SIBLING CONCORDANCE IN SYMPTOM ONSET AND ATROPHY GROWTH RATES IN STARGARDT DISEASE USING ULTRA-WIDEFIELD FUNDUS AUTOFLUORESCENCE.

PURPOSE: To investigate concordance in symptom onset, area of dark autofluorescence (DAF), and growth rate (GR) between Stargardt disease siblings at an age-matched time point. METHODS: In this retrospective longitudinal study of sibling pairs with identical biallelic ABCA4 variants, age at symptom onset, best-corrected visual acuity, atrophy area, and effective radius of DAF on ultra-widefield fundus autofluorescence were recorded. Absolute intersibling differences for both eyes were compared with absolute interocular differences using the Mann-Whitney test. RESULTS: Overall 39 patients from 19 families were recruited. In 16 families, age-matched best-corrected visual acuity and DAF were compared between siblings. In 8 families, DAF GR was compared. The median (range) absolute difference in age at symptom onset between siblings was 3 (0-35) years. Absolute intersibling differences in age-matched best-corrected visual acuity were greater than interocular differences ( P = 0.01). Similarly, absolute intersibling differences in DAF area and radius were greater than interocular differences ( P = 0.04 for area and P = 0.001 for radius). Differences between absolute interocular and intersibling GR were not statistically significant ( P = 0.44 for area GR and P = 0.61 for radius GR). CONCLUSION: There was significant discordance in age-matched best-corrected visual acuity and DAF beyond the expected limits of interocular asymmetry. Lack of significant intersibling differences in GR warrants further investigation.

A novel phenotype in a family with autosomal dominant retinal dystrophy due to c.1430A > G in retinoid isomerohydrolase (RPE65) and c.37C > T in bestrophin 1 (BEST1).

PURPOSE: The c.1430A > G (Asp477Gly) variant in RPE65 has been reported in Irish and Scottish families with either an autosomal dominant retinal dystrophy (adRD) that resembles choroideremia, a vitelliform macular dystrophy or an isolated macular atrophy. We report novel features on multimodal imaging and the natural history of a family harbouring this variant in combination with the BEST1 c.37C > T (Arg13Cys) variant. METHODS: Members of a family with an adRD were examined clinically to ascertain phenotype and underwent genetic testing. Multimodal imaging included widefield colour fundus photography, quantitative autofluorescence (qAF) and spectral domain optical coherence tomography. Electrophysiology and microperimetry were also performed. RESULTS: Vision loss was attributed to foveal atrophy in the proband and choroidal neovascularisation and a vitello-eruptive lesion in one affected son. Peripheral retinal white dots corresponding to subretinal deposits were seen in three patients. The median qAF8 values in the proband (I:1) were low (40 and 101 in OD and OS) at age 79. Similarly, the qAF8 values for the middle son (II:2) were also low (100 and 87 in ODS and OS) at age 60. Electrophysiology showed disproportionate reduction in Arden ratio prior to the gradual loss of full-field responses. Microperimetry demonstrated an enlarging scotoma in the proband. CONCLUSIONS: The coexistence of the pathogenic BEST1 c.37C > T variant may modify clinical features observed in RPE65 adRD. This study expands our understanding of RPE65 adRD as a retinoid cycle disorder supported by the reduced qAF, fine white retinal dots and corresponding subretinal deposits on OCT in affected members.

CLASSIFYING ABCA4 MUTATION SEVERITY USING AGE-DEPENDENT ULTRA-WIDEFIELD FUNDUS AUTOFLUORESCENCE-DERIVED TOTAL LESION SIZE.

PURPOSE: To establish a mutation-specific age-dependent ultra-widefield fundus autofluorescence (UWF-FAF) trajectory in a large Stargardt disease (STGD1) cohort using total lesion size (TLS) and to develop a clinical method for variant classification. METHODS: A retrospective study of patients with biallelic ABCA4 mutations that were evaluated with UWF-FAF. Boundaries of TLS, defined by stippled hyper/hypoautofluorescence, were outlined manually. Pathogenicity was assessed according to ACMG/AMP criteria, and mutation severities were classified based on the current literature. Age-dependent trajectories in TLS were examined in patients with nullizygous, mild, and intermediate mutations. Mutations of uncertain severities were classified using a clinical criterion based on age of symptom onset and TLS. RESULTS: Eighty-one patients with STGD1 (mean age = 42 ± 20 years and mean visual acuity = 20/200) were recruited from 65 unrelated families. Patients with biallelic null/severe variants (n = 6) demonstrated an increase in TLS during their second decade reaching a mean ± SD of 796 ± 29 mm2 by age 40. Those harboring mild mutations c.5882G>A or c.5603A>T had lesions confined to the posterior pole with a mean ± SD TLS of 30 ± 39 mm2. Intermediate mutations c.6079C>T or c.[2588G>C;5603A>T] in trans with a null/severe mutation had a mean ± SD TLS of 397 ± 29 mm2. Thirty-two mutations were predicted to cause severe (n = 22), intermediate (n = 6), and mild (n = 5) impairment of ABCA4 function based on age of symptom onset and TLS. CONCLUSION: Age-dependent TLS showed unique ABCA4 mutation-specific trajectories. Our novel clinical criterion using age of symptom onset and TLS to segregate ABCA4 mutations into three severity groups requires further molecular studies to confirm its validity.

