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Aims: This study aimed to evaluate the effectiveness of somatostatin analogues (SA) for cystoid maculopathy (CM) in retinitis pigmentosa (RP) patients., Materials and Methods: In this retrospective case series, clinical and imaging characteristics of 28 RP patients with CM, unresponsive to carbonic anhydrase inhibitors, were collected from medical charts. All patients received SA treatment as an alternative (octreotide long-acting release at 20 mg/month or 30 mg/month, or lanreotide at 90 mg/month or 120 mg/month). Outcome measures were mean reduction in foveal thickness (FT) and foveal volume (FV) and mean increase in best-corrected visual acuity at 3, 6 and 12 months of treatment initiation. Linear mixed models were used to calculate the effectiveness over time., Results: 52 eyes of 28 RP patients were included; 39% were male. The median age at the start of treatment was 39 years (IQR 30-53). Median follow-up was 12 months (range 6-12). From baseline to 12 months, the mean FT decreased from 409±136 µm to 334±119 µm and the mean FV decreased from 0.31±0.10 mm 3 to 0.25±0.04 mm 3 . Linear mixed model analyses showed a significant decrease in log FT and log FV at 3, 6 and 12 months after the start of treatment compared with baseline measurements (p<0.001, p<0.001, p<0.001). Mean best-corrected visual acuity did not increase significantly (0.46±0.35 logMAR to 0.45±0.38 logMAR after 12 months)., Discussion: SA may be an effective alternative treatment to reduce CM in RP patients., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Purpose: Gene-based therapies for inherited retinal dystrophies (IRDs) are upcoming. Treatment before substantial vision loss will optimize outcomes. It is crucial to identify common phenotypes and causative genes in children. This study investigated the frequency of these in pediatric IRD with the aim of highlighting relevant groups for future therapy., Methods: Diagnostic, genetic, and demographic data, collected from medical charts of patients with IRD aged up to 20 years (n = 624, 63% male), registered in the Dutch RD5000 database, were analyzed to determine frequencies of phenotypes and genetic causes. Phenotypes were categorized as nonsyndromic (progressive and stationary IRD) and syndromic IRD. Genetic causes, mostly determined by whole-exome sequencing (WES), were examined. Additionally, we investigated the utility of periodic reanalysis of WES data in genetically unresolved cases., Results: Median age at registration was 13 years (interquartile range, 9-16). Retinitis pigmentosa (RP; n = 123, 20%), Leber congenital amaurosis (LCA; n = 97, 16%), X-linked retinoschisis (n = 64, 10%), and achromatopsia (n = 63, 10%) were the most frequent phenotypes. The genetic cause was identified in 76% of the genetically examined patients (n = 473). The most frequently disease-causing genes were RS1 (n = 32, 9%), CEP290 (n = 28, 8%), CNGB3 (n = 21, 6%), and CRB1 (n = 17, 5%). Diagnostic yield after reanalysis of genetic data increased by 7%., Conclusions: As in most countries, RP and LCA are the most prominent pediatric IRDs in the Netherlands, and variants in RS1 and CEP290 were the most prominent IRD genotypes. Our findings can guide therapy development to target the diseases and genes with the greatest needs in young patients.

Purpose: Gyrate atrophy of the choroid and retina (GACR) is an autosomal recessive inherited metabolic disorder (IMD) characterised by progressive retinal degeneration, leading to severe visual impairment. The rapid developments in ophthalmic genetic therapies warrant knowledge on clinical phenotype of eligible diseases such as GACR to define future therapeutic parameters in clinical trials., Methods: Retrospective chart analysis was performed in nineteen patients. Data were analysed using IBM SPSS Statistics version 28.0.1.1., Results: Nineteen patients were included with a mean age of 32.6 years (range 8-58). Mean age at onset of ophthalmic symptoms was 7.9 years (range 3-16). Median logMAR of visual acuity at inclusion was 0.26 (range -0.18-3.00). Mean age at cataract surgery was 28.8 years (n = 11 patients). Mean spherical equivalent of the refractive error was -8.96 (range -20.87 to -2.25). Cystoid maculopathy was present in 68% of patients, with a loss of integrity of the foveal ellipsoid zone (EZ) in 24/38 eyes. Of the 14 patients treated with dietary protein restriction, the four patients who started the diet before age 10 showed most benefit., Conclusion: This study demonstrates the severe ophthalmic disease course associated with GACR, as well as possible benefit of early dietary treatment. In addition to visual loss, patients experience severe myopia, early-onset cataract, and CME. There is a loss of foveal EZ integrity at a young age, emphasising the need for early diagnosis enabling current and future therapeutic interventions., (© 2024. The Author(s).)

Purpose: We investigated the natural history of retinal dystrophy owing to variants in the MYO7A gene., Methods: Fifty-three patients (mean age, 33.6 ± 16.7 years) with Usher syndrome owing to biallelic, mostly pathogenic, variants in MYO7A underwent baseline and two annual follow-up visits. Best-corrected visual acuity (BCVA), semiautomatic kinetic visual field, full-field electroretinogram, color fundus imaging, microperimetry, spectral-domain optical coherence tomography, and fundus autofluorescence were assessed., Results: At baseline, all patients presented with decreased BCVA (66.4 ± 17.9 Early Treatment Diabetic Retinopathy score and 59.5 ± 21.7 Early Treatment Diabetic Retinopathy score, in the better- and worse-seeing eyes, respectively), restricted semiautomatic kinetic visual field (III4e area, 3365.8 ± 4142.1°2; 4176.4 ± 4400.3°2) and decreased macular sensitivity (9.7 ± 9.9 dB; 9.0 ± 10.2 dB). Spectral-domain optical coherence tomography revealed reduced central macular thickness (259.6 ± 63.0 µm; 250.7 ± 63.3 µm) and narrowed ellipsoid zone band width (2807.5 ± 2374.6 µm; 2615.5 ± 2370.4 µm). Longitudinal analyses (50 patients) showed a significant decrease of BCVA in better-seeing eyes, whereas no changes were observed in worse-seeing eyes for any parameter. BCVA, semiautomatic kinetic visual field (III4e and V4e) and macular sensitivity were related significantly to age at baseline. Hyperautofluorescent foveal patch (16 eyes [31.4%]) and abnormal central hypoautofluorescence (9 eyes [17.6%]) were significantly associated with worse morphological and functional read-outs compared with the hyperautofluorescent ring pattern (22 eyes [43.1%])., Conclusions: Our European multicentric study offers the first prospective longitudinal analysis in one of the largest cohorts of MYO7A patients described to date, confirming the slow disease progression. More important, this study emphasizes the key role of fundus autofluorescence patterns in retinal impairment staging and advocates its adoption as an objective biomarker in patient selection for future gene therapy clinical trials.

