Retinal Dystrophies Associated With Peripherin-2: Genetic Spectrum and Novel Clinical Observations in 241 Patients.

Purpose: To describe the clinical, electrophysiological and genetic spectrum of inherited retinal diseases associated with variants in the PRPH2 gene. Methods: A total of 241 patients from 168 families across 15 sites in 9 countries with pathogenic or likely pathogenic variants in PRPH2 were included. Records were reviewed for age at symptom onset, visual acuity, full-field ERG, fundus colour photography, fundus autofluorescence (FAF), and SD-OCT. Images were graded into six phenotypes. Statistical analyses were performed to determine genotype-phenotype correlations. Results: The median age at symptom onset was 40 years (range, 4-78 years). FAF phenotypes included normal (5%), butterfly pattern dystrophy, or vitelliform macular dystrophy (11%), central areolar choroidal dystrophy (28%), pseudo-Stargardt pattern dystrophy (41%), and retinitis pigmentosa (25%). Symptom onset was earlier in retinitis pigmentosa as compared with pseudo-Stargardt pattern dystrophy (34 vs 44 years; P = 0.004). The median visual acuity was 0.18 logMAR (interquartile range, 0-0.54 logMAR) and 0.18 logMAR (interquartile range 0-0.42 logMAR) in the right and left eyes, respectively. ERG showed a significantly reduced amplitude across all components (P < 0.001) and a peak time delay in the light-adapted 30-Hz flicker and single-flash b-wave (P < 0.001). Twenty-two variants were novel. The central areolar choroidal dystrophy phenotype was associated with 13 missense variants. The remaining variants showed marked phenotypic variability. Conclusions: We described six distinct FAF phenotypes associated with variants in the PRPH2 gene. One FAF phenotype may have multiple ERG phenotypes, demonstrating a discordance between structure and function. Given the vast spectrum of PRPH2 disease our findings are useful for future clinical trials.

A multinational survey of potential participant perspectives on ocular gene therapy.

Amidst rapid advancements in ocular gene therapy, understanding patient perspectives is crucial for shaping future treatment choices and research directions. This international cross-sectional survey evaluated knowledge, attitudes, and perceptions of ocular genetic therapies among potential recipients with inherited retinal diseases (IRDs). Survey instruments included the Attitudes to Gene Therapy-Eye (AGT-Eye), EQ-5D-5L, National Eye Institute Visual Functioning Questionnaire (NEI-VFQ-25), and Patient Attitudes to Clinical Trials (PACT-22) instruments. This study included 496 participant responses (89% adults with IRDs; 11% parents/guardians/carers) from 35 countries, with most from the United States of America (USA; 69%) and the United Kingdom (11%). Most participants (90%) indicated they would likely accept gene therapy if it was available, despite only 45% agreeing that they had good knowledge of gene therapy. The main sources of information were research registries (60% of participants) and the internet (61%). Compared to data from our recently published Australian national survey of people with IRDs (n = 694), USA respondents had higher knowledge of gene therapy outcomes, and Australian respondents indicated a higher perceived value of gene therapy treatments. Addressing knowledge gaps regarding outcomes and financial implications will be central to ensuring informed consent, promoting shared decision-making, and the eventual clinical adoption of genetic therapies.

Natural history and biomarkers of KCNV2-associated retinopathy.

BACKGROUND: KCNV2-associated retinopathy is an autosomal recessive inherited retinal disease classically named cone dystrophy with supernormal rod response (CDSRR). This study aims to identify the best biomarker for evaluating the condition. METHODS: A retrospective review of eight patients from seven families with genetically confirmed KCNV2-associated retinopathy was performed. The best corrected visual acuity (BCVA), full-field electroretinogram (ffERG), pattern ERG (pERG), fundus imaging: retinal photograph and fundus autofluorescence (FAF), and optical coherence tomography (OCT) were analysed. RESULTS: There was a disproportionate increase in b-wave amplitude with a relatively small light intensity increase, especially between the two dimmest stimuli of DA 0.002 and 0.01 (-2.7 and -2.0 log cd.s/m2). The a-wave amplitude was normal. The a-wave peak time was delayed in all stimuli. The b-wave peak time was delayed compared to normal, but the gap tightened as intensity increased. The b:a wave ratio was above or at the upper limit for the reference values. FAF bull's eye maculopathy pattern was prominent and variable foveal disruption on OCT was apparent in all patients. Legal blindness was reached before the age of 25. CONCLUSIONS: We identified three potential electrophysiology biomarkers to assist in evaluating future therapies: the disproportionate b-wave amplitude jump, delayed a-wave and b-wave peak time, and the higher than normal b:a wave ratio. Any of these biomarkers found with photoreceptor ellipsoid zone foveal-perifoveal disruption should prompt consideration for KCNV2 retinopathy. The BCVA natural history data suggests the probable optimum therapeutic window in the first three decades of life.

