A combination treatment based on drug repurposing demonstrates mutation-agnostic efficacy in pre-clinical retinopathy models.

Inherited retinopathies are devastating diseases that in most cases lack treatment options. Disease-modifying therapies that mitigate pathophysiology regardless of the underlying genetic lesion are desirable due to the diversity of mutations found in such diseases. We tested a systems pharmacology-based strategy that suppresses intracellular cAMP and Ca2+ activity via G protein-coupled receptor (GPCR) modulation using tamsulosin, metoprolol, and bromocriptine coadministration. The treatment improves cone photoreceptor function and slows degeneration in Pde6ßrd10 and RhoP23H/WT retinitis pigmentosa mice. Cone degeneration is modestly mitigated after a 7-month-long drug infusion in PDE6A-/- dogs. The treatment also improves rod pathway function in an Rpe65-/- mouse model of Leber congenital amaurosis but does not protect from cone degeneration. RNA-sequencing analyses indicate improved metabolic function in drug-treated Rpe65-/- and rd10 mice. Our data show that catecholaminergic GPCR drug combinations that modify second messenger levels via multiple receptor actions provide a potential disease-modifying therapy against retinal degeneration.

Perimacular Atrophy Following Voretigene Neparvovec-Rzyl Treatment in the Setting of Previous Contralateral Eye Treatment With a Different Viral Vector.

Purpose: To report on cases of unilateral perimacular atrophy after treatment with voretigene neparvovec-rzyl, in the setting of previous contralateral eye treatment with a different viral vector. Design: Single-center, retrospective chart review. Methods: In this case series, four patients between the ages of six and 11 years old with RPE65-related retinopathy were treated unilaterally with rAAV2-CB-hRPE65 as part of a gene augmentation clinical trial (NCT00749957). Six to 10 years later the contralateral eyes were treated with the Food and Drug Administration-approved drug, voretigene neparvovec-rzyl. Best-corrected visual acuity (BCVA), fundus photos, ocular coherence tomography, two-color dark-adapted perimetry, full field stimulus threshold testing (FST), and location of subretinal bleb and chorioretinal atrophy were evaluated. Results: Three out of four patients showed unilateral perimacular atrophy after treatment with voretigene, ranging from five to 22 months after treatment. Areas of robust visual field improvement were followed by areas of chorioretinal atrophy. Despite perimacular changes, BCVA, FST, and subjective improvements in vision and nyctalopia were maintained. Perimacular atrophy was not observed in the first eye treated with the previous viral vector. Conclusions: We observed areas of robust visual field improvement followed by perimacular atrophy in voretigene treated eyes, as compared to the initially treated contralateral eyes. Translational Relevance: Caution is advised when using two different viral vectors between eyes in gene therapy. This may become an important issue in the future with increasing gene therapy clinical trials for inherited retinal dystrophies.

Ablation of Fatty Acid Transport Protein-4 Enhances Cone Survival, M-cone Vision, and Synthesis of Cone-Tropic 9-cis-Retinal in rd12 Mouse Model of Leber Congenital Amaurosis.

The canonical visual cycle employing RPE65 as the retinoid isomerase regenerates 11-cis-retinal to support both rod- and cone-mediated vision. Mutations of RPE65 are associated with Leber congenital amaurosis that results in rod and cone photoreceptor degeneration and vision loss of affected patients at an early age. Dark-reared Rpe65-/- mouse has been known to form isorhodopsin that employs 9-cis-retinal as the photosensitive chromophore. The mechanism regulating 9-cis-retinal synthesis and the role of the endogenous 9-cis-retinal in cone survival and function remain largely unknown. In this study, we found that ablation of fatty acid transport protein-4 (FATP4), a negative regulator of 11-cis-retinol synthesis catalyzed by RPE65, increased the formation of 9-cis-retinal, but not 11-cis-retinal, in a light-independent mechanism in both sexes of RPE65-null rd12 mice. Both rd12 and rd12;Fatp4-/- mice contained a massive amount of all-trans-retinyl esters in the eyes, exhibiting comparable scotopic vision and rod degeneration. However, expression levels of M- and S-opsins as well as numbers of M- and S-cones surviving in the superior retinas of rd12;Fatp4-/ - mice were at least twofold greater than those in age-matched rd12 mice. Moreover, FATP4 deficiency significantly shortened photopic b-wave implicit time, improved M-cone visual function, and substantially deaccelerated the progression of cone degeneration in rd12 mice, whereas FATP4 deficiency in mice with wild-type Rpe65 alleles neither induced 9-cis-retinal formation nor influenced cone survival and function. These results identify FATP4 as a new regulator of synthesis of 9-cis-retinal, which is a "cone-tropic" chromophore supporting cone survival and function in the retinas with defective RPE65.

