Clinical and Molecular Analysis of Stargardt Disease With Preserved Foveal Structure and Function




Purpose


To describe a cohort of patients with Stargardt disease who show a foveal-sparing phenotype.


Design


Retrospective case series.


Methods


The foveal-sparing phenotype was defined as foveal preservation on autofluorescence imaging, despite a retinopathy otherwise consistent with Stargardt disease. Forty such individuals were ascertained and a full ophthalmic examination was undertaken. Following mutation screening of ABCA4, the molecular findings were compared with those of patients with Stargardt disease but no foveal sparing.


Results


The median age of onset and age at examination of 40 patients with the foveal-sparing phenotype were 43.5 and 46.5 years. The median logMAR visual acuity was 0.18. Twenty-two patients (22/40, 55%) had patchy parafoveal atrophy and flecks; 8 (20%) had numerous flecks at the posterior pole without atrophy; 7 (17.5%) had mottled retinal pigment epithelial changes; 2 (5%) had multiple atrophic lesions, extending beyond the arcades; and 1 (2.5%) had a bull’s-eye appearance. The median central foveal thickness assessed with spectral-domain optical coherence tomographic images was 183.0 μm (n = 33), with outer retinal tubulation observed in 15 (45%). Twenty-two of 33 subjects (67%) had electrophysiological evidence of macular dysfunction without generalized retinal dysfunction. Disease-causing variants were found in 31 patients (31/40, 78%). There was a higher prevalence of the variant p.Arg2030Gln in the cohort with foveal sparing compared to the group with foveal atrophy (6.45% vs 1.07%).


Conclusions


The distinct clinical and molecular characteristics of patients with the foveal-sparing phenotype are described. The presence of 2 distinct phenotypes of Stargardt disease (foveal sparing and foveal atrophy) suggests that there may be more than 1 disease mechanism in ABCA4 retinopathy.


Stargardt disease is an autosomal recessive disorder caused by mutations in the ABCA4 gene. It is the most common single-gene retinal degeneration, with a reported prevalence of 1:10 000. Most cases typically present with central visual loss within the first 2 decades of life, and during the course of the disorder there is macular atrophy with yellow-white flecks in the posterior pole, at the level of the retinal pigment epithelium (RPE). However, Stargardt disease is associated with a variable phenotype and severity. Autofluorescence (AF) imaging and electroretinography may assist the diagnosis, and parameters such as age of onset, visual acuity, AF pattern, and the nature of the electrophysiological findings assist both in the determination of disease severity and in the provision of prognostic information. Increasingly, high-resolution imaging using spectral-domain optical coherence tomography (SDOCT) is providing insights into the retinal architectural changes that occur in Stargardt disease.


There is wide phenotypic variability in ABCA4 retinopathy and sequence variants in ABCA4 have also been implicated in cone dystrophy, cone-rod dystrophy, and “retinitis pigmentosa” in addition to Stargardt disease. There is also extensive allelic heterogeneity, with more than 700 sequences in the ABCA4 gene having been reported to date. The phenotypic variability and the high genetic heterogeneity have confounded attempts to examine genotype-phenotype correlations comprehensively.


A cohort of Stargardt disease patients who had better visual acuity than “typical” Stargardt disease patients, and who showed sparing of the fovea on funduscopy, has been described. There are also reports of patients with “late-onset” Stargardt disease, including individuals with a foveal-sparing phenotype, who harbor ABCA4 variants. In those reports, SDOCT images demonstrated a well-preserved foveal structure including the neurosensory retina, which differed from previous observations of early foveal photoreceptor damage in “typical” Stargardt disease with central retinal atrophy. Foveal-sparing forms of Stargardt disease may thus reflect a distinct pathogenesis.


The present study describes the clinical findings and molecular genetic characteristics of “foveal-sparing” Stargardt disease in a large cohort from a single center. The molecular data of the cohort with the foveal-sparing phenotype are compared with ABCA4 variants observed in patients with Stargardt disease but without foveal sparing.


