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Age-related cataracts is a highly prevalent eye disorder that results in the clouding of the crystalline lens and is one of the leading causes of visual impairment and blindness. The disease is influenced by multiple factors including genetics, prolonged exposure to ultraviolet radiation, and a history of diabetes. However, the extent to which each of these factors contributes to the development of cataracts remains unclear. Our study identified 101 independent genome-wide significant loci, 57 of which are novel. We identified multiple genes and biological pathways associated with the cataracts, including four drug-gene interactions. Our results suggest a causal association between type 1 diabetes and cataracts. Also, we highlighted a surrogate measure of UV light exposure as a marker of cataract risk in adults., (© 2024. The Author(s).)

Zebrafish are an outstanding model for assessing the involvement of genes in paediatric cataracts. Gene discovery for cataracts is enhanced by manipulation of the genome of zebrafish embryos and comparing the phenotypes of mutant progeny with the wildtype embryos. However, wildtype laboratory fish can also develop cataracts, potentially confounding the results. In this study, we compared the baseline cataract rate between two commonly used wildtype laboratory strains, AB and TL, and also an outbred transgenic line with mCherry reporter. We assessed a total of 805 lens images of fish at 4 days post-fertilisation for cataracts and scored each cataract observed as mild, moderate or severe. We found that the AB strain had a cataract rate of 16.2%, TL had 8.9%, and mCherry had 0.7% and these rates were significantly different. We found that TL strain had a lower rate of mild cataracts than AB fish, however, the rate of moderate and severe phenotypes in the AB and the TL strain was similar. Overall, we showed that the baseline cataract rate varies significantly between the strains housed in a single facility and conclude that baseline rates of cataracts should be assessed when planning experiments to assess the genetic causes of cataracts., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Prostate cancer is one of the most commonly diagnosed cancers in men and unfortunately, disease will progress in up to a third of patients despite primary treatment. Currently, there is a significant lack of prognostic tests that accurately predict disease course; however, the acquisition of somatic chromosomal variation in the form of DNA copy number variants may help understand disease progression. Notably, studies have found that a higher burden of somatic copy number alterations (SCNA) correlates with more aggressive disease, recurrence after surgery and metastasis. Here we will review the literature surrounding SCNA formation, including the roles of key tumour suppressors and oncogenes (PTEN, BRCA2, NKX3.1, ERG and AR), and their potential to inform diagnostic and prognostic clinical testing to improve predictive value. Ultimately, SCNAs, or inherited germline alterations that predispose to SCNAs, could have significant clinical utility in diagnostic and prognostic tests, in addition to guiding therapeutic selection., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

Background: There are currently no specific biomarkers for multiple sclerosis (MS). Identifying robust biomarkers for MS is crucial to improve disease diagnosis and management., Methods: This study first used six Mendelian randomisation methods to assess causal relationship of 174 metabolites with MS, incorporating data from European-ancestry metabolomics (n=8569-86 507) and MS (n=14 802 MS cases, 26 703 controls) genomewide association studies. Genetic scores for identified causal metabolite(s) were then computed to predict MS disability progression in an independent longitudinal cohort (AusLong study) of 203 MS cases with up to 15-year follow-up., Results: We found a novel genetic causal effect of serine on MS onset (OR=1.67, 95% CI 1.51 to 1.84, p=1.73×10 -20 ), such that individuals whose serine level is 1 SD above the population mean will have 1.67 times the risk of developing MS. This is robust across all sensitivity methods (OR ranges from 1.49 to 1.67). In an independent longitudinal MS cohort, we then constructed time-dynamic and time-fixed genetic scores based on serine genetic instrument single-nucleotide polymorphisms, where higher scores for raised serum serine level were associated with increased risk of disability worsening, especially in the time-dynamic model (RR=1.25, 95% CI 1.10 to 1.42, p=7.52×10 -4 )., Conclusions: These findings support investigating serine as an important candidate biomarker for MS onset and disability progression., Competing Interests: Competing interests: BVT has received compensation for consulting, talks, and advisory/steering board activities for Merck, Novartis, Biogen, and Roche. He receives research funding support from MS Research Australia, Medical Research Future Fund Australia and the National Health & Medical Research Council Australia., (© Author(s) (or their employer(s)) 2023. No commercial re-use. See rights and permissions. Published by BMJ.)

