Mutation Spectrum of the ABCA4 Gene in 335 Stargardt Disease Patients From a Multicenter German Cohort—Impact of Selected Deep Intronic Variants and Common SNPs

Abstract

PurposeStargardt disease (STGD1) is an autosomal recessive retinopathy, caused by mutations in the retina-specific ATP-binding cassette transporter (ABCA4) gene. To establish the mutational spectrum and to assess effects of selected deep intronic and common genetic variants on disease, we performed a comprehensive sequence analysis in a large cohort of German STGD1 patients.MethodsDNA samples of 335 STGD1 patients were analyzed for ABCA4 mutations in its 50 coding exons and adjacent intronic sequences by resequencing array technology or next generation sequencing (NGS). Parts of intron 30 and 36 were screened by Sanger chain-terminating dideoxynucleotide sequencing. An in vitro splicing assay was used to test selected variants for their splicing behavior. By logistic regression analysis we assessed the association of common ABCA4 alleles while a multivariate logistic regression model calculated a genetic risk score (GRS).ResultsOur analysis identified 148 pathogenic or likely pathogenic mutations, of which 48 constitute so far unpublished ABCA4-associated disease alleles. Four rare deep intronic variants were found once in 472 alleles analyzed. In addition, we identified six risk-modulating common variants. Genetic risk score estimates suggest that defined common ABCA4 variants influence disease risk in carriers of a single pathogenic ABCA4 allele.ConclusionsOur study adds to the mutational spectrum of the ABCA4 gene. Moreover, in our cohort, deep intronic variants in intron 30 and 36 likely play no or only a minor role in disease pathology. Of note, our findings demonstrate a possible modifying effect of common sequence variants on ABCA4-associated disease.

Cost‐effective molecular inversion probe‐based ABCA4 sequencing reveals deep‐intronic variants in Stargardt disease

“…The other novel variant c.3191–19G>A, tested in BA16, did not show a splice defect (Figure S3). In addition to this novel splice variant, the previously reported variant c.6148G>C (Schulz et al, ) did not show a splice defect using BA31 (Figure S3).…”

“…First, variants were prioritized by using the criteria “Quality by Depth” >500, which represents the overall coverage of the region, an Allele frequency (AF) < 0.005 in the dbSNP database and in an in‐house whole exome data set of 21,559 persons, and an AF < 0.01 in control population datasets, such as the Genome Aggregation Database (gnomAD; http://gnomad.broadinstitute.org/), and passing standard quality filters including gene components such as exons and canonical splice sites were prioritized. Previously reported pathogenic variants with high AFs such as c.2588G>C and c.5603A>T (AFs in non‐Finnish European [nFE] in gnomAD 0.00784 and 0.06647, respectively) (Cornelis et al, ; F. P. Cremers et al, ; Schulz et al, ; Zernant et al, , ), were selected separately. Known deep‐intronic variants were selected based on prior knowledge from literature (Albert et al, ; Bauwens et al, , ; Braun et al, ; Sangermano et al, ; Schulz et al, ; Zernant et al, ).…”

“…Previously reported pathogenic variants with high AFs such as c.2588G>C and c.5603A>T (AFs in non‐Finnish European [nFE] in gnomAD 0.00784 and 0.06647, respectively) (Cornelis et al, ; F. P. Cremers et al, ; Schulz et al, ; Zernant et al, , ), were selected separately. Known deep‐intronic variants were selected based on prior knowledge from literature (Albert et al, ; Bauwens et al, , ; Braun et al, ; Sangermano et al, ; Schulz et al, ; Zernant et al, ).…”

The CommonABCA4Variant p.Asn1868Ile Shows Nonpenetrance and Variable Expression of Stargardt Disease When Present intransWith Severe Variants

“…25,26 This variant was found to be phenotypically expressed only when in trans with a deleterious mutation (i.e., presence of the variants on different alleles) and explained >50% of all monoallelic cases and~80% of late-onset cases in a US cohort. 26 In addition, the c.2588G>C (p.[Gly863Ala, Gly863del]) variant was proposed to act as a mild pathogenic variant only when in cis with p.Asn1868Ile (i.e., presence of the variants on the same allele).…”

Choroidal Flow Signal in Late-Onset Stargardt Disease and Age-Related Macular Degeneration: An OCT-Angiography Study

“…At least two pathogenic or likely pathogenic ABCA4 mutations were identified in three STGD1 patients (23%), one pathogenic or likely pathogenic ABCA4 mutation in combination with a variant of unknown significance was found in seven STGD1 patients (54%), and two variants of unknown significance were detected in one STGD1 patient (8%) whereas only one likely pathogenic ABCA4 mutation was found in two STGD1 patients (15%) (Supplementary Table S1). The frequent c.5603A>T (p.Asn1868Ile) variant recently shown to be associated with STGD1 development, in particular with the late-onset phenotype, 50,51 was identified in six STGD1 patients. Eleven STGD1 eyes and eight AMD eyes revealed foveal involvement of the RPE atrophy.…”