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

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.

Grant Number: 5R01HL162919-04
NIH Institute/Center: NIH
Principal Investigator: Robert Blanton