Understanding how variations in the plasma and brain proteome contribute to multiple sclerosis susceptibility can provide important insights to guide drug repurposing and therapeutic development for the disease. However, the role of genetically predicted protein abundance in multiple sclerosis remains largely unknown. Integrating plasma proteomics (n = 3301) and brain proteomics (n = 376 discovery; n = 152 replication) into multiple sclerosis genome-wide association studies (n = 14 802 cases and 26 703 controls), we employed summary-based methods to identify candidate proteins involved in multiple sclerosis susceptibility. Next, we evaluated associations of the corresponding genes with multiple sclerosis at tissue-level using large gene expression quantitative trait data from whole-blood (n = 31 684) and brain (n = 1194) tissue. Further, to assess transcriptional profiles for candidate proteins at cell-level, we examined gene expression patterns in immune cell types (Dataset 1: n = 73 cases and 97 controls; Dataset 2: n = 31 cases and 31 controls) for identified plasma proteins, and in brain cell types (Dataset 1: n = 4 cases and 5 controls; Dataset 2: n = 5 cases and 3 controls) for identified brain proteins. In a longitudinal multiple sclerosis cohort (n = 203 cases followed up to 15 years), we also assessed the corresponding gene-level associations with the outcome of disability worsening. We identified 39 novel proteins associated with multiple sclerosis risk. Based on five identified plasma proteins, four available corresponding gene candidates showed consistent associations with multiple sclerosis risk in whole-blood, and we found TAPBPL upregulation in multiple sclerosis B cells, CD8+ T cells and natural killer cells compared with controls. Among the 34 candidate brain proteins, 18 were replicated in a smaller cohort and 14 of 21 available corresponding gene candidates also showed consistent associations with multiple sclerosis risk in brain tissue. In cell-specific analysis, six identified brain candidates showed consistent differential gene expression in neuron and oligodendrocyte cell clusters. Based on the 39 protein-coding genes, we found 23 genes that were associated with disability worsening in multiple sclerosis cases. The findings present a set of candidate protein biomarkers for multiple sclerosis, reinforced by high concordance in downstream transcriptomics findings at tissue-level. This study also highlights the heterogeneity of cell-specific transcriptional profiles for the identified proteins and that numerous candidates were also implicated in disease progression. Together, these findings can serve as an important anchor for future studies of disease mechanisms and therapeutic development., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Intravitreal anti-vascular endothelial growth factor (anti-VEGF) injections are the standard of care for diabetic macular edema (DME), a common complication of diabetes. This study aimed to identify factors influencing DME intravitreal anti-VEGF treatment outcomes in real-world practice., Methods: This was a multi-center retrospective observational study using medical chart review of participants receiving anti-VEGF injections for DME (N = 248). Demographic and clinical variables were assessed for association with best corrected visual acuity (BCVA) and central macular thickness (CMT) outcomes using regression models., Results: There was a significant improvement in BCVA (p < 0.001) and CMT (p < 0.001) after 12 months of treatment, although 21% of participants had decreased BCVA, and 41% had a < 10% CMT reduction at 12 months. Higher baseline BCVA (p = 0.022, OR=-0.024, 95% CI=-0.046,-0.004) and longer duration of diabetic retinopathy (p = 0.048, OR=-0.064, 95% CI=-0.129,-0.001) were negative predictors for BCVA response, whereas Aflibercept treatment (p = 0.017, OR = 1.107, 95% CI = 0.220,2.051) compared with other drugs and a positive "early functional response" (p < 0.001, OR=-1.393, 95% CI=-1.946,-0.857) were positive predictors. A higher baseline CMT (p < 0.001, OR = 0.019, 95% CI = 0.012,0.0261) and an "early anatomical response", (p < 0.001, OR=-1.677, 95% CI=-2.456, -0.943) were predictors for greater reduction in CMT. Overall, the variables could predict only 23% of BCVA and 52% of CMT response., Conclusions: The study shows a significant proportion of DME patients do not respond to anti-VEGF therapy and identifies several clinical predictors for treatment outcomes., Trial Registration: The study was approved through the Human Research Ethics Committee, University of Tasmania (approval number H0012902), and the Southern Adelaide Clinical Human Research Ethics Committee (approval number 86 - 067)., (© 2023. The Author(s).)

Diabetic retinopathy (DR) is the most common microvascular complication related to diabetes. There is evidence that genetics play an important role in DR pathogenesis, but the complexity of the disease makes genetic studies a challenge. This chapter is a practical overview of the basic steps for genome-wide association studies with respect to DR and its associated traits. Also described are approaches that can be adopted in future DR studies. This is intended to serve as a guide for beginners and to provide a framework for further in-depth analysis., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

The standardization of variant curation criteria is essential for accurate interpretation of genetic results and clinical care of patients. The variant curation guidelines developed by the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP) in 2015 are widely used but are not gene specific. To address this issue, the Clinical Genome Resource (ClinGen) Variant Curation Expert Panels (VCEP) have been tasked with developing gene-specific variant curation guidelines. The Glaucoma VCEP was created to develop rule specifications for genes associated with primary glaucoma, including myocilin (MYOC), the most common cause of Mendelian glaucoma. Of the 28 ACMG/AMP criteria, the Glaucoma VCEP adapted 15 rules to MYOC and determined 13 rules not applicable. Key specifications included determining minor allele frequency thresholds, developing an approach to counting probands and segregations, and reviewing functional assays. The rules were piloted on 81 variants and led to a change in classification in 40% of those that were classified in ClinVar, with functional evidence influencing the classification of 18 variants. The standardized variant curation guidelines for MYOC provide a framework for the consistent application of the rules between laboratories, to improve MYOC genetic testing in the management of glaucoma., (© 2022 The Authors. Human Mutation published by Wiley Periodicals LLC.)

Corneal dystrophies describe a clinically and genetically heterogeneous group of inherited disorders. The International Classification of Corneal Dystrophies (IC3D) lists 22 types of corneal dystrophy, 17 of which have been demonstrated to result from pathogenic variants in 19 identified genes. In this study, we investigated the diagnostic yield of genetic testing in a well-characterised cohort of 58 individuals from 44 families with different types of corneal dystrophy. Individuals diagnosed solely with Fuchs endothelial corneal dystrophy were excluded. Clinical details were obtained from the treating ophthalmologist. Participants and their family members were tested using a gene candidate and exome sequencing approach. We identified a likely molecular diagnosis in 70.5% families (31/44). The detection rate was significantly higher among probands with a family history of corneal dystrophy (15/16, 93.8%) than those without (16/28, 57.1%, p = .015), and among those who had undergone corneal graft surgery (9/9, 100.0%) compared to those who had not (22/35, 62.9%, p = .041). We identified eight novel variants in five genes and identified five families with syndromes associated with corneal dystrophies. Our findings highlight the genetic heterogeneity of corneal dystrophies and the clinical utility of genetic testing in reaching an accurate clinical diagnosis., (© 2022 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals LLC.)

