP Synthesis Pathway; Actins; Acute; Award; Binding; Cardiac; Cardiac Myocytes; Cardiotonic Agents; Cell Line; Cell Survival; Cell Therapy; Cell Transplantation; Cells; Chronic; Depressed mood; Devices; Dilated Cardiomyopathy; Drug Delivery Systems; Engineering; Environment; Enzymes; Gap Junctions; Heart; Heart Contractilities; Heart failure; Hip region structure; Ischemia; Modeling; Muscle Contraction; Muscular Dystrophies; Mutation; Myocardial Infarction; Myocardial Ischemia; Myocardium; Myosin ATPase; Nucleotides; Nude Rats; Parents; Power stroke; Pump; Rattus; Research Project Grants; Rest; Ribonucleotide Reductase; Testing; Tissues; Transgenic Model; Translational Research; Transplantation; Variant; adeno-associated viral vector; base; cardiac repair; cell replacement therapy; engineered stem cells; experimental study; gene therapy; graft function; heart function; improved; improved functioning; improved outcome; in vivo; mouse model; novel; overexpression; parent project; small molecule

ABSTRACT. The parent project is built around 20 years of mechanistic and translational research based on two fundamental discoveries: 1) 2-deoxy ATP (dATP) is a potent natural nucleotide stimulant of cardiac contractility (via improved myosin binding to actin & faster detachment after the power stroke), and 2) hiPSC- CMs that overexpress the rate-limiting enzyme for dATP synthesis, ribonucleotide reductase (RNR), have both increased contractility and deliver dATP to the rest of the heart via gap junctions. Thus we are testing the hypothesis that engineering hiPSC-CMs to elevate RNR (RNR-hiPSC-CMs) will improve outcomes in cell replacement therapy for MI (compared with control hiPSC-CMs), improving contractility of both graft and native myocardium. There are several highly novel aspects to our approach. 1) It is the first proposed use of cellular nucleotide manipulation to improve in vivo cardiac function. 2) The approach is not limited to replacement of lost tissue (with hiPSC-CMs) with a better functioning graft, but may also substantially benefit the post-MI depressed function of native myocardium. 3) The first use of engineered hiPSC-CMs to deliver what is effectively a small molecule therapy (dATP), a natural compound that improves heart muscle contraction. This effectively makes hiPSC-CMs a drug delivery device with cardiac specific delivery and effects. Aim 1 develops and test engineered mutations in RNR that increase it’s stability and activity in cardiomyocytes and their ability to titrate increasing levels of dATP produced in hiPSC-CMs. Aim 2 uses AAV vectors for RNR variants, selected from Aim 1, to investigate their capacity to improve cardiac function in a mouse model of myocardial infarct and heart failure. Aim 3 will produce engineered hiPS cell lines that will act as dATP ‘donor cells’ following differentiation, for transplantation into acute MI and more challenging chronic MI athymic rat models to determine their capacity to improve function beyond transplantation of non-engineered hiPSC-CMs. We will evaluate the persistence of these effects and determine the long-term stability and viability of these cell lines. We expect significant contractile improvement of both the graft and native myocardium with RNR-hiPSC-CMs vs. hiPSC-CMs and this effect will be modulated by the dATP producing capacity of the transplanted cells. Results from these studies will elucidate the potential of this combination cell- and small molecule therapy to ameliorate or even improve pump function in failing hearts. This supplement, as the candidates research project will extend the project with 2 aims. Aim 1 will investigate the mechanism by which cardiac muscle using dATP is less susceptible to reductions in contractile strength when pH is reduced, such as occurs in ischemia. Aim 2 will determine whether elevation (rescue) of cardiac function can occur in a different model of dilated cardiomyopathy (than MI), that occurring in Deuchenne’s Muscular Dystrophy (DMD) using a rat transgenic model.

抽象的。家长项目建立了大约20年的机械和翻译研究 基于两个基本发现：1）2-脱氧ATP（DATP）是一种潜在的自然核卫星刺激剂 收缩力（通过改善肌球蛋白与肌动蛋白的结合，并在动力中风后更快的脱离），以及2）Hipsc- 过表达DatP合成速率限制酶的CMS，核糖核苷酸还原酶（RNR），都具有两个 通过间隙连接增加了收缩性并将DATP运送到心脏的其余部分。我们正在测试 假设工程HIPSC-CMS提升RNR（RNR-HIPSC-CMS）将改善细胞的预后 MI的替代疗法（与对照HIPSC-CM相比），提高了移植物和天然的收缩力 心肌。我们的方法有几个新颖的方面。 1）这是第一次提出的细胞使用 核苷酸操纵以改善体内心脏功能。 2）方法不仅限于替代丢失 组织（带有HIPSC-CMS）具有更好功能的移植物，但也可能有益于MI后抑郁症 本地心肌的功能。 3）首次使用工程的HIPSC-CMS来提供有效的小 分子疗法（DATP），一种天然化合物，可改善心肌收缩。这有效地做到了 HIPSC-CMS具有心脏特异性递送和影响的药物输送装置。 AIM 1 RNR中的开发和测试工程突变，以提高其稳定性和活动 心肌细胞及其在HIPSC-CMS中滴定增加的DATP水平的能力。 AIM 2使用AAV 从AIM 1中选择的RNR变体的向量，以研究其改善心脏功能的能力 心肌梗塞和心力衰竭的小鼠模型。 AIM 3将产生工程的臀部细胞系，以行动 随着Datp分化后的Datp“供体细胞”，将其移植到急性MI中，并挑战更多慢性MI 无胸腺大鼠模型，以确定其提高功能的能力超出非工程的移植 HIPSC-CMS。我们将评估这些影响的持久性，并确定长期稳定性和生存能力 这些细胞系。我们预计，通过 RNR-IHIPSC-CMS与HIPSC-CMS，此效果将由DATP产生能力调节 移植细胞。这些研究的结果将阐明该组合细胞和小的潜力 分子疗法可以改善甚至改善心脏失败的泵功能。 这种补充，因为候选人研究项目将以2个目标扩展该项目。 AIM 1将调查 使用DATP的心脏肌肉不易降低收缩力的机制 当pH减少时，例如缺血中发生。 AIM 2将确定是否高程（救援）心脏 功能可能发生在不同的膨胀心肌病模型（而不是MI）中，该模型发生在Deuchenne的 使用大鼠转基因模型的肌肉营养不良（DMD）。