Generation of two induced pluripotent stem cell lines (LVPEIi004-A and LVPEIi005-A) from probands with Leber Congenital Amaurosis 2 (LCA2) and harboring mutations in RPE65.

Leber Congenital Amaurosis 2 is an early onset retinal dystrophy that occurs due to mutation in RPE65 gene. Here, we report the generation of two patient specific induced pluripotent stem cell lines harboring nonsense mutations in exon 7 (c.646A > T) and exon 9 (c.992G > A) of RPE65 gene, respectively, which leads to premature translational termination and formation of defective protein. These lines were generated by the reprogramming of human dermal fibroblast cells using integration-free, episomal constructs expressing stemness genes. The stable lines maintained a normal karyotype, expressed the key stemness factors, underwent trilineage differentiation, and maintained their genetic identity and genomic integrity.

Multi-luminance mobility testing after gene therapy in the context of retinal functional diagnostics.

BACKGROUND: Voretigene neparvovec (Luxturna®) is the first approved gene therapy for RPE65-linked Leber congenital amaurosis (LCA). Though individual effects are highly variable, most recipients report improved vision in everyday life. To describe such effects, visual navigation tests are now frequently used in clinical trials. However, it is still unclear how their results should be interpreted compared to conventional parameters of visual function. METHODS: Seven LCA patients underwent a multi-luminance visual navigation test (Ora-VNCTM) before and 3 months after receiving Luxturna gene therapy. Their performance was rated based on the luminance level at which they passed the course. Differences between the first and second test were correlated to changes in visual acuity, full-field stimulus thresholds, chromatic pupil campimetry, and dark-adapted perimetry. RESULTS: A few patients displayed notable improvements in conventional measures of visual function whereas patients with advanced retinal degeneration showed no relevant changes. Independent of these results, almost all participants improved in the visual navigation task by one or more levels. The improvement in the mobility test was best correlated to the change in full-field stimulus thresholds. Other measures of visual functions showed no clear correlation with visual navigation. DISCUSSION: In patients who passed the test's more difficult levels, improved visual navigation can be attributed to the reactivation of rods. However, the performance of patients with low vision seemed to depend much more on confounding factors in the easier levels. In sum, such tests might only be meaningful for patients with better preserved visual functions.

Development of a novel prediction model based on protein structure for identifying RPE65-associated inherited retinal disease (IRDs) of missense variants.

Purpose: This study aimed to develop a prediction model to classify RPE65-mediated inherited retinal disease (IRDs) based on protein secondary structure and to analyze phenotype-protein structure correlations of RPE65 missense variants in a Chinese cohort. Methods: Pathogenic or likely pathogenic missense variants of RPE65 were obtained from UniProt, ClinVar, and HGMD databases. The three-dimensional structure of RPE65 was retrieved from the Protein Data Bank (PDB) and modified with Pymol software. A novel prediction model was developed using LASSO regression and multivariate logistic regression to identify RPE65-associated IRDs. A total of 21 Chinese probands with RPE65 variants were collected to analyze phenotype-protein structure correlations of RPE65 missense variants. Results: The study found that both pathogenic and population missense variants were associated with structural features of RPE65. Pathogenic variants were linked to sheet, ß-sheet, strands, ß-hairpins, Fe2+ (iron center), and active site cavity, while population variants were related to helix, loop, helices, and helix-helix interactions. The novel prediction model showed accuracy and confidence in predicting the disease type of RPE65 variants (AUC = 0.7531). The study identified 25 missense variants in Chinese patients, accounting for 72.4% of total mutations. A significant correlation was observed between clinical characteristics of RPE65-associated IRDs and changes in amino acid type, specifically for missense variants of F8 (H68Y, P419S). Conclusion: The study developed a novel prediction model based on the protein structure of RPE65 and investigated phenotype-protein structure correlations of RPE65 missense variants in a Chinese cohort. The findings provide insights into the precise diagnosis of RPE65-mutated IRDs.

Gene Supplementation in Mice Heterozygous for the D477G RPE65 Variant Implicated in Autosomal Dominant Retinitis Pigmentosa.