Objective: Paediatric (childhood or congenital) cataract is an opacification of the normally clear lens of the eye and has a genetic basis in at least 18% of cases in Australia. This study aimed to replicate clinical gene screening to identify variants likely to be causative of disease in an Australian patient cohort., Methods and Analysis: Sixty-three reported isolated cataract genes were screened for rare coding variants in 37 Australian families using genome sequencing., Results: Disease-causing variants were confirmed in eight families with variant classification as 'likely pathogenic'. This included novel variants PITX3 p.(Ter303LeuextTer100), BFSP1 p.(Glu375GlyfsTer2), and GJA8 p.(Pro189Ser), as well as, previously described variants identified in genes GJA3, GJA8, CRYAA, BFSP1, PITX3, COL4A1 and HSF4 . Additionally, eight variants of uncertain significance with evidence towards pathogenicity were identified in genes: GJA3, GJA8, LEMD2, PRX, CRYBB1, BFSP2, and MIP ., Conclusion: These findings expand the genotype-phenotype correlations of both pathogenic and benign variation in cataract-associated genes. They further emphasise the need to develop additional evidence such as functional assays and variant classification criteria specific to paediatric cataract genes to improve interpretation of variants and molecular diagnosis in patients., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Multiple sclerosis (MS) is a complex neuroinflammatory/degenerative disease of the central nervous system (CNS) that results in the formation of demyelinated lesions and axon degeneration. MS aetiology is complex, with genetics estimated to account for ∼48% of MS risk (International Multiple Sclerosis Genetics Consortium, 2019). Despite this, families with a high incidence of MS are rare. We have generated four induced pluripotent stem cell (iPSC) lines from individuals with relapsing-remitting and secondary progressive MS within a single family. The generation of disease-specific iPSC lines from multiple members of a single family will facilitate MS genetic and functional studies., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Advances in genome sequencing technologies have unlocked new possibilities in identifying disease-associated and causative genetic markers, which may in turn enhance disease diagnosis and improve prognostication and management strategies. With the capability of examining genetic variations ranging from single-nucleotide mutations to large structural variants, whole-genome sequencing (WGS) is an increasingly adopted approach to dissect the complex genetic architecture of neurologic diseases. There is emerging evidence for different structural variants and their roles in major neurologic and neurodevelopmental diseases. This review first describes different structural variants and their implicated roles in major neurologic and neurodevelopmental diseases, and then discusses the clinical relevance of WGS applications in neurology. Notably, WGS-based detection of structural variants has shown promising potential in enhancing diagnostic power of genetic tests in clinical settings. Ongoing WGS-based research in structural variations and quantifying mutational constraints can also yield clinical benefits by improving variant interpretation and disease diagnosis, while supporting biomarker discovery and therapeutic development. As a result, wider integration of WGS technologies into health care will likely increase diagnostic yields in difficult-to-diagnose conditions and define potential therapeutic targets or intervention points for genome-editing strategies., Competing Interests: B.V.T. has received compensation for consulting, talks, and advisory/steering board activities for Merck, Novartis, Biogen, and Roche. He receives research funding support from MS Research Australia, Medical Research Future Fund Australia, and the National Health & Medical Research Council Australia. Go to Neurology.org/NG for full disclosure., (Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.)

Intraocular anti-vascular endothelial growth factor (VEGF) therapies are the front-line treatment for diabetic macular edema (DME); however, treatment response varies widely. This study aimed to identify genetic determinants associated with anti-VEGF treatment response in DME. We performed a genome-wide association study on 220 Australian patients with DME treated with anti-VEGF therapy, genotyped on the Illumina Global Screening Array, and imputed to the Haplotype Reference Consortium panel. The primary outcome measures were changes in central macular thickness (CMT in microns) and best-corrected visual acuity (BCVA in ETDRS letters) after 12 months. Association between single nucleotide polymorphism (SNP) genotypes and DME outcomes were evaluated by linear regression, adjusting for the first three principal components, age, baseline CMT/BCVA, duration of diabetic retinopathy, and HbA1c. Two loci reached genome-wide significance (p < 5 × 10−8) for association with increased CMT: a single SNP on chromosome 6 near CASC15 (rs78466540, p = 1.16 × 10−9) and a locus on chromosome 12 near RP11-116D17.1 (top SNP rs11614480, p = 2.69 × 10−8). Four loci were significantly associated with reduction in BCVA: two loci on chromosome 11, downstream of NTM (top SNP rs148980760, p = 5.30 × 10−9) and intronic in RP11-744N12.3 (top SNP rs57801753, p = 1.71 × 10−8); one near PGAM1P1 on chromosome 5 (rs187876551, p = 1.52 × 10−8); and one near TBC1D32 on chromosome 6 (rs118074968, p = 4.94 × 10−8). In silico investigations of each locus identified multiple expression quantitative trait loci and potentially relevant candidate genes warranting further analysis. Thus, we identified multiple genetic loci predicting treatment outcomes for anti-VEGF therapies in DME. This work may potentially lead to managing DME using personalized treatment approaches.

