Whole-Genome Sequencing in Diagnostics of Unsolved Cases of Inherited Retinal Diseases

The study purpose was to assess the diagnostic value of whole-genome sequencing (WGS) in identifying candidate genetic variants in patients with inherited retinal diseases (IRDs). We investigated 107 unrelated Russian patients with retinal diseases who carried single pathogenic variants in any of nine common autosomal recessive IRD genes (ABCA4, USH2A, EYS, CRB1, RPE65, CNGB3, CNGA3, CEP290, and GUCY2D). WGS was performed for all patients; candidate splice variants were functionally validated using minigene assays. The total diagnostic yield was 40.2% (43/107). In 25.2% of cases (27/107), the diagnosis was established by identifying a second variant in one of the following candidate genes: EYS (n = 9), USH2A (n = 8), ABCA4 (n = 5), CNGB3 (n = 3), CRB1 and CEP290 (n = 1 each). In 15.0% of cases (16/107), the diagnosis was reclassified by variants in alternative IRD genes (autosomal dominant, X-linked, mitochondrial, or syndromic). The majority of second variants detected in AR cases were CNVs (55.5%, 15 patients), followed by non-coding variants (26.0%, seven), structural rearrangements (14.8%, four) and exonic point variant (3.7%, one). Two recurrent CNVs were observed: a gross deletion of exons 22-24 in USH2A (n = 4) and a deletion partially affecting exon 19 of EYS (n = 4). Structural variants were mostly represented by delINV rearrangements (n = 5). Minigene assays confirmed that three novel deep intronic variants (in CHM, EYS, and CEP290) and one non-canonical splice-acceptor variant in USH2A which disrupt splicing, causes pseudoexon inclusion with premature termination or in-frame exon skipping respectively. Thus, WGS improves the diagnostic yield in IRDs by detecting variants missed by targeted sequencing, supporting its clinical utility in ophthalmologic practice.