Methods


Patients


AF images of the right eyes of 438 individuals with a clinical diagnosis of retinopathy compatible with Stargardt disease were surveyed and 40 patients were identified with an apparently normal AF signal at the fovea (foveal sparing). After informed consent, blood samples were collected and genomic DNA was extracted from the peripheral blood leukocytes. The protocol of the study adhered to the provisions of the Declaration of Helsinki and was approved by the local Ethics Committee of Moorfields Eye Hospital.


For the purposes of this report, patients presenting to the hospital with signs of atrophy within the macula, bilaterally, with or without surrounding flecks were potentially included as having Stargardt disease. Patients with a stationary visual dysfunction were excluded. A careful drug history was taken to allow exclusion of those with retinotoxic maculopathy. Patients with a dominant family history were excluded. Where a family history was not extensive, or whenever 2 generations were affected, the RDS/PRPH2 gene, with its coding region and intron-exon boundaries, was sequenced. In those patients over 50 years of age, care was taken not to include cases of atrophic age-related macular degeneration in which there were soft drusen, or patients with maternally inherited diabetes and deafness in whom the distribution of atrophy and autofluorescence appearance had a distinctive appearance. The m.3243A>G variant was assayed if this phenotype was in any way suggested.


Clinical Assessment


A detailed medical history was obtained and a comprehensive ophthalmologic examination was performed for all 40 patients. The age of onset was defined as the age at which visual loss was first noted by the patient or as the age at the latest examination for asymptomatic patients. The duration of the disease was calculated as the difference between age at onset and age at the latest examination. Assessment included best-corrected visual acuity, dilated ophthalmoscopy, color fundus photography, AF imaging, SDOCT imaging, and electrophysiological assessment. Best-corrected Snellen visual acuity was converted to equivalent logMAR visual acuity.


Color fundus photography was performed with a TRC-50IA retinal fundus camera (Topcon, Tokyo, Japan). AF images before 2009 were obtained with an HRA 2 (Heidelberg Engineering, Heidelberg, Germany; excitation light 488 nm; barrier filter 500 nm; field of view 30 × 30 degres), and images after 2009 were undertaken using the Spectralis with viewing module version 5.1.2.0 (Heidelberg Engineering; excitation light 488 nm; barrier filter 500 nm; fields of view 30 × 30 degrees and 55 × 55 degrees).


SDOCT was undertaken with the Spectralis with viewing module version 5.1.2.0. The SDOCT protocol included a horizontal linear scan (100 B-scans averaged to improve the signal-to-noise ratio) centered on the fovea, where possible, and a volume scan (minimum of 19 B-scan slices, 20 × 20 degrees). The HEYEX software interface (version 1.6.2.0; Heidelberg Engineering) was used for retinal thickness measurement. Central foveal thickness was defined as the distance between the inner retinal surface and the inner border of the RPE. Evidence of outer retinal tubulation was assessed from all the B-scan slices of each eye by 2 authors (K.F. and A.R.W.).


Electrophysiology


Electrophysiological assessment included full-field electroretinogram (ERG) and pattern electroretinogram (PERG) recorded with gold foil electrodes that incorporated the standards of the International Society for Clinical Electrophysiology of Vision. The full-field ERGs were used to assess generalized rod and cone system function and included: (1) dark-adapted dim flash 0.01 candela-seconds per square meter (cd·s·m −2 ); (2) dark-adapted bright flash 11.0 cd·s·m −2 ; (3) light-adapted 3.0 cd·s·m −2 30 Hz flicker; and (4) light-adapted 3.0 cd·s·m −2 at 2 Hz. The PERG P50 component and multifocal electroretinogram (mfERG) were used to assess macular function. Some patients had mfERG recording (RETIscan System; Roland Consult, Wiesbaden, Germany) with a stimulus consisting of 61 scaled hexagons covering in total a visual field of 55 degrees, at a viewing distance of 33 cm. All main components of the ERG and the PERG P50 component were used to classify patients into 3 groups; this is a partial modification of a previous report : (1) patients with normal ERGs with or without a PERG P50 abnormality; (2) subjects with a PERG P50 abnormality and additional generalized cone-mediated ERG abnormality (assessed with photopic ERGs); and (3) individuals with a PERG P50 abnormality and generalized cone system electrophysiological abnormality and additional generalized rod-mediated ERG abnormality (assessed using scotopic ERGs).