Objectives: To assess whether insulin therapy impacts the effectiveness of anti-vascular endothelial growth factor (anti-VEGF) injection for the treatment of diabetic macular edema (DME) in type 2 diabetes mellitus., Methods: This was a retrospective multi-center analysis. The best-corrected visual acuity (BCVA) at 12 months, BCVA change, central macular thickness (CMT), CMT change, and cumulative injection number were compared between the insulin and the oral hypoglycemic agent (OHA) groups., Results: The mean final BCVA and CMT improved in both the insulin (N = 137; p < 0.001; p < 0.001, respectively) and the OHA group (N = 61; p = 0.199; p < 0.001, respectively). The two treatment groups were comparable for final BCVA (p = 0.263), BCVA change (p = 0.184), final CMT (p = 0.741), CMT change (p = 0.458), and the cumulative injections received (p = 0.594). The results were comparable between the two groups when stratified by baseline vision (p > 0.05) and baseline HbA1c (p > 0.05)., Conclusion: Insulin therapy does not alter treatment outcomes for anti-VEGF therapy in DME., (© 2022. The Author(s).)

To identify functionally related genes associated with diabetic retinopathy (DR) risk using gene set enrichment analyses applied to genome-wide association study meta-analyses., Methods: We analyzed DR GWAS meta-analyses performed on 3246 Europeans and 2611 African Americans with type 2 diabetes. Gene sets relevant to 5 key DR pathophysiology processes were investigated: tissue injury, vascular events, metabolic events and glial dysregulation, neuronal dysfunction, and inflammation. Keywords relevant to these processes were queried in 4 pathway and ontology databases. Two GSEA methods, Meta-Analysis Gene set Enrichment of variaNT Associations (MAGENTA) and Multi-marker Analysis of GenoMic Annotation (MAGMA), were used. Gene sets were defined to be enriched for gene associations with DR if the P value corrected for multiple testing (Pcorr) was <.05., Results: Five gene sets were significantly enriched for numerous modest genetic associations with DR in one method (MAGENTA or MAGMA) and also at least nominally significant (uncorrected P < .05) in the other method. These pathways were regulation of the lipid catabolic process (2-fold enrichment, Pcorr = .014); nitric oxide biosynthesis (1.92-fold enrichment, Pcorr = .022); lipid digestion, mobilization, and transport (1.6-fold enrichment, P = .032); apoptosis (1.53-fold enrichment, P = .041); and retinal ganglion cell degeneration (2-fold enrichment, Pcorr = .049). The interferon gamma (IFNG) gene, previously implicated in DR by protein-protein interactions in our GWAS, was among the top ranked genes in the nitric oxide pathway (best variant P = .0001)., Conclusions: These GSEA indicate that variants in genes involved in oxidative stress, lipid transport and catabolism, and cell degeneration are enriched for genes associated with DR risk. NOTE: Publication of this article is sponsored by the American Ophthalmological Society., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Multiple sclerosis (MS) is a chronic autoimmune and neurodegenerative disease that results in immune cell infiltration of the central nervous system (CNS) and demyelination in young adults. Substantial progress has been made in developing disease modifying therapies for people with relapsing-remitting MS, but options remain limited for people with primary progressive MS (PPMS). PPMS accounts for ∼15% of MS diagnoses. Herein, we generated a human induced pluripotent stem cell line (hiPSC) from a person with clinically definite PPMS. This disease-specific hiPSC line will be useful for studying PPMS in vitro, allowing the generation of immune and CNS cell types., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Cataract is the leading cause of blindness worldwide. Congenital or paediatric cataract can result in permanent visual impairment or blindness even with best attempts at treatment. A significant proportion of paediatric cataract has a genetic cause. Therefore, identifying the genes that lead to cataract formation is essential for understanding the pathological process of inherited paediatric cataract as well as to the development of new therapies. Despite clear progress in genomics technologies, verification of the biological effects of newly identified candidate genes and variants is still challenging. Here, we provide a step-by-step pipeline to evaluate cataract candidate genes in F0 zebrafish using CRISPR-Cas9 ribonucleoprotein complexes (RNP). Detailed descriptions of CRISPR-Cas9 RNP design and formulation, microinjection, optimization of CRISPR-Cas9 RNP reagent dose and delivery route, editing efficacy analysis as well as cataract formation evaluation are included. Following this protocol, any cataract candidates can be readily and efficiently evaluated within 2 weeks using basic laboratory supplies., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: Age-related cataract is the leading cause of blindness worldwide. Variants in the EPHA2 gene increase the disease risk, and its knockout in mice causes cataract. We investigated whether age, sex, and genetic background, risk factors for age-related cataract, and Epha2 genotype influence Epha2-related cataract development in mice., Methods: Cataract development was monitored in Epha2+/+, Epha2+/-, and Epha2-/- mice (Epha2Gt(KST085)Byg) on C57BL/6J and FVB:C57BL/6J (50:50) backgrounds. Cellular architecture of lenses, endoplasmic reticulum (ER) stress, and redox state were determined using histological, molecular, and analytical techniques., Results: Epha2-/- and Epha2+/- mice on C57BL/6J background developed severe cortical cataracts by 18 and 38 weeks of age, respectively, compared to development of similar cataract significantly later in Epha2-/- mice and no cataract in Epha2+/- mice in this strain on FVB background, which was previously reported. On FVB:C57BL/6J background, Epha2-/- mice developed severe cortical cataract by 38 weeks and Epha2+/- mice exhibited mild cortical cataract up to 64 weeks of age. Progression of cataract in Epha2-/- and Epha2+/- female mice on C57BL/6J and mixed background, respectively, was slower than in matched male mice. N-cadherin and β-catenin immunolabeling showed disorganized lens fiber cells and disruption of lens architecture in Epha2-/- and Epha2+/- lenses, coinciding with development of severe cataracts. EPHA2 immunolabeling showed intracellular accumulation of the mutant EPHA2-β-galactosidase fusion protein that induced a cytoprotective ER stress response and in Epha2+/- lenses was also accompanied by glutathione redox imbalance., Conclusions: Both, Epha2-/- and Epha2+/- mice develop age-related cortical cataract; age as a function of Epha2 genotype, sex, and genetic background influence Epha2-related cataractogenesis in mice.