Retinal pigment epithelium and age-related macular degeneration: A review of major disease mechanisms.

Age-related macular degeneration (AMD) is a progressive degenerative disease that is the leading cause of vision loss in the elderly population. Degeneration/dysregulation of the retinal pigment epithelium (RPE), a supportive monolayer of cells underlying the photoreceptors, is commonly seen in patients with AMD. While treatment exists for the neovascular/wet form of AMD, there is currently no cure for the non-exudative/dry form of AMD, making it imperative to understand the pathogenesis of this disease. Although our understanding of the aetiology of AMD has increased over the years, the underlying disease mechanism has not yet been identified, mainly due to the multifactorial nature of this disease. Herein, we review some of the commonly proposed degeneration pathways of RPE cells and their role in the pathogenesis of AMD; including activation of the complement cascade, oxidative stress-induced cell death mechanisms, dysfunctional mitochondria and the role of crystallins in AMD disease progression.

Effect of Ciliary Neurotrophic Factor on Retinal Neurodegeneration in Patients with Macular Telangiectasia Type 2: A Randomized Clinical Trial.

PURPOSE: To test the effects of an encapsulated cell-based delivery of a neuroprotective agent, ciliary neurotrophic factor (CNTF), on progression of macular telangiectasia type 2, a neurodegenerative disease with no proven effective therapy. DESIGN: Randomized sham-controlled clinical trial. PARTICIPANTS: Ninety-nine study eyes of 67 eligible participants were enrolled. METHODS: Single-masked randomized clinical trial of 24 months' duration conducted from May 2014 through April 2017 in 11 clinical centers of retinal specialists in the United States and Australia. Participants were randomized 1:1 to surgical implantation of intravitreal sustained delivery of human CNTF versus a sham procedure. MAIN OUTCOME MEASURES: The primary outcome was the difference in the area of neurodegeneration as measured in the area of the ellipsoid zone disruption (or photoreceptor loss) measured on spectral-domain (SD) OCT images at 24 months from baseline between the treated and untreated groups. Secondary outcomes included comparison of visual function changes between treatment groups. RESULTS: Among the 67 participants who were randomized (mean age, 62±8.9 years; 41 women [61%]; 58 white persons [86%]), 65 (97%) completed the study. Two participants (3 study eyes) died and 3 participants (4 eyes) were found ineligible. The eyes receiving sham treatment had 31% greater progression of neurodegeneration than the CNTF-treated eyes. The difference in mean area of photoreceptor loss was 0.05±0.03 mm2 (P = 0.04) at 24 months. Retinal sensitivity changes, measured using microperimetry, were correlated highly with the changes in the area of photoreceptor loss (r = 0.86; P < 0.0001). The mean retinal sensitivity loss of the sham group was 45% greater than that of the treated group (decrease, 15.81±8.93 dB; P = 0.07). Reading speed deteriorated in the sham group (-13.9 words per minute) with no loss in the treated group (P = 0.02). Serious adverse ocular effects were found in 2 of 51 persons (4%) in the sham group and 2 of 48 persons (4%) in the treated group. CONCLUSIONS: In participants with macular telangiectasia type 2, a surgical implant that released CNTF into the vitreous cavity, compared with a sham procedure, slowed the progression of retinal degeneration. Further research is needed to assess longer-term clinical outcomes and safety.

Gene therapy with recombinant adeno-associated vectors for neovascular age-related macular degeneration: 1 year follow-up of a phase 1 randomised clinical trial.