The use of AAV-RPE65 vectors for gene supplementation has achieved spectacular success as a treatment for individuals with autosomal recessive retinal disease caused by biallelic mutations in the visual cycle gene RPE65. However, the efficacy of this approach in treating autosomal dominant retinitis pigmentosa (adRP) associated with a monoallelic mutation encoding a rare D477G RPE65 variant has not been studied. Although lacking a severe phenotype, we now find that knock-in mice heterozygous for D477G RPE65 (D477G KI mice) can be used to evaluate outcomes of AAV-RPE65 gene supplementation. Total RPE65 protein levels, which are decreased in heterozygous D477G KI mice, were doubled following subretinal delivery of rAAV2/5.hRPE65p.hRPE65. In addition, rates of recovery of the chromophore 11-cis retinal after bleaching were significantly increased in eyes that received AAV-RPE65, consistent with increased RPE65 isomerase activity. While dark-adapted chromophore levels and a-wave amplitudes were not affected, b-wave recovery rates were modestly improved. The present findings establish that gene supplementation enhances 11-cis retinal synthesis in heterozygous D477G KI mice and complement previous studies showing that chromophore therapy results in improved vision in individuals with adRP associated with D477G RPE65.

Surgical Observations From the First 120 Cases of Subretinal Gene Therapy for Inherited Retinal Diseases.

PURPOSE: To report surgical observations formulated during the first 120 cases of subretinal gene therapy in patients with inherited retinal degenerations (IRDs). METHODS: A two-surgeon team compiled surgical observations and formulated surgical pearls based on the consecutive cases of subretinal viral vector injection in patients enrolled in clinical trials focusing on choroideremia, achromatopsia, and RP GTPase regulator associated retinitis pigmentosa, as well as patients with retinal pigment epithelium-specific-65-kDa (RPE65) associated Leber congenital amaurosis receiving Food and Drug Administration-approved voretigene neparvovec-rzyl therapy. RESULTS: One hundred twenty subretinal surgeries were performed by a two-surgeon team. Key anatomical features pertinent to surgical management were noted and are described in this article. Surgical decision making for successful subretinal administration of viral vectors and management of potential surgical challenges were formulated. CONCLUSION: Lessons learned during subretinal gene therapy cases may be helpful to other surgeons entering clinical trials or performing postapproval gene therapy administration. Surgical pearls outlined in this article may also be helpful for other targeted subretinal therapies, such as cellular transplantation or retinal prosthesis implantation.

Subretinal deposits in young patients treated with voretigene neparvovec-rzyl for RPE65-mediated retinal dystrophy.

We report a series of three young patients (ages: 22 months, 2 years, and 5 years) who developed subretinal deposits at post-operative week one following subretinal voretigene neparvovec-rzyl treatment for RPE65-mediated retinal dystrophy. In the 5-year-old, subretinal deposits were also observed in the inferior periphery of both eyes. All three patients experienced improved visual function with treatment, and both the macular and inferior subretinal deposits have improved or resolved over the follow-up period. These findings may inform the delivery parameters and safety profile of AAV-based gene therapy as the number of retinal gene therapy trials continues to grow.

Pathogenicity reclassification of the RPE65 c.1580A>G (p.His527Arg) - a case report.

BACKGROUND: It is of utmost importance to define the molecular diagnosis of patients with retinitis pigmentosa (RP) due to existing targeted therapeutic option: voretigene neparvovec.We provide clinical evidence for pathogenicity reclassification of variants of uncertain significance (VUSs) RPE65 c.1580A>G (p.His527Arg). MATERIALS AND METHODS: A case report of a 10-year-old boy with progressive vision loss. The patient manifested disease highly suggestive of RPE65 retinal dystrophy: nyctalopia, fairly good central vision, severely depressed full-field electroretinography responses and complete loss of peripheral fundus aut ofluorescence. RESULTS: Invitae Inherited Retinal Disorders Panel identified likely pathogenic mutation RPE65 c.499G>T (p.Asp167Tyr) and RPE65 c.1580A>G (p.His527Arg), variant of uncertain significance. Segregation analysis confirmed that these variants are in trans. CONCLUSIONS: We conclude that the variant RPE65 c.1580A>G (p.His527Arg) has contributed to the pathologic phenotype, demonstrating its significance clearly in the case presented, and should be reclassified according to the criteria of evidence as pathogenic. Therefore, patients with this specific variant in homozygous or compound heterozygous form would likely benefit from genetic treatment based on recombinant adeno-associated virus vector, providing a working RPE65 gene to act in place of a mutated RPE65 gene.

Chorioretinal atrophy following voretigene neparvovec despite the presence of fundus autofluorescence.

