Abstract 4366989: Pathogenic variants in myoferlin ( <i>MYOF)</i> cause arrhythmogenic cardiomyopathy

Background: Arrhythmogenic cardiomyopathy (ACM) is an inherited disease characterized by fibrofatty ventricular remodeling and arrhythmias. ACM patients are at risk for sudden cardiac death (SCD). Yet, 40% of ACM cases are genetically unexplained. Myoferlin (MYOF) is a transmembrane protein enriched with calcium (Ca2+)-binding C2 domains, highly expressed in skeletal and cardiac muscle. Pathogenic variants in MYOF have recently been linked to arrhythmogenic right ventricular cardiomyopathy (ARVC) in case reports, however, their mechanistic role is unknown. Hypothesis: We hypothesized that MYOF deficiency promotes Ca2+ handling deficits and consequent arrhythmogenesis in ACM. Methods: Via whole-exome sequencing, we identified a heterozygous missense variant in MYOF (c.4912G&gt;A; p.G1654S) that segregated in a family with two male siblings with left- (ALVC) and right-ventricular (ARVC) ACM. Three other individuals with a diagnosis of arrhythmia and cardiomyopathy harboring MYOF p.G1654S were identified in biobanks. Inducible pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) from affected family members and a heterozygous MYOF knockout mouse model were used to investigate mechanisms of MYOF loss-of-function, as well as the MYOF p.G1654S models. Results: MYOF p.G1654S protein degraded faster than WT through a lysosomal pathway. In patient-specific iPSC-CMs harboring MYOF p.G1654S, live Ca2+ imaging identified disrupted Ca2+ handling and increased arrhythmia burden. These findings were confirmed in neonatal rat ventricular myocytes (NRVM) with silenced MYOF expression. Isogenic CRISPR/Cas9 correction of MYOF p.G1654S in iPSC-CMs rescued depleted MYOF levels and abnormal Ca2+ handling. Using computational modeling, proximity ligation assay, and co-immunoprecipitation, we identified an interaction between MYOF and L-type voltage-gated Ca2+ channel CaV1.2. Ca2+ handling deficits and arrhythmias were rescued with verapamil. A cardiac-specific heterozygous MYOF knockout mouse model showed systolic dysfunction at 3 months of age and altered electrical phenotypes. Conclusion: We identified at least six cases with disruptions in MYOF: two previously published case studies, one newly identified family, and three unrelated cases. We found that loss of MYOF alters Ca2+ handling, contributing to an arrhythmogenic substrate via disruption of MYOF-CaV1.2 interaction. These results establish MYOF as a novel causative gene in ACM and a new therapeutic target.