EQ-5D-5L health utility scores in Australian adults with inherited retinal diseases: A cross-sectional survey.

PURPOSE: Economic evaluations of interventions for ocular disease require utility scores that accurately represent quality of life in the target population. This study aimed to describe the distribution of EQ-5D-5L utility values among Australian adults with symptomatic inherited retinal diseases (IRDs) and to assess the relationship between these scores and vision-related quality of life. METHODS: A survey was administered predominantly online in 2021. Participants completed the EQ-5D-5L general health utility instrument, the EQ vertical visual analogue scale (EQ-VAS) and the National Eye Institute Visual Functioning Questionnaire (NEI-VFQ-25). Self-reported IRD diagnoses were classified as being associated with central or widespread retinal involvement. RESULTS: Responses from 647 participants aged 18-93 years were included, 50.1% were men and 77.6% had an IRD associated with widespread retinal involvement. The majority reported no problems with self-care and no pain/discomfort but did report anxiety/depression and problems with work, study, housework, or family/leisure activities. Most people with widespread involvement reported problems with mobility. Median EQ-5D-5L utility was 0.88 and 0.91 among people with widespread and central involvement, respectively (age and sex-adjusted p = 0.029); and median EQ-VAS was 75 and 80, respectively (adjusted p = 0.003). A moderate curvilinear correlation was observed between EQ-5D-5L and NEI-VFQ-25 composite score (Spearman's ρ 0.69), but not all people with poor vision-related quality of life had low EQ-5D-5L utility values. CONCLUSIONS: EQ-5D-5L health utility values are correlated with vision-related quality of life among adults with IRDs. However, the EQ-5D-5L may not be sensitive to the full impact of vision impairment on quality of life.

Managing the apparently blind child presenting in the first year of life: A review.

Severe vision impairment and blindness in childhood have a significant health burden on the child, family and society. This review article seeks to provide a structured framework for managing the apparently blind child presenting in the first year of life, starting from a comprehensive history and examination. Different investigation modalities and the increasingly important role of genetics will also be described, in addition to common causes of severe vision impairment. Crucially, a systematic approach to the blind infant is key to correct diagnoses and timely management. Incorrect diagnoses can be costly to all involved, however it is important to note that diagnoses can change with ongoing follow-up and investigations. Furthermore, the modern age of ophthalmology requires a multi-disciplinary approach and close collaboration with specialists including paediatricians, neurologists and geneticists, in addition to rehabilitation and low vision services, to ensure the best care for these vulnerable infants.

Suspected case of benign familial fleck retina with functional loss.

Key Clinical Message: Inherited retinal dystrophies typically affect vision in early childhood; however, this case highlights a late onset retinal dystrophy presenting in midlife and the need for extended visual electrophysiology testing to determine the etiology. Abstract: A 53-year-old female was referred for visual electrophysiology following a routine optometric eye examination in which yellow flecks were noted in both fundi and the patient had reported a recent near accident whilst driving at night. There was no reported family history of eye disease. Retinal examination identified bilateral yellow punctate and irregularly shaped lesions throughout the posterior poles sparing the macula region. Fundus autofluorescence showed coinciding hyperfluorescence with the lesions and bilateral hypofluorescent crescents superior to the macular with corresponding retinal thinning. Visual fields and color vision were normal. ISCEV standard 20 min and extended 60-min dark adapted electroretinograms were recorded. Recovery to normal b-wave amplitudes was noted in the DA0.01 flash but reduced a-wave amplitudes were noted in the DA3 and DA10 flash following both dark adapted periods. Cone function was reduced but within normal limits. Genetic screening revealed a previously unreported variant of unknown significance in the gene PLA2G5:c.40 + 5del (rs1364254561) which is a member of the phospholipase A2 family and is associated with familial benign flecked retina.