Purpose: To date, there is no standard treatment regimen for carbonic anhydrase inhibitors (CAIs) in X-linked retinoschisis (XLRS) patients. This retrospective study aims to evaluate the efficacy of CAIs on visual acuity and cystoid fluid collections (CFC) in XRLS patients in Dutch and Belgian tertiary referral centers., Design: Retrospective cohort study., Participants: Forty-two patients with XLRS., Methods: In total, 42 patients were enrolled. To be included, patients had to have previous treatment with an oral CAI (acetazolamide), a topical CAI (brinzolamide/dorzolamide), or a combination of an oral and a topical CAI for at least 4 consecutive weeks. We evaluated the effect of the CAI on best-corrected visual acuity (BCVA) and central foveal thickness (CFT) on OCT., Main Outcome Measures: Central foveal thickness and BCVA., Results: The median age at the baseline visit of the patients in this cohort study was 14.7 (range, 43.6) years, with a median (interquartile range [IQR]) follow-up period of 4.0 (2.2-5.2) years. During the follow-up period, 25 patients were treated once with an oral CAI (60%), 24 patients were treated once with a topical CAI (57%), and 11 patients were treated once with a combination of both topical and oral CAI (26%). We observed a significant reduction of CFT for oral CAI by 14.37 μm per 100 mg per day (P < 0.001; 95% confidence interval [CI], -19.62 to -9.10 μm) and for topical CAI by 7.52 μm per drop per day (P = 0.017; 95% CI, -13.67 to -1.32 μm). The visual acuity changed significantly while on treatment with oral CAI by -0.0059 logMAR per 100 mg (P = 0.008; 95% CI, -0.010 to -0.0013 logMAR). Seven patients (17%) had side effects leading to treatment discontinuation., Conclusions: Our data indicate that treatment with (oral) CAI may be beneficial for short-term management of CFC in patients with XLRS. Despite a significant reduction in CFT, the change in visual acuity was modest and not of clinical significance. Nonetheless, the anatomic improvement of the central retina in these patients may be of value to create an optimal retinal condition for future potential treatment options such as gene therapy., Financial Disclosure(s): The authors have no proprietary or commercial interest in any materials discussed in this article., (Copyright © 2023 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Purpose: Inherited retinal diseases (IRDs) are a group of monogenic conditions that can lead to progressive blindness. Their missing heritability is still considerable, due in part to the presence of disease genes that await molecular identification. The purpose of this work was to identify novel genetic associations with IRDs., Methods: Patients underwent a comprehensive ophthalmological evaluation using standard-of-care tests, such as detailed retinal imaging (macular optical coherence tomography and short-wavelength fundus autofluorescence) and electrophysiological testing. Exome and genome sequencing, as well as computer-assisted data analysis were used for genotyping and detection of DNA variants. A minigene-driven splicing assay was performed to validate the deleterious effects of 1 of such variants., Results: We identified 8 unrelated families from Hungary, the United States, Israel, and The Netherlands with members presenting with a form of autosomal recessive and nonsyndromic retinal degeneration, predominantly described as rod-cone dystrophy but also including cases of cone/cone-rod dystrophy. Age of disease onset was very variable, with some patients experiencing first symptoms during their fourth decade of life or later. Myopia greater than 5 diopters was present in 5 of 7 cases with available refractive data, and retinal detachment was reported in 2 cases. All ascertained patients carried biallelic loss-of-function variants in UBAP1L (HGNC: 40028), a gene with unknown function and with homologies to UBAP1, encoding a protein involved in ubiquitin metabolism. One of these pathogenic variants, the intronic NM&#95;001163692.2:c.910-7G>A substitution, was identified in 5 unrelated families. Minigene-driven splicing assays in HEK293T cells confirmed that this DNA change is responsible for the creation of a new acceptor splice site, resulting in aberrant splicing., Conclusion: We identified UBAP1L as a novel IRD gene. Although its function is currently unknown, UBAP1L is almost exclusively expressed in photoreceptors and the retinal pigment epithelium, hence possibly explaining the link between pathogenic variants in this gene and an ocular phenotype., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: To assess the longitudinal vision-related quality of life among patients with CRB1-associated inherited retinal dystrophies., Methods: In this longitudinal questionnaire study, the National Eye Institute Visual Function Questionnaire (39 items, NEI VFQ-39) was applied at baseline, two-year follow-up, and 4-year follow-up in patients with pathogenic CRB1 variants. [Correction added on 20 November 2023, after first online publication: The preceding sentence has been updated in this version.] Classical test theory was performed to obtain subdomain scores and in particular 'near activities' and 'total composite' scores. The Rasch analysis based on previous calibrations of the NEI VFQ-25 was applied to create visual functioning and socio-emotional subscales., Results: In total, 22 patients with a CRB1-associated retinal dystrophy were included, […] with a median age of 25.0 years (interquartile range: 13-31 years) at baseline and mean follow-up of 4.0 ± 0.3 years. [Correction added on 20 November 2023, after first online publication: The preceding sentence has been updated in this version.] A significant decline at 4 years was observed for 'near activities' (51.0 ± 23.8 vs 35.4 ± 14.7, p = 0.004) and 'total composite' (63.0 ± 13.1 vs 52.0 ± 12.1, p = 0.001) subdomain scores. For the Rasch-scaled scores, the 'visual functioning' scale significantly decreased after 2 years (-0.89 logits; p = 0.012), but not at 4-year follow-up (+0.01 logits; p = 0.975). [Correction added on 20 November 2023, after first online publication: In the preceding sentence, "…after 4 years…" has been corrected to "…after 2 years…" in this version.] The 'socio-emotional' scale also showed a significant decline after 2 years (-0.78 logits, p = 0.033) and 4 years (-0.83 logits, p = 0.021)., Conclusion: In the absence of an intervention, a decline in vision-related quality of life is present in patients with pathogenic CRB1 variants at 4-year follow-up. Patient-reported outcome measures should be included in future clinical trials, as they can be a potential indicator of disease progression and treatment efficacy., (© 2023 The Authors. Acta Ophthalmologica published by John Wiley & Sons Ltd on behalf of Acta Ophthalmologica Scandinavica Foundation.)

Importance: Previous studies indicated that female sex might be a modifier in Stargardt disease, which is an ABCA4-associated retinopathy., Objective: To investigate whether women are overrepresented among individuals with ABCA4-associated retinopathy who are carrying at least 1 mild allele or carrying nonmild alleles., Data Sources: Literature data, data from 2 European centers, and a new study. Data from a Radboudumc database and from the Rotterdam Eye Hospital were used for exploratory hypothesis testing., Study Selection: Studies investigating the sex ratio in individuals with ABCA4-AR and data from centers that collected ABCA4 variant and sex data. The literature search was performed on February 1, 2023; data from the centers were from before 2023., Data Extraction and Synthesis: Random-effects meta-analyses were conducted to test whether the proportions of women among individuals with ABCA4-associated retinopathy with mild and nonmild variants differed from 0.5, including subgroup analyses for mild alleles. Sensitivity analyses were performed excluding data with possibly incomplete variant identification. χ2 Tests were conducted to compare the proportions of women in adult-onset autosomal non-ABCA4-associated retinopathy and adult-onset ABCA4-associated retinopathy and to investigate if women with suspected ABCA4-associated retinopathy are more likely to obtain a genetic diagnosis. Data analyses were performed from March to October 2023., Main Outcomes and Measures: Proportion of women per ABCA4-associated retinopathy group. The exploratory testing included sex ratio comparisons for individuals with ABCA4-associated retinopathy vs those with other autosomal retinopathies and for individuals with ABCA4-associated retinopathy who underwent genetic testing vs those who did not., Results: Women were significantly overrepresented in the mild variant group (proportion, 0.59; 95% CI, 0.56-0.62; P < .001) but not in the nonmild variant group (proportion, 0.50; 95% CI, 0.46-0.54; P = .89). Sensitivity analyses confirmed these results. Subgroup analyses on mild variants showed differences in the proportions of women. Furthermore, in the Radboudumc database, the proportion of adult women among individuals with ABCA4-associated retinopathy (652/1154 = 0.56) was 0.10 (95% CI, 0.05-0.15) higher than among individuals with other retinopathies (280/602 = 0.47)., Conclusions and Relevance: This meta-analysis supports the likelihood that sex is a modifier in developing ABCA4-associated retinopathy for individuals with a mild ABCA4 allele. This finding may be relevant for prognosis predictions and recurrence risks for individuals with ABCA4-associated retinopathy. Future studies should further investigate whether the overrepresentation of women is caused by differences in the disease mechanism, by differences in health care-seeking behavior, or by health care discrimination between women and men with ABCA4-AR.

Inherited macular dystrophies (iMDs) are a group of genetic disorders, which affect the central region of the retina. To investigate the genetic basis of iMDs, we used single-molecule Molecular Inversion Probes to sequence 105 maculopathy-associated genes in 1352 patients diagnosed with iMDs. Within this cohort, 39.8% of patients were considered genetically explained by 460 different variants in 49 distinct genes of which 73 were novel variants, with some affecting splicing. The top five most frequent causative genes were ABCA4 (37.2%), PRPH2 (6.7%), CDHR1 (6.1%), PROM1 (4.3%) and RP1L1 (3.1%). Interestingly, variants with incomplete penetrance were revealed in almost one-third of patients considered solved (28.1%), and therefore, a proportion of patients may not be explained solely by the variants reported. This includes eight previously reported variants with incomplete penetrance in addition to CDHR1 :c.783G>A and CNGB3 :c.1208G>A. Notably, segregation analysis was not routinely performed for variant phasing-a limitation, which may also impact the overall diagnostic yield. The relatively high proportion of probands without any putative causal variant (60.2%) highlights the need to explore variants with incomplete penetrance, the potential modifiers of disease and the genetic overlap between iMDs and age-related macular degeneration. Our results provide valuable insights into the genetic landscape of iMDs and warrant future exploration to determine the involvement of other maculopathy genes.

