Kieran Paddock

Introduction: Familial hypertrophic cardiomyopathy (HCM) is a genetic disease characterized by hypertrophy of the ventricular wall, causing obstruction of the left ventricular outflow tract (LVOT).^1–3 It is the most common cause of sudden cardiac death in athletes.² It is predominantly due to mutations in MYBPC3 for myosin binding protein C (MyBPC), which normally inhibits the crossbridge cycle.^2,3 Calcium and phosphorylation levels regulate the activity of MyBPC in the sarcomere,⁴ and it is proposed that MyBPC helps stabilize the energy-conserving “super-relaxed” (SRX) state of myosin.^5,6 Standard treatment for HCM aims to reduce symptoms and sudden cardiac death risk,^1–3,7 though recent research elucidates myosin ATPase and gene expression dysfunction as potential therapeutic targets.^5,8,9

Methods: Myofilament sliding assays involved using total internal reflectance microscopy to visualize sliding velocity in the base sarcomere as well as after adding several molecules of interest, including a MyBPC fragment, a phosphomimetic MyBPC fragment, and a myosin ATPase inhibitor, mavacamten.^4,10 MyBPC fragment conformation was visualized with electron microscopy.¹⁰ Fluorescence decay assays were used to quantify the DRX and SRX states of myosin.^5,6 RNA sequencing was performed in induced pluripotent stem cells in both HCM and control cell lines.⁹ Mavacamten was administered to obstructive HCM participants over a 30-week trial controlled by placebo.⁸

Results: Myofilament sliding assays revealed decreased calcium sensitivity in HCM tissue at physiologic calcium levels and increased calcium sensitivity above physiologic calcium levels.⁴ Myofilament sliding assays also showed phosphorylation of MyBPC enhanced sliding velocity, reversed by a conformation change in MyBPC caused by high calcium levels.¹⁰ HCM tissues had higher levels of dephosphorylation than healthy tissues.⁴ Mavacamten recovered shortening/relaxation kinetics in HCM tissue.⁵ SRX myosin was found to be decreased in HCM tissue specific to MYBPC3 mutations,^5,6 and mavacamten recovered the SRX content.⁵ RNA sequencing revealed a unique gene signature to HCM MYBPC3 cell lines, including upregulation of the RNA metabolism (NMD) pathway, calcium signaling, and hypertrophy genes, and downregulation of cytosolic calcium export pumps.⁹ NMD inhibition normalized gene expression and calcium handling in HCM cell lines.⁹ Mavacamten in HCM participants resulted in greater functional improvements as compared to placebo, including LVOT gradients & NYHA classification.⁸

Conclusions: Decreased calcium sensitivity and compensatory dephosphorylation in HCM tissue represent dysfunction of regulatory mechanisms on MyBPC. Decreased SRX content conveys a more highly active myosin ATPase, which after inhibition with mavacamten recovered shortening/relaxation kinetics and SRX content. A dysfunctional gene signature could represent the base dysfunction in HCM, correctable by NMD inhibition. Mavacamten shows promise in improving functional status in individuals with HCM.

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