A natural history study of autosomal dominant GUCY2D-associated cone-rod dystrophy.

PURPOSE: To describe the natural history of autosomal dominant (AD) GUCY2D-associated cone-rod dystrophies (CRDs), and evaluate associated structural and functional biomarkers. METHODS: Retrospective analysis was conducted on 16 patients with AD GUCY2D-CRDs across two sites. Assessments included central macular thickness (CMT) and length of disruption to the ellipsoid zone (EZ) via optical coherence tomography (OCT), electroretinography (ERG) parameters, best corrected visual acuity (BCVA), and fundus autofluorescence (FAF). RESULTS: At first visit, with a mean age of 30 years (range 5-70 years), 12 patients had a BCVA below Australian driving standard (LogMAR ≥ 0.3 bilaterally), and 1 patient was legally blind (LogMAR ≥ 1). Longitudinal analysis demonstrated a deterioration of LogMAR by - 0.019 per year (p < 0.001). This accompanied a reduction in CMT of - 1.4 µm per year (p < 0.0001), lengthened EZ disruption by 42 µm per year (p = < 0.0001) and increased area of FAF by 0.05 mm2 per year (p = 0.027). Similarly, cone function decreased with increasing age, as demonstrated by decreasing b-wave amplitude of the light-adapted 30 Hz flicker and fused flicker (p = 0.005 and p = 0.018, respectively). Reduction in CMT and increased EZ disruption on OCT were associated with functional changes including poorer BCVA and decreased cone function on ERG. CONCLUSION: We have described the natural long-term decline in vision and cone function associated with mutations in GUCY2D and identified a set of functional and structural biomarkers that may be useful as outcome parameters for future therapeutic clinical trials.

Prevalence of Glaucoma Following Paediatric Cataract Surgery in an Australian Tertiary Referral Centre.

Purpose: Secondary glaucoma following childhood cataract surgery remains the most common complication in the paediatric population. This study aimed to determine the incidence, time to progression and risk factors associated with the development of secondary glaucoma following childhood cataract surgery in a paediatric population. Outcome measures were the detection of secondary glaucoma, postoperative time frame to development of glaucoma and risk factors in its development. Patients and Methods: A retrospective case series was conducted between 2003 and 2017 at a tertiary children's hospital in Sydney. The patient population included those 16 years or less of age who underwent congenital cataract extraction, with or without an intraocular lens implantation and who had been followed up for a minimum of six months following surgery. Patients were excluded if they had cataract aetiology other than congenital idiopathic cataract. Multivariate Cox Regression analysis was used to determine relevant risk factors. Results: A total of 320 eyes in 216 patients were included in the study. Secondary glaucoma developed in 11.9% of eyes. In those that developed secondary glaucoma, the average time to onset from surgery was 3.2 years (median 2.75 years). The mean age of diagnosis of secondary glaucoma was 4.58 years (median 3.5 years, range 2.5 months to 13.23 years). Microcornea was the only adverse characteristic significantly associated with an increased risk of secondary glaucoma (HR 6.30, p 0.003). Conclusion: Despite modern surgical techniques, glaucoma remains a significant long-term sequela in children following cataract surgery.

The health care and societal costs of inherited retinal diseases in Australia: a microsimulation modelling study.

