Targeted sequencing using single-molecule Molecular Inversion Probes reveals severe bi-allelic ABCA4 alleles in probands diagnosed with Leber congenital amaurosis

Abstract

Purpose : The cost-effective identification of genetic variants underlying retinitis pigmentosa (RP) and Leber congenital amaurosis (LCA) is challenging as 108 genes can be implicated. We sequenced the coding regions of these genes in a cohort of 4,000 probands in a targeted manner. Using a genotype-first analysis approach, all variants were assessed irrespective of the currently associated phenotypes and inheritance patterns of the mutated genes. In this manner, we searched for novel genotype-phenotype correlations.

Methods : We sequenced all exons, splice sites and known deep-intronic variants of 113 genes associated with RP, LCA, and congenital stationary night blindness in 4,000 probands using 16,812 single-molecule Molecular Inversion Probes (smMIPs) as described elsewhere (https://www.medrxiv.org/content/ 10.1101/2022.11.24.22282656v2). Sequencing libraries were prepared in series of 384 samples and analyzed using Illumina NovaSeq6000. All variants with an allele frequency of ≤0.5% with a class 3, 4, or 5 Franklin-ACMG classification were selected for further interpretation. For ABCA4, only severe and moderately severe alleles were considered using the severity scores published by Cornelis et al. 2022 (PMID: 35120629).

Results : Ninety-one of 163 LCA probands were considered genetically solved with variants in 22 genes. Interestingly, at least four probands were considered genetically solved by bi-allelic severe variants in ABCA4. No pathogenic variants in other RP or LCA-associated genes were detected in these patients.

Conclusions : Here, we describe a new genotype-phenotype correlation identified in at least four LCA probands that we consider genetically solved by bi-allelic severe variants in ABCA4. While there are strong indications that late-onset STGD1 can be caused by a combination of bi-allelic variants in ABCA4 and non-ABCA4 modifiers, onset before the first year of life has not yet been described. These results suggest that non-ABCA4 modifiers play an important role in the severe and mild ends of the ABCA4-disease spectrum. Our results expand the phenotypic spectrum of ABCA4-associated diseases and may have implications for the diagnosis and management of these disorders.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.