Mutation Screening and Molecular Genetic Analysis


Blood samples were collected in EDTA tubes and DNA was extracted with a Nucleon Genomic DNA extraction kit (BACC2; Tepnel Life Sciences, Manchester, United Kingdom), or the Qiagen Gentra Puregene blood kit (Qiagen, Venlo, Netherlands). Mutation screening of ABCA4 was performed with the arrayed primer extension (APEX) microarray (ABCR400 chip, Asper Ophthalmics, Tartu, Estonia) in all probands. The term “variants” used herein includes those sequence changes previously shown to be enriched in patients with Stargardt disease from prior studies. Null variants are those that would be expected to affect splicing, or to introduce a premature truncating codon in the protein if translated. Non-null variants (missense and in-frame deletions or insertions) were analyzed using 3 software prediction programs: SIFT (Sorting Intolerant from Tolerance; sift.jcvi.org/ , accessed March 1, 2013), PolyPhen2 ( genetics.bwh.harvard.edu/pph/index.html , accessed March 1, 2013), and the Human Splicing finder program version 2.4.1 ( www.umd.be/HSF/ , accessed March 1, 2013). Minor allele frequency for each allele was estimated with reference to the Exome Variant Server (NHLBI Exome Sequencing Project, Seattle, Washington, USA; snp.gs.washington.edu/EVS/ , accessed March 1, 2013).


To investigate potential molecular genetic differences between patients with the foveal-sparing phenotype and those with typical disease (without foveal sparing/with foveal atrophy), the molecular data of patients with typical Stargardt disease ascertained at Moorfields Eye Hospital were reviewed. This comparison group consisted of all patients without evidence of foveal sparing on AF imaging and also harbored at least 1 ABCA4 disease-causing variant following screening with the APEX microarray. One hundred and forty subjects from a total cohort of 438 individuals fulfilled these criteria, and the allele frequency of the most prevalent variants was compared between the group of patients with the foveal-sparing phenotype (n = 31) and the group of patients with typical Stargardt disease (n = 140).




Results


The clinical findings in the 40 patients with foveal-sparing Stargardt disease are summarized in Table 1 . There were 22 female patients (55%) and 18 male patients (45%). The panel included 2 sibships; a sibling pair (Patients 16 and 32) and 1 set of 3 siblings (Patients 33, 34, and 35). Twenty-five patients (63%) complained of central visual loss and 2 subjects (5%) presented with diplopia, with 13 individuals (32%) having no visual symptoms. The median age of onset was 43.5 years (range, 25-75 years), and the median age at the examination was 46.5 years (range, 25-75 years). Nineteen patients (47.5%) had onset at ≥45 years of age. The mean duration of disease was 4.1 years (range, 0-25 years). The median logMAR visual acuity was 0.18 in the right eye and 0.18 in the left eye (range, −0.08 to 2.0 and −0.08 to 3.0). Two patients (4 and 26) had a logMAR visual acuity of less than 1.0 in the right eye at the most recent review, with those patients having been diagnosed with the foveal-sparing phenotype of Stargardt disease 4 years earlier, when visual acuity was 0.18 in the right eye and 0.48 in the left eye.



