Association Between Genotype and Phenotype Severity in ABCA4-Associated Retinopathy.

Importance: ABCA4-associated retinopathy is a common inherited retinal disease, and its phenotype spans from late-onset macular dystrophy to extensive cone-rod degeneration. Over 2000 disease-causing variants in the ABCA4 gene have been identified. Objective: To investigate genotype-phenotype correlations in ABCA4-associated retinopathy. Design, Setting, and Participants: This cohort study took place at a single referral center for inherited retinal diseases in Italy. Data were prospectively acquired from January 2015 to June 2022. Patients diagnosed with an inherited retinal disease related to biallelic ABCA4 variants were included for analysis. Exposure: Genotype, classified into 4 groups according to the presence of the (1) p.Gly1961Glu allele, (2) a hypomorphic allele, (3) at least 1 moderate variant (moderate genotypes), or (4) 2 biallelic severe variants (severe genotypes). Main Outcomes and Measures: Total decreased autofluorescence (TDAF) and definitely decreased autofluorescence (DDAF) areas, inner and outer retinal volumes, and the respective progression rate. Results: A total of 71 patients (median [IQR] age, 34 [22.4-47.2] years; 40 [56%] female) were included in the study, and 54 (76%) were followed up for a median (IQR) of 3.5 (1.6-4.7) years. Compared with moderate genotypes, those with the p.Gly1961Glu allele had smaller TDAF lesions by 61% (95% CI, -78% to -33%; P < .001) and DDAF lesions by 77% (95% CI, -93% to -18%; P = .02), along with slower growth rates for both TDAF (0.05 mm/y; 95% CI, 0.01-0.07; P < .001) and DDAF (0.06 mm/y; 95% CI, 0-0.12; P = .004). Hypomorphic alleles were associated with a thicker inner (+0.19 mm3; 95% CI, +0.02 to +0.36; P = .03) and outer retinal volume (+0.16 mm3; 95% CI, +0.03 to +0.28; P = .01) compared with moderate genotypes as well as a slower TDAF growth rate (0.05 mm/y; 95% CI, 0.01-0.08; P = .007). Severe genotypes had a 7-fold larger TDAF area (95% CI, 3.4-14.7; P < .001) and 11-fold larger DDAF area (95% CI, 2.9-42.1; P < .001) compared with moderate genotypes, along with faster growth rates estimated at 0.16 mm/y for TDAF (95% CI, 0.12-0.20; P < .001) and 0.17 mm/y for DDAF (95% CI, 0.12-0.23; P < .001). Conclusions and Relevance: In this study of ABCA4-associated retinopathy, a 4-tier classification of genotypes was found to capture substantial variation in disease phenotype severity. These findings could prove beneficial for the prognostication of patients and warrant consideration of genotype in the design of future clinical trials.

Optical Coherence Tomography Angiography in CRB1-Associated Retinal Dystrophies.

AIM OF THE STUDY: To report optical coherence tomography angiography (OCTA) findings in patients affected by CRB1-associated retinal dystrophies. METHOD: Patients affected by a genetically confirmed CRB1-associated retinal dystrophy were prospectively enrolled in an observational study, along with age- and sex-matched healthy volunteers as control subjects. All study and control subjects received a complete ophthalmic examination and multimodal retinal imaging, including OCTA. RESULT: A total of 12 eyes from 6 patients were included in the study. The mean BCVA of patients was 0.42 ± 0.25 logMAR. Two patients showed large central atrophy, with corresponding definite hypo-autofluorescence on fundus autofluorescence (FAF). Another four patients disclosed different degrees of RPE mottling, with uneven FAF. On OCTA, the macular deep capillary plexus and choriocapillaris had a lower vessel density in eyes affected by CRB1-associated retinopathy when compared to healthy controls. On the other hand, vessel density at the peripapillary radial capillary plexus, superficial capillary plexus, and deep capillary plexus was significantly altered with respect to control eyes. Statistical analyses disclosed a negative correlation between the deep capillary plexus and both LogMAR best corrected visual acuity and central retinal thickness. CONCLUSION: Our study reveals that CRB1-associated retinal dystrophies are characterized by vascular alterations both in the macular and peripapillary region, as assessed by OCTA.

