Assessment and Validation of a Phenotype-Enhanced Variant Classification Framework to Promote or Demote RYR2 Missense Variants of Uncertain Significance

Background Many rare, potentially pathogenic, RYR2 variants identified in individuals with clinically definite catecholaminergic polymorphic ventricular tachycardia are classified ambiguously as variants of uncertain significance (VUS). We aimed to determine if a phenotype-enhanced variant classification approach could reduce the burden of RYR2 VUS encountered during clinical genetic testing. Methods This retrospective study was conducted in 84 RYR2-positive individuals from the Mayo Clinic (Rochester, MN) and validated in 149 RYR2-positive individuals from Amsterdam University Medical Center (Amsterdam, NL). Using a newly developed diagnostic scorecard, the pretest clinical probability of catecholaminergic polymorphic ventricular tachycardia was determined for all RYR2-positive individuals. Each RYR2 variant was then readjudicated using a phenotype-enhanced American College of Medical Genetics approach that incorporates new criteria that reflect the phenotypic strength associated with each individual RYR2 variant. Results Overall, 72 distinct RYR2 variants were identified among the 84 Mayo Clinic (39 unique) and 149 Amsterdam University Medical Center (30 unique) cases. Three variants were present in both cohorts. American College of Medical Genetics guidelines classified 47% of all RYR2 variants as VUS. In the Mayo Clinic cohort, readjudication using amended phenotype-enhanced American College of Medical Genetics standards dropped the VUS rate significantly (20/42 [48%] versus 3/42 [7%]; P<0.001) with 13/20 (65%) RYR2 VUS promoted to likely pathogenic and 4/20 (20%) demoted to likely benign. A similar drop in VUS rate (14/33 [42%] versus 3/33 [9%]; P=0.001) was observed in the Amsterdam University Medical Center validation cohort with 10/14 (71%) RYR2 VUS promoted to likely pathogenic and 1/14 (7%) demoted to likely benign. Conclusions This multicenter study illustrates the potential utility of phenotype-enhanced variant classification in catecholaminergic polymorphic ventricular tachycardia.