INTRODUCTION: Leber congenital amaurosis (LCA) type 2, due to disease-causing variants in RPE65, is characterized by severe visual loss in early infancy. Current treatments include voretigene neparvovec-rzyl (VN) for RPE65-associated LCA. Herein, we present the long-term follow-up of a patient treated with VN using quantitative autofluorescence (488 nm excitation). CASE REPORT: A 9-year-old girl with a diagnosis of LCA with biallelic variants in RPE65 presented for evaluation. The patient underwent VN treatment at the age of 11. The patient returned to clinic at age of 19 at which time imaging revealed evidence of chorioretinal atrophy. Quantitative autofluorescence performed prior to gene therapy and at 6- and 8-year follow-up revealed a central area of fundus autofluorescence. DISCUSSION: This case report demonstrates acquisition of fundus autofluorescence at 6- and 8-year follow-up despite the development of chorioretinal atrophy.

RPE65 c.353G>A, p.(Arg118Lys): A Novel Point Mutation Associated with Retinitis Pigmentosa and Macular Atrophy

Precise genetic diagnosis in RPE65-mediated retinitis pigmentosa (RP) is necessary to establish eligibility for genetic treatment with voretigene neparvovec: a recombinant adeno-associated viral vector providing a functional RPE65 gene. This case report aims to report a novel RP-related point mutation RPE65 c.353G>A, p.(Arg118Lys), a variant of uncertain significance associated with a severe clinical presentation and the striking phenotypic feature of complete macular atrophy. We report the case of a 40-year-old male with inherited retinal dystrophy, all features typical for the RPE65-associated RP, and marked macular atrophy. Genetic testing identified that the patient was a compound heterozygote in trans form with two heterozygous variants: RPE65 c.499G>T, p.(Asp167Tyr) and RPE65 c.353G>A, p.(Arg118Lys). Furthermore, short-wavelength and near-infrared autofluorescence patterns exhibited deficiencies specific to mutations in the visual cycle genes. To the best of our knowledge, RPE65 c.353G>A, p.(Arg118Lys) is the first described point mutation on this locus, among all other reported insertional mutations, currently classified as likely benign and of uncertain significance. We concluded that this variant contributed to the pathological phenotype, demonstrating its significance clearly to be reclassified as likely pathogenic. This being the case, patients with this specific variant in homozygous or compound heterozygous form would be likely candidates for genetic treatment with voretigene neparvovec.

Emerging Gene Manipulation Strategies for the Treatment of Monogenic Eye Disease.

Genetic eye diseases, representing a wide spectrum of simple and complex conditions, are one of the leading causes of visual loss in children and working adults, and progress in the field has led to changes in disease investigation, diagnosis, and management. The past 15 years have seen the emergence of novel therapies for these previously untreatable conditions to the extent that we now have a licensed therapy for one form of genetic eye disease and many more in clinical trial. This is a systematic review of published and ongoing clinical trials of gene therapies for monogenic eye diseases. Databases of clinical trials and the published literature were searched for interventional studies of gene therapies for eye diseases. Standard methodological procedures were used to assess the relevance of search results. A total of 59 registered clinical trials are referenced, showing the significant level of interest in the potential for translation of these therapies from bench to bedside. The breadth of therapy design is encouraging, providing multiple possible therapeutic mechanisms. Some fundamental questions regarding gene therapy for genetic eye diseases remain, such as optimal dosing, the relative benefits of adeno-associated virus (AAV)-packaging and the potential for a significant inflammatory response to the therapy itself. As a result, despite the promise of the eye as a target, it has proven difficult to deliver clinically effective gene therapies to the eye. Despite setbacks, the licensing of Luxturna (voretigene neparvovec, Novartis) for the treatment of RPE65-mediated Leber congenital amaurosis (LCA) is a major advance in efforts to treat these rare, but devastating, causes of visual loss.

[Inherited retinal dystrophy: first results of RPE65 gene replacement therapy in Russia]. / Nasledstvennaya distrofiya setchatki: pervye rezul'taty posle RPE65-genozamestitel'noi terapii v Rossii.