BACKGROUND: Neovascular, or wet, age-related macular degeneration causes central vision loss and represents a major health problem in elderly people, and is currently treated with frequent intraocular injections of anti-VEGF protein. Gene therapy might enable long-term anti-VEGF therapy from a single treatment. We tested the safety of rAAV.sFLT-1 in treatment of wet age-related macular degeneration with a single subretinal injection. METHODS: In this single-centre, phase 1, randomised controlled trial, we enrolled patients with wet age-related macular degeneration at the Lions Eye Institute and the Sir Charles Gairdner Hospital (Nedlands, WA, Australia). Eligible patients had to be aged 65 years or older, have age-related macular degeneration secondary to active subfoveal choroidal neovascularisation, with best corrected visual acuity (BCVA) of 3/60-6/24 and 6/60 or better in the other eye. Patients were randomly assigned (3:1) to receive either 1 × 10(10) vector genomes (vg; low-dose rAAV.sFLT-1 group) or 1 × 10(11) vg (high-dose rAAV.sFLT-1 group), or no gene-therapy treatment (control group). Randomisation was done by sequential group assignment. All patients and investigators were unmasked. Staff doing the assessments were masked to the study group at study visits. All patients received ranibizumab at baseline and week 4, and rescue treatment during follow-up based on prespecified criteria including BCVA measured on the Early Treatment Diabetic Retinopathy Study (EDTRS) scale, optical coherence tomography, and fluorescein angiography. The primary endpoint was ocular and systemic safety. This trial is registered with ClinicalTrials.gov, number NCT01494805. FINDINGS: From Dec 16, 2011, to April 5, 2012, we enrolled nine patients of whom eight were randomly assigned to receive either intervention (three patients in the low-dose rAAV.sFLT-1 group and three patients in the high-dose rAAV.sFLT-1 group) or no treatment (two patients in the control group). Subretinal injection of rAAV.sFLT-1 was highly reproducible. No drug-related adverse events were noted; procedure-related adverse events (subconjunctival or subretinal haemorrhage and mild cell debris in the anterior vitreous) were generally mild and self-resolving. There was no evidence of chorioretinal atrophy. Clinical laboratory assessments generally remained unchanged from baseline. Four (67%) of six patients in the treatment group required zero rescue injections, and the other two (33%) required only one rescue injection each. INTERPRETATION: rAAV.sFLT-1 was safe and well tolerated. These results support ocular gene therapy as a potential long-term treatment option for wet age-related macular degeneration. FUNDING: National Health and Medical Research Council of Australia, Richard Pearce Bequest, Lions Save Sight Foundation, Brian King Fellowship, and Avalanche Biotechnologies, Inc.

Angiographic biomarkers are significant predictors of treatment response to intravitreal aflibercept in diabetic macular edema.

This prospective single-center study aims to identify biomarkers that predict improvement in best-corrected visual acuity (BCVA) and central retinal thickness (CRT) at 6 months, in 76 eyes with diabetic macular edema (DME) treated monthly with intravitreal aflibercept. At baseline, all patients underwent standardized imaging with color photography, optical coherence tomography (OCT), fluorescein angiography (FA) and OCT angiography (OCTA). Glycosylated hemoglobin, renal function, dyslipidemia, hypertension, cardiovascular disease and smoking were recorded. Retinal images were graded in a masked fashion. Baseline imaging, systemic and demographic variables were investigated to detect associations to BCVA and CRT change post aflibercept. Predictors of BCVA improvement included greater macular vessel density quantified using OCTA (p = 0.001) and low-density lipoprotein (LDL) ≥ 2.6 mmol/L (p = 0.017). Lower macular vessel density eyes showed a significant reduction in CRT but no BCVA improvement. Predictors of CRT reduction included peripheral non-perfusion seen on ultrawide-field FA (p = 0.005) and LDL ≥ 2.6 mmol/L (p < 0.001). Retinal angiographic biomarkers derived from OCTA and ultrawide-field FA may help predict functional and anatomic response to anti-vascular endothelial growth factor (VEGF) therapy in patients with DME. Elevated LDL is associated with treatment response in DME. These results may be used to better-select patients who will benefit from intravitreal aflibercept for treatment of DME.

Longitudinal Analysis of Functional and Structural Outcome Measures in PRPH2-Associated Retinal Dystrophy.

