Neuropathy target esterase activity predicts retinopathy among PNPLA6 disorders.

Biallelic pathogenic variants in the PNPLA6 gene cause a broad spectrum of disorders leading to gait disturbance, visual impairment, anterior hypopituitarism, and hair anomalies. PNPLA6 encodes Neuropathy target esterase (NTE), yet the role of NTE dysfunction on affected tissues in the large spectrum of associated disease remains unclear. We present a clinical meta-analysis of a novel cohort of 23 new patients along with 95 reported individuals with PNPLA6 variants that implicate missense variants as a driver of disease pathogenesis. Measuring esterase activity of 46 disease-associated and 20 common variants observed across PNPLA6 -associated clinical diagnoses unambiguously reclassified 10 variants as likely pathogenic and 36 variants as pathogenic, establishing a robust functional assay for classifying PNPLA6 variants of unknown significance. Estimating the overall NTE activity of affected individuals revealed a striking inverse relationship between NTE activity and the presence of retinopathy and endocrinopathy. This phenomenon was recaptured in vivo in an allelic mouse series, where a similar NTE threshold for retinopathy exists. Thus, PNPLA6 disorders, previously considered allelic, are a continuous spectrum of pleiotropic phenotypes defined by an NTE genotype:activity:phenotype relationship. This relationship and the generation of a preclinical animal model pave the way for therapeutic trials, using NTE as a biomarker.

IMI-Management and Investigation of High Myopia in Infants and Young Children.

Purpose: The purpose of this study was to evaluate the epidemiology, etiology, clinical assessment, investigation, management, and visual consequences of high myopia (≤-6 diopters [D]) in infants and young children. Findings: High myopia is rare in pre-school children with a prevalence less than 1%. The etiology of myopia in such children is different than in older children, with a high rate of secondary myopia associated with prematurity or genetic causes. The priority following the diagnosis of high myopia in childhood is to determine whether there is an associated medical diagnosis that may be of greater overall importance to the health of the child through a clinical evaluation that targets the commonest features associated with syndromic forms of myopia. Biometric evaluation (including axial length and corneal curvature) is important to distinguishing axial myopia from refractive myopia associated with abnormal development of the anterior segment. Additional investigation includes ocular imaging, electrophysiological tests, genetic testing, and involvement of pediatricians and clinical geneticists is often warranted. Following investigation, optical correction is essential, but this may be more challenging and complex than in older children. Application of myopia control interventions in this group of children requires a case-by-case approach due to the lack of evidence of efficacy and clinical heterogeneity of high myopia in young children. Conclusions: High myopia in infants and young children is a rare condition with a different pattern of etiology to that seen in older children. The clinical management of such children, in terms of investigation, optical correction, and use of myopia control treatments, is a complex and often multidisciplinary process.

Retinopathy and optic atrophy in a case of COQ2-related primary coenzyme Q10 deficiency.

PURPOSE: To describe a case of primary coenzyme Q10 deficiency in a child manifesting as early-onset renal failure, retinal dystrophy, and optic atrophy leading to progressive vision loss. METHODS: Clinical presentation and workup including visual fields, electroretinogram, and optical coherence tomography are presented. Genetic testing was performed. RESULTS: An eight-year-old female with nephropathy requiring renal transplantation subsequently developed progressive cone-rod dystrophy and optic atrophy. The patient had negative results on a targeted next-generation sequencing retinal dystrophy panel but whole-exome sequencing revealed two variants in COQ2 (likely biallelic), consistent with a diagnosis of primary coenzyme Q10 deficiency. CONCLUSIONS: Primary coenzyme Q10 deficiency is a rare disorder with variable systemic and ocular findings; there is also genetic heterogeneity. Genetic testing aids in the diagnosis of this condition, and variants in the COQ2 and PDSS1 genes appear to have the strongest association with ocular manifestations. Oral supplementation of coenzyme Q10 may slow progression of disease. This case highlights the utility of whole-exome sequencing in the diagnosis of a rare syndromic form of ocular disease and reports a novel phenotypic association for this condition.

Multidisciplinary team directed analysis of whole genome sequencing reveals pathogenic non-coding variants in molecularly undiagnosed inherited retinal dystrophies.