Table 1

Summary of Clinical Findings and Molecular Status of 40 Patients With a Foveal-Sparing Phenotype a of Stargardt Disease




























































































































































































































































































































































































































































































































































































































































































Patient Onset b (y) Age (y) LogMAR Visual Acuity Fundus Pattern c OCT ERG e Mutation Status
CFT d (μm) ORT Group PERG mfERG
OD OS OD OS OD OS OD OS
1 45 45 0 0 3 219 223 NA NA NA NA NA [c.1411 G>A, p.Glu471Lys/c.2588 G>C, p. Gly863Ala/c.4594 G>A, p.Asp1532Asn/c.5693 G>A, p.Arg1898His]
2 33 33 0.18 0.48 1 NA NA 3 ND ND NA NA [c.1622 T>C, p.Leu541Pro/c.3113 C>T, p.Ala1038Val/c.6089 G>A, p.Arg2030Gln]
3 53 66 0.18 0.18 1 NA NA 2 A A NA NA [c.768 G>T, Splice site/c. 6320 G>A, p. Arg2107His ]
4 37 54 1.48 0.18 1 32 39 3 ND ND 2 2 [c.1760 +1 G>T, Splice site/c.4594 G>T, p.Asg1532Tyr ]
5 57 57 0.3 0.18 1 NA NA 1 ND ND NA NA [c. 1805G>A, p. Arg602Gln/c.3898 C>T, p.Arg1300*]
6 65 65 0.18 0 1 211 187 1 N N NA NA [c.5461-10 T>C, Splice site/c. 6089 G>A, p.Arg2030Gln]
7 54 54 0 0 1 189 198 1 A A NA NA [c. 6089 G>A, p.Arg2030Gln/c.6118 C>T, p.Arg2040*]
8 39 44 −0.1 −0.1 4 297 230 3 A A NA NA [c.71 G>A, p.Arg24His/c.4577 C>T, p. Thr1526Met]
9 35 35 0.18 0.18 2 142 154 3 ND ND NA NA [c.658 C>T, p.p.Arg220Cys/c.2588 G>C, p. Gly863Ala]
10 45 54 0.48 0.18 1 102 116 3 ND A NA NA [c.1957 C>T, p.Arg653Cys/c.5693 G>A, p.Arg1898His]
11 43 43 −0.1 0 2 170 185 1 A A 2 2 [c.2588 G>C, p. Gly863Ala/c.4139 C>T, p.Ala1038Val]
12 36 ∗∗ 38 0.3 0 1 220 212 1 A A 1 1 [c.4139 C>T, p.Ala1038Val/c.4594 G>T, p.Asp1532Asn]
13 62 68 −0.1 0.48 1 196 189 1 N N 2 2 [c.4222 T>C, p.Trp1408Arg/c.4918 C>T, p.Arg1640Trp]
14 36 44 0.48 0.48 3 79 89 1 A A NA NA [c.4222 T>C, p.Trp1408Arg/c.4918 C>T, p.Arg1640Trp]
15 46 46 −0.1 −0.1 3 NA NA 1 A A NA NA [c.4469 G>A, p.Cys1490Tyr/c. 6089 G>A, p.Arg2030Gln]
16 44 44 0.18 0 2 NA NA 1 A A NA NA [c.6079 C>T, p.Leu2027Phe/c.6079 C>T, p.Leu2027Phe]
17 48 73 0.18 3 4 135 86 2 A ND NA NA [c.4956 T>G, p.Tyr1652*]
18 56 57 0 0 2 254 273 1 ND A NA NA [c.5018+2 T>C, Splice site]
19 53 53 0.48 0.18 1 137 133 1 A A NA NA [c.5461-10 T>C, Splice site]
20 49 58 0.18 0 1 256 222 1 A N 1 1 [c.5461-10 T>C, Splice site]
21 47 ∗∗ 47 0.3 0.3 1 239 202 1 A A 1 1 [c.1805 G>A, p.Arg602Gln]
22 50 50 0.48 0.18 1 263 261 1 N N NA NA [c.1957 C>T, p.Arg653Cys]
23 39 39 0 −0.1 2 225 228 1 N N NA NA [c.2588 G>C, p. Gly863Ala]
24 55 57 0.48 0.48 1 117 74 1 ND ND NA NA [c.3602 T>G, p.Leu1201Arg]
25 50 54 0.48 0.18 1 147 144 3 ND ND NA NA [c.3602 T>G, p.Leu1201Arg]
26 43 47 2 0.18 1 70 52 1 ND ND NA NA [c.4319 T>C, p.Phe1440Ser]
27 30 51 0.3 0.3 1 75 79 3 A A NA NA [c.4685 T>C, p.Ile1562Thr]
28 29 34 0.18 0.18 3 132 107 1 A A NA NA [c.4926 C>G, p.Ser1642Arg]
29 52 52 0.18 0.18 3 180 200 1 ND ND 2 2 [c.5882 G>A, p.Gly1961Glu]
30 28 28 −0.1 −0.1 2 NA NA 1 N ND NA NA [c.6079 C>T, p.Leu2027Phe]
31 40 40 −0.1 −0.1 2 222 223 NA NA NA NA NA [c.6079 C>T, p.Leu2027Phe]
32 45 48 0.18 3 1 237 252 NA NA NA NA NA NA
33 39 40 0.18 0 5 126 140 NA NA NA NA NA Not detected
34 25 25 0.18 0 3 147 150 NA NA NA NA NA Not detected
35 29 36 0 0.3 3 172 181 2 A A NA NA Not detected
36 40 45 0.3 0 1 NA NA 1 A N NA NA Not detected
37 35 41 0.18 0.18 1 150 131 1 ND ND NA NA Not detected
38 29 29 −0.1 0 2 237 236 NA NA NA NA NA Not detected
39 25 35 −0.1 0 1 237 179 3 ND ND 2 2 Not detected
40 75 75 0.18 0.18 1 194 186 NA NA NA NA NA Not detected

