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，十分之一的 65 岁以上老人死于心力衰竭。我们最近发现了四种细胞的组合。 循环调节剂，诱导成体心肌细胞稳定的胞质分裂，以取代心肌细胞的损失 具体来说，腺病毒过度表达细胞周期蛋白依赖性激酶 1 (CDK1)、CDK4、细胞周期蛋白 B1 和 cyclin D1（统称为4F，即四个因子），有效诱导有丝分裂后小鼠、大鼠、 和人类心肌细胞的过度表达通过蛋白酶体进行自我限制。 介导心肌细胞中蛋白质产物的降解，使用马赛克进行谱系追踪。 双标记 (MADM) 小鼠模型分析表明，15-20% 的成年心肌细胞表达 4F进行稳定的细胞分裂，急性或亚急性后心功能明显改善 目前，我们的方法是诱导心肌细胞增殖的最有效方法； 然而，由于担心其他器官的致瘤潜力，其在人类中的临床适用性受到限制。 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 因此，在本提案中，我们将重点关注进行细胞周期的方法。 心肌细胞诱导更具有临床应用性，并且避免任何致癌潜力。 我们发现需要一种短暂的心肌细胞特异性表达来诱导一个周期 我们的初步数据显示，心肌细胞增殖可以避免对其他组织产生任何潜在的不利影响。 TNNT2 衍生的 4F 在 NIL 中的表达在体外和体内强烈诱导增殖，并改善 在这里，我们将严格证明这一点的有效性和安全性。 多顺反子 NIL 编码 4F 表达，由 TNNT2 的心脏特异性启动子 (TNNT-4F-NIL) 驱动 诱导大鼠和猪心肌梗塞后心肌细胞分裂并改善心功能 模型以及来自健康和梗塞心脏的原位心脏切片，我们的目标首先是充分利用。 证明 TNNT-4F-NIL 仅在体内诱导心肌细胞增殖的功效和特异性。 其次，我们将测试TNNT-4F-NIL在大鼠和猪中的功能功效并进行初步安全性研究 最后，我们将证明 TNNT-4F-NIL 在诱导成人心力衰竭模型中的功效。 心肌细胞在人体心脏切片中原位增殖并改善衰竭人体心脏的收缩功能 这项研究将解决所有临床前功效测试，并对最重要的之一进行初步安全性研究。 这项研究的成功完成将开启第一个心脏再生的前景。 在人体临床试验中。