PURPOSE: To establish disease progression rates in total lesion size (TLS), decreased autofluorescence (DAF) area, total macular volume (TMV), and mean macular sensitivity (MMS) in PRPH2-associated retinal dystrophy. DESIGN: Single-center, retrospective chart review. PARTICIPANTS: Patients with heterozygous pathogenic or likely pathogenic PRPH2 variants. METHODS: Patients who underwent serial ultrawide-field (UWF) fundus autofluorescence (FAF), OCT, and Macular Integrity Assessment microperimetry with at least 1 year of follow-up were included. Linear correlation was performed in eyes of all patients to determine the rate of change over time. MAIN OUTCOME MEASURES: Outcome measures included changes in TLS, DAF area, TMV, and MMS. RESULTS: Twelve patients (mean age, 55) from 10 unrelated families attended 100 clinic visits, which spanned over a mean (SD) of 4.7 (2.0) years. Mean (SD) TLS and DAF radius expansion were 0.14 (0.12) and 0.10 (0.08) mm/year, respectively. Mean (SD) TMV change was -0.071 (0.040) mm3/year with no interocular difference (P = 0.20) and strong interocular correlation (r2 = 0.88, P < 0.01). Mean (SD) MMS change was -0.10 (1.25) dB/year. Mean macular sensitivity declined in 4 and improved in 6 patients. Mean macular sensitivity was subnormal despite a TMV within the normal range. CONCLUSIONS: Serial measurements of UWF-FAF-derived TLS and DAF showed slow expansion. Total macular volume might be a more sensitive measure than MMS in detecting disease progression.

Factors promoting success and influencing complications in laser-induced central vein bypass.

PURPOSE: To evaluate the factors influencing the successful creation of a laser-induced chorioretinal venous anastomosis (L-CRA) and those involved in the development of complications. DESIGN: Interventional cohort study. PARTICIPANTS: Fifty-five patients with a nonischemic central retinal vein occlusion (CRVO) who were randomized to receive an L-CRA from the total of 108 who completed the follow-up period of the Central Vein Bypass Study. METHODS: Patients who were randomized to L-CRA were followed up for an 18-month period. They were stratified in 2 sets of 2 cohorts: those who did or did not demonstrate an L-CRA and those who did or did not demonstrate neovascular complications at the site of the L-CRA. Subgroup analysis was performed to determine what factors influenced the creation of an L-CRA and the development of complications at each individual laser site. MAIN OUTCOME MEASURES: Identification of systemic and local ocular factors associated with increased success rates of L-CRA creation and those involved with an increased risk of neovascular complications. RESULTS: Younger age (P = 0.03), better baseline visual acuity (P = 0.04), and the absence of hypertension (P = 0.001) were systemic features associated with an increased chance of demonstrating a successful L-CRA at each site, whereas sex and duration of the CRVO were not. The position of the L-CRA site did not influence the outcome; however, evidence of rupture of the vein wall at the time of the attempt was associated with a higher chance of success (P = 0.008). Increased risk of neovascularization, which occurred at 12 sites in 10 eyes, was associated with higher central venous pressure before treatment (P = 0.03), prolonged fluorescein transit time (P = 0.0001), and the presence of some capillary nonperfusion (P = 0.01). CONCLUSIONS: Younger age, better baseline visual acuity, and the absence of hypertension were associated with an improved success rate, as was evidence of rupture of the vein wall. High baseline central venous pressure, prolonged fluorescein transit time, and the presence of any retinal ischemia were associated with a higher incidence of neovascular complications. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references.

Retinal video recording a new way to image and diagnose diabetic retinopathy.

PURPOSE: To validate the use of retinal video recording for diabetic retinopathy screening by comparing with standard retinal photography and slit-lamp examination. DESIGN: Evaluation of a new diagnostic technique. PARTICIPANTS: One hundred patients. METHODS: All fundus images were captured using standard retinal still photography (FF 450 plus; Carl Zeiss) and retinal video (EyeScan; Ophthalmic Imaging System, Sacramento, CA), followed by a gold standard slit-lamp biomicroscopy examination. All videos and still images were de-identified, randomized, and interpreted by 2 senior consultant ophthalmologists (M.L.T-K. and L.L.). Kappa statistics, sensitivity, and specificity for all the diabetic retinopathy signs and grades were calculated with reference to slit-lamp examination results as the gold standard. MAIN OUTCOME MEASURES: Sensitivity and specificity of video recording for detecting diabetic retinopathy signs and grades. RESULTS: The mean age (± standard deviation [SD]) of participants was 52.8 ± 15.1 years, mean duration of diabetes (± SD) was 13.7 ± 9.7 years, and the mean glycosylated hemoglobin level was 8.0 ± 1.7%. Compared with the gold standard slit-lamp examination results, the sensitivity and specificity of video recording for detecting the presence of any diabetic retinopathy was 93.8% and 99.2%, respectively (ophthalmologist 1), and 93.3% and 95.2%, respectively (ophthalmologist 2). In contrast, the sensitivity and specificity of retinal photography was 91.8% and 98.4%, respectively (ophthalmologist 1), and 92.1% and 96.8%, respectively (ophthalmologist 2), for detection of any diabetic retinopathy. Both imaging methods had 100% sensitivity and specificity in detecting sight-threatening diabetic retinopathy. For overall diabetic retinopathy grading by both ophthalmologists, the measurements of agreement (Cohen's κ coefficient) between the overall grading obtained from the retinal video versus slit-lamp examination and retinal photography versus slit-lamp examination were more than 0.90. Technical failure rate for retinal video recording and retinal photography were 7.0% and 5.5%, respectively. CONCLUSIONS: This study demonstrated that retinal video recording was equally as effective as retinal photography in the subjects evaluated in this study. It is a novel alternative diabetic retinopathy screening technique that not only offers primary eye care providers the opportunity to view numerous retinal fields within a short period but also is easy to learn by nonexperienced personnel with minimal training. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references.

