Experiments proposed in Project 1 test the hypothesis that altered signaling at the level of sarcomeric proteins is a significant element in maladaptation triggered by genetic and acquired hemodynamic stressors and an important target for biased ligands as a therapeutic intervention. Strong synergistic interactions of Project 1 continue with other projects focused on sarcomere growth and remodeling (Project 2), and maladaptive sarcomeric mechanisms related to control of cardiac dynamics (Project 3). Project 1 relies critically on the support from Core B (Human Cell and Tissue) and Core C (Proteomics and Analytical Biochemistry). We focus on maladaptive responses to stresses inducing dilated cardiomyopathy and the effects and mechanisms of treatment with biased ligands, which signal specifically through ?-arrestin via the angiotensin II receptor (AT1R). Our lab is the first to report that unlike commonly used angiotensin receptor blockers (ARBs), eg. Losartan, a biased ligand acting as an ARB is able to promote contractility via ?-arrestin dependent sarcomere signaling, which alters protein phosphorylation. In view of the potential for this mechanism to ameliorate the progression to DCM, we propose to determine the effects and mechanisms of action of biased ligands as a therapy for familial and acquired DCM.
The Specific Aims are:
Aim #1 To compare the ability of losartan and biased ligands to reverse established maladaptations developed in mouse models of acquired and familial DCM and in preparations from non-failing and failing human preparations (Core B).
Aim #2 To determine the mechanism(s) of the effects by which signaling via ?-arrestin alters myofilament Ca-response and contractility.
Aim #3 To test the effectiveness of biased ligands on restoring structure and function of patient derived human cardiac myocytes generated from inducible pluripotent stem cells (iPSC-CM) obtained from controls (unaffected family members) and patients expressing DCM-linked mutant cardiac TnT-R173W. Our approaches include readouts of EKG, echo-cardiography, P-V loops, blood pressure, apoptosis, biomarkers, and tension/ intra-cellular Ca2+ transients; extensive proteomic analysis in conjunction with Core C; and, established culture techniques and determination of intracellular [Ca2+], shortening and shortening velocity of normal and cTnT mutant iPSC-CM cells. Our proposed studies investigate a novel mechanism of signaling to sarcomeres with a strong potential for translation to therapies for DCM.
Dilated cardiomyopathy is a serious, prevalent (1 in 250) cardiac disorder in which the cause is either unknown, acquired by lifestyle, or linked to mutations, many of which encode proteins in sarcomeres, the molecular machine responsible for contraction of the heart. We have discovered a novel mechanism of for repairing the defect in function of sarcomeres involving therapeutic agents with a realistic chance of clinical development and application to personalized medicine. In this project we expand our studies to include human heart tissue, and patient specific heart cells derived from pluripotent stem cells induces from skin fibroblasts.
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