BENIGN FOVEAL DEPIGMENTATION: A MULTIMODAL IMAGING INVESTIGATION.

PURPOSE: To describe the multimodal imaging characteristics of benign foveal depigmentation. METHODS: The study was designed as prospective observational case series. Patients with benign foveal depigmentation were prospectively investigated by means of multimodal imaging, including blue-light and near-infrared fundus autofluorescence, optical coherence tomography (OCT), OCT angiography, color testing, microperimetry, and electrophysiology. The main outcome measures were vessel density and retinal pigment epithelium (RPE)/photoreceptor complex OCT reflectivity. RESULTS: Overall, 4 patients were identified, with bilateral and unilateral involvement in 1 case and 3 cases, respectively. Fundus autofluorescence provided variable results, showing more impairment on near-infrared fundus autofluorescence. Structural OCT revealed slight attenuation of the outer retinal bands in the area affected by benign foveal depigmentation, associated with choroidal hypertransmission, whereas enface OCT better delineated the attenuation of the reflectivity signal. The mean reflectivity intensity of RPE/photoreceptor complex was statistically significantly reduced in patients with respect to control subjects in the benign foveal depigmentation area. Optical coherence tomography angiography, color testing, microperimetry, electrooculogram, and electroretinogram findings were normal. CONCLUSION: Benign foveal depigmentation may represent a focal RPE disease. The limited alterations within the RPE band, as visualized on enface OCT and confirmed on near-infrared fundus autofluorescence, suggest an impairment in melanin production or distribution within the RPE cells.

Multimodal imaging in Schubert-Bornschein congenital stationary night blindness.

BACKGROUND: Schubert-Bornschein (SB) is the most common type of people with congenital stationary night blindness (CSNB). The aim of the study is to describe the optical coherence tomography (OCT) and optical coherence tomography angiography (OCTA) findings in patients with SB CSNB. METHODS: Prospective, observational case series including three patients with genetically confirmed CSNB along with matched controls, who underwent complete ophthalmic examination and multimodal imaging. RESULTS: On SD-OCT, a significant focal outer plexiform layer (OPL) thickening and a corresponding focal outer nuclear layer (ONL) thinning were identified in the macular area (p < 0.001). OCTA analysis overall showed decreased density of macular deep capillary plexus (mDCP) and macular choriocapillaris (mCC) (p = 0.008 and p = 0.033, respectively). DCP vessel density in the area corresponding to OPL thickening was significantly increased compared to the remaining retina (p < 0.001). CONCLUSION: SB CSNB is characterized by retinal vascular impairment, as detected on OCTA.

Gene Therapy in Inherited Retinal Diseases: An Update on Current State of the Art.

Background: Gene therapy cannot be yet considered a far perspective, but a tangible therapeutic option in the field of retinal diseases. Although still confined in experimental settings, the preliminary results are promising and provide an overall scenario suggesting that we are not so far from the application of gene therapy in clinical settings. The main aim of this review is to provide a complete and updated overview of the current state of the art and of the future perspectives of gene therapy applied on retinal diseases. Methods: We carefully revised the entire literature to report all the relevant findings related to the experimental procedures and the future scenarios of gene therapy applied in retinal diseases. A clinical background and a detailed description of the genetic features of each retinal disease included are also reported. Results: The current literature strongly support the hope of gene therapy options developed for retinal diseases. Although being considered in advanced stages of investigation for some retinal diseases, such as choroideremia (CHM), retinitis pigmentosa (RP), and Leber's congenital amaurosis (LCA), gene therapy is still quite far from a tangible application in clinical practice for other retinal diseases. Conclusions: Gene therapy is an extremely promising therapeutic tool for retinal diseases. The experimental data reported in this review offer a strong hope that gene therapy will be effectively available in clinical practice in the next years.