ACUTE RETINAL NECROSIS ASSOCIATED WITH HERPES ZOSTER VACCINATION.

PURPOSE: To report a case of Zostavax-associated acute retinal necrosis in a patient with chronic lymphocytic leukemia. METHODS: Case report. PATIENTS: A 76-year-old white man. RESULTS: Unilateral acute retinal necrosis with obliterative angiopathy developed in close proximity of a Zostavax vaccine. Treatment with valacyclovir hydrochloride (1 g orally three times a day) and intravitreal ganciclovir (4 mg/0.1 mL) was initiated on presentation. Because of continuous increase of the retinal necrosis, patient was switched to intravenous acyclovir (7.5 mg/kg body weight, adapted to reduced glomerular filtration rate) and given intravitreal foscarnet (2.4 mg/0.1 mL). Despite being on maximal antiviral therapy, the patient suffered a central retinal artery occlusion. DISCUSSION: Acute retinal necrosis is a severe complication and potentially blinding disease of herpes zoster, and can occur in association with herpes zoster immunization, in particular, in immune suppressed patients.

Atrophy Expansion Rates in Stargardt Disease Using Ultra-Widefield Fundus Autofluorescence.

Purpose: To investigate atrophy expansion rate (ER) using ultra-widefield (UWF) fundus autofluorescence (FAF) in Stargardt disease (STGD1). Design: Retrospective, longitudinal study. Participants: Patients with biallelic ABCA4 mutations who were evaluated with UWF FAF and Heidelberg 30° × 30° and 55° × 55° FAF imaging. Methods: Patients with atrophy secondary to STGD1 were classified into genotype groups: group A, biallelic severe or null-like variants with early-onset disease; group B, 1 intermediate variant in trans with severe or null-like variant; and group C, 1 mild variant in trans with severe or null-like variant or late-onset disease. The boundaries of definitely decreased autofluorescence (DDAF) were outlined manually and areas (in square millimeters) were recorded at baseline and follow-up. Bland-Altman analysis was conducted to examine agreement between observers and devices. Linear mixed modeling was used to evaluate predictors of ER in DDAF area and square root area (SRA). Main Outcome Measures: Patient and ocular predictors of DDAF area ER and DDAF SRA ER included age at onset, duration of symptoms, genotype group, baseline visual acuity, and baseline atrophy size. Results: A total of 138 eyes from 69 patients (33 men [47%]; mean age ± standard deviation, 41 ± 20 years; range, 10-83 years) carrying 61 unique ABCA4 variants were recruited. Ultra-widefield FAF measurements were equivalent to Heidelberg 30° × 30° imaging. Baseline DDAF area was the only significant predictor of DDAF area ER (P < 0.001). Age at baseline and genotype group were predictors for DDAF SRA ER. Definitely decreased autofluorescence area ER ranged from 4.65 mm2/year (group A) to 0.62 mm2/year (group C). Conclusions: Ultra-widefield FAF is a feasible and reliable method for assessing atrophy ER in STGD1. The value of ABCA4 mutation severity in predicting atrophy ER warrants further investigation.

Genotype-Specific Lesion Growth Rates in Stargardt Disease.