PURPOSE: To present the main aspects of interdisciplinary diagnostics of patients with hereditary retinal diseases and the first results of the follow-up of patients with inherited retinal dystrophies (IRD) caused by biallelic mutations in the gene RPE65 after gene replacement therapy in Russia. MATERIAL AND METHODS: The cohort of patients consisted of six children (5-15 years old) with the diagnosis of Leber amaurosis type 2. All patients underwent a multi-disciplinary examination using conventional clinical, instrumental and molecular-genetic methods. Genetic diagnosis was established based on the results of two-stage DNA diagnostics using high-performance parallel sequencing of a custom panel and family segregation analysis by Sanger sequencing. RESULTS: In the Research Centre for Medical Genetics the first group of Russian patients with an orphan inherited retinal disease was verified, they underwent subretinal injection of the gene replacement drug Voretigene neparvovec (12 eyes) in the Helmholtz National Medical Research Center of Eye Diseases. According to the regulated terms of monitoring gene therapy patients, they were examined in the Research Centre for Medical Genetics after 1, 3, 6 and 12 months, and then once per year. Thus, the available data allows us to analyze the first results 3 months after the treatment. CONCLUSION: The presented data on inherited retinal dystrophies caused by biallelic mutations in the RPE65 gene emphasize the need to change the diagnostic algorithm in the ophthalmic practice. The use of clinical instrumental and molecular genetic diagnostic methods makes it possible to apply etiotropic treatment to patients with a disabling disease that was previously considered untreatable. The gene replacement drug Voretigene neparvovec registered in Russia showed irrefutable first positive results in all targeted patients.

Short-Term Outcomes of the First in Vivo Gene Therapy for RPE65-Mediated Retinitis Pigmentosa.

Here, we report early treatment outcomes of gene therapy for early onset retinitis pigmentosa (RP) (Leber congenital amaurosis) associated with biallelic RPE65 mutation in a 30-year-old female patient. Initially, her visual acuity (VA) was 20/200, and her visual field (VF) was severely constricted to the center in the left eye. Her electroretinography showed nearly extinct signals. Full-field stimulus threshold test (FST) revealed diminished dark-adapted light sensitivity. Voretigene neparvovec-rzyl (VN) is the first in vivo viral gene therapy agent to be approved. At 3 months after subretinal injection of VN in the left eye, VA, VF, and FST showed sustained improvement. She did not exhibit any signs of adverse effects from the treatment. Gene therapy for RP proved to be an effective and safe treatment in an advanced case of RPE65-associatied early onset RP.

In vivo base editing rescues cone photoreceptors in a mouse model of early-onset inherited retinal degeneration.

Leber congenital amaurosis (LCA) is the most common cause of inherited retinal degeneration in children. LCA patients with RPE65 mutations show accelerated cone photoreceptor dysfunction and death, resulting in early visual impairment. It is therefore crucial to develop a robust therapy that not only compensates for lost RPE65 function but also protects photoreceptors from further degeneration. Here, we show that in vivo correction of an Rpe65 mutation by adenine base editor (ABE) prolongs the survival of cones in an LCA mouse model. In vitro screening of ABEs and sgRNAs enables the identification of a variant that enhances in vivo correction efficiency. Subretinal delivery of ABE and sgRNA corrects up to 40% of Rpe65 transcripts, restores cone-mediated visual function, and preserves cones in LCA mice. Single-cell RNA-seq reveals upregulation of genes associated with cone phototransduction and survival. Our findings demonstrate base editing as a potential gene therapy that confers long-lasting retinal protection.

The first gene therapy for RPE65 biallelic dystrophy with voretigene neparvovec-rzyl in Brazil.

PURPOSE: To report the first Brazilian patient with RPE65 deficiency-inherited retinal dystrophy (RPE65-IRD) treated with voretigene neparvovec-rzyl (VN). METHODS: An adult patient with Leber congenital amaurosis-2 with a homozygous mutation in the RPE65 gene (p.Phe83Leu) was treated bilaterally with VN. The clinical and surgical aspects are described. The baseline and 4-month postoperative ophthalmologic examinations included measurement of the best-corrected visual acuity (BCVA), full-field stimulus threshold (FST) test, Octopus 900 semiautomated kinetic visual fields (VFs), and microperimetry. RESULTS: No complications developed in this patient. The BCVA remained stable. The full-field stimulus threshold test (FST) and VFs showed clinically significant improvements bilaterally. The patient reported significant improvements in the ability to perform daily activities, mainly for those requiring the VFs and vision in a low-luminescence environment. CONCLUSIONS: The treatments were beneficial for this patient who was homozygous for RPE65 p.Phe83Leu. The first VN treatments in an adult Brazilian patient in clinical practice showed measurable improvements in visual outcomes that were meaningful for the patient's daily activities. TRANSLATIONAL RELEVANCE: This case reinforces the clinical trial results and proves that the procedure is feasible in countries such as Brazil.

