Basic/Translational Science -> Cell Physiology, Pharmacology, and Signaling D-PO02 - Poster Session II (ID 47) Poster

D-PO02-026 - Characterization Of The Calcium Release Channel Deficiency Syndrome Secondary To A Homozygous, Multi-exon Duplication In Ryr2 (ID 985)


Background: We recently discovered Calcium Release Channel Deficiency Syndrome (CRCDS) stemming from a homozygous multi-exon RYR2 duplication mutation associated with exertion-related sudden cardiac arrest (SCA) in two large Amish pedigrees. The homozygous duplication precipitates a dramatic reduction in RyR2 protein translation, a loss-of-function in calcium handling, and a calcium-induced calcium release apparatus that is insensitive to catecholamines and caffeine.
Objective: To characterize functionally patient-derived induced pluripotent stem cell-cardiomyocytes (iPSC-CMs) from two unrelated patients.
Methods: Homozygous RYR2-duplication (RYR2-DUP) iPSC-CMs were generated from two unrelated patients (12-year-old female and a 10-year-old male) with exertional SCA and an unrelated control. Patch-clamp electrophysiology was used to assess action potential duration (APD) before and after 100 nM Isoproterenol (ISO). Arrhythmic activity before and after ISO and response to drug-therapy (nadolol [10µM], propranolol [1µM], and flecainide [6µM]) was assessed using an xCELLigence® RTCA CardioECR instrument.
Results: There were no differences among control and both patient’s iPSC-CMs in terms of APD at 50% of repolarization. However, one of the patient’s iPSC-CMs revealed a significantly prolonged APD90 compared with control iPSC-CMs (612 ± 69 ms vs. 421 ± 46 ms, p<0.05). Delayed afterdepolarization (DAD) was observed in patient iPSC-CMs but not in controls. Compared to control-iPSC-CMs (4%), there was a significantly elevated arrhythmic activity in the RYR2-DUP-iPSC-CMs (patient 1, 10%, p<0.001 and patient 2, 17%, p<0.0001) in response to ISO. Nadolol, propranolol, and flecainide reduced erratic activity by 8.5 fold, 6.8 fold, and 2.4 fold, respectively.
Conclusion: We demonstrate that the homozygous multi-exon RYR2 duplication-mediated CRCDS leads to a pathogenic substrate that is prone to DAD arrhythmic activity. While flecainide alone partially rescued the phenotype, both nadolol and propranolol were able to fully rescue the ISO treated patient iPSC-CMs by significantly reducing the arrhythmic activity of the cells below the baseline arrhythmic level of the control iPSC-CMs.