Fuchs' endothelial corneal dystrophy (FECD) is a progressive vision impairing disease caused by thickening of Descemet's membrane and gradual degeneration and loss of corneal endothelial cells. The aim of this study was to identify differentially expressed genes between FECD-affected and unaffected corneal endothelium to gain insight into the pathophysiological mechanisms underlying this disease. Microarray gene expression analysis was performed on total RNA from FECD-affected and unaffected corneal endothelium-Descemet's membrane (CE-DM) specimens using the Illumina HumanHT-12 v4.0 expression array. RNA from pools of FECD-affected (n = 3 per pool) and individual unaffected (n = 3) specimens was used for comparison. Altered expression of a sub-set of differentially expressed genes was validated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) in independent specimens. Bioinformatics analysis was performed using InnateDB to reveal functional relationships among the differentially expressed genes and molecular pathways involved in the disease. A total of 16,513 genes were found expressed in the corneal endothelium of which 142 genes were differentially expressed between FECD-affected and unaffected endothelium (log 2 fold-change ≥1.5, corrected p-value ≤0.05). Most of the genes were up-regulated (126) and a small proportion down-regulated (16) in affected corneal endothelium. Of the twelve genes prioritised for validation, differential expression of 10 genes, including those ranked 57th and 81st by significance validated by qRT-PCR (8 up-regulated and 2 downregulated, corrected p ≤ 0.05), one gene showed a trend for up-regulation in affected endothelium, consistent with the microarray analysis and another was up-regulated in an independent study indicating robustness of the differential expression dataset. Bioinformatic analysis revealed significant over-representation of differentially expressed genes in extracellular matrix reorganisation, cellular remodelling, immune response, and inflammation. Network analysis showed functional inter-relatedness of the majority of the dysregulated genes and revealed known direct functional relationships between 20 of the genes; many of these genes have roles in macrophage differentiation, phagocytosis and inflammation. This is the second report of microarray gene expression analysis in FECD. This study revealed a set of highly dysregulated genes in the corneal endothelium in FECD. More than a third of the dysregulated genes in the disease have been discovered for the first time and thus are novel. The dysregulated genes strongly suggest the presence of phagocytic cells, most likely immune cells, and inflammation in corneal endothelium in the disease. This study provides a molecular framework for delineating the mechanisms underlying these cellular processes in FECD., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Genomic testing assesses many genes in one test. It is often used in the diagnosis of heterogeneous single gene disorders where pathogenic variation in one of many genes are known to cause similar phenotypes, or where a clinical diagnosis is difficult to reach. In the ophthalmic setting, genomic testing can be used to diagnose several groups of diseases, including inherited retinal dystrophies, paediatric cataract, glaucoma and anterior segment dysgenesis and other syndromic developmental disorders with eye involvement. The testing can encompass several modalities ranging from whole genome sequencing to exome sequencing or targeted gene panels. The advantages to the patient of receiving a molecular diagnosis include an end to the diagnostic odyssey, determination of prognosis and clarification of treatment, access to accurate genetic counselling, and confirming eligibility for clinical trials or genetic specific therapies. Genomic testing is a powerful addition to the diagnosis and management of inherited eye disease., (© 2021 Royal Australian and New Zealand College of Ophthalmologists.)

Inherited paediatric cataract is a rare Mendelian disease that results in visual impairment or blindness due to a clouding of the eye's crystalline lens. Here we report an Australian family with isolated paediatric cataract, which we had previously mapped to Xq24. Linkage at Xq24-25 (LOD = 2.53) was confirmed, and the region refined with a denser marker map. In addition, two autosomal regions with suggestive evidence of linkage were observed. A segregating 127 kb deletion (chrX:g.118373226&#95;118500408del) in the Xq24-25 linkage region was identified from whole-genome sequencing data. This deletion completely removed a commonly deleted long non-coding RNA gene LOC101928336 and truncated the protein coding progesterone receptor membrane component 1 (PGRMC1) gene following exon 1. A literature search revealed a report of two unrelated males with non-syndromic intellectual disability, as well as congenital cataract, who had contiguous gene deletions that accounted for their intellectual disability but also disrupted the PGRMC1 gene. A morpholino-induced pgrmc1 knockdown in a zebrafish model produced significant cataract formation, supporting a role for PGRMC1 in lens development and cataract formation. We hypothesise that the loss of PGRMC1 causes cataract through disrupted PGRMC1-CYP51A1 protein-protein interactions and altered cholesterol biosynthesis. The cause of paediatric cataract in this family is the truncating deletion of PGRMC1, which we report as a novel cataract gene., (© 2021. The Author(s).)