The purpose of this paper is to identify likely pathogenic non-coding variants in inherited retinal dystrophy (IRD) genes, using genome sequencing (GS). Patients with IRD were recruited to the study and underwent comprehensive ophthalmological evaluation and GS. The results of GS were investigated through virtual gene panel analysis, and plausible pathogenic variants and clinical phenotype evaluated by the multidisciplinary team (MDT) discussion. For unsolved patients in whom a specific gene was suspected to harbor a missed pathogenic variant, targeted re-analysis of non-coding regions was performed on GS data. Candidate variants were functionally tested by messenger RNA analysis, minigene or luciferase reporter assays. Previously unreported, likely pathogenic, non-coding variants in 7 genes (PRPF31, NDP, IFT140, CRB1, USH2A, BBS10 and GUCY2D), were identified in 11 patients. These were shown to lead to mis-splicing (PRPF31, IFT140, CRB1 and USH2A) or altered transcription levels (BBS10 and GUCY2D). MDT-led, phenotype-driven, non-coding variant re-analysis of GS is effective in identifying the missing causative alleles.

Dominant Cone Rod Dystrophy, Previously Assigned to a Missense Variant in RIMS1, Is Fully Explained by Co-Inheritance of a Dominant Allele of PROM1.

Purpose: Autosomal dominant cone rod dystrophy 7 (CORD7) was initially linked to the gene RIMS1 and reported in a 4-generation British family in 1998. The purpose of this study was to investigate the legitimacy of this association, and to correctly characterize the genetic cause of this condition. Methods: The allele frequency of RIMS1 c.2459G>A, p.Arg820His, was investigated in the Genomes Aggregation Dataset (gnomAD) datasets and whole genome sequencing (WGS) was performed for 4 members of the CORD7 family with filtering of rare pathogenic variants in a virtual gene panel comprising all genes known to be associated with inherited retinal dystrophy (IRD). Cytogenetic analysis was performed to rule out interchromosomal translocation. Results: RIMS1 p.Arg820His has a maximal carrier frequency of >1:5000 in Europeans. A previously well-characterized PROM1 variant: c.1118C>T, p.Arg373Cys, was detected in 9 affected members of the CORD7 family who underwent WGS or direct sequencing. One affected family member is now known to have macular dystrophy in the absence of RIMS1 p.Arg820His. Clinical analysis of affected family members and 27 individuals with retinopathy associated with the same - PROM1 - variant showed consistent phenotypes. Conclusions: The case for pathogenicity of RIMS1 p.Arg820His is not strong based on its presence on 10 alleles in the gnomAD dataset and absence from additional CORD affected individuals. The finding of a known pathogenic variant in PROM1 correlates well with the phenotypic characteristics of the affected individuals, and is likely to account for the condition. Clear evidence of association between RIMS1 and a retinal dystrophy is yet to be described.

Optogenetic approaches to therapy for inherited retinal degenerations.

Inherited retinal degenerations such as retinitis pigmentosa (RP) affect around one in 4000 people and are the leading cause of blindness in working age adults in several countries. In these typically monogenic conditions, there is progressive degeneration of photoreceptors; however, inner retinal neurons such as bipolar cells and ganglion cells remain largely structurally intact, even in end-stage disease. Therapeutic approaches aiming to stimulate these residual cells, independent of the underlying genetic cause, could potentially restore visual function in patients with advanced vision loss, and benefit many more patients than therapies directed at the specific gene implicated in each disorder. One approach investigated for this purpose is that of optogenetics, a method of neuromodulation that utilises light to activate neurons engineered to ectopically express a light-sensitive protein. Using gene therapy via adeno-associated viral vectors, a range of photosensitive proteins have been expressed in remaining retinal cells in advanced retinal degeneration with in vivo studies demonstrating restoration of visual function. Developing an effective optogenetic strategy requires consideration of multiple factors, including the light-sensitive protein that is used, the vector and method for gene delivery, and the target cell for expression because these in turn may affect the quality of vision that can be restored. Currently, at least four clinical trials are ongoing to investigate optogenetic therapies in patients, with the ultimate aim of reversing visual loss in end-stage disease.