Multimodal imaging in pediatric arterial macroaneurysm: A case report.

Our aim is to report a case of asymptomatic retinal arterial macroaneurysm in a 9-year-old female patient. She was referred to our Ophthalmology clinic for a routine ophthalmologic examination and, after the detection of a juxtafoveal saccular vascular enlargement in indirect ophthalmoscopy, underwent a multimodal imaging assessment. Optical coherence tomography-angiography and fluorescein angiography were important to determine the nature of the lesion, identified as a congenital retinal arterial macroaneurysm. Retinal vascular abnormalities represent a rare finding in pediatric patients and must be carefully explored to establish the correct diagnosis. A multimodal imaging approach was very useful to thoroughly reach this target. Vascular abnormalities represent a rare finding in pediatric patients and must be carefully explored to establish the correct diagnosis. A multimodal imaging approach is very useful to study in deep the reported arterial macroaneurysm in a non-invasive way.

Reviewing the Role of Ultra-Widefield Imaging in Inherited Retinal Dystrophies.

Inherited retinal dystrophies (IRD) are a heterogeneous group of rare chronic disorders caused by genetically determined degeneration of photoreceptors and retinal pigment epithelium cells. Ultra-widefield (UWF) imaging is a useful diagnostic tool for evaluating retinal integrity in IRD, including Stargardt disease, retinitis pigmentosa, cone dystrophies, and Best vitelliform dystrophy. Color or pseudocolor and fundus autofluorescence images obtained with UWF provide previously unavailable information on the retinal periphery, which correlates well with visual field measurement or electroretinogram. Despite unavoidable artifacts of the UWF device, the feasibility of investigations in infants and in patients with poor fixation makes UWF imaging a precious resource in the diagnostic armamentarium for IRD.

OPTICAL COHERENCE TOMOGRAPHY AND OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY EVALUATION OF COMBINED HAMARTOMA OF THE RETINA AND RETINAL PIGMENT EPITHELIUM.

PURPOSE: To study multimodal imaging features of combined hamartoma of the retina and retinal pigment epithelium (CHRRPE). METHODS: Six patients (3 males, mean age 11 years) and a healthy age-matched control group made up of 15 healthy subjects (8 males, mean age 12.6 years) were included in the analysis. Complete ophthalmologic examination was performed, including best-corrected visual acuity, anterior and posterior segment slit-lamp evaluation, and tonometry. The multimodal imaging protocol included fundus images, structural optical coherence tomography (OCT), and swept-source OCT angiography (OCTA). The main outcome measures included the qualitative evaluation of both OCT and OCTA features of CHRRPE, retinal and choroidal thickness measurements, and the quantitative analysis of superficial capillary plexus, deep capillary plexus, and choriocapillaris vessel densities. RESULTS: Optical coherence tomography features of CHRRPE were examined extensively. Multiple little hyperreflective triangular outer retinal alterations were found at the CHRRPE edges in all patients; these were dubbed the "shark-teeth" sign. Optical coherence tomography angiography showed rarefaction and morphologic alterations of all retinal plexa. Moreover, quantitative analysis revealed a statistically significant decrease in superficial capillary plexus, deep capillary plexus, and choriocapillaris vessel densities in patients affected by CHRRPE compared with the control group. CONCLUSION: Optical coherence tomography and OCTA analyses allowed the accurate qualitative and quantitative analyses of CHRRPE features. Further studies are needed to better define OCTA changes of CHRRPE better and to improve our understanding of the possible causes of the shark-teeth sign.

CAPILLARY NETWORK ALTERATIONS IN X-LINKED RETINOSCHISIS IMAGED ON OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY.