A = abnormal; CFT = central foveal thickness; ERG = electroretinogram; mfERG = multifocal ERG; N = normal; NA = not available; ND = not detectable; OCT = optical coherence tomography; ORT = outer retinal tubulation; PERG = pattern ERG.

a The foveal-sparing phenotype was defined as foveal preservation on autofluorescence imaging, despite a retinopathy otherwise consistent with Stargardt disease.


b The age of onset was defined as the age at which visual loss was first noted by the patient or as the age at the latest examination for patients (labeled with*) who are not aware of any visual symptom. Two patients complained of diplopia (labeled with**).


c Color fundus photography identified 4 patterns: pattern 1, patchy parafoveal atrophy surrounded by numerous yellow-white flecks; pattern 2, numerous yellow-white flecks at the posterior pole without atrophy; pattern 3, mottled retinal pigment epithelial changes and/or localized parafoveal yellow-white flecks; pattern 4, multiple patchy atrophic lesions, extending beyond the arcades. One patient had a bull’s-eye maculopathy appearance (Patient 33, labeled as pattern 5).


d The CFT was defined as the distance between inner retinal surface and inner border of retinal pigment epithelium.


e Patients were classified on the basis of electrophysiological findings: group 1 – normal ERGs with or without PERG P50 abnormality; group 2 – PERG P50 abnormality and additional generalized cone system abnormality; group 3 – PERG P50 abnormality and additional generalized cone and rod system abnormality. mfERG findings were categorized based on the responses from central and paracentral hexagons into 2 subgroups: 1 – preserved central response surrounded by paracentral reduction; 2 – central and paracentral loss of responses.



Color fundus photography was performed in all 40 patients. Four patterns were identified: pattern 1 (n = 22, 55%) showed patchy parafoveal atrophy surrounded by numerous yellow-white flecks; pattern 2 (n = 8, 20%) had numerous yellow-white flecks at the posterior pole without atrophy; pattern 3 (n = 7, 17.5%) had mottled RPE changes and/or localized parafoveal yellow-white flecks; and pattern 4 (n = 2, 5%) had multiple patchy atrophic lesions, extending beyond the arcades. One patient (2.5%) had a bull’s-eye maculopathy (Patient 33). These data are summarized in Table 1 . The median ages of onset of patterns 1, 2, 3, and 4 were 46.0, 39.5, 36.0, and 43.5 years, respectively; the mean duration of disease was 5.1, 0.1, 2.9, and 15.0 years. The median logMAR visual acuity of patterns 1, 2, 3, and 4 was 0.18, 0.00, 0.18, and 0.05, respectively. Color fundus photographs and AF images of 4 representative cases are shown in Figure 1 , with associated OCT images in Figure 2 .




