Calcium Release Channel Deficiency Syndrome

PROJECT SUMMARY Potentially lethal cardiac channelopathies associated with pathogenic variants in the RYR2-encoded cardiac ryanodine receptor type 2 (RyR2)/calcium release channel (CRC) are the pathogenic basis for a significant portion of autopsy-negative sudden unexplained death in the young (SUDY). RYR2 gain- of-function (GOF) pathogenic variants account for 60% of autosomal dominant catecholaminergic polymorphic ventricular tachycardia (CPVT1), a potentially lethal heritable arrhythmia syndrome that classically manifests as exercise-induced syncope, sudden cardiac arrest (SCA), or sudden cardiac death (SCD). In 2020, we discovered a novel RYR2 loss-of-function (LOF) mechanism that we have termed calcium release channel deficiency syndrome (CRCDS). We identified a novel homozygous duplication (RYR2-DUP) involving ~26,000 bp of intergenic sequence, RYR2’s 5’UTR/promoter region, and exons 1-4 of RYR2 that is responsible for highly penetrant, exertion-related SCA/SCD in the Amish community without an overt phenotype to suggest RYR2-mediated CPVT1. Unlike typical CPVT1, individuals homozygous for the RYR2 duplication have displayed typically only intermittently prolonged QT intervals or prominent U-waves and typically had completely normal exercise/epinephrine stress tests and normal 24-hour Holter monitoring. Thus, cardiologic tests, such as ECG, stress testing, and echocardiogram, are currently unable to reliably identify or further risk stratify family members likely to be homozygous for the RYR2 duplication. Given the potentially lethal nature of these inheritable RYR2- CRCDS variants and the lack of a robust and measurable clinical phenotype, it is vitally important to better understand the spectrum and contribution of CRCDS-associated RYR2 variants in SUDY and to determine the underlying disease-associated mechanisms in patient-specific re-engineered heart cell models using induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). Here, we propose to 1) determine the frequency of CRCDS due to RYR2 coding region, non-coding region, and structural variants (SVs) in SUDY and 2) develop an international registry for CRCDS to identify genotype/phenotype correlates to assist in clinical diagnosis and management of patients with CRCDS. 3) determine the relative contribution of CRCDS (LOF)-associated RyR2 variants versus CPVT1 (GOF)- variants in SUDY using functional studies, 4) develop a function-based iPSC-CM platform for RyR2 VUS resolution, and 5) determine the compensatory mechanisms related to loss of RYR2 transcript/RyR2 protein expression and function as it relates to calcium handling components and heart cell function during early cardiac development.