Purpose: Eye inflammation may occur in patients with inherited retinal dystrophies (IRDs) and is seen frequently in IRDs associated with mutations in the CRB1 gene. The purpose of this study was to determine the types of inflammatory cells involved in IRDs, by deep profiling the composition of peripheral blood mononuclear cells of patients with a CRB1-associated IRD., Methods: This study included 33 patients with an IRD with confirmed CRB1 mutations and 32 healthy controls. A 43-parameter flow cytometry analysis was performed on peripheral blood mononuclear cells isolated from venous blood. FlowSOM and manual Boolean combination gating were used to identify and quantify immune cell subsets., Results: Comparing patients with controls revealed a significant increase in patients in the abundance of circulating CD4+ T cells and CD8+ T cells that express sialyl Lewis X antigen. Furthermore, we detected a decrease in plasmacytoid dendritic cells and an IgA+CD24+CD38+ transitional B-cell subset in patients with an IRD., Conclusions: Patients with a CRB1-associated IRD show marked changes in blood leukocyte composition, affecting lymphocyte and dendritic cell populations. These results implicate inflammatory pathways in the disease manifestations of IRDs.

Purpose: Structural variants (SVs) play an important role in inherited retinal diseases (IRD). Although the identification of SVs significantly improved upon the availability of genome sequencing, it is expected that involvement of SVs in IRDs is higher than anticipated. We revisited short-read genome sequencing data to enhance the identification of gene-disruptive SVs., Methods: Optical genome mapping was performed to improve SV detection in short-read genome sequencing-negative cases. In addition, reanalysis of short-read genome sequencing data was performed to improve the interpretation of SVs and to re-establish SV prioritization criteria., Results: In a monoallelic USH2A case, optical genome mapping identified a pericentric inversion (173 megabase), with 1 breakpoint disrupting USH2A. Retrospectively, the variant could be observed in genome sequencing data but was previously deemed false positive. Reanalysis of short-read genome sequencing data (427 IRD cases) was performed which yielded 30 pathogenic SVs affecting, among other genes, USH2A (n = 15), PRPF31 (n = 3), and EYS (n = 2). Eight of these (>25%) were overlooked during previous analyses., Conclusion: Critical evaluation of our findings allowed us to re-establish and improve our SV prioritization and interpretation guidelines, which will prevent missing pathogenic events in future analyses. Our data suggest that more attention should be paid to SV interpretation and the current contribution of SVs in IRDs is still underestimated., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

A significant number of individuals with a rare disorder such as Usher syndrome (USH) and (non-)syndromic autosomal recessive retinitis pigmentosa (arRP) remain genetically unexplained. Therefore, we assessed subjects suspected of USH2A -associated disease and no or mono-allelic USH2A variants using whole genome sequencing (WGS) followed by an improved pipeline for variant interpretation to provide a conclusive diagnosis. One hundred subjects were screened using WGS to identify causative variants in USH2A or other USH/arRP-associated genes. In addition to the existing variant interpretation pipeline, a particular focus was put on assessing splice-affecting properties of variants, both in silico and in vitro . Also structural variants were extensively addressed. For variants resulting in pseudoexon inclusion, we designed and evaluated antisense oligonucleotides (AONs) using minigene splice assays and patient-derived photoreceptor precursor cells. Biallelic variants were identified in 49 of 100 subjects, including novel splice-affecting variants and structural variants, in USH2A or arRP/USH-associated genes. Thirteen variants were shown to affect USH2A pre-mRNA splicing, including four deep-intronic USH2A variants resulting in pseudoexon inclusion, which could be corrected upon AON treatment. We have shown that WGS, combined with a thorough variant interpretation pipeline focused on assessing pre-mRNA splicing defects and structural variants, is a powerful method to provide subjects with a rare genetic condition, a (likely) conclusive genetic diagnosis. This is essential for the development of future personalized treatments and for patients to be eligible for such treatments., Competing Interests: The authors declare no competing interests., (© 2023 The Author(s).)

Introduction: An increasing number of gene-specific therapies are being developed for inherited retinal degenerations (IRDs). Identification of well-characterized patients is an emerging need. We conducted the second multinational survey among the www.evicr.net and ERN-EYE members to understand the management and treatment of IRDs in Europe and compared it to the 2019 survey., Methods: An electronic survey questionnaire was developed and sent to 124 clinical centers (25 countries) by June/July 2021. Statistical analysis was performed with Excel and R., Results: The overall response rate was 44% but varied among countries. Only 9% of responding centers do not see IRD patients (2019 survey 14%), 42% follow at least 200 patients per year, 18% follow 500-999, and 2% more than 1,000. Databases exist in 86% of the centers (local 86%; national web based 12%). IRD patients are referred to www.evicr.net and ERN-EYE centers mainly by general ophthalmologists, patient self-referral, or medical retina specialists. Most IRD patients are first seen as adults. Signs and symptoms depend on age of onset: in infancy, nystagmus; at older age, night blindness and reduced visual field; reduced visual acuity is described at any age. Comprehensive ophthalmic examination always includes visual acuity and almost always visual field multimodal retinal imaging, electrophysiology, color vision testing, and refraction. Identification of genotypes is successful in 72% of centers in 40-80% of cases (2019 survey 69% of centers). The time for confirmation of the genetic diagnosis varies from 2-4 weeks to 24 months (2019 survey &gt;4 weeks ≤10 years). Genetic testing is covered by public health service in 83%, private health insurance in 29%, research funds in 24%; 5% do not have access to genetic testing (2019 survey 15%). The most striking result is the high increase in the involvement of centers in natural history and gene therapy trials that more than doubled for the latter., Discussion: This second multinational survey on management of IRDs in Europe highlights persistent important differences in the number of IRD patients managed per center, comparable diagnostic work-up, and increasing genotyping in diagnostic laboratories. The important increase in involvement of centers in natural history and gene therapy trials reflects the rapidly evolving field of gene therapy development. The survey provides important follow-up data for researchers, clinicians, caregivers, patient advocate groups, pharmaceutical companies, and investors., (© 2023 The Author(s). Published by S. Karger AG, Basel.)

Introduction: The aim of this study was to evaluate the current management of RPE65 biallelic mutation-associated inherited retinal degeneration (RPE65-IRD) in Europe since market authorization of voretigene neparvovec (VN, LuxturnaTM) in 2018. By July 2022, over 200 patients have been treated outside the USA, of whom about 90% in Europe. We conducted among all centers of the European Vision Institute Clinical Research Network www.evicr.net and health care providers (HCPs) of the European Reference Network dedicated to Rare Eye Diseases (ERN-EYE) the second multinational survey on management of IRDs in Europe elaborated by www.evicr.net with a special focus on RPE65-IRD., Methods: An electronic survey questionnaire with 48 questions specifically addressing RPE65-IRD (2019 survey 35) was developed and sent by June 2021 to 95 www.evicr.net centers and 40 ERN-EYE HCPs and affiliated members. Of note, 11 centers are members of both networks. Statistical analysis was performed with Excel and R., Results: The overall response rate was 44% (55/124); 26 centers follow RPE65 biallelic mutation-associated IRD patients. By June 2021, 8/26 centers have treated 57 RPE65-IRD cases (1-19/center, median 6) and 43 planned for treatment (range 0-10/center, median 6). The overall age range was 3-52 years, and on average 22% of the patients did not (yet) qualify for treatment (range 2-60%/center, median 15%). Main reasons were too advanced (range 0-100, median 75%) or mild disease (range 0-100, median 0). Eighty-three percent of centers (10/12) that follow RPE65 mutation-associated IRD patients treated with VN participate in the PERCEIVE registry (EUPAS31153, www.encepp.eu. Quality of life and full-field stimulus test improvements had the highest scores of the survey-reported outcome parameters in VN treatment follow-up., Conclusion: This second multinational survey on management of RPE65-IRD by www.evicr.net centers and ERN-EYE HCPs in Europe indicates that RPE65-IRD might be diagnosed more reliably in 2021 compared to 2019. By June 2021, 8/26 centers reported detailed results including VN treatment. Main reasons for non-treatment were too advanced or mild disease, followed by absence of 2 class 4 or 5 mutations on both alleles or because of a too young age. Patient satisfaction with treatment was estimated to be high by 50% of the centers., (© 2023 The Author(s). Published by S. Karger AG, Basel.)