OBJECTIVES: To estimate the health care and societal costs of inherited retinal diseases (IRDs) in Australia. DESIGN, SETTING, PARTICIPANTS: Microsimulation modelling study based on primary data - collected in interviews of people with IRDs who had ophthalmic or genetic consultations at the Children's Hospital at Westmead or the Save Sight Institute (both Sydney) during 1 January 2019 - 31 December 2020, and of their carers and spouses - and linked Medicare Benefits Schedule (MBS) and Pharmaceutical Benefits Schedule (PBS) data. MAIN OUTCOME MEASURES: Annual and lifetime costs for people with IRDs and for their carers and spouses, grouped by payer (Australian government, state governments, individuals, private health insurance) and type (health care costs; societal costs: social support, National Disability Insurance Scheme (NDIS), income and taxation, costs associated with caring for family members with IRDs); estimated annual national cost of IRDs. RESULTS: Ninety-four people (74 adults, 20 people under 18 years; 55 girls and women [59%]) and 30 carers completed study surveys (participation rate: adults, 66%; children, 66%; carers, 63%). Total estimated lifetime cost was $5.2 million per person with an IRD, of which 87% were societal and 13% health care costs. The three highest cost items were lost income for people with IRDs ($1.4 million), lost income for their carers and spouses ($1.1 million), and social spending by the Australian government (excluding NDIS expenses: $1.0 million). Annual costs were twice as high for people who were legally blind as for those with less impaired vision ($83 910 v $41 357 per person). The estimated total annual cost of IRDs in Australia was $781 million to $1.56 billion. CONCLUSION: As the societal costs associated with IRDs are much larger than the health care costs, both contributors should be considered when assessing the cost-effectiveness of interventions for people with IRDs. The increasing loss of income across life reflects the impact of IRDs on employment and career opportunities.

IMPDH1-associated autosomal dominant retinitis pigmentosa: natural history of novel variant Lys314Gln and a comprehensive literature search.

BACKGROUND: Inosine monophosphate dehydrogenase (IMPDH) is a key regulatory enzyme in the de novo synthesis of the purine base guanine. Mutations in the inosine monophosphate dehydrogenase 1 gene (IMPDH1) are causative for RP10 autosomal dominant retinitis pigmentosa (adRP). This study reports a novel variant in a family with IMPDH1-associated retinopathy. We also performed a comprehensive review of all reported IMPDH1 disease causing variants with their associated phenotype. MATERIALS AND METHODS: Multimodal imaging and functional studies documented the phenotype including best-corrected visual acuity (BCVA), fundus photograph, fundus autofluorescence (FAF), full field electroretinogram (ffERG), optical coherence tomography (OCT) and visual field (VF) data were collected. A literature search was performed in the PubMed and LOVD repositories. RESULTS: We report 3 cases from a 2-generation family with a novel heterozygous likely pathogenic variant p. (Lys314Gln) (exon 10). The ophthalmic phenotype showed diffuse outer retinal atrophy with mild pigmentary changes with sparse pigmentary changes. FAF showed early macular involvement with macular hyperautofluorescence (hyperAF) surrounded by hypoAF. Foveal ellipsoid zone island can be found in the youngest patient but not in the older ones. The literature review identified a further 56 heterozygous, 1 compound heterozygous, and 2 homozygous variant. The heterozygous group included 43 missense, 3 in-frame, 1 nonsense, 2 frameshift, 1 synonymous, and 6 intronic variants. Exon 10 was noted as a hotspot harboring 18 variants. CONCLUSIONS: We report a novel IMPDH1 variant. IMPDH1-associated retinopathy presents most frequently in the first decade of life with early macular involvement.

Spectrum of ocular disease in children aged between 0 and 3 years at an Australian paediatric tertiary hospital.

BACKGROUND: Childhood ocular disease can be a significant health burden to the child, family and society. Previous studies have examined the spectrum of paediatric ocular disease presenting to tertiary hospitals; however, these studies have broader age ranges, smaller sample sizes, and are largely based in developing countries. This study aims to assess the spectrum of ocular disease in the first 3 years of life presenting to the eye department of an Australian tertiary paediatric hospital. METHODS: The records of 3337 children who had their initial presentation at the eye clinic between the age of 0 and 36 months were reviewed, spanning 6.5 years from 1st July 2012 to 31st December 2018. RESULTS: The most common primary diagnoses overall were strabismic amblyopia (6.0%), retinopathy of prematurity (5.0%) and nasolacrimal duct obstruction (4.5%). Bilateral visual impairment was more common in younger children, while unilateral visual impairment was more common in older children. The proportion of all children presenting with visual impairment was 10.3%, with 5.7% of all children presenting with bilateral visual impairment and 4.6% presenting with unilateral visual impairment. In children with visual impairment, the most common sites of primary abnormality were lens (21.4%), retina (17.3%), and cerebral and visual pathways (12.1%). The most common primary diagnoses in children with visual impairment were cataract (21.4%), strabismic amblyopia (9.3%) and retinoblastoma (6.5%). CONCLUSIONS: The spectrum of eye disease and vision impairment presenting in the first 3 years of life facilitates health care planning, greater community education about vision impairment and importance of early intervention, and guidance for appropriate resource allocation. Health systems can apply these findings to aid in early identification and intervention to reduce preventable blindness and institute appropriate rehabilitation services.