Intravitreal Delivery of rAAV2tYF-CB-hRS1 Vector for Gene Augmentation Therapy in Patients with X-Linked Retinoschisis: 1-Year Clinical Results.

PURPOSE: To evaluate the safety and efficacy of rAAV2tYF-CB-hRS1, a recombinant adeno-associated virus vector expressing retinoschisin (RS1), in individuals with retinal disease caused by mutations in the RS1 gene. DESIGN: Open-label, phase I/II dose-escalation clinical trial. SUBJECTS: Twenty-two adults and 5 children with X-linked retinoschisis (XLRS), aged 10 to 79 years, were enrolled. METHODS: The participants received an intravitreal (IVT) injection of rAAV2tYF-CB-hRS1, at 1 of 3 dose levels, in the poorer-seeing eye and were followed up for a minimum of 1 year after treatment. MAIN OUTCOME MEASURES: The primary safety measures were local (ocular) or systemic (nonocular) adverse events (AEs) during the 12-month period after study agent administration. Efficacy was assessed based on measures of best-corrected visual acuity (BCVA), schisis cavity volume, static perimetry visual field testing, and electroretinography (ERG). RESULTS: The IVT administration of rAAV2tYF-CB-hRS1 was generally safe at each of the dose levels. There were no AEs resulting in early termination, and no dose-limiting toxicities were reported. The most common ocular AEs observed were related to ocular inflammation (blurred vision, visual impairment, and the presence of vitreous cells, keratic precipitates, vitreous floaters, anterior chamber cells, and vitreous haze). Ocular inflammation was generally either mild or moderate in severity and responsive to standard immunosuppressive therapy, except in 3 participants (all in the highest-dose group) who developed chronic uveitis, which required prolonged therapy. Two patients experienced retinal detachments. There was no overall improvement in BCVA, visual fields, or ERG in the study eye compared with that in the fellow eye for any dose group. Variable changes in the cystic cavity volume over time were similar in the study and fellow eyes. CONCLUSIONS: Gene augmentation therapy with rAAV2tYF-CB-hRS1 for XLRS was generally safe and well tolerated but failed to demonstrate a measurable treatment effect. The clinical trial is ongoing through 5 years of follow-up to assess its long-term safety.

Variability of retinopathy consequent upon novel mutations in LAMA1.

PURPOSE: Bi-allelic mutations in LAMA1 (laminin 1) (OMIM # 150320) cause Poretti-Boltshauser Syndrome (PTBHS), a rare non-progressive cerebellar dysplasia disorder with ophthalmic manifestations including oculomotor apraxia, high myopia, and retinal dystrophy. Only 38 variants, nearly all loss of function have been reported. Here, we describe novel LAMA1 variants and detailed retinal manifestations in two unrelated families. METHODS: Whole-genome sequencing was conducted on three siblings of a consanguineous family with myopia and retinal dystrophy and on a child from an unrelated non-consanguineous couple. Clinical evaluation included full ophthalmic examination, detailed colour, autofluorescence retinal imaging, retinal optical coherence tomography (OCT), fluorescein angiography under anesthesia, and pattern and full-field electroretinography. RESULTS: Genetic analysis revealed a novel homozygous LAMA1 frameshift variant, c.1492del p.(Arg498Glyfs *25), in the affected siblings in family 1 and a novel frameshift c.3065del p.(Gly1022Valfs *2) and a deletion spanning exons 17-23 in an unrelated individual in family 2. Two of the three siblings and the unrelated child had oculomotor apraxia in childhood; none of the siblings had symptoms of other neurological dysfunction as adults. All four had myopia. The affected siblings had a qualitatively similar retinopathy of wide-ranging severity. The unrelated patient had a severe abnormality of retinal vascular development, which resulted in vitreous haemorrhage and neovascular glaucoma in the left eye and a rhegmatogenous retinal detachment in the right eye. CONCLUSIONS: This report describes the detailed retinal structural and functional consequences of LAMA1 deficiency in four patients from two families, and these exhibit significant variability with evidence of both retinal dystrophy and abnormal and incomplete retinal vascularisation.

WFS1-Associated Optic Neuropathy: Genotype-Phenotype Correlations and Disease Progression.

