Cardiac Myosin Binding Protein-C in Development and Reversal of Heart Failure

2017年，心力衰竭（HF）困扰着650万美国人，5年死亡率约为50%，并且可能会增加 到2030年达到800万。在过去的46年里，死亡率没有显著改善， 由于射血分数降低的心力衰竭（HFrEF）中的低血压而停用有效药物， 以及射血分数保留性心力衰竭（HFpEF）的有效治疗方法的缺乏，联合收割机 这表明需要新的治疗方法。心肌肌球蛋白结合蛋白-C（cMyBPC）位于 心脏肌肉的细丝。cMyBPC在其M结构域的磷酸化增加了跨桥循环速率。 由于增加跨桥循环率可以改善收缩性和lustropy，我们假设， cMyBPC的磷酸化提供了一种新的中枢机制，可以预防和治疗由于不同的 可使.使用模拟3个丝氨酸（S）位点去磷酸化和磷酸化cMyBPC的小鼠模型 在M-结构域，我们最近发现cMyBPC磷酸化可以减轻与年龄相关的发育， 的HFpEF和经主动脉缩窄（TAC）手术诱导的HFrEF，如通过改善生存率和更好的 保持舒张功能。因此，cMyBPC磷酸化具有治疗HFrEF和HFrBPC的潜力。 HFpEF。然而，我们发现的证据表明，2个新的S磷酸化位点以外，我们的研究3S网站添加 显著的功能效应。因此，我们建立了2种新的基因敲入小鼠模型， 阐明最大磷酸化cMyBPC（5SD）和去磷酸化cMyBPC（5SA）作用的位点 M域。我们还发现压力应激可以激活粘着斑激酶（FAK）磷酸化 cMyBPC中的酪氨酸（Y）残基。这一发现导致了一个新的支持假说，即压力应力 触发FAK磷酸化cMyBPC以增加收缩性作为代偿反应。与这些 考虑到这些发现，我们打算确定最大M结构域磷酸化的功效，并阐明 新的FAK-cMyBPC机制。目的#1：确定磷酸化的功效和伴随机制 cMyBPC在衰老、压力应激和肥胖的HF诱导条件下保护心脏功能。我们 将用2年老化、TAC或高脂肪饮食来攻击小鼠。我们将使用功能和生物化学的结合 技术来确定潜在的机制。cMyBPC（5SD）小鼠抵抗退化和 使用cMyBPC（5SD）基因治疗逆转衰竭WT心脏的有效性将量化cMyBPC的疗效 磷酸化分别用于预防和治疗。目标#2：阐明信号传导机制， 压力应激诱导的cMyBPC酪氨酸磷酸化的功能结果。我们将确定Y研究中心， 证实FAK-cMyBPC相互作用，并阐明FAK磷酸化cMyBPC对模型的影响， 从完整的乳头肌到新的基因敲入小鼠。影响：疗效结果和机制性见解 从这项研究提供了证据，翻译cMyBPC磷酸化的新的HF治疗。

Heart failure (HF) afflicted 6.5 million Americans in 2017, carries 5-year mortality of ~50%, and will likely increase to > 8 million by 2030. The lack of significant improvement in mortality for the last 46 years, the reality of needing to discontinue effective medications due to hypotension in heart failure with reduced ejection fraction (HFrEF), and the paucity of effective treatment for heart failure with preserved ejection fraction (HFpEF) combine to suggest that new treatments are needed. Cardiac myosin binding protein-C (cMyBPC) resides on the thick filament of the heart muscle. Phosphorylation of cMyBPC at its M-domain increases cross-bridge cycling rate. Because increasing cross-bridge cycling rate can improve both contractility and lusitropy, we hypothesize that phosphorylation of cMyBPC provides a novel central mechanism that can prevent and treat HF due to different causes. Using mouse models that mimic de-phosphorylated and phosphorylated cMyBPC at 3 serine (S) sites in the M-domain, we recently discover that cMyBPC phosphorylation mitigates both aged-related development of HFpEF and trans-aortic constriction (TAC) surgery induced HFrEF, as seen by improved survival and better preservation of diastolic function. Thus, cMyBPC phosphorylation holds potential to treat both HFrEF and HFpEF. However, we find evidence that 2 new S phosphorylation sites outside our study of 3 S sites add significant functional effects. Consequently, we have made 2 new knock-in mouse models to include the new sites for elucidating the effects of maximally phosphorylated cMyBPC(5SD) and dephosphorylated cMyBPC(5SA) M-domain. We also find evidence that pressure stress can activate focal adhesion kinase (FAK) to phosphorylate tyrosine (Y) residue(s) in cMyBPC. This discovery leads to a new supporting hypothesis that pressure stress triggers FAK to phosphorylate cMyBPC to increase contractility as a compensatory response. With these discoveries in mind, we intend to determine the efficacy of maximal M-domain phosphorylation and elucidate the novel FAK-cMyBPC mechanism. Aim #1: Determine the efficacy and companion mechanisms of phosphorylated cMyBPC to preserve cardiac function under HF inducing conditions of aging, pressure stress, and obesity. We will challenge mice with 2-yr aging, TAC, or high fat diet. We will use a combination of functional and biochemistry techniques to determine the underlying mechanisms. Ability of cMyBPC(5SD) mouse to resist deterioration and effectiveness of using cMyBPC(5SD) gene therapy to reverse failing WT hearts will quantify the efficacy of cMyBPC phosphorylation for prevention and treatment respectively. Aim #2: Elucidate signaling mechanism and functional results of pressure stress induced tyrosine phosphorylation of cMyBPC. We will identify Y site(s), confirm FAK-cMyBPC interaction, and elucidate effects of FAK phosphorylating cMyBPC on models ranging from intact papillary muscle to new knock-in mouse. Impact: Efficacy results and mechanistic insights stemming from this study provide evidence for translating cMyBPC phosphorylation to new HF treatment.