Figure 1


Color fundus photographs and autofluorescence images of 4 representative cases with a foveal-sparing phenotype of Stargardt disease (Patients 6, 11, 29, and 8), showing 4 fundus phenotypes identified in this study, respectively. Images show examples of each of 4 fundus phenotypes identified in this study: Patient 6 shows pattern 1 (patchy parafoveal atrophy surrounded by numerous yellow-white flecks); Patient 11 shows pattern 2 (numerous yellow-white flecks at the posterior pole without atrophy); Patient 29 shows pattern 3 (mottled retinal pigment epithelium changes and localized parafoveal yellow-white flecks); and Patient 8 shows pattern 4 (multiple patchy atrophic lesions, extending beyond the arcades).



Figure 2


Spectral-domain optical coherence tomographic images of the 4 representative cases with the foveal-sparing phenotype of Stargardt disease illustrated in Figure 1 (Patients 6, 11, 29, and 8), demonstrating relatively preserved foveal structure and outer retinal tubulation. There is relatively preserved foveal structure in all 4 patients, with 2 showing outer retinal tubulation (Patients 6 and 8). Arrows in the magnified images identify outer retinal tubulation.


SDOCT images were obtained in 33 individuals ( Table 1 ). The median central foveal thickness of the right and left eyes was 180.0 μm and 185.0 μm, respectively (range, 32-219 μm and 39-273 μm). The median central foveal thickness for each fundus pattern in the right eye was 179.5 μm for pattern 1 (18 patients); 223.5 μm for pattern 2 (6 subjects); 159.5 μm for pattern 3 (6 individuals); and 216.0 μm for pattern 4 (2 patients). Fifteen patients (45%) showed evidence of outer retinal tubulation. The median age of onset and the mean duration of disease of these 15 patients with outer retinal tubulation were 45.0 and 7.2 years, compared to 43.0 and 2.1 years for the 18 subjects without outer retinal tubulation. The median visual acuity was the same in both groups. AF and SDOCT images of 2 representative cases (Patients 9 and 17) demonstrate slow progression over time ( Figures 3 and 4 ).




Figure 3


Serial autofluorescence and spectral-domain optical coherence tomographic images of Patient 9, with the foveal-sparing phenotype of Stargardt disease, showing increasing of atrophy and architectural disruption. Patient 9 presented without any visual symptoms 5 years prior to the most recent examination. LogMAR visual acuity was 0.0 in each eye. Autofluorescence images show numerous foci of high or low signal at the posterior pole at baseline, with increasing atrophy over time. Optical coherence tomographic images at baseline show relatively preserved foveal structure with an intact junction between the inner and outer photoreceptor segments (IS/OS) in each eye; IS/OS disruption is present at follow-up examination.



Figure 4


Serial autofluorescence and spectral-domain optical coherence tomographic images of Patient 17, with the foveal-sparing phenotype of Stargardt disease, illustrating enlargement of atrophy and progression of a snowball-like lesion to a recognizable outer retinal tubulation. Autofluorescence images at baseline show multiple patchy asymmetrical atrophic lesions in each eye. LogMAR visual acuity was −0.08 in the right eye but 1.0 in the left eye at baseline. Enlargement of the areas of atrophy occurs over time, with relative preservation of the foveal autofluorescence signal in the right eye (logMAR visual acuity 0.18). Disruption of outer retinal structure, with a snowball-like lesion (arrow) at the fovea in the right eye and foveal atrophy in the left eye, is present in baseline optical coherence tomographic images. The snowball-like lesion progresses over time, with outer retinal tubulation clearly present at follow-up in the right eye.


