Comprehensive analysis of two hotspot codons in the TUBB4B gene and associated phenotypes.

Our purpose was to elucidate the genotype and ophthalmological and audiological phenotype in TUBB4B-associated inherited retinal dystrophy (IRD) and sensorineural hearing loss (SNHL), and to model the effects of all possible amino acid substitutions at the hotspot codons Arg390 and Arg391. Six patients from five families with heterozygous missense variants in TUBB4B were included in this observational study. Ophthalmological testing included best-corrected visual acuity, fundus examination, optical coherence tomography, fundus autofluorescence imaging, and full-field electroretinography (ERG). Audiological examination included pure-tone and speech audiometry in adult patients and auditory brainstem response testing in a child. Genetic testing was performed by disease gene panel analysis based on genome sequencing. The molecular consequences of the substitutions of residues 390 and 391 on TUBB4B and its interaction with α-tubulin were predicted in silico on its three-dimensional structure obtained by homology modelling. Two independent patients had amino acid exchanges at position 391 (p.(Arg391His) or p.(Arg391Cys)) of the TUBB4B protein. Both had a distinct IRD phenotype with peripheral round yellowish lesions with pigmented spots and mild or moderate SNHL, respectively. Yet the phenotype was milder with a sectorial pattern of bone spicules in one patient, likely due to a genetically confirmed mosaicism for p.(Arg391His). Three patients were heterozygous for an amino acid exchange at position 390 (p.(Arg390Gln) or p.(Arg390Trp)) and presented with another distinct retinal phenotype with well demarcated pericentral retinitis pigmentosa. All showed SNHL ranging from mild to severe. One additional patient showed a variant distinct from codon 390 or 391 (p.(Tyr310His)), and presented with congenital profound hearing loss and reduced responses in ERG. Variants at codon positions 390 and 391 were predicted to decrease the structural stability of TUBB4B and its complex with α-tubulin, as well as the complex affinity. In conclusion, the twofold larger reduction in heterodimer affinity exhibited by Arg391 substitutions suggested an association with the more severe retinal phenotype, compared to the substitution at Arg390.

Comparison of Full-Field Stimulus Threshold Measurements in Patients With Retinitis Pigmentosa and Healthy Subjects With Dilated and Nondilated Pupil.

Purpose: The common protocol of full-field stimulus threshold (FST) testing recommends pupil dilation. The aim of this study is to investigate the difference between FST measurements with dilated and nondilated pupils in healthy subjects and patients with retinitis pigmentosa (RP). Methods: Twenty healthy subjects and 20 RP patients were selected. One pupil of each subject was dilated; the other eye was measured in physiological width of the pupil. The FST was conducted using Diagnosys Espion E2/E3 with white, blue, and red stimuli. Statistical analysis was conducted with a mixed-model analysis of variance and a paired t-test. Results: The statistical analysis revealed a significant difference between measurements of dilated and nondilated pupils with the following: blue stimuli for all subjects and groups except those with highly progressed RP; white stimuli for all tested subjects in total, for RP patients with better-preserved visual field (VF), and rod-mediated FST response; and red stimuli for RP patients with better-preserved VF and rod-mediated FST response. On average, the difference between the FST values for RP patients were -3.2 ± 3 dB for blue, -2.3 ± 2.9 dB for white, and -0.83 ± 3 dB for red stimuli. The correlation between the FST values of dilated and nondilated pupils with all three stimuli was linear. Conclusions: Current recommendations are to perform FST with dilated pupils. However, based on this study's findings, pupil dilation can be omitted for clinical diagnostics or rough follow-ups. Translational Relevance: Our data provide useful information for the clinical use of FST.

RPE65-Associated Retinal Dystrophies: Phenotypes and Treatment Effects with Voretigene Neparvovec.