OBJECTIVE: To evaluate the pattern of vision loss and genotype-phenotype correlations in WFS1-associated optic neuropathy (WON). DESIGN: Multicenter cohort study. METHODS: The study involved 37 patients with WON carrying pathogenic or candidate pathogenic WFS1 variants. Genetic and clinical data were retrieved from the medical records. Thirteen patients underwent additional comprehensive ophthalmologic assessment. Deep phenotyping involved visual electrophysiology and advanced psychophysical testing with a complementary metabolomic study. MAIN OUTCOME MEASURES: WFS1 variants, functional and structural optic nerve and retinal parameters, and metabolomic profile. RESULTS: Twenty-two recessive and 5 dominant WFS1 variants were identified. Four variants were novel. All WFS1 variants caused loss of macular retinal ganglion cells (RGCs) as assessed by optical coherence tomography (OCT) and visual electrophysiology. Advanced psychophysical testing indicated involvement of the major RGC subpopulations. Modeling of vision loss showed an accelerated rate of deterioration with increasing age. Dominant WFS1 variants were associated with abnormal reflectivity of the outer plexiform layer (OPL) on OCT imaging. The dominant variants tended to cause less severe vision loss compared with recessive WFS1 variants, which resulted in more variable phenotypes ranging from isolated WON to severe multisystem disease depending on the WFS1 alleles. The metabolomic profile included markers seen in other neurodegenerative diseases and type 1 diabetes mellitus. CONCLUSIONS: WFS1 variants result in heterogenous phenotypes influenced by the mode of inheritance and the disease-causing alleles. Biallelic WFS1 variants cause more variable, but generally more severe, vision and RGC loss compared with heterozygous variants. Abnormal cleftlike lamination of the OPL is a distinctive OCT feature that strongly points toward dominant WON.

Variants in PAX6, PITX3 and HSF4 causing autosomal dominant congenital cataracts.

BACKGROUND: Lens development is orchestrated by transcription factors. Disease-causing variants in transcription factors and their developmental target genes are associated with congenital cataracts and other eye anomalies. METHODS: Using whole exome sequencing, we identified disease-causing variants in two large British families and one isolated case with autosomal dominant congenital cataract. Bioinformatics analysis confirmed these disease-causing mutations as rare or novel variants, with a moderate to damaging pathogenicity score, with testing for segregation within the families using direct Sanger sequencing. RESULTS: Family A had a missense variant (c.184 G>A; p.V62M) in PAX6 and affected individuals presented with nuclear cataract. Family B had a frameshift variant (c.470-477dup; p.A160R*) in PITX3 that was also associated with nuclear cataract. A recurrent missense variant in HSF4 (c.341 T>C; p.L114P) was associated with congenital cataract in a single isolated case. CONCLUSIONS: We have therefore identified novel variants in PAX6 and PITX3 that cause autosomal dominant congenital cataract.

Association of ambient air pollution with age-related macular degeneration and retinal thickness in UK Biobank.

AIM: To examine the associations of air pollution with both self-reported age-related macular degeneration (AMD), and in vivo measures of retinal sublayer thicknesses. METHODS: We included 115 954 UK Biobank participants aged 40-69 years old in this cross-sectional study. Ambient air pollution measures included particulate matter, nitrogen dioxide (NO2) and nitrogen oxides (NOx). Participants with self-reported ocular conditions, high refractive error (< -6 or > +6 diopters) and poor spectral-domain optical coherence tomography (SD-OCT) image were excluded. Self-reported AMD was used to identify overt disease. SD-OCT imaging derived photoreceptor sublayer thickness and retinal pigment epithelium (RPE) layer thickness were used as structural biomarkers of AMD for 52 602 participants. We examined the associations of ambient air pollution with self-reported AMD and both photoreceptor sublayers and RPE layer thicknesses. RESULTS: After adjusting for covariates, people who were exposed to higher fine ambient particulate matter with an aerodynamic diameter <2.5 µm (PM2.5, per IQR increase) had higher odds of self-reported AMD (OR=1.08, p=0.036), thinner photoreceptor synaptic region (ß=-0.16 µm, p=2.0 × 10-5), thicker photoreceptor inner segment layer (ß=0.04 µm, p=0.001) and thinner RPE (ß=-0.13 µm, p=0.002). Higher levels of PM2.5 absorbance and NO2 were associated with thicker photoreceptor inner and outer segment layers, and a thinner RPE layer. Higher levels of PM10 (PM with an aerodynamic diameter <10 µm) was associated with thicker photoreceptor outer segment and thinner RPE, while higher exposure to NOx was associated with thinner photoreceptor synaptic region. CONCLUSION: Greater exposure to PM2.5 was associated with self-reported AMD, while PM2.5, PM2.5 absorbance, PM10, NO2 and NOx were all associated with differences in retinal layer thickness.