Reported growth rates (GR) of atrophic lesions in Stargardt disease (STGD1) vary widely. In the present study, we report the longitudinal natural history of patients with confirmed biallelic ABCA4 mutations from five genotype groups: c.6079C>T, c.[2588G>C;5603A>T], c.3113C>T, c.5882G>A and c.5603A>T. Fundus autofluorescence (AF) 30° × 30° images were manually segmented for boundaries of definitely decreased autofluorescence (DDAF). The primary outcome was the effective radius GR across five genotype groups. The age of DDAF formation in each eye was calculated using the x-intercept of the DDAF effective radius against age. Discordance between age at DDAF formation and symptom onset was compared. A total of 75 eyes from 39 STGD1 patients (17 male [44%]; mean ± SD age 45 ± 19 years; range 21-86) were recruited. Patients with c.3113C>T or c.6079C>T had a significantly faster effective radius GR at 0.17 mm/year (95% CI 0.12 to 0.22; p < 0.001 and 0.14 to 0.21; p < 0.001) respectively, as compared to those patients harbouring c.5882G>A at 0.06 mm/year (95% CI 0.03-0.09), respectively. Future clinical trial design should consider the effect of genotype on the effective radius GR and the timing of DDAF formation relative to symptom onset.

Progressive sector retinitis pigmentosa due to c.440G>T mutation in SAG in an Australian family.

BACKGROUND: Heterozygous c.440 G > T mutation in the S-antigen visual arrestin (SAG) gene has been described as a cause of autosomal dominant retinitis pigmentosa (adRP) in a series of patients of Hispanic origin. This study presents the early and late clinical features and disease progression rates in an Australian family with SAG adRP. MATERIALS AND METHODS: An observational case series of four family members with adRP. They were examined clinically, with multi-modal retinal imaging and electroretinography (ERG) to ascertain phenotype. Disease progression rate was measured using optical coherence tomography (OCT) and fundus autofluorescence (FAF). A retinal dystrophy panel was used for the proband and cascade testing with targeted Sanger sequencing was conducted in other available family members. RESULTS: The proband presented at 36 years of age with profoundly reduced full-field ERG responses despite a sector RP phenotype. This progressed to a classic RP pattern over several decades leaving a small residual island of central visual field. The horizontal span of the residual outer nuclear layer and the area of hyperautofluorescent ring contracted at a rate of 8-11% and 9-14% per year, respectively. DNA sequencing confirmed the segregation of SAG c.440 G > T mutation with disease. CONCLUSION: SAG adRP presents with a reduced full-field ERG response consistent with a rod-cone dystrophy in mid-life despite a sector RP phenotype. Centripetal progression of the disease into the macula can be tracked by OCT and FAF imaging.

The Central Retinal Vein Bypass Study: a trial of laser-induced chorioretinal venous anastomosis for central retinal vein occlusion.

PURPOSE: To evaluate the effectiveness of a laser-induced chorioretinal venous anastomosis (L-CRA) as a treatment for nonischemic central retinal vein occlusion (CRVO). DESIGN: Prospective, randomized, controlled, multicenter clinical trial. PARTICIPANTS: A total of 113 consecutive patients with a nonischemic CRVO of >3 months' duration and visual acuity of < or =20/50. METHODS: Patients were randomized to L-CRA (58 patients) or conventional care (55 patients). They underwent standardized retinal photography, fluorescein angiography, and ophthalmic examinations, together with standardized assessments of best-corrected visual acuity, performed by masked visual acuity assessors using Early Treatment Diabetic Retinopathy Study protocols. Analysis was performed by intention-to-treat. MAIN OUTCOME MEASURES: The primary outcome measure was change in visual acuity at 18 months. Secondary outcomes were progression of retinal ischemia and rates of adverse events. RESULTS: A total of 53 control patients and 55 treatment patients completed the study. The 2 groups were comparable for age, age- and gender-adjusted mean visual acuity, and most other parameters. In the treated group of 55 patients, 42 (76.4%) developed an L-CRA. Over the 18-month follow-up period, treated eyes had an 8.3 letter mean improvement from baseline compared with control eyes (P = 0.03). Treated eyes that developed a functional L-CRA achieved an 11.7 letter mean improvement from baseline over the control group after 18 months (P = 0.004). Conversion to the ischemic CRVO category occurred in 20.8% of control eyes and in 9.6% of treated eyes overall (P = 0.33). Of the treated group who developed an L-CRA where the retinal ischemia was due to progression of the CRVO, 4.9% progressed to the ischemic category (P = 0.03). Neovascularization developed at the site of the L-CRA in 10 of 55 treated eyes (18.2%). Vitrectomy surgery was required by 5 of 55 treated eyes (9.1%) because of macular traction or nonresolving vitreous hemorrhage. CONCLUSIONS: Chorioretinal venous anastomosis was created in 76.4% of eyes with nonischemic CRVO in this study. Eyes that developed an anastomosis had a significant improvement (11.7 letters) in final visual acuity after 18 months, compared with eyes in the control group (P = 0.004). Complications were managed successfully with careful follow-up and early intervention.