Retinal dystrophies linked to the RPE65 gene are mostly fast-progressing retinal diseases, with childhood onset of night blindness and progressive visual loss up to the middle adult age. Rare phenotypes linked to this gene are known with congenital stationary night blindness or slowly progressing retinitis pigmentosa, as well as an autosomal dominant c.1430A>G (p.Asp477Gly) variant. This review gives an overview of the current knowledge of the clinical phenotypes, as well as experience with the efficacy and safety of the approved gene augmentation therapy voretigene neparvovec.

Chorioretinal Atrophy Growth After Voretigene Neparvovec Retinotopically Is Connected to Retinal Functional Rescue.

Purpose: Chorioretinal atrophy growth after voretigene neparvovec has been reported recently with its positive correlation with successful treatment. This finding raised the question on long-term effects and the etiology of the chorioretinal atrophy. Methods: Using local retinal functional diagnostics, we tested whether the atrophy growth is connected to the initial local functional improvement after the therapy. Results: The results describe factors predicting the development of atrophy. First, the atrophy emerges after approximately 3 months in an area with local functional rescue before. The areas of the greatest gain in the number of functionally rescued rods are prone to be the initial spots of atrophy growth in almost one-half of the cases and the retinotopy corresponds with the area of a high number of post-treatment functioning rods. Second, the dark-adapted perimetry shows that the atrophy growth is in the area with functioning rescued rods. However, the rods with the greatest sensitivity gain are not the parts of the growing atrophy in the first 2 years after intervention. This preservation of rods with the greatest sensitivity seems to explain the excellent profile of rods rescue over the long term measured by full-field stimulus threshold and reported earlier. Conclusions: A disbalance between the increase of functional rods and their threshold shortly after treatment could be an indicator for a metabolic origin of chorioretinal atrophy after voretigene neparvovec. Translational Relevance: A basic understanding of the photoreceptor rescue aspects after gene therapy can demonstrate a metabolic causal influence of the efficacy on the development of side effects, such as chorioretinal atrophy.

Influence of open-source virtual-reality based gaze training on navigation performance in Retinitis pigmentosa patients in a crossover randomized controlled trial.

METHODS: A group of RP patients (n = 8, aged 20-60) participated in a study consisting of two 4-week-phases, both carried out by the same patient group in randomized order: In the 'training phase', participants carried out a Virtual-Reality gaze training for 30 minutes per day; In the 'control phase', no training occurred. Before and after each phase, participants were tasked to move through a randomized real-world obstacle course. Navigation performance in the obstacle course as well as eye-tracking data during the trials were evaluated. The study is registered at the German Clinical Trials Register (DRKS) with the ID DRKS00032628. RESULTS: On average, the time required to move through the obstacle course decreased by 17.0% after the training phase, the number of collisions decreased by 50.0%. Both effects are significantly higher than those found in the control phase (p < 0.001 for required time, p = 0.0165 for number of collisions), with the required time decreasing by 5.9% and number of collisions decreasing by 10.4% after the control phase. The average visual area observed by participants increases by 4.41% after training, however the effect is not found to be significantly higher than in the control phase (p = 0.394). CONCLUSION: The performance increase over the training phase significantly surpasses the natural learning effect found in the control phase, suggesting that Virtual-Reality based gaze training can have a positive effect on real-world navigation tasks for patients with RP. The training is available as work-in-progress open-source software.

Splicing defects and CRISPR-Cas9 correction in isogenic homozygous photoreceptor precursors harboring clustered deep-intronic ABCA4 variants.