Objective: To compare the visual outcomes of intravitreal antivascular endothelial growth factor (anti-VEGF) injections in neovascular age-related macular degeneration (nAMD), diabetic macular oedema (DMO) and retinal vein occlusion (RVO) in a real-world setting., Methods and Analysis: Retrospective analysis of data from the Tasmanian Ophthalmic Biobank database. The median change in best-corrected visual acuity (BCVA) between baseline and 12 months post initiating intravitreal anti-VEGF treatment were compared between the three diseases. Final BCVA, central macular thickness (CMT), cumulative number of injections and overall predictors of change in BCVA and CMT were also determined., Results: At 12 months, change in BCVA was significantly different between nAMD, DMO and RVO cohorts (p=0.032), with lower median change for DMO (2 letters, range -5 to 20) than for RVO (11 letters, range -20 to 35). Likewise, CMT change was significantly different between the three cohorts (p=0.022), with a smaller reduction in CMT in DMO (-54 µm, range -482 to 50) than RVO patients (-137 µm, range -478 to 43; p=0.033). Total number of injections received (p=0.028) and final BCVA score (p=0.024) were also significantly different between the groups. Baseline BCVA was a negative predictor (p=0.042) and baseline CMT a positive predictor (p<0.001) of outcome. After adjusting for baseline BCVA and CMT, diagnosis of nAMD or RVO was a predictor of visual improvement compared with the DMO., Conclusions: At the end of 12 months, nAMD and RVO cohorts had the greatest improvement in BCVA, however the final BCVA for DMO was significantly better than for nAMD., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Purpose: Over a third of patients with Acanthamoeba keratitis (AK) experience severe inflammatory complications (SICs). This study aimed to determine if some contact lens (CL) wearers with AK were predisposed to SICs due to variations in key immune genes., Methods: CL wearers with AK who attended Moorfields Eye Hospital were recruited prospectively between April 2013 and October 2014. SICs were defined as scleritis and/or stromal ring infiltrate. Genomic DNA was processed with an Illumina Low Input Custom Amplicon assay of 58 single nucleotide polymorphism (SNP) targets across 18 genes and tested for association in PLINK., Results: Genomic DNA was obtained and analyzed for 105 cases of AK, 40 (38%) of whom experienced SICs. SNPs in the CXCL8 gene encoding IL-8 was significantly associated with protection from SICs (chr4: rs1126647, odds ratio [OR] = 0.3, P = 0.005, rs2227543, OR = 0.4, P = 0.007, and rs2227307, OR = 0.4, P = 0.02) after adjusting for age, sex, steroids prediagnosis, and herpes simplex keratitis (HSK) misdiagnosis. Two TLR-4 SNPs were associated with increased risk of SICs (chr9: rs4986791 and rs4986790, both OR = 6.9, P = 0.01). Th-17 associated SNPs (chr1: IL-23R rs11209026, chr2: IL-1β rs16944, and chr12: IL-22 rs1179251) were also associated with SICs., Conclusions: The current study identifies biologically relevant genetic variants in patients with AK with SICs; IL-8 is associated with a strong neutrophil response in the cornea in AK, TLR-4 is important in early AK disease, and Th-17 genes are associated with adaptive immune responses to AK in animal models. Genetic screening of patients with AK to predict severity is viable and this would be expected to assist disease management.

Keratoconus is characterised by reduced rigidity of the cornea with distortion and focal thinning that causes blurred vision, however, the pathogenetic mechanisms are unknown. It can lead to severe visual morbidity in children and young adults and is a common indication for corneal transplantation worldwide. Here we report the first large scale genome-wide association study of keratoconus including 4,669 cases and 116,547 controls. We have identified significant association with 36 genomic loci that, for the first time, implicate both dysregulation of corneal collagen matrix integrity and cell differentiation pathways as primary disease-causing mechanisms. The results also suggest pleiotropy, with some disease mechanisms shared with other corneal diseases, such as Fuchs endothelial corneal dystrophy. The common variants associated with keratoconus explain 12.5% of the genetic variance, which shows potential for the future development of a diagnostic test to detect susceptibility to disease.

Nuclear cataract is the most common type of age-related cataract and a leading cause of blindness worldwide. Age-related nuclear cataract is heritable (h 2  = 0.48), but little is known about specific genetic factors underlying this condition. Here we report findings from the largest to date multi-ethnic meta-analysis of genome-wide association studies (discovery cohort N = 14,151 and replication N = 5299) of the International Cataract Genetics Consortium. We confirmed the known genetic association of CRYAA (rs7278468, P = 2.8 × 10 -16 ) with nuclear cataract and identified five new loci associated with this disease: SOX2-OT (rs9842371, P = 1.7 × 10 -19 ), TMPRSS5 (rs4936279, P = 2.5 × 10 -10 ), LINC01412 (rs16823886, P = 1.3 × 10 -9 ), GLTSCR1 (rs1005911, P = 9.8 × 10 -9 ), and COMMD1 (rs62149908, P = 1.2 × 10 -8 ). The results suggest a strong link of age-related nuclear cataract with congenital cataract and eye development genes, and the importance of common genetic variants in maintaining crystalline lens integrity in the aging eye.

Diabetic retinopathy (DR) is the most common microvascular complication of diabetes mellitus (DM). DR is complex and the term encompasses several clinical subtypes of diabetic eye disease, including diabetic macular edema (DME), the most frequent cause of central vision loss in DR patients. Both genetic and environmental factors contribute to the pathophysiology of DR and its subtypes. While numerous studies have identified several susceptibility genes for DR, few have investigated the impact of genetics on DME susceptibility. This review will focus on the current literature surrounding genetic risk factors associated with DME. We will also highlight the small number of studies investigating the genetics of response to antivascular endothelial growth factor (anti-VEGF) injection, which is used to treat DME., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2020 Rajya L. Gurung et al.)

