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

cGMP依赖性蛋白激酶1 α（PKG 1 α）对抗病理性左心室（LV）肥大 通过在心肌细胞（CM）中的作用和重塑，并通过促进血管生成来调节血压。 平滑肌细胞（VSMC）舒张和血管舒张。激活PKG 1的药物，包括硝酸盐， 沙库巴曲/缬沙坦、vericiguat和其他药物可改善心力衰竭伴左室射血减少的死亡率 因此，这是HFrEF治疗的核心进展。然而，这些疗法仍然存在。 HFrEF疗效不完全。此外，PKG 1诱导的血管舒张引起的低血压严重 限制了这些药物的实际应用。这个项目的首要假设是， LV中的下游PKG 1 α抗重塑底物可以揭示新的治疗候选物， 目前PKG 1激活药物的这些关键局限性。混合谱系激酶3（MLK 3） 作为一种新的PKG 1 α相互作用蛋白和抗重塑分子。我们建议探索以下令人兴奋的 这些发现将MLK 3鉴定为HFrEF中的免疫相关分子。1)PKG 1 α-MLK 3相互作用 衰竭LV的下降，MLK 3是PKG 1 α介导的西地那非对LV的治疗作用所必需的 压力超负荷后的功能，从而确定PKG对心肌MLK 3调节的破坏是促进作用 HF患者的LV重塑和PKG 1激活药物疗效降低。2)MLK 3激酶功能与 病理性CM和LV功能障碍和重塑，但不影响体内血压。3)MLK 3缺失 在体内促进高血压，但MLK 3通过MLK 3激酶独立地调节血压 与PKG 1 α的信号转导无关。我们建议测试一个两部分的新模型，其中1） PKG 1 α激活MLK 3通过MLK 3激酶依赖性途径促进LV对压力超负荷的代偿 2）MLK 3通过激酶非依赖性作用对抗高血压， 刚度通过在VSMC中的作用。SA 1将使用我们实验室开发的新型MLK 3细胞特异性缺失模型 确定MLK 3在LV功能和血压的基础调节中的CM和SMC特异性作用， 慢性左心室对压力超负荷或心肌梗死的反应。SA 2将决定激酶 MLK 3对LV功能和血压的依赖性与激酶非依赖性作用。SA 3将确定 通过检测MLK 3对LV治疗效果的要求，确定MLK 3与HF治疗的翻译相关性 与目前可用的PKG 1激活药物的血压效应相比。这些研究将定义新的 MLK 3调节血压的机制，以及MLK 3调节血压的机制- 独立的功能介导目前PKG 1激活药物的治疗效果。这些研究有 有可能将MLK 3激酶激活确定为促进PKG 1治疗效果的新型治疗策略 对LV功能和重塑的影响，但避免不必要的低血压，这限制了PKG 1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.