Induction of Cardiomyocyte Proliferation via Transient Expression of Cell Cycle Factors as a Promising Therapy for Heart Failure

通过细胞周期因子的瞬时表达诱导心肌细胞增殖作为心力衰竭的一种有前景的治疗方法

• 批准号: 10365990
• 负责人: Tamer M A Mohamed
• 金额: $ 76.96万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-22 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10365990
• 关键词: Acute; Address; Adenoviruses; Adult; Adverse effects; Area; Blood; CDC2 gene; CDK4 gene; Calcium; Cardiac; Cardiac Myocytes; Cell Cycle; Cell Proliferation; Cell division; Cells; Clinical Trials; Contracts; Cyclin D1; Cytokinesis; Data; Daughter; Disease; Electrophysiology (science); Event; Family suidae; Gene Expression Profile; Generations; Heart; Heart failure; Hip region structure; Human; In Situ; In Vitro; Infarction; Ischemia; Kidney; Lentivirus; Liver; Mediating; Methods; Mitotic; Modeling; Mosaicism; Mus; Muscle Cells; Myocardial Infarction; Natural regeneration; Neurons; Oncogenic; Organ; Pancreas; Parents; Persons; Proliferation Marker; Property; Rattus; Recording of previous events; Reperfusion Therapy; Reporter; Rest; Rodent; Safety; Slice; Specificity; System; Technology; Testing; Therapeutic; Time; Tissues; Transgenic Organisms; Treatment Failure; Troponin T; cell type; clinical application; cyclin B1; dosage; efficacy evaluation; efficacy testing; first-in-human; gene therapy; healing; heart function; human old age (65+); improved; in vivo; mouse model; multicatalytic endopeptidase complex; novel; novel strategies; overexpression; porcine model; preclinical efficacy; promoter; protein degradation; safety study; tool; transcriptome sequencing; translational potential; tumorigenic

Heart failure kills 1 in 10 people over the age of 65 in the USA. We recently identified a combination of four cell- cycle regulators that induces stable cytokinesis in adult cardiomyocytes to replace their loss following myocardial infarction. Specifically, adenoviral overexpression of cyclin-dependent kinase 1 (CDK1), CDK4, cyclin B1, and cyclin D1 (collectively known as 4F, i.e. four factors), efficiently induced cell division in post-mitotic mouse, rat, and human cardiomyocytes. Overexpression of the cell-cycle regulators was self-limiting through proteasome- mediated degradation of the protein products in cardiomyocytes. In vivo, lineage tracing using the Mosaic Analysis of Double Marker (MADM) mouse model revealed that 15–20% of adult cardiomyocytes expressing the 4F underwent stable cell division, with significant improvement in cardiac function after acute or subacute myocardial infarction. Currently, our approach is the most robust method to induce cardiomyocyte proliferation; however, clinical applicability in humans is limited by concerns for tumorigenic potential in other organs. Our preliminary data in vitro shows that in other cell types (e.g. neurons), 4F induce cell proliferation continuously for 5-6 successive rounds over 7 days. Therefore, in this proposal we will focus on approaches to make cell cycle induction in cardiomyocytes more clinically applicable and avoid any oncogenic potential. We hypothesize that a transient and cardiomyocyte-specific expression is needed to induce one cycle of cardiomyocyte proliferation to avoid any potential adverse effects on other tissues. Our preliminary data shows that TNNT2 derived 4F expression in NIL is robustly inducing proliferation in vitro and in vivo and improved cardiac function after myocardial infarction. Here, we will rigorously demonstrate the efficacy and safety of this polycistronic NIL encoding 4F expression driven by the cardiac specific promoter of TNNT2 (TNNT-4F-NIL) in inducing cardiomyocyte division and improving cardiac function after myocardial infarction in vivo in rat and pig models as well as in situ in human heart slices from healthy and infarcted hearts. Here, we aim first to fully demonstrate the efficacy and specificity of TNNT-4F-NIL to induce proliferation only in cardiomyocytes in vivo. Secondly, we will test the functional efficacy and perform initial safety studies for TNNT-4F-NIL in rat and pig models of heart failure. Lastly, we will demonstrate the efficacy of TNNT-4F-NIL in inducing adult human cardiomyocyte proliferation in situ in human heart slices and improving contractile function of failing human heart slices. This study will address all preclinical efficacy testing and perform initial safety studies for one of the most promising approaches to regenerate the heart. The successful completion of this study will allow the start of first in human clinical trial.

在美国，心力衰竭杀死了65岁以上10人中的1人。我们最近确定了四个细胞的组合 影响成年心肌细胞中稳定细胞因子的循环调节剂，以替代心肌后损失 梗塞，特定于细胞周期蛋白依赖性激酶1（CDK1），CDK4，细胞周期蛋白B1和 细胞周期蛋白D1（统称为4F，即四个因素），有效诱导了有效的小鼠，大鼠， 和人类心肌细胞。细胞周期调节剂的过表达是通过蛋白酶体进行自限制的 心肌细胞中蛋白质产物的介导降解。在体内，使用马赛克进行谱系跟踪 双标记（MADM）小鼠模型的分析表明，表达的成年心肌细胞中有15-20％ 4F进行了稳定的细胞分裂，急性或亚急性后心脏功能显着改善 心肌梗塞。目前，我们的方法是诱导心肌细胞增殖的最强大方法。 然而，人类中的临床适用性受到其他器官中肿瘤潜力的关注的限制。我们的 体外初步数据表明，在其他细胞类型（例如神经元）中，4F持续诱导细胞增殖 在7天内成功进行了5-6个成功的回合。因此，在此提案中，我们将重点介绍使细胞周期的方法 心肌细胞的诱导更适用，并避免任何致癌潜力。 我们假设需要一种瞬态和心肌细胞特异性表达来诱导一个周期 心肌细胞增殖，以避免对其他组织的任何潜在不利影响。我们的初步数据显示 TNNT2在NIL中衍生的4F表达在体外和体内强烈诱导并改善 心肌梗塞后心脏功能。在这里，我们将严格证明这一点的效率和安全性 由TNNT2的心脏特异性启动子（TNNT-4F-NIL）驱动的4F表达的多源NIL编码4F表达 大鼠和猪体内心肌梗死后诱导心肌细胞分裂和改善心脏功能 由健康和梗塞心脏的人类心脏切片中的模型以及原位。在这里，我们首先要完全 证明TNNT-4F-NIL的效率和特异性仅在体内诱导心肌细胞中诱导增殖。 其次，我们将测试功能效率，并在大鼠和猪中对TNNT-4F-NIL进行初步安全研究 心力衰竭的模型。最后，我们将证明TNNT-4F-nil在诱导的成年人中的效率 心脏切片中的心肌细胞增殖并改善了失败的人心的收缩功能 切片。这项研究将解决所有临床前效率测试，并对最初的安全研究进行最初的安全研究之一 有前途的方法来再生心脏。这项研究的成功完成将允许首次开始 在人类临床试验中。