Early recognition of CLN3 disease facilitated by visual electrophysiology and multimodal imaging.

BACKGROUND: Neuronal ceroid lipofuscinosis is a group of neurodegenerative disorders with varying visual dysfunction. CLN3 is a subtype which commonly presents with visual decline. Visual symptomatology can be indistinct making early diagnosis difficult. This study reports ocular biomarkers of CLN3 patients to assist clinicians in early diagnosis, disease monitoring, and future therapy. METHODS: Retrospective review of 5 confirmed CLN3 patients in our eye clinic. Best corrected visual acuity (BCVA), electroretinogram (ERG), ultra-widefield (UWF) fundus photography and fundus autofluorescence (FAF), and optical coherence tomography (OCT) studies were undertaken. RESULTS: Five unrelated children, 4 females and 1 male, with median age of 6.2 years (4.6-11.7) at first assessment were investigated at the clinic from 2016 to 2021. Four homozygous and one heterozygous pathogenic CLN3 variants were found. Best corrected visual acuities (BCVAs) ranged from 0.18 to 0.88 logMAR at first presentation. Electronegative ERGs were identified in all patients. Bull's eye maculopathies found in all patients. Hyper-autofluorescence ring surrounding hypo-autofluorescence fovea on FAF was found. Foveal ellipsoid zone (EZ) disruptions were found in all patients with additional inner and outer retinal microcystic changes in one patient. Neurological problems noted included autism, anxiety, motor dyspraxia, behavioural issue, and psychomotor regression. CONCLUSIONS: CLN3 patients presented at median age 6.2 years with visual decline. Early onset maculopathy with an electronegative ERG and variable cognitive and motor decline should prompt further investigations including neuropaediatric evaluation and genetic assessment for CLN3 disease. The structural parameters such as EZ and FAF will facilitate ocular monitoring.

Eye Abnormalities in Children with Fetal Alcohol Spectrum Disorders: A Systematic Review.

PURPOSE: Although eye abnormalities are reported in fetal alcohol spectrum disorders (FASD), no systematic review based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines has been undertaken. Our aim was to document the range and prevalence of eye abnormalities reported in children with prenatal alcohol exposure (PAE) and/or FASD. METHODS: Searches of electronic databases and manual searches. Eligible articles were observational studies in children with PAE and/or FASD; peer reviewed journal articles in the English language; and studies reporting quantitative or frequency data on functional/structural eye abnormalities. Pooled prevalence, odds ratio, and mean differences were calculated. RESULTS: Of the 1,068 retrieved articles 36 were eligible, including articles on children with diagnosed fetal alcohol syndrome/FASD (N = 31); PAE (N = 3); and FASD or PAE without FASD (N = 2). Structural and functional eye abnormalities were identified, the most prevalent being short palpebral fissure length (66.1%), visual impairment (55.5%), epicanthus (53.5%), subnormal stereoacuity (53.0%), abnormal retinal tortuosity (50.5%), impaired fixation ability (33.3%), telecanthus (31.7%), optic nerve hypoplasia (30.2%), and small optic discs (27.0%). Compared to non-exposed controls, strabismus, subnormal vision, ptosis, short palpebral fissure length, microphthalmos, smaller optic disc area, and retinal vessel tortuosity were more prevalent in children with FASD. CONCLUSIONS: Examination of eyes and vision should be considered in children with PAE and suspected or diagnosed FASD to enable early identification and optimal management. This first comprehensive, systematic literature review demonstrates the variety and frequency of eye abnormalities reported in PAE/FASD.

Survey of perspectives of people with inherited retinal diseases on ocular gene therapy in Australia.

