Mahoney-Schaefer, M. | bioRxiv Science

bioRxiv2025

Myosin modulator Aficamten inhibits force in cardiac muscle by altering myosin's biochemical activity without changing thick filament structure

BackgroundInhibiting contractility by targeting cardiac myosin is an effective treatment for patients with hypertrophic cardiomyopathy (HCM). Aficamten is a second in class myosin inhibitor with promising clinical data showing improvements in hemodynamics and symptoms in patients with HCM. While it is known that Aficamten inhibits force and cardiomyocyte contractility by stabilizing the weak pre-powerstroke conformation, effects on myosin structure and kinetics during loaded contraction are lacking. MethodsPermeabilized porcine cardiac tissue and myofibrils were used for single-molecule imaging of ATP turn over, X-ray diffraction, and mechanical measurements. Engineered heart tissues from human induced pluripotent stem cell cardiomyocytes were used to evaluate effects on force and contraction kinetics. ResultsIn contrast to Mavacamten, Aficamten does not structurally sequester myosin heads along the thick filament. Aficamten inhibits ATPase activity by shifting myosin heads from higher to slower ATPase state, with the emergence of a super slow biochemical nucleotide turnover state. This results in decreased force and calcium sensitivity without altering cross-bridge cycling. These contractile mechanical changes are comparable to Mavacamten. Our myofibril mechanical assay showed inhibition of force with accelerated relaxation. In EHTs, while Mavacamten and Aficamten inhibit cardiac twitch forces, Mavacamten reduces the activation kinetics while both result in faster relaxation. ConclusionsWe used a combination of biochemical and biomechanical assays to show that Aficamten inhibits myosin ATPase without appreciably altering myosin structure. This is different from Mavacamten that strongly affects both. While both compounds inhibit contractility, differences in mechanisms of action and kinetics of force activation and relaxation could allow use in different patient populations.

biophysics↗

bioRxiv2023

Danicamtiv increases myosin recruitment and alters the chemomechanical cross bridge cycle in cardiac muscle

Modulating myosin function is a novel therapeutic approach in patients with cardiomyopathy. Detailed mechanism of action of these agents can help predict potential unwanted affects and identify patient populations that can benefit most from them. Danicamtiv is a novel myosin activator with promising preclinical data that is currently in clinical trials. While it is known danicamtiv increases force and cardiomyocyte contractility without affecting calcium levels, detailed mechanistic studies regarding its mode of action are lacking. Using porcine cardiac tissue and myofibrils we demonstrate that Danicamtiv increases force and calcium sensitivity via increasing the number of myosin in the "on" state and slowing cross bridge turnover. Our detailed analysis shows that inhibition of ADP release results in decreased cross bridge turnover with cross bridges staying on longer and prolonging myofibril relaxation. Using a mouse model of genetic dilated cardiomyopathy, we demonstrated that Danicamtiv corrected calcium sensitivity in demembranated and abnormal twitch magnitude and kinetics in intact cardiac tissue. Significance StatementDirectly augmenting sarcomere function has potential to overcome limitations of currently used inotropic agents to improve cardiac contractility. Myosin modulation is a novel mechanism for increased contraction in cardiomyopathies. Danicamtiv is a myosin activator that is currently under investigation for use in cardiomyopathy patients. Our study is the first detailed mechanism of how Danicamtiv increases force and alters kinetics of cardiac activation and relaxation. This new understanding of the mechanism of action of Danicamtiv can be used to help identify patients that could benefit most from this treatment.

physiology↗