Keratoconus, a progressive corneal ectasia, is a complex disease with both genetic and environmental risk factors. The exact etiology is not known and is likely variable between individuals. Conditions such as hay fever and allergy are associated with increased risk, while diabetes may be protective. Behaviors such as eye rubbing are also implicated, but direct causality has not been proven. Genetics plays a major role in risk for some individuals, with many large pedigrees showing autosomal inheritance patterns. Several genes have been implicated using linkage and follow-up sequencing in these families. Genome-wide association studies for keratoconus and for quantitative traits such as central corneal thickness have identified several genetic loci that contribute to a cumulative risk for keratoconus, even in people without a family history of the disease. Identification of risk genes for keratoconus is improving our understanding of the biology of this complex disease.

Purpose: Developmental abnormalities of the ocular anterior segment in some cases can lead to ocular hypertension and glaucoma. CPAMD8 is a gene of unknown function recently associated with ocular anterior segment dysgenesis, myopia, and ectopia lentis. We sought to assess the contribution of biallelic CPAMD8 variants to childhood and juvenile open-angle glaucoma., Design: Retrospective, multicenter case series., Participants: A total of 268 probands and their relatives with a diagnosis of childhood or juvenile open-angle glaucoma., Purpose: Developmental abnormalities of the ocular anterior segment in some cases can lead to ocular hypertension and glaucoma. CPAMD8 is a gene of unknown function recently associated with ocular anterior segment dysgenesis, myopia, and ectopia lentis. We sought to assess the contribution of biallelic CPAMD8 variants to childhood and juvenile open-angle glaucoma., Methods: Patients underwent a comprehensive ophthalmic assessment, with DNA from patients and their relatives subjected to genome, exome, or capillary sequencing. CPAMD8 RNA expression analysis was performed on tissues dissected from cadaveric human eyes., Main Outcome Measures: Diagnostic yield within a cohort of childhood and juvenile open-angle glaucoma, prevalence and risk of ophthalmic phenotypes, and relative expression of CPAMD8 in the human eye., Results: We identified rare (allele frequency < 4×10 -5 ) biallelic CPAMD8 variants in 5.7% (5/88) of probands with childhood glaucoma and 2.1% (2/96) of probands with juvenile open-angle glaucoma. When including family members, we identified 11 individuals with biallelic variants in CPAMD8 from 7 unrelated families. Nine of these individuals were diagnosed with glaucoma (9/11, 81.8%), with a mean age at diagnosis of 9.22±14.89 years, and all individuals with glaucoma required 1 or more incisional procedures to control high intraocular pressure. Iris abnormalities were observed in 9 of 11 individuals, cataract was observed in 8 of 11 individuals (72.7%), and retinal detachment was observed in 3 of 11 individuals (27.3%). CPAMD8 expression was highest in neural crest-derived tissues of the adult anterior segment, suggesting that CPAMD8 variation may cause malformation or obstruction of key drainage structures., Conclusions: Biallelic CPAMD8 variation was associated with a highly heterogeneous phenotype and in our cohorts was the second most common inherited cause of childhood glaucoma after CYP1B1 and juvenile open-angle glaucoma after MYOC. CPAMD8 sequencing should be considered in the investigation of both childhood and juvenile open-angle glaucoma, particularly when associated with iris abnormalities, cataract, or retinal detachment., (Copyright © 2020 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Nanophthalmos and posterior microphthalmos are ocular abnormalities in which both eyes are abnormally small, and typically associated with extreme hyperopia. We recruited 40 individuals from 13 kindreds with nanophthalmos or posterior microphthalmos, with 12 probands subjected to exome sequencing. Nine probands (69.2%) were assigned a genetic diagnosis, with variants in MYRF, TMEM98, MFRP, and PRSS56. Two of four PRSS56 families harbored the previously described c.1066dupC variant implicated in over half of all reported PRSS56 kindreds, with different surrounding haplotypes in each family suggesting a mutational hotspot. Individuals with a genetic diagnosis had shorter mean axial lengths and higher hyperopia than those without, with recessive forms associated with the most extreme phenotypes. These findings detail the genetic architecture of nanophthalmos and posterior microphthalmos in a cohort of predominantly European ancestry, their relative clinical phenotypes, and highlight the shared genetic architecture of rare and common disorders of refractive error., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Most genetic susceptibility to cutaneous melanoma remains to be discovered. Meta-analysis genome-wide association study (GWAS) of 36,760 cases of melanoma (67% newly genotyped) and 375,188 controls identified 54 significant (P < 5 × 10 -8 ) loci with 68 independent single nucleotide polymorphisms. Analysis of risk estimates across geographical regions and host factors suggests the acral melanoma subtype is uniquely unrelated to pigmentation. Combining this meta-analysis with GWAS of nevus count and hair color, and transcriptome association approaches, uncovered 31 potential secondary loci for a total of 85 cutaneous melanoma susceptibility loci. These findings provide insights into cutaneous melanoma genetic architecture, reinforcing the importance of nevogenesis, pigmentation and telomere maintenance, together with identifying potential new pathways for cutaneous melanoma pathogenesis.