Objective: To describe the spectrum of Leber congenital amaurosis (LCA) and cone-rod dystrophy (CORD) associated with the GUCY2D gene and to identify potential end points and optimal patient selection for future therapeutic trials., Design: International, multicenter, retrospective cohort study., Subjects: Eighty-two patients with GUCY2D-associated LCA or CORD from 54 families., Methods: Medical records were reviewed for medical history, best-corrected visual acuity (BCVA), ophthalmoscopy, visual fields, full-field electroretinography, and retinal imaging (fundus photography, spectral-domain OCT [SD-OCT], fundus autofluorescence)., Main Outcomes Measures: Age of onset, evolution of BCVA, genotype-phenotype correlations, anatomic characteristics on funduscopy, and multimodal imaging., Results: Fourteen patients with autosomal recessive LCA and 68 with autosomal dominant CORD were included. The median follow-up times were 5.2 years (interquartile range [IQR] 2.6-8.8 years) for LCA and 7.2 years (IQR 2.2-14.2 years) for CORD. Generally, LCA presented in the first year of life. The BCVA in patients with LCA ranged from no light perception to 1.00 logarithm of the minimum angle of resolution (logMAR) and remained relatively stable during follow-up. Imaging for LCA was limited but showed little to no structural degeneration. In patients with CORD, progressive vision loss started around the second decade of life. The BCVA declined annually by 0.022 logMAR (P < 0.001) with no difference between patients with the c.2513G>A and the c.2512C>T GUCY2D variants (P = 0.798). At the age of 40 years, the probability of being blind or severely visually impaired was 32%. The integrity of the ellipsoid zone (EZ) and that of the external limiting membrane (ELM) on SD-OCT correlated significantly with BCVA (Spearman ρ = 0.744, P = 0.001, and ρ = 0.712, P < 0.001, respectively) in those with CORD., Conclusions: Leber congenital amaurosis associated with GUCY2D caused severe congenital visual impairment with relatively intact macular anatomy on funduscopy and available imaging, suggesting long preservation of photoreceptors. Despite large variability, GUCY2D-associated CORD generally presented during adolescence, with a progressive loss of vision, and culminated in severe visual impairment during mid-to-late adulthood. The integrity of the ELM and EZ may be suitable structural end points for therapeutic studies of GUCY2D-associated CORD., (Copyright © 2022 American Academy of Ophthalmology. All rights reserved.)

Purpose: To assess the incidence of Stargardt disease (STGD1) and to evaluate demographics of incident cases., Methods: For this retrospective cohort study, demographic, clinical and genetic data of patients with a clinical diagnosis of STGD1 were registered between September 2010 and January 2020 in a nationwide disease registry. Annual incidence (2014-2018) and point prevalence (2018) were assessed on the basis of this registry., Results: A total of 800 patients were registered, 56% were female and 83% were of European ancestry. The incidence was 1.67-1.95:1,000,000 per year and the point prevalence in 2018 was approximately 1:22,000-1:19,000 (with and without 10% of potentially unregistered cases). Age at onset was associated with sex (p = 0.027, Fisher's exact); 1.9x more women than men were observed (140 versus 74) amongst patients with an age at onset between 10 and 19 years, while the sex ratio in other age-at-onset categories approximated one. Late-onset STGD1 (≥45 years) constituted 33% of the diagnoses in 2014-2018 compared to 19% in 2004-2008. Diagnostic delay (≥2 years between the first documentation of macular abnormalities and diagnosis) was associated with older age of onset (p = 0.001, Mann-Whitney). Misdiagnosis for age-related macular degeneration (22%) and incidental STGD1 findings (14%) was common in patients with late-onset STGD1., Conclusion: The observed prevalence of STGD1 in real-world data was lower than expected on the basis of population ABCA4 allele frequencies. Late-onset STGD1 was more frequently diagnosed in recent years, likely due to higher awareness of its phenotype. In this pretherapeutic era, mis- and underdiagnosis of especially late-onset STGD1 and the role of sex in STGD1 should receive special attention., (© 2021 The Authors. Acta Ophthalmologica published by John Wiley & Sons Ltd on behalf of Acta Ophthalmologica Scandinavica Foundation.)

Background: Inherited retinal diseases (IRDs) can be caused by variants in >270 genes. The Bardet-Biedl syndrome 1 ( BBS1 ) gene is one of these genes and may be associated with syndromic and non-syndromic autosomal recessive retinitis pigmentosa (RP). Here, we identified a branchpoint variant in BBS1 and assessed its pathogenicity by in vitro functional analysis., Methods: Whole genome sequencing was performed for three unrelated monoallelic BBS1 cases with non-syndromic RP. A fourth case received MGCM 105 gene panel analysis. Functional analysis using a midigene splice assay was performed for the putative pathogenic branchpoint variant in BBS1 . After confirmation of its pathogenicity, patients were clinically re-evaluated, including assessment of non-ocular features of Bardet-Biedl syndrome., Results: Clinical assessments of probands showed that all individuals displayed non-syndromic RP with macular involvement. Through detailed variant analysis and prioritisation, two pathogenic variants in BBS1 , the most common missense variant, c.1169T>G (p.(Met390Arg)), and a branchpoint variant, c.592-21A>T, were identified. Segregation analysis confirmed that in all families, probands were compound heterozygous for c.1169T>G and c.592-21A>T. Functional analysis of the branchpoint variant revealed a complex splicing defect including exon 8 and exon 7/8 skipping, and partial in-frame deletion of exon 8., Conclusion: A putative severe branchpoint variant in BBS1 , together with a mild missense variant, underlies non-syndromic RP in four unrelated individuals. To our knowledge, this is the first report of a pathogenic branchpoint variant in IRDs that results in a complex splice defect. In addition, this research highlights the importance of the analysis of non-coding regions in order to provide a conclusive molecular diagnosis., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. No commercial re-use. See rights and permissions. Published by BMJ.)

Purpose: To investigate the natural disease course of retinal dystrophies associated with crumbs cell polarity complex component 1 (CRB1) and identify clinical end points for future clinical trials., Design: Single-center, prospective case series., Methods: An investigator-initiated nationwide collaborative study that included 22 patients with CRB1-associated retinal dystrophies. Patients underwent ophthalmic assessment at baseline and 2 years after baseline. Clinical examination included best-corrected visual acuity (BCVA) using Early Treatment Diabetic Retinopathy Study charts, Goldmann kinetic perimetry (V4e isopter seeing retinal areas), microperimetry, full-field electroretinography, full-field stimulus threshold (FST), fundus photography, spectral-domain optical coherence tomography, and fundus autofluorescence imaging., Results: Based on genetic, clinical, and electrophysiological data, patients were diagnosed with retinitis pigmentosa (19 [86%]), cone-rod dystrophy (2 [9%]), or isolated macular dystrophy (1 [5%]). Analysis of the entire cohort at 2 years showed no significant changes in BCVA (P = .069) or V4e isopter seeing retinal areas (P = .616), although signs of clinical progression were present in individual patients. Macular sensitivity measured on microperimetry revealed a significant reduction at the 2-year follow-up (P < .001). FST responses were measurable in patients with nonrecordable electroretinograms. On average, FST responses remained stable during follow-up., Conclusion: In CRB1-associated retinal dystrophies, visual acuity and visual field measures remain relatively stable over the course of 2 years. Microperimetry showed a significant decrease in retinal sensitivity during follow-up and may be a more sensitive progression marker. Retinal sensitivity on microperimetry may serve as a functional clinical end point in future human treatment trials for CRB1-associated retinal dystrophies., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: To describe the natural course, phenotype, and genotype of patients with X-linked retinoschisis (XLRS)., Design: Retrospective cohort study., Participants: Three hundred forty patients with XLRS from 178 presumably unrelated families., Methods: This multicenter, retrospective cohort study reviewed medical records of patients with XLRS for medical history, symptoms, visual acuity (VA), ophthalmoscopy, full-field electroretinography, and retinal imaging (fundus photography, spectral-domain [SD] OCT, fundus autofluorescence)., Main Outcome Measures: Age at onset, age at diagnosis, severity of visual impairment, annual visual decline, and electroretinography and imaging findings., Results: Three hundred forty patients were included with a mean follow-up time of 13.2 years (range, 0.1-50.1 years). The median ages to reach mild visual impairment and low vision were 12 and 25 years, respectively. Severe visual impairment and blindness were observed predominantly in patients older than 40 years, with a predicted prevalence of 35% and 25%, respectively, at 60 years of age. The VA increased slightly during the first 2 decades of life and subsequently transitioned into an average annual decline of 0.44% (P < 0.001). No significant difference was found in decline of VA between variants that were predicted to be severe and mild (P = 0.239). The integrity of the ellipsoid zone (EZ) as well as the photoreceptor outer segment (PROS) length in the fovea on SD OCT correlated significantly with VA (Spearman's ρ = -0.759 [P < 0.001] and -0.592 [P = 0.012], respectively). Fifty-three different RS1 variants were found. The most common variants were the founder variant c.214G→A (p.(Glu72Lys)) (101 patients [38.7%]) and a deletion of exon 3 (38 patients [14.6%])., Conclusions: Large variabilities in phenotype and natural course of XLRS were seen in this study. In most patients, XLRS showed a slow deterioration starting in the second decade of life, suggesting an optimal window of opportunity for treatment within the first 3 decades of life. The integrity of EZ as well as the PROS length on SD OCT may be important in choosing optimal candidates for treatment and as potential structural end points in future therapeutic studies. No clear genotype-phenotype correlation was found., (Copyright © 2021 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Mutations in PRPH2, encoding peripherin-2, are associated with the development of a wide variety of inherited retinal diseases (IRDs). To determine the causality of the many PRPH2 variants that have been discovered over the last decades, we surveyed all published PRPH2 variants up to July 2020, describing 720 index patients that in total carried 245 unique variants. In addition, we identified seven novel PRPH2 variants in eight additional index patients. The pathogenicity of all variants was determined using the ACMG guidelines. With this, 107 variants were classified as pathogenic, 92 as likely pathogenic, one as benign, and two as likely benign. The remaining 50 variants were classified as variants of uncertain significance. Interestingly, of the total 252 PRPH2 variants, more than half (n = 137) were missense variants. All variants were uploaded into the Leiden Open source Variation and ClinVar databases. Our study underscores the need for experimental assays for variants of unknown significance to improve pathogenicity classification, which would allow us to better understand genotype-phenotype correlations, and in the long-term, hopefully also support the development of therapeutic strategies for patients with PRPH2-associated IRD., (© 2021 The Authors. Human Mutation published Wiley Periodicals LLC.)