PURPOSE: To assess foveal and parafoveal vasculature at the superficial capillary plexus, deep capillary plexus, and choriocapillaris of patients with X-linked retinoschisis by means of optical coherence tomography angiography. METHODS: Six patients with X-linked retinoschisis (12 eyes) and seven healthy controls (14 eyes) were recruited and underwent complete ophthalmologic examination, including best-corrected visual acuity, dilated fundoscopy, and 3 × 3-mm optical coherence tomography angiography macular scans (DRI OCT Triton; Topcon Corp). After segmentation and quality review, optical coherence tomography angiography slabs were imported into ImageJ 1.50 (NIH; Bethesda) and digitally binarized. Quantification of vessel density was performed after foveal avascular zone area measurement and exclusion. Patients were additionally divided into "responders" and "nonresponders" to dorzolamide therapy. RESULTS: Foveal avascular zone area resulted markedly enlarged at the deep capillary plexus (P < 0.001), particularly in nonresponders. Moreover, patients disclosed a significant deep capillary plexus rarefaction, when compared with controls (P: 0.04); however, a subanalysis revealed that this damage was limited to the fovea (P: 0.006). Finally, the enlargement of foveal avascular zone area positively correlated with a decline in best-corrected visual acuity (P: 0.01). CONCLUSION: Prominent foveal vascular impairment is detectable in the deep capillary plexus of patients with X-linked retinoschisis. Our results correlate with functional outcomes, suggesting a possible vascular role in X-linked retinoschisis clinical manifestations.

VASCULAR ALTERATIONS REVEALED WITH OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY IN PATIENTS WITH CHOROIDEREMIA.

PURPOSE: Choroideremia is a rare degenerative retinal disease that causes incurable blindness. It occurs as a result of the deficiency of the X-linked CHM gene, which encodes the Rab escort protein 1 (REP1). Gene therapy has been developed to treat CHM using adeno-associated viral vectors and is currently undergoing clinical trials. Expression of the CHM gene is ubiquitous throughout the retina, and it is therefore important to identify which retinal layers are affected in the disease process. The purpose of this study was to assess in particular the choriocapillaris using optical coherence tomography angiography because this layer is difficult to see with conventional imaging techniques. METHODS: Six men with choroideremia were identified and underwent standardized optical coherence tomography angiography as part of an ethics-approved clinical study and were compared with age-matched control subjects. RESULTS: The choriocapillaris appeared normal in regions where the retinal pigment epithelium remained intact, but it was deficient elsewhere. The outer retinal vasculature showed significant changes peripherally but also some changes centrally. The inner retinal vasculature appeared unaffected by the disease process. CONCLUSION: Choroideremia is a disease in which the choriocapillaris maintains a normal structure until the loss of the overlying retinal pigment epithelium. The inner retina also appears not to be affected at the vascular level. Although this study is limited by the small number of patients eligible for inclusion in the study, the observations support the concept of targeting gene therapy to the retinal pigment epithelium and outer retina because there is no evidence of independent degeneration of the choriocapillaris.

Integration of multigene panels for the diagnosis of hereditary retinal disorders using Next Generation Sequencing and bioinformatics approaches.

In recent years, Next-Generation Sequencing (NGS) opened a new way for the study of pathogenic mechanisms and for molecular diagnosis of inherited disorders. In the present work, we focused our attention on the inherited retinal dystrophies (IRDs), a group of specific disorders of the retina, displaying a very high clinical and genetic heterogeneity, whose genetic diagnosis is not easily feasible. It represents a paradigmatic example for the integration of clinical and molecular examination toward precision medicine. In this paper, we discuss the use of targeted NGS resequencing of selected gene panels in a cohort of patients affected by IRDs. We tested the hypothesis to apply a selective approach based on a careful clinical examination. By this approach we reached a 66% overall detection rate for pathogenic variants, with a 52% diagnostic yield. Reduction of the efforts for validation and classification of variants is a clear advantage for the management of genetic testing in a clinical setting.

Resolution of cystoid macular edema following arginine-restricted diet and vitamin B6 supplementation in a case of gyrate atrophy.

