Engineered Stem Cells for Cardiac Repair

用于心脏修复的工程干细胞

基本信息

批准号：
10544645
负责人：
Charles E Murry
金额：
$ 10.7万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-02-01 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10544645
关键词：
ATP Synthesis Pathway Actins Acute Award Binding Cardiac Cardiac Muscle Contraction Cardiac Myocytes Cardiomyopathies Cardiotonic Agents Cell Line Cell Therapy Cell Transplantation Cells Chronic Depressed mood Dilated Cardiomyopathy Duchenne muscular dystrophy Elements Engineering Environment Enzymes Exhibits Gap Junctions Heart Heart Contractilities Heart failure Hip region structure Investigation Ischemia Light Mechanics Modeling Modification Mutation Myocardial Infarction Myocardial Ischemia Myocardium Myosin ATPase Nucleotides Parents Performance Pharmaceutical Preparations Power stroke Production Pump Rattus Research Project Grants Rest Ribonucleotide Reductase Span 20 System Techniques Testing Time Tissues Transgenic Organisms Translational Research Variant adeno-associated viral vector cardiac repair cell replacement therapy engineered stem cells experimental study gene therapy heart function improved improved outcome in vivo innovation mouse model novel overexpression parent project small molecule

项目摘要

ABSTRACT: The parent project is built on two fundamental discoveries, spanning 20 years of mechanistic and translational research: 1) 2-deoxy ATP (dATP) is a potent natural nucleotide stimulant of cardiac contractility (via increased myosin binding to actin and faster detachment following the power stroke), and 2) hiPSC-CMs overexpressing the rate-limiting enzyme for dATP synthesis, ribonucleotide reductase (RNR), exhibit both increased contractility and dATP delivery to the rest of the heart via gap junctions. Consequently, we are investigating the hypothesis that altering hiPSC-CMs to increase RNR (RNR-hiPSC-CMs) improves outcomes in cell replacement treatment for MI (as compared to control hiPSC-CMs), by increasing the contractility of both the graft and native myocardium. Our technique incorporates several highly innovative elements. 1) This is the first time that cellular nucleotide modification has been offered as a means of improving in vivo heart function. 2) The technique is not restricted to replacing lost tissue (with hiPSC-CMs) with a more functional graft but may also significantly benefit the native myocardium's post-MI depressed function. 3) The first application of modified hiPSC-CMs to deliver a small molecule treatment (dATP) that enhances cardiac muscle contraction. This essentially transforms hiPSC-CMs into a cardiac-specific medication delivery system. Aim 1 is to generate and characterize engineered mutations in RNR that enhance its stability and activity in cardiomyocytes, as well as their ability to titrate increasing quantities of dATP produced in hiPSC-CMs. Aim 2 investigates the ability of RNR variations identified in Aim 1 to improve cardiac function in a mouse model of myocardial infarction and heart failure using AAV vectors. Aim 3 will generate engineered hiPS cell lines that, upon differentiation, will act as dATP 'donor cells' for transplantation into acute MI and more problematic chronic MI arrhythmic rat models to test their capacity to increase function beyond that of non-designed hiPSC-CMs. We will assess the persistence of these effects and the cell lines' long-term survival and stability. We anticipate a significant contractile improvement in both the graft and native myocardium using RNR-hiPSC-CMs vs. hiPSC- CMs, which will be controlled by the transplanted cells' dATP producing capacity. These investigations will shed light on the potential for this combination cell- and small-molecule therapy to improve or perhaps restore pump performance in failing hearts. This addition, as the candidate's research project, will expand the scope of the study by pursuing two objectives. Aim 1 will examine the mechanical and structural mechanisms that allow cardiac muscle utilizing dATP to be less sensitive to contractile strength losses when the pH is decreased, as occurs during ischemia. Aim 2 will investigate whether cardiac function can be improved in a different cardiomyopathy model (dilated instead of MI), utilizing a Duchenne’s Muscular Dystrophy (DMD) dilated cardiomyopathy model expressed in a transgenic rat.

摘要：母项目建立在两项基本发现之上，跨越了 20 年的机械和转化研究：1) 2-脱氧 ATP (dATP) 是一种有效的心肌收缩力天然核苷酸兴奋剂（通过肌球蛋白与肌动蛋白的结合增加，动力冲程后分离速度更快），2) hiPSC-CM 过表达 dATP 合成的限速酶核糖核苷酸还原酶 (RNR)，表现出两者我们检查了收缩力的增加以及 dATP 通过间隙连接传递到心脏其他部分的情况。研究改变 hiPSC-CM 以增加 RNR (RNR-hiPSC-CM) 改善结果的假设在 MI 的细胞替代治疗中（与对照 hiPSC-CM 相比），通过增加两者的收缩性我们的技术融合了几个高度创新的元素 1) 这就是。首次将细胞核苷酸修饰作为改善体内心脏功能的手段2)。该技术不仅限于用功能更强的移植物替代丢失的组织（使用 hiPSC-CM），还可以也显着有益于心肌梗死后的低下功能 3) 改良的首次应用。 hiPSC-CM 提供小分子治疗 (dATP)，增强心肌收缩。本质上将 hiPSC-CM 转变为心脏特异性药物输送系统。目标 1 是生成并表征 RNR 中的工程突变，以增强其稳定性和活性心肌细胞，以及它们滴定 hiPSC-CM 中产生的 dATP 数量的能力 Aim 2。研究目标 1 中确定的 RNR 变异改善小鼠模型心脏功能的能力使用 AAV 载体治疗心肌梗塞和心力衰竭，Aim 3 将产生工程化的 hiPS 细胞系，该细胞系：分化后，将作为 dATP“供体细胞”移植到急性心肌梗死和更有问题的慢性心肌梗死中 MI 心律失常大鼠模型，测试其增强功能的能力，超过非设计的 hiPSC-CM。我们预计将评估这些影响的持续性以及细胞系的长期存活和稳定性。使用 RNR-hiPSC-CM 与 hiPSC- 相比，移植心肌和天然心肌的收缩能力显着改善 CM，将由移植细胞的 dATP 产生能力控制。这些研究将揭示。揭示这种细胞和小分子组合疗法改善或恢复泵的潜力失败的心的表现。作为候选人的研究项目，这一补充将通过追求两个目标来扩大研究范围目标 1 将研究允许心肌利用的机械和结构机制。当 pH 值降低时（如缺血期间发生的情况），dATP 对收缩强度损失不太敏感。目标 2 将研究在不同的心肌病模型（扩张型心肌病）中心功能是否可以得到改善而不是 MI），利用杜氏肌营养不良症 (DMD) 扩张型心肌病模型转基因大鼠。