Inherited retinal diseases (IRDs) are a major cause of visual impairment. These clinically heterogeneous disorders are caused by pathogenic variants in more than 270 genes. As 30-40% of cases remain genetically unexplained following conventional genetic testing, we aimed to obtain a genetic diagnosis in an IRD cohort in which the genetic cause was not found using whole-exome sequencing or targeted capture sequencing. We performed whole-genome sequencing (WGS) to identify causative variants in 100 unresolved cases. After initial prioritization, we performed an in-depth interrogation of all noncoding and structural variants in genes when one candidate variant was detected. In addition, functional analysis of putative splice-altering variants was performed using in vitro splice assays. We identified the genetic cause of the disease in 24 patients. Causative coding variants were observed in genes such as ATXN7, CEP78, EYS, FAM161A, and HGSNAT. Gene disrupting structural variants were also detected in ATXN7, PRPF31, and RPGRIP1. In 14 monoallelic cases, we prioritized candidate noncanonical splice sites or deep-intronic variants that were predicted to disrupt the splicing process based on in silico analyses. Of these, seven cases were resolved as they carried pathogenic splice defects. WGS is a powerful tool to identify causative variants residing outside coding regions or heterozygous structural variants. This approach was most efficient in cases with a distinct clinical diagnosis. In addition, in vitro splice assays provide important evidence of the pathogenicity of rare variants., (© 2021. The Author(s).)

Background: Leber hereditary optic neuropathy (LHON) is an orphan disease which leads to painless subacute loss of central vision in both eyes. It develops mainly in young adults and is more common in males. It most often leads to lifelong blindness. Idebenone has shown to have a favourable effect in promoting vision recovery in LHON-patients with recent visual impairement., Case Description: Two male LHON patients, aged 27 and 54 years of age were misdiagnosed during one year with optic neuritis and conversion disorder. The delay caused unnecessary emotional suffering and took away the opportunity of idebenone treatment. This can be prevented by greater awareness of disease characteristcs and OCT-scanning., Conclusion: Therapy for LHON requires a timely diagnosis.

A substantial proportion of subjects with autosomal recessive retinitis pigmentosa (arRP) or Usher syndrome type II (USH2) lacks a genetic diagnosis due to incomplete USH2A screening in the early days of genetic testing. These cases lack eligibility for optimal genetic counseling and future therapy. USH2A defects are the most frequent cause of USH2 and are also causative in individuals with arRP. Therefore, USH2A is an important target for genetic screening. The aim of this study was to assess unscreened or incompletely screened and unexplained USH2 and arRP cases for (likely) pathogenic USH2A variants. Molecular inversion probe (MIP)-based sequencing was performed for the USH2A exons and their flanking regions, as well as published deep-intronic variants. This was done to identify single nucleotide variants (SNVs) and copy number variants (CNVs) in 29 unscreened or partially pre-screened USH2 and 11 partially pre-screened arRP subjects. In 29 out of these 40 cases, two (likely) pathogenic variants were successfully identified. Four of the identified SNVs and one CNV were novel. One previously identified synonymous variant was demonstrated to affect pre-mRNA splicing. In conclusion, genetic diagnoses were obtained for a majority of cases, which confirms that MIP-based sequencing is an effective screening tool for USH2A . Seven unexplained cases were selected for future analysis with whole genome sequencing.

Purpose: The purpose of the study was to demonstrate whether photodynamic therapy in patients with acute central serous chorioretinopathy, with the leakage point within one optic disk diameter from the fovea, can be safely deferred., Methods: A single-center, randomized, controlled trial was conducted. Patients were randomized to photodynamic therapy within a week after presentation (Group I, 26 patients) or observation during 3 months (Group II, 26 patients). If leakage or subretinal fluid was observed during any control visit, photodynamic therapy was performed (again) within a week., Primary Outcome: Primary outcome was change of visual acuity (Early Treatment Diabetic Retinopathy Study) after 12 months. Secondary outcomes were visual acuity, central foveal thickness, metamorphopsia, and color discrimination., Results: Photodynamic therapy procedures: group I, 26 at baseline, 2 retreatments at 3 months; group II, 10 at 3 months, 1 at 6 months (2 subjects refusing treatment), 2 retreatments at 6 months. At 12 months, mean visual acuity of all patients had improved by 6.5 letters (P < 0.001), mean central foveal thickness was 172 µm less (P < 0.001). After photodynamic therapy, visual acuity recovered faster and metamorphopsia significantly improved (3 months, P < 0.001). Differences between groups at 12 months were not significant., Conclusion: The (intended) number of photodynamic therapy (re)treatments in group II (n = 15) was 46% less than in group I (n = 28). Visual acuity and central foveal thickness at 12 months were similar. Therefore, the preferred management of acute central serous chorioretinopathy at presentation appears to be observation for 3 months.

Purpose: To investigate the retinal structure and function in patients with CRB1-associated retinal dystrophies (RD) and to explore potential clinical endpoints., Methods: In this prospective cross-sectional study, 22 patients with genetically confirmed CRB1-RD (aged 6-74 years), and who had a decimal best-corrected visual acuity (BCVA) ≥ 0.05 at the last visit, were studied clinically with ETDRS BCVA, corneal topography, spectral-domain optical coherence tomography (SD-OCT), fundus autofluorescence, Goldmann visual field (VF), microperimetry, full-field electroretinography (ERG) and full-field stimulus testing (FST). Ten patients were from a genetic isolate (GI)., Results: Patients had retinitis pigmentosa (n = 19; GI and non-GI), cone-rod dystrophy (n = 2; GI) or macular dystrophy (n = 1; non-GI). Median age at first symptom onset was 3 years (range 0.8-49). Median decimal BCVA in the better and worse-seeing eye was 0.18 (range 0.05-0.83) and 0.08 (range light perception-0.72), respectively. Spectral-domain optical coherence tomography (SD-OCT) showed cystoid maculopathy in 8 subjects; inner retinal thickening (n = 20), a well-preserved (para)foveal outer retina (n = 7) or severe (para)foveal outer retinal atrophy (n = 14). All retinal layers were discernible in 13/21 patients (62%), with mild to moderate laminar disorganization in the others. Nanophthalmos was observed in 8 patients (36%). Full-field stimulus testing (FST) provided a subjective outcome measure for retinal sensitivity in eyes with (nearly) extinguished ERG amplitudes., Conclusions: Despite the generally severe course of CRB1-RDs, symptom onset and central visual function are variable, even at advanced ages. Phenotypes may vary within the same family. Imaging and functional studies in a prospective longitudinal setting should clarify which endpoints may be most appropriate in a clinical trial., (© 2021 The Authors. Acta Ophthalmologica published by John Wiley & Sons Ltd on behalf of Acta Ophthalmologica Scandinavica Foundation.)