We report the outcome of 3 years of arginine-restricted diet and vitamin B6 supplementation in a boy who presented with gyrate atrophy of the choroid and retina and bilateral cystoid macular edema. The diagnosis of gyrate atrophy was made on the basis of clinical findings and increased plasma ornithine levels. Molecular genetic testing revealed a disease-causing homozygous mutation in the ornithine aminotransferase (OAT) gene. After 3 months of dietary modification and pyridoxine supplementation, visual acuity improved, and optical coherence tomography showed resolution of cystoid macular edema in both eyes. This anatomical and functional improvement was maintained during 3 years of follow-up.

Regressive Retinal Flecks in CRX-Mutated Early-Onset Retinal Dystrophy.

PURPOSE: To describe a peculiar flecked-retina phenotype in a young female affected by early-onset retinal dystrophy due to a heterozygous mutation in the cone-rod transcription factor CRX gene. CASE REPORT: A 5-year-old girl presented with poor vision and nystagmus from the first month after birth. Opththalmologic examination at baseline revealed an altered foveal reflex, epiretinal membrane, and yellow fleck-like retinal deposits in the mid- and extreme periphery bilaterally that disappeared after 3 years of follow-up. Electoretinogram was non-recordable in both rods and cones components bilaterally. Genomic sequencing identified a heterozygous missense mutation -c.425A > G (Tyr142Cys) in CRX. CONCLUSIONS: We identified a novel early-onset retinal dystrophy-related heterozygous CRX mutation associated with early and severe rod and cone dysfunction and regressive flecked-retina appearance on ophthalmoscopy.

Morpho-functional correlation of fundus autofluorescence in Stargardt disease.

BACKGROUND: To correlate patterns in short-wavelength (SW) and near-infrared (NIR) fundus autofluorescence (FAF) with morpho-functional outcomes in eyes affected by Stargardt disease. METHODS: Fifty-four eyes of 27 patients were prospectively enrolled. All patients underwent a complete ophthalmologic examination including SW-FAF, NIR-FAF, microperimetry and spectral-domain optical coherence tomography (SD-OCT). The main outcome measures were identification of a correlation between NIR-FAF and SW-FAF patterns within the foveal region and best corrected visual acuity (BCVA) values. Secondary outcome measures were correlation of FAF patterns with SD-OCT findings and retinal sensitivity on microperimetry. RESULTS: Eyes showing a pattern of foveal hyper-FAF on NIR-FAF had a higher BCVA than eyes with a reduced FAF signal (0.44±0.23 LogMAR vs 1.08±0.19, p<0.001). Similarly, mean sensitivity within 2° of the foveal region was significantly better (6.45±2.39 dB) in eyes with hyper-FAF than in eyes with hypo-FAF (0.23±0.45 dB, p<0.001). Moreover, eyes with hyper-FAF on SW-FAF did not present a significant difference in BCVA (0.73±0.31 vs 0.83±0.43, p=0.335) and mean retinal sensitivity (4.34±3.91 dB vs 2.33±2.96, p=0.07) compared with the subgroup with foveal hypo-FAF. The integrity of both the photoreceptor inner/outer segment junction and the photoreceptor outer segment/retinal pigmented epithelium junction was significantly correlated with a preserved BCVA and a foveal hyper-FAF pattern on NIR-FAF. CONCLUSIONS: Our data suggest that NIR-FAF patterns correlate with morpho-functional outcomes in eyes affected by Stargardt disease. Longitudinal investigations are warranted to assess more precisely the actual contribution of NIR-FAF in the clinical characterisation of Stargardt disease.

Posterior polymorphous corneal dystrophy concomitant to large colloid drusen.