Electrophysiological assessment was performed in 33 patients. ERG and PERG were recorded in all; mfERG was obtained in 8 individuals. Twenty-two of the 33 patients (67%) were in ERG group 1, 3 (9%) in ERG group 2, and 8 (24%) in ERG group 3. PERG was normal in both eyes of 3 patients (9%), abnormal responses in either eye of 16 subjects (48%), and undetectable responses in either eye of 14 individuals (42%). The median PERG P50 amplitude of PERG was 0.5 μV (range, 0.0-3.6 μV). The mfERG showed preservation of the response to the central hexagon surrounded by reduced responses to the paracentral hexagons in 3 of 8 patients (38%). These 3 patients had normal full-field ERGs. Five of 8 subjects (62%) had severely reduced central and paracentral responses, including 2 with normal ERGs and 3 with the group 3 ERG phenotype. The electrophysiological findings are summarized in Table 1 ; representative traces appear in Figure 5 .




Figure 5


Full-field electroretinograms, pattern electroretinograms, and multifocal electroretinograms from 2 representative cases with the foveal-sparing phenotype of Stargardt disease, showing localized spared foveal function and marked macular dysfunction with generalized rod and cone system involvement, respectively (Patients 12 and 8). Full-field electroretinograms (ERGs) included: (1) dark-adapted dim flash 0.01 candela-seconds per square meter (cd·s·m −2 ) (DA 0.01); (2) dark-adapted bright flash 11.0 cd·s·m −2 (DA 11.0); (3) light-adapted 3.0 cd·s·m −2 30 Hz flicker (LA 3.0 30 Hz); and (4) light-adapted 3.0 cd·s·m −2 at 2 Hz (LA 3.0 2 Hz). Patient 12 demonstrates normal ERGs and subnormal P50 components of pattern electroretinogram (PERG), consistent with macular dysfunction; multifocal electroretinograms (mfERGs) reveal sparing of localized foveal function with severe paracentral macular dysfunction bilaterally. Patient 8 has abnormal ERGs, consistent with generalized rod and cone system dysfunction, with marked PERG P50 reduction indicating marked macular involvement. Normal traces are shown for comparison.


Molecular Genetics


Likely disease-causing ABCA4 variants were detected in 31 of 40 patients: 2 or more variants were identified in 16 patients and 1 variant in 15 subjects ( Table 1 ). Detailed results, including in silico analysis to assist in the prediction of pathogenicity of the variants, are shown in Table 2 . Thirty variants were found in 31 patients: 7 null variants, with 3 predicted to affect splicing and 1 uncertain effect (c.5461-10T>C); and 23 non-null variants ( Table 2 ). Twenty-nine previously reported disease-causing variants and 1 novel putative disease-causing variant were identified (c.1760+1G>T). The most common variants identified were p.Gly863Ala, c.5461-10T>C, p.Leu2027Phe, and p.Arg2030Gln, occurring, respectively, in 4, 3, 3, and 4 patients with the foveal-sparing phenotype of Stargardt disease. One patient was identified to be homozygous for the p.Leu2027Phe variant; none had 2 or more null variants.



Table 2

Investigation of the Pathogenicity of ABCA4 Variants Identified in 40 Patients With the Foveal-Sparing Phenotype of Stargardt Disease
























































































































































































































































































































































































































































































































