NEW GENES AND DISEASES / NGS & RELATED TECHNIQUES: P.328 Neuromuscular disease variant of unknown significance (VUS) resolution using muscle biopsy evaluation: The Iowa experience

Abstract

Whole exome sequencing and large next generation sequencing panels are being performed frequently in muscular dystrophy (MD) and other neuromuscular disease patients. These tests often reveal numerous variants of unknown significance (VUS) that result in diagnostic uncertainty. A potential means to resolve the pathogenicity of VUS is to evaluate muscle biopsy pathology. The purpose of this study was to investigate the efficacy of our extended muscle biopsy workup in interpreting VUS. Muscle biopsies accessioned at the University of Iowa between November 2015 and December 2019 for the purpose of evaluating VUS were reviewed. The 144 cases included in the study came from 31 states or provinces of the USA and Canada. A total of 91 different genes had a VUS. Individual patients had 1 to 8 VUS, with an average of 1.8 VUS/patient. Thirty-three MD genes had a VUS, 9 of which were dystroglycanopathy-related. Genes causing non-dystrophic myopathies (17) and mitochondrial diseases (9) were also commonly represented. The most frequent VUS involved RYR1 (18) and CAPN3 (15) with COL6A3, DYSF, and TTN having 14 each. Variants in the three genes encoding components of collagen VI made up the largest single disease group with 33 cases. The workup to evaluate VUS was tailored to the individual patient history, muscle biopsy histopathology, and genetic testing data, and included histochemistry, immunostaining, electron microscopy, and/or western blotting. Using these techniques, 33% of cases showed evidence that one of the VUS was likely to be disease-causing. Overall, VUS in CAPN3, DMD, and RYR1 were the most frequently solved as pathogenic (9, 7, and 4 cases, respectively). In 35% of cases, the workup suggested that the VUS were not likely to be pathogenic. Interestingly, in 12% of cases, a genetic MD or other myopathy was suggested that was completely unrelated to the VUS provided. For example, dystrophinopathy was diagnosed in 4 cases, despite molecular genetics reports of VUS in other unrelated genes. In the remaining 20% of cases, the workup was unable to provide guidance about the VUS. Our data suggest that muscle biopsy testing can provide information critical to determining pathogenicity in 80% of neuromuscular disease cases with a VUS.