SSBP1-Disease Update: Expanding the Genetic and Clinical Spectrum, Reporting Variable Penetrance and Confirming Recessive Inheritance.

Purpose: To report novel genotypes and expand the phenotype spectrum of SSBP1-disease and explore potential disease mechanism. Methods: Five families with previously unsolved optic atrophy and retinal dystrophy underwent whole genome sequencing as part of the National Institute for Health Research BioResource Rare-Diseases and the UK's 100,000 Genomes Project. In silico analysis and protein modelling was performed on the identified variants. Deep phenotyping including retinal imaging and International Society for Clinical Electrophysiology of Vision standard visual electrophysiology was performed. Results: Seven individuals from five unrelated families with bilateral optic atrophy and/or retinal dystrophy with extraocular signs and symptoms in some are described. In total, 6 SSBP1 variants were identified including the previously unreported variants: c.151A>G, p.(Lys51Glu), c.335G>A p.(Gly112Glu), and c.380G>A, p.(Arg127Gln). One individual was found to carry biallelic variants (c.380G>A p.(Arg127Gln); c.394A>G p.(Ile132Val)) associated with likely autosomal recessive SSBP1-disease. In silico analysis predicted all variants to be pathogenic and Three-dimensional protein modelling suggested possible disease mechanisms via decreased single-stranded DNA binding affinity or impaired higher structure formation. Conclusions: SSBP1 is essential for mitochondrial DNA replication and maintenance, with defects leading to a spectrum of disease that includes optic atrophy and/or retinal dystrophy, occurring with or without extraocular features. This study provides evidence of intrafamilial variability and confirms the existence of an autosomal recessive inheritance in SSBP1-disease consequent upon a previously unreported genotype.

Allele-specific gene editing to rescue dominant CRX-associated LCA7 phenotypes in a retinal organoid model.

Cases of Leber congenital amaurosis caused by mutations in CRX (LCA7) exhibit an early form of the disease and show signs of significant photoreceptor dysfunction and eventual loss. To establish a translational in vitro model system to study gene-editing-based therapies, we generated LCA7 retinal organoids harboring a dominant disease-causing mutation in CRX. Our LCA7 retinal organoids develop signs of immature and dysfunctional photoreceptor cells, providing us with a reliable in vitro model to recapitulate LCA7. Furthermore, we performed a proof-of-concept study in which we utilize allele-specific CRISPR/Cas9-based gene editing to knock out mutant CRX and saw moderate rescue of photoreceptor phenotypes in our organoids. This work provides early evidence for an effective approach to treat LCA7, which can be applied more broadly to other dominant genetic diseases.

Beyond factor H: The impact of genetic-risk variants for age-related macular degeneration on circulating factor-H-like 1 and factor-H-related protein concentrations.

Age-related macular degeneration (AMD) is a leading cause of vision loss; there is strong genetic susceptibility at the complement factor H (CFH) locus. This locus encodes a series of complement regulators: factor H (FH), a splice variant factor-H-like 1 (FHL-1), and five factor-H-related proteins (FHR-1 to FHR-5), all involved in the regulation of complement factor C3b turnover. Little is known about how AMD-associated variants at this locus might influence FHL-1 and FHR protein concentrations. We have used a bespoke targeted mass-spectrometry assay to measure the circulating concentrations of all seven complement regulators and demonstrated elevated concentrations in 352 advanced AMD-affected individuals for all FHR proteins (FHR-1, p = 2.4 × 10-10; FHR-2, p = 6.0 × 10-10; FHR-3, p = 1.5 × 10-5; FHR-4, p = 1.3 × 10-3; FHR-5, p = 1.9 × 10-4) and FHL-1 (p = 4.9 × 10-4) when these individuals were compared to 252 controls, whereas no difference was seen for FH (p = 0.94). Genome-wide association analyses in controls revealed genome-wide-significant signals at the CFH locus for all five FHR proteins, and univariate Mendelian-randomization analyses strongly supported the association of FHR-1, FHR-2, FHR-4, and FHR-5 with AMD susceptibility. These findings provide a strong biochemical explanation for how genetically driven alterations in circulating FHR proteins could be major drivers of AMD and highlight the need for research into FHR protein modulation as a viable therapeutic avenue for AMD.

