Investigating Mixed Lineage Kinase 3 as a blood pressure-independent protein kinase G1 effector in heart failure

CGMP依赖的蛋白激酶1α(PKG_1α)对抗病理性左室肥厚 通过在心肌细胞(CM)中的作用而重塑，并通过促进血管来调节血压 平滑肌细胞(VSMC)的松弛和血管扩张。激活PKG1的药物，包括硝酸盐， 萨舒比利/valsartan、vericuat和其他药物通过减少左心室射血来改善心力衰竭的死亡率。 分数(HFrEF)，因此代表了HFrEF治疗的中心进展。然而，这些疗法仍然存在 受限于HFrEF的不完全疗效。此外，PKG1诱导的血管扩张导致的低血压已经严重 限制了这些制剂的实际使用。这一计划的首要假设是，识别 LV中的下游PKG1α抗重构底物可以揭示需要克服的新的治疗方案 这些是当前PKG1激活药物的关键局限性。我们已经鉴定出混合血统激酶3(MLK3) 作为一种新的PKG1型α相互作用蛋白和抗重构分子。我们打算探索以下令人兴奋的事情 发现MLK3在HFrEF中是一个翻译相关的分子。1)PKG1MLK3与α的相互作用 西地那非的治疗作用与PKG1MK3在α介导下的治疗作用有关 压力超负荷后的功能，从而将PKG对心肌MLK3调节的破坏识别为促进 心衰时左室重构及PKG1激动剂疗效降低。2)MLK3激酶功能相反 病理性CM和LV功能障碍和重塑，但不影响体内血压。3)MLK3缺失 在体内促进高血压，但MLK3对血压的调节是通过MLK3非依赖性的 机制，不依赖于PK1α的信号传导。我们建议测试一个由两部分组成的新模型，其中1) PKG1MLK3激活通过依赖MLK3的α途径促进左心室对压力超负荷的代偿 CM的作用机制；2)MLK3通过对血管的非依赖性作用对抗高血压 在VSMC中扮演一个角色的僵硬。SA1将使用我们实验室开发的新的MLK3细胞特异性缺失模型 为了确定MLK3在基础调节左心功能和血压中的CM和SMC特异性作用以及 在慢性左心室对压力超负荷或心肌梗死的反应。SA2将决定该激酶 MLK3对左心功能和血压的依赖性和非依赖性效应。SA3将决定 从MLK3对LV治疗效果的要求看MLK3与心力衰竭治疗的翻译相关性 对比目前可用的PKG1激活药物对血压的影响。这些研究将定义小说 MLK3调节血压和MLK3血压的机制- 独立的功能介导了当前PKG1激活药物的治疗效果。这些研究具有 确认MLK3激活作为促进PKG1治疗效果的新治疗策略的可能性 对左心功能和重构，但避免不良的低血压，它有有限的PKG1激活药物。

The cGMP-dependent protein kinase 1 alpha (PKG1α) opposes pathological left ventricular (LV) hypertrophy and remodeling via roles in the cardiac myocyte (CM) and regulates blood pressure by promoting vascular smooth muscle cell (VSMC) relaxation and vasodilation. Drugs which activate PKG1, including nitrates, sacubitril/valsartan, vericiguat, and others have improved mortality in heart failure with reduced LV ejection fraction (HFrEF) and thus represent a central advance in HFrEF treatment. However, these therapies remain limited by incomplete efficacy in HFrEF. Furthermore, hypotension from PKG1-induced vasodilation has severely limited the practical use of these agents. The overarching hypothesis of this program is that identifying downstream PKG1α anti-remodeling substrates in the LV can reveal novel therapeutic candidates to overcome these critical limitations of current PKG1-activating drugs. We have identified Mixed Lineage Kinase 3 (MLK3) as a novel PKG1α-interacting protein and anti-remodeling molecule. We propose to explore the following exciting findings which identify MLK3 as a translationally relevant molecule in HFrEF. 1) PKG1α-MLK3 interaction declines in the failing LV, and MLK3 is required for PKG1α-mediated therapeutic effects of sildenafil on LV function after pressure overload, thus identifying disruption of myocardial MLK3 regulation by PKG as promoting LV remodeling and decreasing the efficacy of PKG1-activating drugs in HF. 2) MLK3 kinase function opposes pathological CM and LV dysfunction and remodeling but does not affect blood pressure in vivo. 3) MLK3 deletion promotes hypertension in vivo, but MLK3 regulation of blood pressure occurs through MLK3 kinase independent mechanisms and independently of signaling by PKG1α. We propose to test a two-part novel model in which 1) PKG1α activation of MLK3 promotes LV compensation to pressure overload through MLK3 kinase-dependent mechanisms in the CM; and 2) MLK3 opposes hypertension through kinase-independent effects on vascular stiffness through a role in the VSMC. SA1 will use novel MLK3 cell-specific deletion models developed in our lab to determine the CM and SMC-specific roles of MLK3 in basal regulation of LV function and blood pressure and in the chronic LV response to pressure overload or myocardial infarction. SA2 will determine the kinase dependent versus kinase independent effects of MLK3 on LV function and blood pressure. SA3 will determine the translational relevance of MLK3 to HF treatment by testing the requirement of MLK3 for LV therapeutic effects versus blood pressure effects of currently available PKG1-activating drugs. These studies will define novel mechanisms through which MLK3 regulates blood pressure and through which MLK3 blood pressure- independent functions mediate the therapeutic effect of current PKG1-activating drugs. These studies have the potential to identify MLK3 kinase activation as a novel therapeutic strategy to promote PKG1 therapeutic effects on LV function and remodeling but avoid undesired hypotension which has limited PKG1 activating drugs.