Purpose: To investigate the natural history of RHO-associated retinitis pigmentosa (RP)., Methods: A multicenter, medical chart review of 100 patients with autosomal dominant RHO-associated RP., Results: Based on visual fields, time-to-event analysis revealed median ages of 52 and 79 years to reach low vision (central visual field <20°) and blindness (central visual field <10°), respectively. For the best-corrected visual acuity (BCVA), the median age to reach mild impairment (20/67 ≤ BCVA < 20/40) was 72 years, whereas this could not be computed for lower acuities. Disease progression was significantly faster in patients with a generalized RP phenotype (n = 75; 75%) than that in patients with a sector RP phenotype (n = 25; 25%), in terms of decline rates of the BCVA (P < 0.001) and V4e retinal seeing areas (P < 0.005). The foveal thickness of the photoreceptor-retinal pigment epithelium (PR + RPE) complex correlated significantly with BCVA (Spearman's ρ = 0.733; P < 0.001)., Conclusion: Based on central visual fields, the optimal window of intervention for RHO-associated RP is before the 5th decade of life. Significant differences in disease progression are present between generalized and sector RP phenotypes. Our findings suggest that the PR + RPE complex is a potential surrogate endpoint for the BCVA in future studies., (Copyright © 2020 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the Opthalmic Communications Society, Inc.)

Purpose: An increasing number of gene therapies are developed for Inherited Retinal Degenerations (IRD). To date, 1 treatment has been approved for clinical use (FDA USA 2017, EMA Europe 2018, MoHAP UAE 2019, SFDA Saudi Arabia 2019, Swiss Medic Switzerland 2020, TGA Australia 2020, and BFR Brazil 2020). While such therapies do not provide complete cure, they may halt degeneration or partially restore function. Identification of well-characterized patients is an emerging need. We conducted the first multinational survey to understand the management of IRDs in Europe., Methods: An electronic survey questionnaire containing 112 questions was developed and sent to the 101 EVICR.net clinical centers (14 European countries and Israel)., Results: The overall response rate was 49%. Only 14% of responding centers do not see IRD patients; 52% that manage IRD patients follow ≥200 patients, 16% > 1,000. Databases exist in 86% of the centers; of these, 75% are local files, 28% local Web-based database, and 19% national Web-based. IRD patients are referred to EVICR.net centers mainly by general ophthalmologists, patient self-referrals, and medical retina specialists. Most IRD patients are first seen in adulthood. Most prominent signs and symptoms depend on the age of onset, for example, nystagmus in infancy, or night blindness, and reduced visual acuity at older age. The time from inquiring for first appointment and clinical diagnosis varies among countries: in 29% of centers, the mean time is <4 weeks, although can be up to 35 months in others. The time to genetic diagnosis is ≥4 weeks, the maximum 10 years, likely depending on access to genetic testing, and the improvement of the tests available. Comprehensive eye examination always includes autofluorescence imaging and perimetry (86% static, 76% kinetic, and 21% microperimetry), and frequently optical coherence tomography (OCT) (95%), electroretinography (93%), and fundus photography (93%). Identified genotypes were reported in 40-80% patients by 69% of centers, and in 80-100% by 5%. Genetic testing is provided by public health insurance in 77% of centers, private health insurance in 38%, center budget in 13%, research funds in 18%; and 15% of centers do not have access to genetic testing., Conclusion: At the start of this era of ocular gene therapy for IRD patients, this first international survey on management of IRDs in Europe highlights significant heterogeneity between centers and across countries and provides important baseline data for researchers, clinicians, pharmaceutical companies, and investors., (© 2021 The Author(s) Published by S. Karger AG, Basel.)

Introduction: Treatment of exudative age-related macular degeneration (ARMD) has shifted to pro re nata and treat-extend-stop strategies. However, a rational discontinuation strategy is lacking. To develop such a strategy, it is important to determine choroidal neovascularization (CNV) recurrence rates after anti-VEGF treatment is discontinued. Here we report prospective data on persistent and recurrent CNV activity after discontinuation of bevacizumab treatment., Methods: This prospective, single-center clinical trial enrolled 191 patients with exudative ARMD. Patients were randomly assigned to receiving intravitreal bevacizumab injections every 4, 6, or 8 weeks for 1 year. CNV activity was determined in the 157 patients who completed the 1-year treatment regimen. Patients with inactive CNV were then followed for signs of CNV reactivation., Results: After 1 year of treatment, 66 (42%) of the 157 patients still had signs of persistent active CNV. Of the remaining 91 (58%) patients, 61 (67%) needed retreatment for active CNV within the first year after discontinuation of treatment (mean 4.28 ± 0.29 months). CNV was reactivated in 50 (80%) of the 61 patients within 6 months after their final treatment for CNV., Conclusion: Based on quiescent disease, anti-VEGF therapy was discontinued in 58% of the patients after they received bevacizu-mab injections every 4, 6, or 8 weeks for 1 year; 67% showed reactivated CNV within a year after discontinuation. The high reactivation rate of CNV shown in this study should help clinicians to develop rational discontinuation protocols., Trial Registration: This study is registered as NTR1174 at http://www.trialregister.nl., (© 2021 S. Karger AG, Basel.)

Purpose: The first ocular gene augmentation therapy, voretigene neparvovec (VN) (Luxturna®), has been approved for clinical use in an increasing number of countries (FDA USA 2017, EMA Europe 2018, MoHAP United Arab Emirates 2019, SFDA Saudi Arabia 2019, Swiss Medic Switzerland 2020, TGA Australia 2020, BFR Brazil 2020). Among the EVICR.net clinical centers, we conducted the first multinational survey to understand distribution, diagnostic work-up, and management of inherited retinal degeneration (IRD) cases in Europe with a special focus on RPE65 mutation-associated IRDs., Methods: An electronic survey questionnaire including 35 questions specifically addressing RPE65 mutation-associated IRDs was developed and sent to the 101 EVICR.net clinical centers., Results: The overall response rate was 49%. Forty-two centers see IRD patients, and 22/42 follow patients with confirmed biallelic RPE65 mutations. Fifteen of the 22 centers (68%) and 3/22 (14%) follow 1-5 and 6-10 patients with homozygous RPE65 mutations, respectively. Additionally, 15/22 (68%) and 3/22 (14%) follow 1-5 and >20 patients with compound heterozygous RPE65 mutations, respectively. Fifty-nine percent of mutations were ACMG Class 4 and 5 (at least 1 allele), 82.8% reported previously and 17.2% novel. Referral diagnoses (the mean per center) were Leber congenital amaurosis (38.2%), early-onset severe retinal degeneration (16.8%), rod-cone-dystrophy/retinitis pigmentosa (RP) (28.1%), and unclassified visual impairment (17.0%). Twenty-five percent of the centers changed the referral diagnosis in >47.5% of cases; 32% follow a specific referral process for RPE65 mutation-associated IRD patients. Annual follow-up visits are done in 55% of the centers and biannual visits in 23%. In 32%, other centers also follow the patients. Kinetic perimetry is done in 82%, static perimetry in 45%, and microperimetry in 18% of the centers. Full-field light stimulus threshold testing with blue and red stimuli to quantify the rod and cone function is used in 6/22 centers (27%). A mobility course is available in one center (5%)., Conclusion: This first multinational survey on management of patients with RPE65 mutation-associated IRDs in Europe shows that about half of the responding EVICR.net centers have such patients under care. There is heterogeneity in diagnoses and management practices. At the start of clinical practice experience with VN, these data provide a useful baseline and highlight the need for consensus/guidelines to inform standard of care in this new era of gene therapy., (© 2021 The Author(s) Published by S. Karger AG, Basel.)

