A Novel in-vitro Patient-Specific Re-Engineered Cardiomyocyte Model of Multifocal Ectopic Purkinje-Related Premature Contractions

Introduction: Multifocal ectopic Purkinje-related premature contractions (MEPPC), the most recent addition to the spectrum of SCN5A -mediated cardiac channelopathies, is characterized by frequent premature ventricular complexes (PVCs) originating from the Purkinje system, atrial arrhythmias, and a predilection for PVC-mediated dilated cardiomyopathy. Hypothesis: To investigate the electrophysiology phenotype of MEPPC in a novel, patient-specific, re-engineered cardiomyocyte model. Methods: Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) were generated from a 27-year-old male with a history of frequent, multifocal PVCs and atrial tachycardia, positive for a putative MEPPC-causative SCN5A variant (p.Q419P). A CRISPR-Cas9 Q419P variant-corrected isogenic control (IC) was created to establish the normalized electrophysiology recordings for that MEPPC model. The standard whole-cell patch clamp technique was used to measure Q419P-SCN5A- and IC-derived NaV1.5 sodium currents using voltage clamp configuration. In addition, action potential duration (APD) of the iPSC-CMs were measured using FluoVolt optical voltage-sensing dye. Results: Analysis of the current-voltage relationship revealed that neither peak sodium current densities nor late inward sodium current were impacted by Q419P-SCN5A. However, Q419P-SCN5A significantly right-shifted the V 1/2 in inactivation by 8.3 mV from -81.1±1.7 mV (IC, n=5) to -72.8±0.2 mV (Q419P-SCN5A, n=6, p<0.05 vs. IC) and left-shifted the V 1/2 in activation by -4.1 mV from -39.5±0.3 mV (IC, n=5) to -43.6±0.9 mV (Q419P-SCN5A, n=6, p<0.05 vs. IC) leading to a larger hyperpolarized window current. Optical APD90 measurement revealed no difference between the Q419P-containing (476 ± 23 ms) and the IC (474 ± 46 ms, p=0.867) cardiomyocytes. Conclusions: This is the first in-vitro disease model of MEPPC using patient-specific, re-engineered heart cells. Consistent with the findings in the non-cardiac HEK cell system, this MEPPC-causative variant contributes to increased window current in cardiomyocytes without affecting the cardiac APD. This novel cardiac cell model for MEPPC may facilitate new insights into the pathobiology of MEPPC and enable the testing of novel therapies for it.