Exon Nucleotide Substitution and Amino Acid Change Number of Alleles Het/Homo Previous Report SIFT a Polyphen 2 a HSF Matrix a Allelic Frequency Observed by EVS a Reference
Pred. Index (0-1) Pred. Hum Var Score (0-1) Site Affected Wt CV Mt CV CV % Variation
2 c.71G>A, p.Arg24His 1 Het Lewis Tol. NA PRD 0.98 No change ND
6 c.768G>T, Splice site 1 Het Klevering Tol. 0.56 NA Donor 70.4 58 Site broken (−17.51) ND
6 c.658C>T, p.Arg220Cys 1 Het Webster Tol. NA Benign 0.39 No change ND
11 c.1411G>A, p.Glu471Lys 1 Het Allikmets Tol NA Benign 0.01 Acceptor 71.7 43 Site broken (−40.4) 11/13006 db SNP (rs1800548)
12 c.1622T>C, p.Leu541Pro 1 Het Fishman Intol. 0.00 PRD 0.961 No change 2/13006 db SNP (rs61751392)
Int 12 c.1760+1G>T, Splice site 1 Het This study NA NA Donor 84.6 58 WT site broken (−31.72) ND
13 c.1805G>A, p.Arg602Gln 2 Het Webster Tol. NA PRD 0.513 48.9 78 New site (+59.14) 2/13006 db SNP (rs61749410)
14 c.1957C>T, p.Arg653Cys 2 Het Rivera Tol. 0.10 PRD 0.999 No change ND
17 c. 2588G>C, p.Gly863Ala 4 Het Allikmets Intol. 0.01 PRD 0.996 No change 68/13006 db SNP (rs76157638)
21 c.3113C>T, p.Ala1038Val 1 Het Webster Tol. NA Benign 0.014 Donor 43.5 70 New site (+61.72) 22/13006 db SNP (rs61751374)
24 c.3602T>G, p.Leu1201Arg 2 Het Lewis Tol. NA Benign 0.052 Donor 61.3 74 New site (+20.08) 416/13006 db SNP (rs61750126)
27 c.3898C>T, p.Arg1300* 1 Het Rivera NA NA ND
28 c.4139C>T, p.Pro1380Leu 2 Het Lewis Intol. 0.01 Benign 0.377 No change 2/13006 db SNP (rs61750130)
28 c.4222 T>C, p.Trp1408Arg 2 Het Lewis Tol. NA PRD 0.845 No change ND dbSNP (rs61750135)
29 c.4319T>C, p.Phe1440Ser 1 Het Lewis Tol. NA PRD 0.744 No change ND dbSNP (rs61750141)
30 c.4469G>A, p.Cys1490Tyr 1 Het Webster Intol. 0.03 PRD 0.994 No change ND dbSNP (rs61751402)
31 c.4577C>T, p.Thr1526Met 1 Het Lewis Intol. 0.00 PRD 0.91 No change ND db SNP (rs61750152)
31 c.4594G>T, p.Asp1532Asn 3 Het Lewis Tol. NA PRD 0.853 No change ND
33 c.4685T>C, p.Ile1562Thr 1 Het Allikmets Tol. NA Benign 0.034 No change 18/13006 db SNP (rs1762111)
35 c.4956T>G, p.Tyr1652* 1 Het Fumagalli NA NA Acceptor 43 72 New site (+67.36) ND
35 c.4918C>T, p.Arg1640Trp 2 Het Rozet Intol. 0.00 PRD 1 No change ND dbSNP (rs61751404)
35 c.4926C>G, p.Ser1642Arg 1 Het Birch Tol. 0.68 Benign 0.116 No change ND db SNP (rs61753017)
Int 35 c.5018+2T>C, Splice site 1 Het Fumagalli NA NA Donor 81.2 54 WT site broken (−33.07) ND
Int 38 c.5461-10T>C 3 Het Briggs NA NA No change 3/13006 db SNP (rs1800728)
40 c.5693G>A, p.Arg1898His 2 Het Allikmets NA Benign 0.00 No change 25/13006 db SNP (rs1800552)
42 c.5882G>A, p.Gly1961Glu 1 Het Allikmets Tol. 0.18 PRD 1 No change 41/13006 db SNP (rs1800553)
44 c.6079C>T, p.Leu2027Phe 4 Homo Lewis Intol. 0.02 PRD 0.999 No change 4/13006 db SNP (rs61751408)
44 c.6089G>A, p.Arg2030Gln 4 Het Lewis Tol. NA PRD 0.995 No change 8/13006 db SNP (rs61750641)
44 c.6118C>T, p.Arg2040* 1 Het Rosenberg NA NA ND
46 c.6320G>A, p.Arg2107His 1 Het Fishman Intol. 0.00 PRD 0.996 No change 91/13006 db SNP (rs62642564)

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Jan 9, 2017 | Posted by in OPHTHALMOLOGY | Comments Off on Clinical and Molecular Analysis of Stargardt Disease With Preserved Foveal Structure and Function
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