PMC4699863_01
Pedigree of the family illustrating the segregation of four disease-causing ABCA4 alleles with two Stargardt disease (STGD1) phenotypes of varying severity. Filled circles and squares denote affected males and females, respectively, and centrally filled shapes denote heterozygous carriers. ABCA4 variants, and their order on the chromosome, are listed in the key. The symbol (+) denotes the presence of a disease-causing ABCA4 variant while (-) represents the wild-type allele. The mother of the proband (I-2, arrowhead) was affected with pattern macular dystrophy which was phenotypically distinct from the other affected individuals and the father (I-1) was affected with chronic central serous chorioretinopathy.
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F1
Ten members of a two-generation family presented to the clinic with a history of dominant retinal disease (Figure 1, pedigree). Interestingly, both of these mutations originated from carrier mothers (I-2, I-4) of the two STGD1 patients (Figure 1).
Advanced ABCA4 phenotype of the proband, paternal aunt and uncles. (A) The proband, a 43-year-old man of German descent harboring the p.C54Y and c.[302+68C>T;4539+2028C>T] variants, presented with large well-delineated lesions of chorioretinal atrophy and central pigment clumping in both eyes on autofluorescence imaging and color fundus photographs. Surrounding areas of dense hyperautofluorescence preceding a reticular pattern of granular flecks were found extended throughout the periphery. (B) Full-field electroretinogram testing in the proband (II-2) revealed significantly reduced amplitudes in rod and cone responses in the right (blue trace) and left (red trace) eyes when compared to an age-matched control (dotted gray trace). The paternal aunt (c.[302+68C>T;4539+2028C>T]; c.6148-698_c.6670del/insTGTGCACCTCCCTAG) presented with similarly large area of chorioretinal atrophy while the paternal uncle (D) with the same compound heterozygous genotype exhibited generalized atrophy of the posterior pole. Various degrees of sparing of the peripapillary region in each patient are marked with yellow arrowheads.
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F2
The peripapillary region around the optic nerve was notably spared of any apparent disease-associated changes (Figure 2A, yellow arrowheads). Full-field electroretinogram (ERG) testing in the proband revealed significant reductions in both rod and cone function that is indicative of advanced-stage, generalized photoreceptor loss throughout the retina. (Figure 2B). The peripapillary regions in both were affected with flecks or partially spared, respectively. (Figure 2C and 2D, yellow arrowheads).
The paternal cousin (36-year-old son of the paternal uncle) harboring the p.G1961E allele and the c.6148-698_c.6670del/insTGTGCACCTCCCTAG variant presented at a comparatively milder disease stage. (A) Autofluorescence imaging revealed a localized lesion of retinal pigment epithelium mottling and photoreceptor loss in the central macula (B) as correlated with spectral domain-optical coherence tomography. The lesion is surrounded by a confluent pattern of round, pisciform flecks centrifugally-distributed across the midperiphery of the retina.
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F3
A lesion of mottled atrophy and photoreceptor loss was confined to the central macula in each eye (Figure 3A and 3B) with a scattered, confluent pattern of subretinal pisciform flecks centrifugally distributed throughout the mid-periphery.
Macular findings in the mother and father of the proband who each carry a single ABCA4 variant, p.C54Y and c.[302+68C>T;4539+2028C>T], respectively. (A) The mother presented with a confined lesion of atrophy and large hyperfluorescent flecks sparing the fovea in each eye. Notably, atrophy extends into the peripapillary region of the optic nerve in both eyes. (B) The father exhibited a recent onset of central serous chorioretinopathy in the left eye with fluid and pigment epithelial detachment over the fovea in the left eye. Pigmentary changes noted in the macula are likely attributed to early, dry age-related macular degeneration.
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F4
It was noted that atrophy from the macular lesion extended to the peripapillary region around the optic disc (Figure 4A), resulting in a phenotype described as 'pattern dystrophy'. The proband's father (I-1), aged 75 years, also presented with a recent onset of macular findings consistent with non-exudative central serous chorioretinopathy in the left eye (Figure 4B); SD-OCT revealed a minor pigment epithelial detachment in this eye (Figure 4C).
The deletion/insertion variant in the ABCA4 locus in the family. A. aCGH log2 ratio plot showing the heterozygous deletion identified in patient II-3. The breakpoint junction sequences alignment underneath the plot reveals two inserted DNA fragments (green) conjugated by microhomologies (purple). The potential origins of the inserted DNA fragments are marked with underlines labeled with "1" and "2", respectively. DIST/PROX, human genome reference sequences at the distal/proximal end of the junction. B. Potential molecular mechanism explaining the deletion etiology. FoSTeS, fork stalling and template switching. C. Pedigree of the family identified with the deletion/insertion variant allele (c.6148-698_c.6670del/insTGTGCACCTCCCTAG). The gel pictures underneath the pedigree demonstrate the segregation of the deletion allele with three other potential disease-causing alleles identified in this family. wt, wild type allele; del, deletion/insertion variant allele.
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F5
While these reports have been rare, we screened the patients I-3, II-1 and II-3 on the using custom designed high density array CGH, which revealed a ~5 kb heterozygous deletion at the ABCA4 locus (Figure 5A) in patients I-3 and II-3. Breakpoint junction sequencing mapped the precise genomic interval (Chr1: 94463476- 94468246), revealing a 4771 bp deletion (Figure 5B). The insertion could be split into two halves; both were potentially from regions close to the proximal end of the deletion (Figure 5B). Microhomologies were identified at the breakpoint junctions (Figure 5B). Genotyping analysis of c.6148-698_c.6670del/insTGTGCACCTCCCTAG revealed its presence in unaffected individuals I-6 and I-7, who were carriers of this variant, and in affected individuals I-3 and I-5 in combination with a second variant consisting of a complex deep intronic mutant allele c.[302+68C>T;4539+2028C>T] (Figure 5C).