Many gene therapies are in development for treating people with inherited retinal diseases (IRD). We hypothesized that potential recipients of gene therapy would have knowledge gaps regarding treatment. We aimed to assess knowledge, attitudes, and perceptions of genetic therapies among potential recipients with IRD, using a novel instrument we designed (Attitudes to Gene Therapy-Eye (AGT-Eye)) and their associations with demographic data, self-reported visual status, and tools assessing quality of life and attitudes toward clinical trials using a community-based cross-sectional survey of Australian adults with IRD. AGT-Eye, overall quality of life EQ-5D-5L, National Eye Institute Visual Functioning Questionnaire (NEI-VFQ-25) and Patient Attitudes to Clinical Trials (PACT-22) instruments were administered. Six hundred and eighty-one people completed the study, 51.7% women of mean age 53.5 years (SD ± 15.8). Most participants (91.6%) indicated they would likely accept gene therapy if it was available to them or family members. However, only 28.3% agreed that they had good knowledge of gene therapy. Most obtained information about gene therapy from the internet (49.3%). Respondents with post-graduate degrees scored highest compared to other educational levels on methods (p < 0.001) and outcomes (p = 0.003) and were more likely to see economic value of treatment (p = 0.043). Knowledge gaps were present regarding methods and outcomes of gene therapy. This survey has shown high level of interest in the IRD community for gene therapies, and highlights areas for improved clinician and patient education.

Electrophysiological Assessment in Birdshot Chorioretinopathy: Flicker Electroretinograms Recorded With a Handheld Device.

Purpose: The flicker electroretinogram (ERG) is a sensitive indicator of retinal dysfunction in birdshot chorioretinopathy (BCR). We explored recordings from a handheld device in BCR, comparing these with conventional recordings in the same patients and with handheld ERGs from healthy individuals. Methods: Non-mydriatic flicker ERGs, using the handheld RETeval system (LKC Technologies), were recorded with skin electrodes at two centers. At one center (group 1), the stimuli (85 Td·s, 850 Td background) delivered retinal illuminance equivalent to international standards; at the other center (group 2), a different protocol was used (32 Td·s, no background). Patients also underwent international standard flicker ERG recordings with conventional electrodes following mydriasis. Portable ERGs from patients were also compared with those from healthy individuals. Results: Thirty-two patients with BCR (mean age ± SD, 56.4 ± 11.3 years) underwent recordings. Portable and standard ERG parameters correlated strongly (r > 0.75, P < 0.01) in both groups. Limits of agreement for peak times were tighter in group 1 (n = 21; -4.3 to +2.0 ms [right eyes], -3.9 to 1.5 ms [left eyes]) than in group 2 (n = 11; -3.4 to +6.9 ms [right eyes], -4.8 to +9.0 ms [left eyes]). Compared with healthy controls (n = 66 and n = 90 for groups 1 and 2, respectively), patients with BCR showed smaller mean amplitudes and longer peak times. Conclusions: Portable ERGs correlated strongly with conventional recordings, suggesting potential in rapid assessment of cone system function in office settings. Translational Relevance: Flicker ERGs, known to be useful in BCR, can be obtained rapidly with a portable device with skin electrodes and natural pupils.

Whole Genome Sequencing, Focused Assays and Functional Studies Increasing Understanding in Cryptic Inherited Retinal Dystrophies.

The inherited retinal dystrophies (IRDs) are a clinically and genetically complex group of disorders primarily affecting the rod and cone photoreceptors or other retinal neuronal layers, with emerging therapies heralding the need for accurate molecular diagnosis. Targeted capture and panel-based strategies examining the partial or full exome deliver molecular diagnoses in many IRD families tested. However, approximately one in three families remain unsolved and unable to obtain personalised recurrence risk or access to new clinical trials or therapy. In this study, we investigated whole genome sequencing (WGS), focused assays and functional studies to assist with unsolved IRD cases and facilitate integration of these approaches to a broad molecular diagnostic clinical service. The WGS approach identified variants not covered or underinvestigated by targeted capture panel-based clinical testing strategies in six families. This included structural variants, with notable benefit of the WGS approach in repetitive regions demonstrated by a family with a hybrid gene and hemizygous missense variant involving the opsin genes, OPN1LW and OPN1MW. There was also benefit in investigation of the repetitive GC-rich ORF15 region of RPGR. Further molecular investigations were facilitated by focused assays in these regions. Deep intronic variants were identified in IQCB1 and ABCA4, with functional RNA based studies of the IQCB1 variant revealing activation of a cryptic splice acceptor site. While targeted capture panel-based methods are successful in achieving an efficient molecular diagnosis in a proportion of cases, this study highlights the additional benefit and clinical value that may be derived from WGS, focused assays and functional genomics in the highly heterogeneous IRDs.