Splicing defects from deep-intronic variants significantly contribute to the mutational spectrum in ABCA4-associated inherited retinal diseases, necessitating functional validation for their pathological classification. Typically, minigene assays in HEK293(T) can qualitatively assess splicing defects, yet they often fail to quantitatively reproduce the resulting mis-splicing patterns, leaving uncertainty on severity and pathogenicity. As a potential cellular model derived from patient cells, photoreceptor precursor cells (PPCs) play a pivotal role in assessing the severity of specific splicing mutations. Nevertheless, the accessibility of biosamples is commonly constrained, and their establishment is costly and laborious. In this study, we combined and investigated the use of a minigene assay and isogenic PPCs, as superior qualitative and more accessible cellular models for the assessment of splicing defects. Specifically, we focused on the clustered c.5196+1013A>G, c.5196+1056A>G, and c.5196+1216C>A deep-intronic variants in intron 36 of ABCA4, comparing their resulting (mis)splicing patterns in minigene-transfected cells and isogenic CRISPR-Cas9-knocked-in PPCs harboring these pathogenic variants in homozygous state. Moreover, we demonstrate the successful correction of these three splicing defects in homozygous mutant PPCs using a single pair of guide RNAs to target Cas9 cleavage, thereby identifying an efficient gene editing strategy for therapeutic applications.

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.

Diagnostic genome sequencing improves diagnostic yield: a prospective single-centre study in 1000 patients with inherited eye diseases.

PURPOSE: Genome sequencing (GS) is expected to reduce the diagnostic gap in rare disease genetics. We aimed to evaluate a scalable framework for genome-based analyses 'beyond the exome' in regular care of patients with inherited retinal degeneration (IRD) or inherited optic neuropathy (ION). METHODS: PCR-free short-read GS was performed on 1000 consecutive probands with IRD/ION in routine diagnostics. Complementary whole-blood RNA-sequencing (RNA-seq) was done in a subset of 74 patients. An open-source bioinformatics analysis pipeline was optimised for structural variant (SV) calling and combined RNA/DNA variation interpretation. RESULTS: A definite genetic diagnosis was established in 57.4% of cases. For another 16.7%, variants of uncertain significance were identified in known IRD/ION genes, while the underlying genetic cause remained unresolved in 25.9%. SVs or alterations in non-coding genomic regions made up for 12.7% of the observed variants. The RNA-seq studies supported the classification of two unclear variants. CONCLUSION: GS is feasible in clinical practice and reliably identifies causal variants in a substantial proportion of individuals. GS extends the diagnostic yield to rare non-coding variants and enables precise determination of SVs. The added diagnostic value of RNA-seq is limited by low expression levels of the major IRD disease genes in blood.

A morphometric analysis of the retinal arterioles with adaptive optics imaging in RPE65-associated retinal dystrophy after treatment with voretigene neparvovec.

PURPOSE: To investigate the changes in retinal arterial architecture after treatment with voretigene neparvovec in patients with retinal dystrophy caused by bi-allelic mutations in the RPE65 gene. METHODS: Sixteen eyes treated with voretigene neparvovec at the University Eye Clinic in Tuebingen, Germany, underwent adaptive optics ophthalmoscopy (AO) imaging at baseline and 2 weeks, 1, 3, 6 and 12 months after treatment. Follow-up was performed in six eyes of four patients. For each eye, five different positions at arterial vessels were selected and the wall-to-lumen ratio (WLR), the lumen diameter (LD) and the wall cross-sectional area (WCSA) were measured by the manufacturer's software over the observational period. RESULTS: Vast retinal atrophy dominated all gained AO images. WLR fluctuated in the observation period without statistically significant change. LD and WCSA changed significantly after 2 weeks from the baseline examination and returned to values similar to baseline thereafter. There were no signs of inflammation such as macrophages or perivascular accumulated fluid visible. CONCLUSION: AO imaging of the retinal vessels in RPE65-associated retinal dystrophies (IRD) is challenging. There was no change in the retinal arterial vasculature over the observation period of 12 months that would indicate inflammatory changes. Decrease of the LD and WCSA shortly after treatment might be caused by the perioperative prednisolone intake. AO of retinal vessels can be used as a diagnostic module to complement monitoring the disease and effects of genetic treatments if the acquisition is possible in selected cases.

An early onset cone dystrophy due to CEP290 mutation: a case report.