The cause of autosomal-dominant retinitis pigmentosa (adRP), which leads to loss of vision and blindness, was investigated in families lacking a molecular diagnosis. A refined locus for adRP on Chr17q22 (RP17) was delineated through genotyping and genome sequencing, leading to the identification of structural variants (SVs) that segregate with disease. Eight different complex SVs were characterized in 22 adRP-affected families with >300 affected individuals. All RP17 SVs had breakpoints within a genomic region spanning YPEL2 to LINC01476. To investigate the mechanism of disease, we reprogrammed fibroblasts from affected individuals and controls into induced pluripotent stem cells (iPSCs) and differentiated them into photoreceptor precursor cells (PPCs) or retinal organoids (ROs). Hi-C was performed on ROs, and differential expression of regional genes and a retinal enhancer RNA at this locus was assessed by qPCR. The epigenetic landscape of the region, and Hi-C RO data, showed that YPEL2 sits within its own topologically associating domain (TAD), rich in enhancers with binding sites for retinal transcription factors. The Hi-C map of RP17 ROs revealed creation of a neo-TAD with ectopic contacts between GDPD1 and retinal enhancers, and modeling of all RP17 SVs was consistent with neo-TADs leading to ectopic retinal-specific enhancer-GDPD1 accessibility. qPCR confirmed increased expression of GDPD1 and increased expression of the retinal enhancer that enters the neo-TAD. Altered TAD structure resulting in increased retinal expression of GDPD1 is the likely convergent mechanism of disease, consistent with a dominant gain of function. Our study highlights the importance of SVs as a genomic mechanism in unsolved Mendelian diseases., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: To study the disease course of RPE65-associated inherited retinal degenerations (IRDs) as a function of the genotype, define a critical age for blindness, and identify potential modifiers., Methods: Forty-five patients with IRD from 33 families with biallelic RPE65 mutations, 28 stemming from a genetic isolate. We collected retrospective data from medical charts. Coexisting variants in 108 IRD-associated genes were identified with Molecular Inversion Probe analysis., Results: Most patients were diagnosed within the first years of life. Daytime visual function ranged from near-normal to blindness in the first four decades and met WHO criteria for blindness for visual acuity and visual field in the fifth decade. p.(Thr368His) was the most common variant (54%). Intrafamilial variability and interfamilial variability in disease severity and progression were observed. Molecular Inversion Probe analysis confirmed all RPE65 variants and identified one additional variant in LRAT and one in EYS in two separate patients., Conclusion: All patients with RPE65-associated IRDs developed symptoms within the first year of life. Visual function in childhood and adolescence varied but deteriorated inevitably toward blindness after age 40. In this study, genotype was not predictive of clinical course. The variance in severity of disease could not be explained by double hits in other IRD genes.

Purpose: Missing heritability in human diseases represents a major challenge, and this is particularly true for ABCA4-associated Stargardt disease (STGD1). We aimed to elucidate the genomic and transcriptomic variation in 1054 unsolved STGD and STGD-like probands., Methods: Sequencing of the complete 128-kb ABCA4 gene was performed using single-molecule molecular inversion probes (smMIPs), based on a semiautomated and cost-effective method. Structural variants (SVs) were identified using relative read coverage analyses and putative splice defects were studied using in vitro assays., Results: In 448 biallelic probands 14 known and 13 novel deep-intronic variants were found, resulting in pseudoexon (PE) insertions or exon elongations in 105 alleles. Intriguingly, intron 13 variants c.1938-621G>A and c.1938-514G>A resulted in dual PE insertions consisting of the same upstream, but different downstream PEs. The intron 44 variant c.6148-84A>T resulted in two PE insertions and flanking exon deletions. Eleven distinct large deletions were found, two of which contained small inverted segments. Uniparental isodisomy of chromosome 1 was identified in one proband., Conclusion: Deep sequencing of ABCA4 and midigene-based splice assays allowed the identification of SVs and causal deep-intronic variants in 25% of biallelic STGD1 cases, which represents a model study that can be applied to other inherited diseases.

Background: Transcorneal electrical stimulation (TES) has been suggested as a possible treatment for retinitis pigmentosa (RP)., Objective: To expand the safety assessment of repeated applications of an electrical current from a DTL-like electrode in patients with RP., Methods: This single-arm open label interventional safety trial included a total of 105 RP patients from 11 European centers, who received weekly TES for 6 months on 1 eye followed by observation for another 6 months without stimulation. The primary outcome measure was safety, indicated by the frequency and severity of adverse events. Secondary measures included intraocular pressure and central retinal thickness. Visual field and visual acuity were examined using the methods available at each site., Results: Dry eye sensation was the most common adverse event recorded (37.5%). Serious adverse events secondary to TES were not observed. Most adverse events were mild and all resolved without sequelae. The secondary outcome measures revealed no significant or clinically relevant changes., Conclusion: The present results confirm the excellent safety profile of TES. Transient dry eye symptoms were the most common adverse event., (© 2020 S. Karger AG, Basel.)

Background: The phenotypic spectrum of many rare disorders is much wider than previously considered. Mucopolysaccharidosis type III (Sanfilippo syndrome, MPS III), is a lysosomal storage disorder traditionally considered to be characterized by childhood onset, progressive neurocognitive deterioration with a rapidly or slowly progressing phenotype. The presented MPS III case series demonstrates adult onset phenotypes with mild cognitive impairment or non-neuronopathic phenotypes., Methods: In this case series all adult MPS III patients with a mild- or non-neuronopathic phenotype, who attend the outpatient clinic of 3 expert centers for lysosomal storage disorders were included. A mild- or non-neuronopathic phenotype was defined as having completed regular secondary education and attaining a level of independency during adulthood, involving either independent living or a paid job., Results: Twelve patients from six families, with a median age at diagnosis of 43 years (range 3-68) were included (11 MPS IIIA, 1 MPS IIIB). In the four index patients symptoms which led to diagnostic studies (whole exome sequencing and metabolomics) resulting in the diagnosis of MPS III; two patients presented with retinal dystrophy, one with hypertrophic cardiomyopathy and one with neurocognitive decline. The other eight patients were diagnosed by family screening. At a median age of 47 years (range 19-74) 9 out of the 12 patients had normal cognitive functions. Nine patients had retinal dystrophy and 8 patients hypertrophic cardiomyopathy., Conclusion: We show the very mild end of the phenotypic spectrum of MPS III, ranging from late-onset stable neurocognitive impairment to a fully non-neuronopathic phenotype. Awareness of this phenotype could lead to timely diagnosis and genetic counseling.

Purpose: To investigate the role of two deep-intronic ABCA4 variants, that showed a mild splice defect in vitro and can occur on the same allele as the low penetrant c.5603A>T, in Stargardt disease (STGD1)., Methods: Ophthalmic data were assessed of 18 STGD1 patients who harbored c.769-784C>T or c.4253+43G>A in combination with a severe ABCA4 variant. Subjects carrying c.[769-784C>T; 5603A>T] were clinically compared with a STGD1 cohort previously published carrying c.5603A>T noncomplex. We calculated the penetrances of the intronic variants using ABCA4 allele frequency data of the general population and investigated the effect of c.769-784C>T on splicing in photoreceptor progenitor cells (PPCs)., Results: Mostly, late-onset, foveal-sparing STGD1 was observed among subjects harboring c.769-784C>T or c.4253+43G>A (median age of onset, 54.5 and 52.0 years, respectively). However, ages of onset, phenotypes in fundo, and visual acuity courses varied widely. No significant clinical differences were observed between the c.[769-784C>T; 5603A>T] cohort and the c.4253+43G>A or the c.5603A>T cohort. The penetrances of c.769-784C>T (20.5%-39.6%) and c.4253+43G>A (35.8%-43.1%) were reduced, when not considering the effect of yet unidentified or known factors in cis, such as c.5603A>T (identified in 7/7 probands with c.769-784C>T; 1/8 probands with c.4253+43G>A). Variant c.769-784C>T resulted in a pseudo-exon insertion in 15% of the total mRNA (i.e., ∼30% of the c.769-784C>T allele alone)., Conclusions: Two mild intronic ABCA4 variants could further explain missing heritability in late-onset STGD1, distinguishing it from AMD. The observed clinical variability and calculated reduced penetrance urge research into modifiers within and outside of the ABCA4 gene.