Transcription factor overexpression drives reliable differentiation of retinal pigment epithelium from human induced pluripotent stem cells.

Age-related macular degeneration and genetic forms of blindness such as Best Disease and Retinitis Pigmentosa can be caused by degeneration of the Retinal Pigment Epithelium (RPE). RPE generated from patient-derived induced pluripotent stem cells (iPSCs) is valuable for both the study of disease mechanisms and development of therapeutic strategies. However, protocols to produce iPSC-derived RPE in vitro are often inefficient, labor-intensive, low-throughput, and highly variable between cell lines and within batches. Here, we report a robust, scalable method to generate iPSC-RPE using doxycycline-inducible expression of eye field transcription factors OTX2, PAX6 and MITF paired with RPE-permissive culture media. Doxycycline addition induces exogenous expression of these transcription factors in Best Disease patient- and wildtype iPSCs to efficiently produce monolayers of RPE with characteristic morphology and gene expression. Further, these RPE monolayers display functionality features including light absorption via pigmentation, polarity-driven fluid transport, and phagocytosis. With this method, we achieve a highly efficient and easily scalable differentiation without the need for mechanical isolation or enrichment methods, generating RPE cultures applicable for in vitro studies.

Expanding the FDXR-Associated Disease Phenotype: Retinal Dystrophy Is a Recurrent Ocular Feature.

Purpose: The purpose of this study was to report retinal dystrophy as a novel clinical feature and expand the ocular phenotype in patients harboring biallelic candidate FDXR variants. Methods: Patients carrying biallelic candidate FDXR variants were identified by whole genome sequencing (WGS) as part of the National Institute for Health Research BioResource rare-disease and the UK's 100,000 Genomes Project (100KGP) with an additional case identified by exome sequencing. Retrospective clinical data were collected from the medical records. Haplotype reconstruction was performed in families harboring the same missense variant. Results: Ten individuals from 8 unrelated families with biallelic candidate variants in FDXR were identified. In addition to bilateral optic atrophy and variable extra-ocular findings, 7 of 10 individuals manifested retinal dystrophy comprising dysfunction and degeneration of both rod and cone photoreceptors. Five of 10 subjects had sensorineural hearing loss. The previously unreported missense variant (c.1115C > A, p.(Pro372His)) was found in 5 of 8 (62.5%) study families. Haplotype reconstruction using WGS data demonstrated a likely ancestral haplotype. Conclusions: FDXR-associated disease is a phenotypically heterogeneous disorder with retinal dystrophy being a major clinical feature observed in this cohort. In addition, we hypothesize that a number of factors are likely to drive the pathogenesis of optic atrophy, retinal degeneration, and perhaps the associated systemic manifestations.

PHENOTYPIC HETEROGENEITY IN A FAMILY WITH X-LINKED FAMILIAL EXUDATIVE VITREORETINOPATHY WITH PREVENTION OF VISUAL LOSS IN AN AFFECTED MALE CHILD WITH LASER TREATMENT IN INFANCY.