PURPOSE: Biallelic mutations in the CEP290 gene cause early onset retinal dystrophy or syndromic disease such as Senior-Loken or Joubert syndrome. Here, we present an unusual non-syndromic case of a juvenile retinal dystrophy caused by biallelic CEP290 mutations imitating initially the phenotype of achromatopsia or slowly progressing cone dystrophy. METHODS: We present 13 years of follow-up of a female patient who presented first with symptoms and findings typical for achromatopsia. The patient underwent functional and morphologic examinations, including fundus autofluorescence imaging, spectral-domain optical coherence tomography, electroretinography, color vision and visual field testing. RESULTS: Diagnostic genetic testing via whole genome sequencing and virtual inherited retinal disease gene panel evaluation finally identified two compound heterozygous variants c.4452_4455del;p.(Lys1484Asnfs*4) and c.2414T > C;p.(Leu805Pro) in the CEP290 gene. CONCLUSIONS: CEP290 mutation causes a wide variety of clinical phenotypes. The presented case shows a phenotype resembling achromatopsia or early onset slowly progressing cone dystrophy.

Rod and Cone Function Measured Objectively by Chromatic Pupil Campimetry Show a Different Preservation Between Distinct Genotypes in Retinitis Pigmentosa.

Purpose: Verifying whether specific genotypes causing retinitis pigmentosa (RP) show differences in the preservation of rod and cone function measured by chromatic pupil campimetry (CPC). Methods: Sixty-three RP eyes (37 male, 14-58 years) were measured using CPC with specific photopic and scotopic protocols, and the relative maximal constriction amplitudes and latencies to constriction onset were analyzed per genotype (RP due to variants in EYS, n = 14; PDE6A, n = 10; RPE65, n = 15; USH2A, n = 10; and RPGR, n = 14). Correlation analyses between the pupillary responses were performed with age, full-field stimulus threshold (FST), and optical coherence tomography (OCT) for cones and rods, respectively, to the genotype. Results: Pupillary responses were most severely reduced in RPE65-RP. Patients with disease-associated variants in EYS and USH2A were accompanied with better-preserved rod function compared with the other subgroups, reaching statistical significance between EYS and RPE65. Cone function was statistically significantly correlated with age in USH2A-RP with an annual decline of 2.4%. Correlations of pupillary responses were found with FST but barely with the ellipsoid zone area in OCT. Latency was significantly more prolonged in RPE65-RP compared with the other genotypes for cones. Conclusions: Rod and cone function measured objectively by CPC showed a different preservation between genotypes in RP. However, heterogeneity inside the same genotype was present. CPC data correlated with FST, but structural OCT parameters seem to be limited indicators for photoreceptor function in RP. Prolonged time dynamics for cones in RPE65 mutations suggest an impact on cone processing and might provide additional information in the evaluation of therapy effects.

Impact of A-Scan Rate on Image Quality and Acquisition Time in OCT.

PURPOSE: This study aims to examine the impact of the A-scan rate in optical coherence tomography (OCT) on scan quality and acquisition time. METHODS: Two horizontal OCT scans per scan rate (20, 85, 125 kHz) of the right eye were captured with the same OCT device (Spectralis SHIFT, HRA + OCT, Heidelberg Engineering GmbH, Heidelberg, Germany) of patients who presented to the inherited retinal dystrophies consultation, thus predominantly challenging patients due to reduced fixation ability. Scan quality was measured by the Q score, a signal-to-noise-ratio (SNR). Acquisition time was measured in seconds. RESULTS: Fifty-one patients were included in the study. The highest quality was seen for an A-scan rate of 20 kHz (44.49 dB), succeeded by scans of an A-Scan rate of 85 kHz (38.53 dB) and of 125 kHz (36.65 dB). Differences in scan quality between the various A-scan rates were statistically significant. The acquisition time seen for an A-scan rate of 20 kHz (6.45 s) was significantly longer than those seen for an A-Scan rate of 85 kHz (1.51 s) and of 125 kHz (1.69 s). CONCLUSION: An A-scan rate of 20 kHz resulted in a significantly higher scan quality but also a significantly longer acquisition time compared to scan rates of 85 kHz and 125 kHz. Differences between an A-scan rate of 85 kHz and 125 kHz were marginal.

