Detyrosinated microtubules in cardiomyocyte mechanics

项目摘要 心力衰竭的一个常见且目前难治的特征是心脏组织的硬化，其损害了心脏的功能。 心脏放松的能力。微管细胞骨架有助于心肌细胞的内部硬度， 并且在某些条件下可以阻碍心肌细胞收缩和舒张的能力。通过第一 五年的R01，我们发现心肌细胞的硬度受到翻译后的严格调控， 微管的脱酪氨酸，并且脱酪氨酸的微管在人心脏中持续升高， 衰竭，伴随心肌硬度增加。我们还发现， 微管足以降低硬度并改善心肌细胞的收缩和舒张， 来自不同形式心力衰竭患者的心肌组织。我们进一步鉴定了 负责心脏中的脱酪氨酸，并表明靶向这种酶足以有力地 改善衰竭的人类心脏细胞的松弛。因此，去酪氨酸形成了一种有前途的新的治疗方法， 治疗心力衰竭的靶点。这项拟议中的研究将测试遗传或小的假设， "酪氨酸化周期"的分子靶向可以稳定地改善不同的心肌细胞的收缩和舒张功能。 小型和大型心力衰竭动物模型。三个目标下的研究将涉及以下几个组成部分： 这个假设。在目标1中，我们将探索是否有一种基因治疗方法， 酶（TTL）足以改善心力衰竭遗传小鼠模型的收缩功能， 射血分数保留心力衰竭手术模型舒张功能改善Aim 2实验 将专注于一种不同的治疗方式，包括新的和高效的小分子抑制剂， 脱酪氨酸酶（VASH）。我们将评估这些新型抑制剂的药代动力学， 它们在啮齿动物和哺乳动物中减少脱酪氨酸和改善心脏功能的耐受性和功效， 人体细胞和组织。在目标3中，我们将探索更大的动物研究，并测试是否 靶向去酪氨酸足以改善患有肥厚性心肌病的猫的心肌细胞和心肌功能。 心肌病和射血分数正常的心力衰竭。我们的跨物种、多尺度和 多管齐下的方法将平衡我们的简化严谨和综合相关性的目标，最终 进一步临床翻译。总之，这项工作将确定靶向脱酪氨酸微管是否可以稳定地 改善心力衰竭中心脏功能，并鉴定可适用于 进入临床管道。

Project Summary A common and currently intractable feature of heart failure is the stiffening of cardiac tissue that impairs the heart's ability to relax. The microtubule cytoskeleton contributes to the internal stiffness of heart muscle cells, and under certain conditions can impede the ability of cardiomyocytes to both contract and relax. Over the first five years of this R01, we found that cardiomyocyte stiffness is tightly regulated by post-translational detyrosination of microtubules, and that detyrosinated microtubules are consistently elevated in human heart failure, concomitant with increased myocardial stiffness. We also found that reducing detyrosinated microtubules is sufficient to lower stiffness and improve contraction and relaxation in cardiomyocytes and myocardial tissue from patients with diverse forms of heart failure. We further identified the enzyme responsible for detyrosination in the heart, and showed that targeting this enzyme is sufficient to robustly improve relaxation in failing human heart cells. As such, detyrosination forms a promising new therapeutic target for the treatment of heart failure. The proposed research will test the hypothesis that genetic or small molecule targeting of the “tyrosination cycle” can stably improve both systolic and diastolic function in different small and large animal models of heart failure. Studies under three aims will address several components of this hypothesis. In Aim 1, we will explore whether a gene therapy approach overexpressing the tyrosinating enzyme (TTL) is sufficient to improve systolic function in a genetic mouse model of heart failure, and to improve diastolic function in surgical model of heart failure with preserved ejection fraction. Aim 2 experiments will focus on a different therapeutic modality consisting of novel and highly potent small molecule inhibitors of the detyrosinating enzyme (VASH). We will evaluate the pharmacokinetics of these novel inhibitors and test their tolerability and efficacy for reducing detyrosination and improving cardiac function in both rodent and human cells and tissues. In Aim 3, we will move our exploration to larger animal studies and test whether targeting detyrosination is sufficient to improve myocyte and myocardial function in cats with hypertrophic cardiomyopathy and with heart failure with preserved ejection fraction. Our cross-species, multi-scale and multi-pronged approach will balance our goals of reductionist rigor and integrative relevance that ultimately furthers clinical translation. Together, this work will determine if targeting detyrosinated microtubules can stably improve cardiac function in heart failure, and identify therapeutic compounds that may be suitable for progression into a clinical pipeline.