Prevalence And Novelty Of ABCA4 Mutations In French Patients With Stargardt Disease Or Simplex Cone-rod Dystrophy

Abstract

Purpose: : In order to prepare for future gene replacement therapy, we genetically characterized ABCA4 variants in a phenotypically well-documented French cohort of Stargardt and sporadic or autosomal recessive (ar) index cases of cone-rod dystrophy (CRD).

Methods: : Detailed phenotypic characterization was performed on 125 Stargardt and 79 simplex or ar CRD patients. Microarray technology using the ABCA4 Asper chip was first applied. Pathogenic variants were checked by direct sequencing. When only one pathogenic variant was identified, direct sequencing of the 50 coding exons and their flanking intronic regions of ABCA4 was performed to identify the second pathogenic variant. Co-segregation analysis was subsequently performed on available family members.

Results: : Within the Stargardt group, 66.4% were found to carry at least 1 pathogenic variant after ABCA4 microarray screening. Of these, 77% were found to be compound heterozygous or homozygous using direct sequencing. Applying this technique, we identified 9 new variants likely to be pathogenic including 2 nonsense, 5 missense and 2 splice site. Within the cone-rod dystrophy group, only 26.6% were found with at least 1 pathogenic variant after ABCA4 microarray screening. Of these, 57% were compound heterozygous or homozygous using direct sequencing. Based on this technique, we identified 1 new nonsense pathogenic variant.

Conclusions: : This extensive genetic screening will serve as basis for the up coming clinical trial on ABCA4 gene replacement therapy. Furthermore, the results of our screening methods highlighted the following issues: 1) 33.6% of Stargardt and 73.4% of CRD cases did not show any mutation in ABCA4 after microarray screening, which could be due to novel ABCA4 mutations or mutations in other known or yet to be discovered genes. 2) In 23% of Stargardt and 43% of CRD cases, direct sequencing did not reveal a second pathogenic variant and could have missed a large deletion or a variant in a regulatory element, or indicate also the involvment of other known or unknown genes that remains to be identified. To address these issues, we propose to use Next-Generation Sequencing (NGS) of the entire ABCA4 gene and/or a NGS-retinal panel developed in our laboratory.

Keywords: gene screening • retinal degenerations: hereditary • genetics