The VA-CAL Test Quantifies Improvement of Visual Acuity in Achromatopsia by Means of Short-Wave Cutoff Filter Glasses in Daily Living Conditions.

Purpose: To quantify visual performance of patients with achromatopsia at various contrast and luminance combinations typical for daily living conditions, in comparison to controls, and to measure beneficial effects of short-wavelength cutoff filter glasses used by patients with achromatopsia to reduce glare sensation. Methods: Best-corrected visual acuity (BCVA) was tested with Landolt rings using an automated device (VA-CAL test). The visual acuity space was assessed for each participant with and without filter glasses (transmission >550 nm) at 46 contrast-luminance combinations (18%-95%; 0-10,000 cd/m2). The BCVA differences between both conditions were calculated for each combination as absolute values and relative to individual standard BCVA. Results: Fourteen achromats (mean ± SD: 37.9 ± 17.6 years) and 14 normally sighted controls (mean ± SD: 25.2 ± 2.8 years) were included in the study. Without filter glasses, achromats' BCVA was best at 30 cd/m2 (mean ± SEM: 0.76 ± 0.046 logarithm of the minimum angle of resolution [logMAR], contrast = 89%) and worst at 10,000 cd/m2 (mean ± SEM: 1.41 ± 0.08 logMAR, contrast = 18%), a deterioration up to 0.6 logMAR due to increased luminance and decreased contrast. Filter glasses improved achromats' BCVA for almost all luminances by about 0.2 logMAR but lowered controls' BCVA by about 0.1 logMAR. Conclusions: The VA-CAL test provides numerical proof that short-wavelength cutoff filter glasses can help patients with achromatopsia in everyday life, avoiding the common situation of severe visual impairment at certain daily object contrasts and ambient luminances. Translational Relevance: The VA-CAL test discovers losses of spatial resolution in the visual acuity space not seen in standardized BCVA assessment. Filter glasses improve the patients' daily visual performance, rendering them a strongly recommended visual aid in achromatopsia.

Photoreceptor-Specific Temporal Contrast Sensitivities in RP1L1-Associated Occult Macular Dystrophy.

Purpose: The purpose of this study was to compare L-, M-, S-cone-, and rod-driven temporal contrast sensitivities (tCS) in patients with RP1L1-associated autosomal-dominant occult macular dystrophy (OMD), and to investigate how photoreceptor degeneration determines which post-receptoral channels dominate perception. Methods: Photoreceptor isolating stimuli were created with the silent substitution technique. Photoreceptor-selective tCS deviations (D L-cone/M-cone/S-cone/Rod) were obtained as a function of temporal frequency with identical retinal adaptation, by subtracting tCS from age-corrected normal values. A linear-mixed effects model was used for analysis. Results: Eleven genetically confirmed patients were included (7 women, 5 men; age = 52.27 ± 14.44 years). Overall, L- and M-cone-driven sensitivity deviations (DL-cone and DM-cone) were more negative than DS-cone; DRod was normal at frequencies between 8 and 12 Hz in all subjects. Rod-driven tCS functions allowed identification of two subgroups of patients: one with band-pass properties and one with low-pass properties, suggesting dominance of different post-receptoral filters. The same filtering properties were observed in L-cone-driven tCS functions. Furthermore, the two subgroups also differed in clinical parameters (spherical equivalent, BCVA, perimetry, and ocular coherence tomography (OCT) reflectivity of the ellipsoid zone relative to the RPE). Conclusions: OMD was characterized predominantly by deterioration of L- and M-cone-cone driven function in the perifovea. Rod-driven functions were normal. Differences in the photoreceptor signals were further modified by postreceptoral filters.