Edge of Scotoma Sensitivity as a Microperimetry Clinical Trial End Point in USH2A Retinopathy.

Purpose: Microperimetry is commonly used to assess retinal function. We perform cross-sectional and longitudinal analysis on microperimetry parameters in USH2A retinopathy and explore end points suitable for future clinical trials. Methods: Microperimetry was performed using two grids, Grid 1 (18° diameter) and Grid 2 (6° diameter). In Grid 1, four parameters (number of nonscotomatous loci, mean sensitivity [MS], responding point sensitivity [RPS], and edge of scotoma sensitivity [ESS]) were analyzed. In Grid 2, number of nonscotomatous loci and MS were examined. Interocular symmetry was also examined. Longitudinal analysis was conducted in a subset of eyes. Results: Microperimetry could be performed in 16 of 21 patients. In Grid 1 (n = 15; average age, 35.6 years), average number of nonscotomatous loci, MS, RPS, and ESS were 46.6 loci, 10.0 dB, 14.7 and 9.6 dB, respectively. In Grid 2 (n = 13; average age, 37.4 years), 12 eyes had measurable sensitivity across the entire grid. Average MS was 23.8 dB. Interocular analysis revealed large 95% confidence intervals for all parameters. Longitudinally, Grid 1 (n = 12, average follow-up 2.6 years) ESS showed the fastest rate of decline (-1.84 dB/y) compared with MS (-0.34 dB/y) and RPS (-0.90 dB/y). Conclusions: Our data suggest that ESS may be more useful than MS and RPS in test grids that cover a large extent of the macula. We caution the use of contralateral eye as an internal control. Translational Relevance: ESS may decrease the duration or sample size of treatment trials in USH2A retinopathy.

Phenotype-genotype correlations in a pseudodominant Stargardt disease pedigree due to a novel ABCA4 deletion-insertion variant causing a splicing defect.

BACKGROUND: Deletion-insertion (delins) variants in the retina-specific ATP-binding cassette transporter gene, subfamily A, member 4 (ABCA4) accounts for <1% in Stargardt disease. The consequences of these delins variants on splicing cannot be predicted with certainty without supporting in vitro data. METHODS: Candidate ABCA4 variants were revealed by genetic and segregation analysis of a family with pseudodominant Stargardt disease using a commercial panel and Sanger sequencing. RNA extracted from patient-derived fibroblasts was analyzed by RT-PCR to evaluate splicing behavior of the ABCA4 variants. RESULTS: Affected members carrying the novel c.6031_6044delinsAGTATTTAACCAATATTT variant in exon 44 presented with contrasting phenotypes; from early-onset cone-rod dystrophy to late-onset macular dystrophy. This variant resulted in a 56-nucleotide deletion in the mutant allele by activation of a cryptic splice acceptor site which disrupts the reading frame and results in a premature termination codon (p.Ile2003LeufsTer41). If translated, the crucial functional domains near the C-terminus would be truncated from the ABCA4 protein. CONCLUSION: This work demonstrates the intrafamilial phenotypic variability in a pseudodominant Stargardt disease pedigree and the use of patient-derived fibroblasts to evaluate the effect of a novel ABCA4 delins variant on splicing to complement in silico pathogenicity assessment.

Complement factor H Y402H and C-reactive protein polymorphism and photodynamic therapy response in age-related macular degeneration.

PURPOSE: To investigate the association between complement factor H (CFH) and C-reactive protein (CRP) genotypes and response to photodynamic therapy (PDT) treatment for neovascular age-related macular degeneration (AMD). DESIGN: Retrospective cohort study. PARTICIPANTS: The study cohort consisted of 273 neovascular AMD patients treated with PDT. METHODS: Genotypes were determined for the common T-->C single nucleotide polymorphism (SNP) in exon 9 of the CFH gene (rs1061170; Y402H), as well as 9 selected tagging SNPs across the CRP gene (rs2808635, rs1417938, rs1800947, rs1130864, rs1205, rs3093077, rs876538, rs876537, and rs1572970). Visual acuity outcome after PDT was retrospectively calculated and patients were classified as PDT-positive responders or PDT-negative responders. Logistic regression analysis was used to evaluate the association between individual SNPs and PDT treatment response while adjusting for relevant covariates. MAIN OUTCOME MEASURES: Response to PDT treatment defined by visual acuity; genotypes of CFH Y402H and CRP polymorphism. RESULTS: Of the 273 patients, 75 had a positive response after PDT treatment. The frequency of CC genotype of the CFH Y402H polymorphism in the PDT-positive response group was lower than in the negative PDT response group (26.4% vs 31.6%) but this difference failed to reach statistical significance. Two CRP SNPs (rs2808635 and rs876538) were significantly associated with PDT treatment response. Positive treatment response was seen in individuals homozygous for the minor allele of the rs2808635 (GG; odds ratio [OR], 3.92; 95% confidence interval [CI], 1.40-10.97; P = 0.048) and rs876538 (AA; OR, 6.49; 95% CI, 1.65-25.47; P = 0.048) variants after adjusting for relevant covariates. The remaining 7 CRP genetic variants did not reveal any significant association with treatment response. CONCLUSIONS: Our data did not show significant association between the CFH Y402H polymorphism and PDT treatment response for neovascular AMD; however, CRP genetic variants were associated with a positive response to PDT treatment for neovascular AMD.

