Cohort-scale automated patch clamp data improves variant classification and penetrance stratification for SCN5A-Brugada Syndrome

Abstract

BackgroundBrugada Syndrome (BrS) is an inherited arrhythmia disorder that causes an elevated risk of sudden cardiac death. Approximately 20% of patients with BrS have rare variants in SCN5A, which encodes the cardiac sodium channel NaV1.5. Genetic workup of BrS is often complicated by SCN5A variants of uncertain significance (VUS) and/or incomplete penetrance. MethodsWe analyzed all 252 missense and in-frame insertion/deletion SCN5A variants from a previously published large cohort of BrS cases (n=3,335 patients) using a calibrated high- throughput automated patch clamp (APC) assay. Variant functional Z-scores were assigned evidence levels ranging from BS3_moderate (normal function) to PS3_strong (loss-of-function), as defined by American College of Medical Genetics and Genomics criteria. Functional evidence was combined with population frequency, hot-spot, case counts, protein length changes, and in silico predictions. Odds ratios of BrS case-control enrichment and penetrance for BrS were calculated from variant frequencies in the BrS cohort and in gnomAD. ResultsMost variants (146/252) were functionally abnormal (Z [≤] -2), with 100 having severe loss-of-function (Z [≤] -4). Functional evidence enabled the reclassification of 110 of 225 VUS; 104 to likely pathogenic and 6 to likely benign. SCN5A variants with loss-of-function were mainly localized to the transmembrane domains, especially the regions comprising the central pore. SCN5A variant penetrance was proportional to the severity of loss-of-function; variants with Z [≤] -6 had penetrance of 24.5% (15.9 - 37.7% CI) and an odds ratio of 501 for BrS. ConclusionsThis cohort-scale APC dataset stratifies SCN5A variants found in BrS patients into normal function "bystander" variants that have a low risk for BrS and loss-of-function variants that have a high risk for BrS. Functional data can be integrated with other criteria to reclassify a substantial fraction of VUS. The dataset helps clarify the SCN5A-BrS relationship and will improve the diagnosis and clinical management of BrS probands and their families.