Generation of a patient-derived induced pluripotent cell line (SCTCi016-A) carrying a homozygous variant in RPE65.

Leber congenital amaurosis (LCA) can be caused by mutations in more than 20 different genes. One of these, RPE65, encodes a protein essential for the visual cycle that is expressed in retinal pigment epithelium cells. In this work, we describe the generation and characterization of the human iPSC line SCTCi16-A. This hiPSC line was generated from peripheral blood mononuclear cells (PBMCs) from a patient affected with LCA caused by the homozygous c.11+5G>A variant in the RPE65 gene. Reprograming was conducted using episomal vectors containing OCT3/4, SOX2, KLF4, L-MYC, and LIN28.

Epidemiology of Mutations in the 65-kDa Retinal Pigment Epithelium (RPE65) Gene-Mediated Inherited Retinal Dystrophies: A Systematic Literature Review.

INTRODUCTION: Inherited retinal dystrophies (IRDs) represent a genetically diverse group of progressive, visually debilitating diseases. Adult and paediatric patients with vision loss due to IRD caused by biallelic mutations in the 65-kDa retinal pigment epithelium (RPE65) gene are often clinically diagnosed as retinitis pigmentosa (RP), and Leber congenital amaurosis (LCA). This study aimed to understand the epidemiological landscape of RPE65 gene-mediated IRD through a systematic review of the literature, as the current evidence base for its epidemiology is very limited. METHODS: Medline, Embase, and other databases were searched for articles on the epidemiology of RPE65 gene-mediated IRDs from inception until June 2021. Studies were included if they were original research articles reporting the epidemiology of RP and LCA and/or proportion of RPE65 gene mutations in these clinically diagnosed or molecularly confirmed IRDs patients. RESULTS: A total of 100 studies with relevant data were included in this systematic review. The range for prevalence of LCA and RP in the literature was 1.20-2.37 and 11.09-26.43 per 100,000, respectively. The proportion of RPE65 mutations in clinically diagnosed patients with LCA was found to be between ~ 2-16% within the US and major European countries (France, Germany, Italy, Spain, and the UK). This range was also comparable to our findings in the Asian region for RPE65-LCA (1.26-16.67%). Similarly, for these European countries, RPE65-RP was estimated between 0.23 and 1.94%, and RPE65-IRD range was 1.2-14%. Further, in the Americas region, mutations in RPE65 were reported to cause 1-3% of RP and 0.8-3.7% of IRD cases. Lastly, the RPE65-IRD range was 4.81-8% in the Middle East region. CONCLUSIONS: There are significant variations in reporting of RPE65 proportions within countries as well as regions. Generating robust epidemiological evidence on RPE65 gene-mediated IRDs would be fundamental to support rare disease awareness, timely therapeutic intervention, and public health decision-making.

Real-world outcomes of voretigene neparvovec treatment in pediatric patients with RPE65-associated Leber congenital amaurosis.

PURPOSE: To investigate real-world safety and efficacy of voretigene neparvovec gene therapy administration in pediatric patients with biallelic RPE65 disease-causing variants. METHODS: A retrospective study of 27 eyes of 14 patients with RPE65-associated Leber congenital amaurosis examined postoperative complications and longitudinal changes in photoreceptor function following treatment with subretinal injection of voretigene neparvovec. Full-field stimulus threshold testing (FST), Goldmann visual fields (GVF), best-corrected visual acuity (BCVA), and central subfield thickness (CST) on optical coherence tomography (OCT) scans were collected preoperatively and up to 12 months posttreatment. RESULTS: Baseline through 6-12 month follow-up FST and GVF data were obtained for 13 eyes of 7 patients. FST improved for each eye after treatment with a mean improvement of 2.1 log-units (P < 0.001) and GVF improved for each eye with a mean improvement of 221 sum degrees (P < 0.001). BCVA improved from logMAR 0.98 at baseline to logMAR 0.83 at last follow-up (P < 0.001). Across 19 eyes of 10 patients included in CST analysis, there was a small but statistically significant 9-µ decrease in mean CST from baseline to last follow-up (P < 0.001). The most common postoperative issues included elevation in intraocular pressure (59%), persistent intraocular inflammation (15%), and vitreous opacities (26%) that resolved over a period of months. CONCLUSIONS: This report provides some of the earliest longitudinal real-world evidence of the pediatric safety and efficacy of voretigene neparvovec using multiple functional and structural measures of the retina. Outcomes demonstrate significant improvements in visual function consistent with clinical trial results.