Three-Year Follow-Up of Phase 1 and 2a rAAV.sFLT-1 Subretinal Gene Therapy Trials for Exudative Age-Related Macular Degeneration.

PURPOSE: To assess the safety and the 3-year results of combined phase 1 and 2a randomized controlled trials of rAAV.sFLT-1 gene therapy (GT) for wet age-related macular degeneration. DESIGN: Phase 1/2a clinical trial. METHODS: Patients were prospectively randomized into control (n = 13) and GT (n = 24) groups. GT patients received 1X1011vg rAAV.sFLT-1 and were seen every month for 1 year then as needed every 1 to 2 months. They were given retreatment anti-vascular endothelial growth factor injections according to predetermined criteria. At 12 months, GT patients were divided into 2 groups: HD-1 (n = 14), requiring <2, and HD-2 (n = 10), requiring >2 retreatments. RESULTS: Between 1 year and 3 years there were 3 adverse events (AEs) and 33 serious AEs reported. Of these, 15 occurred in the 13 control subjects and 21 in the 24 GT patients. Except for 1 case of transient choroiditis in a control patient, serious AEs were deemed to be unrelated to the study. Control patients received a median of 7.0 retreatments and lost a median of 7.0 Early Treatment Diabetic Retinopathy Study (ETDRS) letters, HD-1 patients received a median of 2.5 retreatments and lost a median of 4.0 ETDRS letters, and HD-2 patients received a median of 11.0 retreatments and lost a median of 7.0 ETDRS letters over 3 years. Center point thickness fluctuated. Thirty-three percent of control subjects, 44% of HD-2 patients, and 51% of HD-1 patients showed maintenance of baseline visual acuity. Four HD-1 patients (34%) maintained significant visual improvement at 3 years. None of these observations were statistically significant. CONCLUSIONS: Given the small number of patients, this study was unable to unequivocally confirm the existence of a biologic efficacy signal; however, it confirmed that rAAV.sFLT-1 gene delivery was well tolerated among the elderly.

Surgical Removal of Internal Limiting Membrane and Layering of AAV Vector on the Retina Under Air Enhances Gene Transfection in a Nonhuman Primate.

Purpose: To determine if the surgical removal of the internal limiting membrane (ILM) in nonhuman primates (NHPs) will result in safe and effective transfection of adeno-associated viral (AAV2) vectors using green fluorescent protein (GFP) as a reporter. Methods: Six Macaca fascicularis NHP eyes underwent vitrectomy, ILM peel with layering of 1.7 × 1013 genome copies per milliliter of AAV2-GFP under air. Four control eyes underwent only vitrectomy and pooling under air. The intensity and area transfected was quantified in vivo with fundus autofluorescence (FAF) imaging. NHPs were euthanized 16 weeks postsurgery and immunohistochemical analysis assessed GFP expression at the cellular level. Results: There was a larger area of fluorescence in ILM peeled eyes then in non-ILM peeled eyes (50.7 [33.1-58.4] pixel2 versus 5.1 [0.6-7.6] pixel2, P < 0.01). The intensity of fluorescence was also higher in ILM peeled eyes (10.3 [2.2-18.5] vs. 1.9 [0.6-4.4], P = 0.05). Non-ILM peeled eyes displayed fluorescence confined to the foveal center. Histological sections showed colocalization in the Müller cell layer, ganglion cell layer, and photoreceptor cell layer in the ILM peeled eyes. In non-ILM peeled eyes GFP expression was only in the ganglion cell layer in three eyes and was confined to the immediate vicinity of the fovea. Conclusions: ILM appears to be the predominate barrier to AAV transfection. An efficacious and safe method of AAV2 gene delivery, taking into account the potential need for repeat treatments, appears to be the surgical removal of ILM and layering of AAV under air.v.