Mechanism-inspired Strategies to Prevent Pathogenic Late Na Current in Cardiac Arrhythmias

PROJECT SUMMARY PI: Manu Ben-Johny, Ph.D. NaV1.5 channels are fundamentally involved in the normal function and pathophysiology of the heart. NaV1.5 dysfunction is linked to a variety of life-threatening cardiac diseases, including congenital and acquired cardiac arrhythmias, cardiomyopathies, and heart failure. An emerging commonality for these pathologies is increased late Na current, that results in sustained Na influx during the plateau phase of the cardiac action potential. Understanding mechanisms that regulate late Na current is pivotal to understanding NaV1.5 dysfunction in cardiac pathophysiology and for developing long-sought pharmacology. Our recent studies and preliminary data suggest that late Na current is powerfully tuned by a Na channel modulator named fibroblast growth factor homologous factor (FHF) that is endogenous to cardiomyocytes. Yet, how FHF accomplishes this important mode of NaV1.5 regulation and its relevance to disease pathogenic mechanisms is not fully determined. In this collaborative project, we seek (1) to dissect the molecular mechanism of FHF regulation of late Na current, and (2) to identify the physiological and pathophysiological relevance of this modulatory scheme. Armed with in depth mechanistic insights, we seek to engineer novel peptide-based inhibitors of late Na current. In particular, as FHF undergoes extensive alternative-splicing, we evaluate isoform-specificity of late Na current regulation using a novel FRET assay and through extensive single-channel analysis. To probe the physiological impact of FHF regulation of late Na current, we virally manipulate FHF levels in cardiomyocytes differentiated from long-QT and mixed-syndrome patient-derived induced pluripotent stem cells as well as transgenic mouse ventricular myocytes. In so doing, this proposal promises new biophysical and physiological insights into modulation of cardiac NaV1.5 and inform upon mechanisms underlying variable clinical manifestations of Na channelopathies.