RPE65 Variant p.(E519K) Causes a Novel Dominant Adult-Onset Maculopathy in 83 Affected Individuals.

Purpose: Recessive RPE65-associated retinopathy is a well-known target for gene therapy, whereas dominant RPE65-associated retinopathy, due to the Irish founder variant p.(D477G), has been reported only once until now and is very rare. Here, we present the discovery of a novel, second dominant RPE65-associated retinopathy caused by variant c.1555G>A, p.(E519K).

Methods: Genomic data was investigated in a Belgian discovery cohort (n = 2873) and an international replication cohort (n = 18,796) with inherited retinal disease (IRD). Heterozygous p.(E519K) individuals underwent extensive phenotyping. Haplotype phasing was based on long-read sequencing and microsatellite analysis. Variant p.(E519K) was assessed in vitro using an enzymatic assay, Western blotting, co-immunoprecipitation, cellular thermal shift assay (CETSA), minigene assays, and in silico using protein modeling (AlphaFold).

Results: The monoallelic p.(E519K) variant was found in 83 affected individuals from Belgium, the Netherlands, France, and Canada, all of European ancestry. A shared region of 464 kilobases (kb) confirmed a founder effect. Variant p.(E519K) lowers RPE65 protein expression and enzymatic activity, with altered protein stability predicted and experimentally confirmed. Genotype-phenotype data support dominant inheritance and phenotypic variability, respectively, characterized by late-onset macular dystrophy with two main subtypes.

Conclusions: The discovery of a dominant RPE65-IRD due to founder variant p.(E519K) reduces the diagnostic gap in dominant IRD and highlights a novel target for therapy.