Purpose: Using exome sequencing, the underlying variants in many persons with autosomal recessive diseases remain undetected. We explored autosomal recessive Stargardt disease (STGD1) as a model to identify the missing heritability., Methods: Sequencing of ABCA4 was performed in 8 STGD1 cases with one variant and p.Asn1868Ile in trans, 25 cases with one variant, and 3 cases with no ABCA4 variant. The effect of intronic variants was analyzed using in vitro splice assays in HEK293T cells and patient-derived fibroblasts. Antisense oligonucleotides were used to correct splice defects., Results: In 24 of the probands (67%), one known and five novel deep-intronic variants were found. The five novel variants resulted in messenger RNA pseudoexon inclusions, due to strengthening of cryptic splice sites or by disrupting a splicing silencer motif. Variant c.769-784C>T showed partial insertion of a pseudoexon and was found in cis with c.5603A>T (p.Asn1868Ile), so its causal role could not be fully established. Variant c.4253+43G>A resulted in partial skipping of exon 28. Remarkably, antisense oligonucleotides targeting the aberrant splice processes resulted in (partial) correction of all splicing defects., Conclusion: Our data demonstrate the importance of assessing noncoding variants in genetic diseases, and show the great potential of splice modulation therapy for deep-intronic variants.

Purpose: To describe the phenotype and clinical course of patients with RPGR-associated retinal dystrophies, and to identify genotype-phenotype correlations., Methods: A multicenter medical records review of 74 male patients with RPGR-associated retinal dystrophies., Results: Patients had retinitis pigmentosa (RP; n = 52; 70%), cone dystrophy (COD; n = 5; 7%), or cone-rod dystrophy (CORD; n = 17; 23%). The median follow-up time was 11.6 years (range 0-57.1). The median age at symptom onset was 5.0 years (range 0-14 years) for patients with RP and 23.0 years (range 0-60 years) for patients with COD/CORD. The probability of being blind (best-corrected visual acuity <0.05) at the age of 40 was 20% and 55% in patients with RP and COD/CORD, respectively. RPGR-ORF15 mutations were associated with high myopia (P = 0.01), which led to a faster best-corrected visual acuity decline in patients with RP (P < 0.001) and COD/CORD (P = 0.03). Patients with RP with RPGR-ORF15 mutations had a faster visual field decline (P = 0.01) and thinner central retina (P = 0.03) than patients with mutations in exon 1 to 14., Conclusion: Based on best-corrected visual acuity survival probabilities, the intervention window for gene therapy for RPGR-associated retinal dystrophies is relatively broad in patients with RP. RPGR-ORF15 mutations were associated with COD/CORD and with a more severe phenotype in RP. High myopia is a risk factor for faster best-corrected visual acuity decline.

Purpose: To assess the phenotypic variability and natural course of inherited retinal diseases (IRDs) caused by EYS mutations., Methods: Multiethnic cohort study (N = 30) with biallelic EYS variants from a clinical IRD database (retinitis pigmentosa [RP], N = 27; cone-rod dystrophy [CRD], N = 1; and macular dystrophy, N = 2). In vitro minigene splice assay was performed to determine the effect on EYS pre-mRNA splicing of the c.1299+5&#95;1299+8del variant in macular dystrophy patients., Results: We found 27 different EYS variants in RP patients and 7 were novel. The rate of visual field loss of the V4e isopter area was -0.84 ± 0.44 ln(deg2) per year, and the rate of visual acuity loss was 0.75 Early Treatment Diabetic Retinopathy Study letters per year. Ellipsoid zone width was correlated with area of the hyperautofluorescent ring, with rs = 0.78 and P < 0.001. Rate of decline in ellipsoid zone width was -57 ± 17 μm per year (P < 0.01) (n = 14) or -3.69% ± 0.51% from baseline per year (P < 0.001). An isolated CRD patient carried a homozygous EYS variant (c.9405T>A), previously identified in RP patients. Two siblings with macular dystrophy carried compound heterozygous EYS variants: c.1299+5&#95;1299+8del and c.6050G>T. The former was novel and shown to result in skipping of exon 8, and the latter was a known RP variant., Conclusions: We report on EYS-associated macular dystrophy, extending the spectrum of EYS-associated IRDs. We observed heterogeneity between RP patients in age of onset and disease progression. Identical EYS variants were found in cases with RP, CRD, and macular dystrophy. Screening for EYS variants in CRD and macular dystrophy patients might increase the diagnostic yield in previously unsolved cases.

Purpose: Intravitreal anti-vascular endothelial growth factor (VEGF) injections are an effective treatment for neovascular age-related macular degeneration (nARMD). Bevacizumab appears to be a cost-effective off-label anti-VEGF alternative to ranibizumab, but an optimal injection schedule has not yet been determined. In this study, we investigate whether on-demand bevacizumab treatment every 8 weeks is non-inferior to on-demand bevacizumab every 4 weeks in treating nARMD., Methods: A total of 120 nARMD patients were randomly assigned to an on-demand regimen of intravitreal bevacizumab (IVB) every 4 (n = 60) or 8 weeks (n = 60). The primary outcome was visual acuity (VA) change after 1 year of treatment., Results: Visual acuity (VA) improved between baseline and 1 year in both treatment groups. The mean change in the VA score at 1 year was not significantly different between bevacizumab administration on-demand every 4 weeks [5.6 ± 10.2 Early Treatment Diabetic Retinopathy Study (ETDRS) letter] or 8 weeks (4.6 ± 12.0 ETDRS letters). A reduction in the central retinal thickness was observed in both groups. At 1 year, the mean decrease in central foveal thickness ranged from 61 ± 90 μm in the 4-week group to 91 ± 83 μm in the 8-week group (p = 0.07). The mean number of IVB treatments during the study period was 8.7 ± 2.3 in the 4-week group and 5.9 ± 1.0 in the 8-week group., Conclusion: At 1 year, bevacizumab administration on-demand every 8 weeks was non-inferior to administration every 4 weeks. The results strongly suggest that bevacizumab acts longer than 4 weeks in ARMD, reducing the burden of injections for patients., (© 2018 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.)

Background: Retinitis pigmentosa (RP) shows substantial genetic heterogeneity. It has been estimated that in approximately 60%-80% of RP cases, the genetic diagnosis can be found using whole exome sequencing (WES). In this study, the purpose was to identify causative variants in individuals with genetically unexplained retinal disease, which included one consanguineous family with two affected siblings and one case with RP., Methods: To identify the genetic defect, WES was performed in both probands, and clinical analysis was performed. To obtain insight into the function of KIAA1549 in photoreceptors, mRNA expression, knockdown and protein localisation studies were performed., Results: Through analysis of WES data, based on population allele frequencies, and in silico prediction tools, we identified a homozygous missense variant and a homozygous frameshift variant in KIAA1549 that segregate in two unrelated families. Kiaa1549 was found to localise at the connecting cilium of the photoreceptor cells and the synapses of the mouse retina. Both variants affect the long transcript of KIAA1549 , which encodes a 1950 amino acid protein and shows prominent brain expression. The shorter transcript encodes a 734 amino acid protein with a high retinal expression and is affected by the identified missense variant. Strikingly, knockdown of the long transcript also leads to decreased expression of the short transcript likely explaining the non-syndromic retinal phenotype caused by the two variants targeting different transcripts., Conclusion: In conclusion, our results underscore the causality of segregating variants in KIAA1549 for autosomal recessive RP. Moreover, our data indicate that KIAA1549 plays a role in photoreceptor function., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2018. No commercial re-use. See rights and permissions. Published by BMJ.)

Purpose: To describe the phenotypic spectrum of retinal disease caused by the c.2991+1655A>G mutation in CEP290 and to compare disease severity between homozygous and compound heterozygous patients., Methods: Medical records were reviewed for best-corrected visual acuity (BCVA), age of onset, fundoscopy descriptions. Foveal outer nuclear layer (ONL) and ellipsoid zone (EZ) presence was assessed using spectral-domain optical coherence tomography (SD-OCT). Differences between compound heterozygous and homozygous patients were analyzed based on visual performance and visual development., Results: A total of 66 patients were included. The majority of patients had either light perception or no light perception. In the remaining group of 14 patients, median BCVA was 20/195 Snellen (0.99 LogMAR; range 0.12-1.90) for the right eye, and 20/148 Snellen (0.87 LogMAR; range 0.22-1.90) for the left. Homozygous patients tended to be more likely to develop light perception compared to more severely affected compound heterozygous patients (P = 0.080) and are more likely to improve from no light perception to light perception (P = 0.022) before the age of 6 years. OCT data were available in 12 patients, 11 of whom had retained foveal ONL and EZ integrity up to 48 years (median 23 years) of age., Conclusions: Homozygous patients seem less severely affected compared to their compound-heterozygous peers. Improvement of visual function may occur in the early years of life, suggesting a time window for therapeutic intervention up to the approximate age of 17 years. This period may be extended by an intact foveal ONL and EZ on OCT.