Human iPSC-Derived Retinal Organoids and Retinal Pigment Epithelium for Novel Intronic RPGR Variant Assessment for Therapy Suitability.

The RPGR gene encodes Retinitis Pigmentosa GTPase Regulator, a known interactor with ciliary proteins, which is involved in maintaining healthy photoreceptor cells. Variants in RPGR are the main contributor to X-linked rod-cone dystrophy (RCD), and RPGR gene therapy approaches are in clinical trials. Hence, elucidation of the pathogenicity of novel RPGR variants is important for a patient therapy opportunity. Here, we describe a novel intronic RPGR variant, c.1415 − 9A>G, in a patient with RCD, which was classified as a variant of uncertain significance according to current clinical diagnostic criteria. The variant lay several base pairs intronic to the canonical splice acceptor site, raising suspicion of an RPGR RNA splicing abnormality and consequent protein dysfunction. To investigate disease causation in an appropriate disease model, induced pluripotent stem cells were generated from patient fibroblasts and differentiated to retinal pigment epithelium (iPSC-RPE) and retinal organoids (iPSC-RO). Abnormal RNA splicing of RPGR was demonstrated in patient fibroblasts, iPSC-RPE and iPSC-ROs, leading to a predicted frameshift and premature stop codon. Decreased RPGR expression was demonstrated in these cell types, with a striking loss of RPGR localization at the ciliary transitional zone, critically in the photoreceptor cilium of the patient iPSC-ROs. Mislocalisation of rhodopsin staining was present in the patient's iPSC-RO rod photoreceptor cells, along with an abnormality of L/M opsin staining affecting cone photoreceptor cells and increased photoreceptor apoptosis. Additionally, patient iPSC-ROs displayed an increase in F-actin expression that was consistent with an abnormal actin regulation phenotype. Collectively, these studies indicate that the splicing abnormality caused by the c.1415 − 9A>G variant has an impact on RPGR function. This work has enabled the reclassification of this variant to pathogenic, allowing the consideration of patients with this variant having access to gene therapy clinical trials. In addition, we have identified biomarkers of disease suitable for the interrogation of other RPGR variants of uncertain significance.

Electronegative electroretinogram in the modern multimodal imaging era.

BACKGROUND: The electronegative electroretinogram (ERG) reflecting inner retinal dysfunction can assist as a diagnostic tool to determine the anatomical location in eye disease. The aim of this study is to determine the frequency and aetiology of electronegative ERG in a tertiary ophthalmology centre and to develop a clinical algorithm to assist patient management. METHODS: Retrospective review of ERGs performed at the Save Sight Institute from January 2011 to December 2020. ERGs were performed according to ISCEV standard. The b:a ratio was analysed in dark adapted (DA) 3.0 or 12.0 recordings. Patients with ratio of ≤1.0 were included. RESULTS: A total of 4421 patients had ERGs performed during study period, of which 139 patients (3.1%) had electronegative ERG. The electronegative ERG patients' median age at referral time was 37 (0.7-90.6) years. The causative aetiologies were photoreceptor dystrophy (48, 34.5%), Congenital Stationary Night Blindness (CSNB) (33, 23.7%), retinal ischemia (18, 12.9%), retinoschisis (15, 10.8%), paraneoplastic autoimmune retinopathy (PAIR) and nonPAIR (14, 10.1%), batten disease (4, 2.9%), and inflammatory retinopathy (4, 2.9%). There were three patients with an unclassified diagnosis. Thirty-two patients (23%) had good vision and a normal fundus appearance. Eleven patients (7.9%) had good vision and normal results in all multimodal imaging. CONCLUSIONS: The frequency of electronegative ERG in our referral centre was 3.1% with photoreceptor dystrophy as the main aetiology. A significant number of the cases had good vision with normal fundus or normal multimodal imaging. This further highlights the value of an ERG in this modern multimodal imaging era.