Current Management of Patients with RPE65 Mutation Associated Inherited Retinal Degenerations in Europe: Results of a 2-Year Follow-Up Multinational Survey.

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 (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 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 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, http://www.encepp.eu/encepp/viewResource.htm?id=37005). 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 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.

The Usher syndrome 1C protein harmonin regulates canonical Wnt signaling.

Human Usher syndrome (USH) is the most common form of hereditary combined deaf-blindness. USH is a complex genetic disorder, and the pathomechanisms underlying the disease are far from being understood, especially in the eye and retina. The USH1C gene encodes the scaffold protein harmonin which organizes protein networks due to binary interactions with other proteins, such as all USH proteins. Interestingly, only the retina and inner ear show a disease-related phenotype, although USH1C/harmonin is almost ubiquitously expressed in the human body and upregulated in colorectal cancer. We show that harmonin binds to ß-catenin, the key effector of the canonical Wnt (cWnt) signaling pathway. We also demonstrate the interaction of the scaffold protein USH1C/harmonin with the stabilized acetylated ß-catenin, especially in nuclei. In HEK293T cells, overexpression of USH1C/harmonin significantly reduced cWnt signaling, but a USH1C-R31* mutated form did not. Concordantly, we observed an increase in cWnt signaling in dermal fibroblasts derived from an USH1C R31*/R80Pfs*69 patient compared with healthy donor cells. RNAseq analysis reveals that both the expression of genes related to the cWnt signaling pathway and cWnt target genes were significantly altered in USH1C patient-derived fibroblasts compared to healthy donor cells. Finally, we show that the altered cWnt signaling was reverted in USH1C patient fibroblast cells by the application of Ataluren, a small molecule suitable to induce translational read-through of nonsense mutations, hereby restoring some USH1C expression. Our results demonstrate a cWnt signaling phenotype in USH establishing USH1C/harmonin as a suppressor of the cWnt/ß-catenin pathway.

Static Perimetry in the Rate of Progression in USH2A-related Retinal Degeneration (RUSH2A) Study: Assessment Through 2 Years.

PURPOSE: To evaluate disease progression using static perimetry (SP) in patients with USH2A-related retinal degeneration, including Usher syndrome type 2 (USH2) and nonsyndromic autosomal recessive retinitis pigmentosa. DESIGN: Prospective, observational cohort study. METHODS: A total of 102 patients with biallelic disease-causing sequence variants in USH2A with baseline best-corrected visual acuity (BCVA) letter score ≥54 were recruited from 16 clinical sites in Europe and North America. SP, BCVA, full-field stimulus thresholds, spectral domain optical coherence tomography macular scans, and fundus-guided mesopic microperimetry were performed at baseline and annually. The main outcome measures were total hill of vision (VTOT), hill of vision in the central 30° (V30), VTOT minus V30 (VPERIPH), and mean sensitivity. RESULTS: The average decline (95% CI) was 2.05 (1.40, 2.70) decibel-steradian (dB-sr)/y for VTOT, 0.48 (0.32, 0.65) dB-sr/y for V30, 1.53 (0.97, 2.08) dB-sr/y for VPERIPH, and 0.55 (0.40, 0.71) dB/y for mean sensitivity. Average percentage decline per year was 8.3 (5.5, 11.1) for VTOT, 5.2 (3.0, 7.4) for V30, 16.0 (9.5, 22.0) for VPERIPH, and 5.1 (3.5, 6.7) for mean sensitivity. Changes from baseline to year 2 in all SP measures were highly correlated (r's ranging from 0.52 [V30 vs VPERIPH] to 0.98 [VTOT vs VPERIPH]). CONCLUSIONS: Quantitative measures of SP declined significantly over 2 years in USH2A-related retinal degeneration. The annual percentage rate of change was greatest for VTOT and VPERIPH, whereas V30 and mean sensitivity changed least, reflecting earlier and more severe peripheral degeneration compared with central loss.