PURPOSE: To present the scope of prenatal diagnosis and early treatment of patients with clinically heterogeneous phenotypic retinal dysplasia associated with NDP gene variants. METHODS: Retrospective. Review of electronic medical records. RESULTS: Twenty-nine-year-old woman known to carry a NDP gene variant presented to the eye clinic for consultation and risk assessment at her second pregnancy. Her 11-year-old son had bilateral retinal detachment, despite surgical treatment. The family declined prenatal testing. The patient was born full term, was examined, and underwent genetic testing after birth. He was found to have bilateral retinal avascular periphery abnormalities and preretinal hemorrhages on the left eye. The patient received bilateral laser treatment at 2 months of age. He was found to be doing well at 16 months after treatment with adequate visual acuity and flat maculae. The asymptomatic mother and maternal grandfather of the proband were found to have retinal periphery abnormalities with unremarkable posterior pole and excellent visual acuity. CONCLUSION: NDP gene variants associated with X-linked familial exudative vitreoretinopathy phenotype benefit from early treatment. Providers who take care of these patients need to monitor closely the pregnancy and delivery of a male child born to a female carrier to offer appropriate and timely treatment.

The X-linked retinopathies: Physiological insights, pathogenic mechanisms, phenotypic features and novel therapies.

X-linked retinopathies represent a significant proportion of monogenic retinal disease. They include progressive and stationary conditions, with and without syndromic features. Many are X-linked recessive, but several exhibit a phenotype in female carriers, which can help establish diagnosis and yield insights into disease mechanisms. The presence of affected carriers can misleadingly suggest autosomal dominant inheritance. Some disorders (such as RPGR-associated retinopathy) show diverse phenotypes from variants in the same gene and also highlight limitations of current genetic sequencing methods. X-linked disease frequently arises from loss of function, implying potential for benefit from gene replacement strategies. We review X-inactivation and X-linked inheritance, and explore burden of disease attributable to X-linked genes in our clinically and genetically characterised retinal disease cohort, finding correlation between gene transcript length and numbers of families. We list relevant genes and discuss key clinical features, disease mechanisms, carrier phenotypes and novel experimental therapies. We consider in detail the following: RPGR (associated with retinitis pigmentosa, cone and cone-rod dystrophy), RP2 (retinitis pigmentosa), CHM (choroideremia), RS1 (X-linked retinoschisis), NYX (complete congenital stationary night blindness (CSNB)), CACNA1F (incomplete CSNB), OPN1LW/OPN1MW (blue cone monochromacy, Bornholm eye disease, cone dystrophy), GPR143 (ocular albinism), COL4A5 (Alport syndrome), and NDP (Norrie disease and X-linked familial exudative vitreoretinopathy (FEVR)). We use a recently published transcriptome analysis to explore expression by cell-type and discuss insights from electrophysiology. In the final section, we present an algorithm for genes to consider in diagnosing males with non-syndromic X-linked retinopathy, summarise current experimental therapeutic approaches, and consider questions for future research.

Genetic Testing Leading to Early Identification of Childhood Ocular Manifestations of Usher Syndrome.

OBJECTIVES: Hearing-loss gene panel testing (HLGPT) is increasingly accessible as a first-line test in determining the etiology of sensorineural hearing loss (SNHL) in children. A major advantage of HLGPT is early identification of syndromic forms of SNHL, especially Usher syndrome, prior to the development of overt syndromic phenotype, which may impact management and counseling. Here, we describe early ocular findings in children with clinically non-syndromic SNHL identified by HLGPT as having two variants associated with Usher Syndrome. METHODS: A total of 184 children, ages 1 month - 15 years of age, evaluated at one tertiary pediatric children's hospital for clinically non-syndromic SNHL, underwent next-generation sequencing of 150 genes involved in hearing loss. Children with two variants in genes associated with Usher syndrome were referred for evaluation by pediatric ophthalmology. RESULTS: A total of 18/184 tested children had two variants in Usher syndrome-associated genes, including MYO7A, GPR98 (ADGRV1), USH2A, and PDZD7. SNHL varied from moderate to profound. 29% of the children who underwent clinical ophthalmology evaluation were found to have previously unidentified retinal abnormalities on retinal imaging or electroretinography consistent with inherited retinal degeneration. CONCLUSION: Among this ethnically and racially diverse pediatric population with apparently non-syndromic SNHL, HLGPT yielded a high proportion (10%) of children with two variants in genes associated with Usher syndrome. Early genetic testing allows early identification of variants conferring a diagnosis of Usher syndrome at a stage prior to visual symptoms. This allows for more informed genetic counseling, reproductive planning, and sensory deficit interventions. LEVEL OF EVIDENCE: 4 Laryngoscope, 131:E2053-E2059, 2021.