PURPOSE: To describe the previously unreported concomitance of 2 uncommon ocular conditions: posterior polymorphous corneal dystrophy (PPCD) and large colloid drusen (LCD). METHODS: A 45-year-old woman underwent a complete ophthalmologic examination with slit-lamp biomicroscopy and blue fundus autofluorescence with spectral-domain optical coherence tomography, as well as complete systemic examination and renal function investigation. RESULTS: On slit-lamp biomicroscopy, a corneal lesion located at Descemet membrane was observed in the right eye. The clinical features of deep posterior stromal-endothelial linear bands with vesicles and irregular opacities of posterior corneal surface were consistent with the diagnosis of PPCD. Fundus biomicroscopy and blue fundus autofluorescence showed LCD. DISCUSSIONS: We report the unusual coexistence of PPCD and LCD in a young, healthy subject. Posterior polymorphous corneal dystrophy and LCD share morphologic similarities and dysfunctions of collagen architecture in the basement membrane layer, which suggests a possible common pathogenic pathway.

Spectral domain optical coherence tomography findings in patients with retinitis pigmentosa.

BACKGROUND: To report the morphological macular findings detected by spectral domain optical coherence tomography (SD-OCT) and to determine their prevalence in patients with retinitis pigmentosa (RP). METHODS: SD-OCT scans of 176 eyes from 90 patients affected by RP were reviewed. A careful evaluation was carried out on photoreceptor inner/outer segment (IS/OS) junction, external limiting membrane (ELM), inner limiting membrane thickening (ILMT), epiretinal membranes (ERMs), retinal micropseudocysts (MPCs), cystoid macular edema (CME), macular holes (MHs) and choroidal neovascularization (CNV). RESULTS: The photoreceptor IS/OS junction was absent in the foveal region of 24 eyes (13.6%) and disrupted in 84 eyes (47.7%). The ELM was absent in 24 eyes (13.6%), whereas the ILMT was found in 118 eyes (67%). The presence of an ERM was detected in 48 eyes (27.3%). Some sort of vitreomacular alteration (ILMT and/or ERM) was identifiable in a total of 94.3% of eyes with RP. The presence of MPCs was detected in 32 eyes (18.2%). An evident CME was found in 22 eyes (12.5%). We also found MHs in 8 eyes (4.5%) and CNV in 3 eyes (1.7%). CONCLUSIONS: Our data indicate that RP is associated with alterations of many retinal layers. In particular, the vitreoretinal interface is affected in 94% of patients, and MPC can be identified in 18% of eyes. SD-OCT may contribute to the understanding of the pathophysiological mechanism involved in RP.

Ocular phenotypes associated with biallelic mutations in BEST1 in Italian patients.

PURPOSE: To report on the phenotype and the genotype of Italian patients carrying BEST1 mutations on both alleles. METHODS: Five Italian patients from four independent pedigrees with retinal dystrophy associated with biallelic BEST1 variants were recruited from different parts of Italy. Molecular genetic analysis of the BEST1 gene was performed with direct sequencing techniques. All the subjects included in the study were clinically evaluated with a standard ophthalmologic examination, fundus photography, optical coherence tomography scan, and electrophysiological investigations. RESULTS: Six BEST1 variants were identified. Three, c.1699del (p.Glu557AsnfsX52), c.625delAAC (p.Asn179del), and c.139C>T (p.Arg47Cys), were novel, and three had already been reported in the literature, c.301C>A(p.Pro101Thr), c.934G>A (p.Asp312Asn), and c.638A>G (p.Glu213Gly). Four were missense mutations, and two were deletions. Only one BEST1 mutation was located within one of the four mutational clusters described in typical autosomal dominant Best vitelliform macular dystrophy (BVMD). Four patients showed a BVMD phenotype while one patient presented a clinical picture consistent with autosomal recessive bestrophinopathy (ARB). CONCLUSIONS: Biallelic BEST1 sequence variants can be associated with at least two different phenotypes: BVMD and ARB. The phenotypic result of the molecular changes probably depends on the characteristics and the combination of the different BEST1 mutations, but unknown modifying factors such as other genes or the environment may also play a role.