Importance: Keratoconus is a condition in which the cornea progressively thins and protrudes in a conical shape, severely affecting refraction and vision. It is a major indication for corneal transplant. To discover new genetic loci associated with keratoconus and better understand the causative mechanism of this disease, we performed a genome-wide association study on patients with keratoconus., Objective: To identify genetic susceptibility regions for keratoconus in the human genome., Design, Setting, and Participants: This study was conducted with data from eye clinics in Australia, the United States, and Northern Ireland. The discovery cohort of individuals with keratoconus and control participants from Australia was genotyped using the Illumina HumanCoreExome single-nucleotide polymorphism array. After quality control and data cleaning, genotypes were imputed against the 1000 Genomes Project reference panel (phase III; version 5), and association analyses were completed using PLINK version 1.90. Single-nucleotide polymorphisms with P < 1.00 × 10-6 were assessed for replication in 3 additional cohorts. Control participants were drawn from the cohorts of the Blue Mountains Eye Study and a previous study of glaucoma. Replication cohorts were from a previous keratoconus genome-wide association study data set from the United States, a cohort of affected and control participants from Australia and Northern Ireland, and a case-control cohort from Victoria, Australia. Data were collected from January 2006 to March 2019., Main Outcomes and Measures: Associations between keratoconus and 6 252 612 genetic variants were estimated using logistic regression after adjusting for ancestry using the first 3 principal components., Results: The discovery cohort included 522 affected individuals and 655 control participants, while the replication cohorts included 818 affected individuals (222 from the United States, 331 from Australia and Northern Ireland, and 265 from Victoria, Australia) and 3858 control participants (2927 from the United States, 229 from Australia and Northern Ireland, and 702 from Victoria, Australia). Two novel loci reached genome-wide significance (defined as P < 5.00 × 10-8), with a P value of 7.46 × 10-9 at rs61876744 in patatin-like phospholipase domain-containing 2 gene (PNPLA2) on chromosome 11 and a P value of 6.35 × 10-12 at rs138380, 2.2 kb upstream of casein kinase I isoform epsilon gene (CSNK1E) on chromosome 22. One additional locus was identified with a P value less than 1.00 × 10-6 in mastermind-like transcriptional coactivator 2 (MAML2) on chromosome 11 (P = 3.91 × 10-7). The novel locus in PNPLA2 reached genome-wide significance in an analysis of all 4 cohorts (P = 2.45 × 10-8)., Conclusions and Relevance: In this relatively large keratoconus genome-wide association study, we identified a genome-wide significant locus for keratoconus in the region of PNPLA2 on chromosome 11.

Glaucoma, a disease characterized by progressive optic nerve degeneration, can be prevented through timely diagnosis and treatment. We characterize optic nerve photographs of 67,040 UK Biobank participants and use a multitrait genetic model to identify risk loci for glaucoma. A glaucoma polygenic risk score (PRS) enables effective risk stratification in unselected glaucoma cases and modifies penetrance of the MYOC variant encoding p.Gln368Ter, the most common glaucoma-associated myocilin variant. In the unselected glaucoma population, individuals in the top PRS decile reach an absolute risk for glaucoma 10 years earlier than the bottom decile and are at 15-fold increased risk of developing advanced glaucoma (top 10% versus remaining 90%, odds ratio = 4.20). The PRS predicts glaucoma progression in prospectively monitored, early manifest glaucoma cases (P = 0.004) and surgical intervention in advanced disease (P = 3.6 × 10 - 6 ). This glaucoma PRS will facilitate the development of a personalized approach for earlier treatment of high-risk individuals, with less intensive monitoring and treatment being possible for lower-risk groups.

Age-related cataract is the major cause of blindness worldwide. Both genetic and environmental factors contribute to the disease. Genetic variation in the Ephrin type-A receptor 2 (EPHA2) gene is associated with the risk of age-related cataract in multiple populations, and exposure to ultraviolet-B (UV-B) radiation is a well-established risk factor for the disease. Epha2 knockout and UV-B radiation independently lead to cataract in mice, and UV-B radiation reportedly alters EPHA2 expression in cultured cells. We hypothesised that an interaction between UV-B radiation exposure and Epha2 signalling may influence cataract development. To test this hypothesis, 5-week-old Epha2 +/+ and Epha2 +/- mice (n = 8 per group) were exposed to repeated below-threshold doses of UV-B radiation (0.0125-0.05 J/cm 2 ), before development of Epha2-mediated cataract. Cataract development was monitored after termination of exposure and at least one month later. Histological analysis of exposed and unexposed lenses was performed to assess pathological changes, and gene expression analysis to investigate the mechanism underlying cataract. Both Epha2 +/+ and Epha2 +/- mice developed UV-B dose-dependent anterior polar cataract; cataract severity in both genotypes of mice exposed to either 0.025 or 0.05 J/cm 2 UV-B was significantly higher than that in matched unexposed mice (p < 0.05). Histological analysis of lenses of both genotypes of mice exposed to 0.025 or 0.05 J/cm 2 UV-B radiation consistently revealed disruption of the lens architecture. A month after the exposure, cataract severity increased in Epha2 +/+ mice treated with the highest dose of UV-B radiation (p = 0.03) but remained unchanged in Epha2 +/- mice. Gene expression analysis of lenses of both genotypes of mice showed significant upregulation of the cell proliferation marker Mki67 in Epha2 +/+ (p = 0.036) but not in Epha2 +/- mice exposed to the highest dose of UV-B radiation compared to matched unexposed mice. In conclusion, this study suggests that repeated exposure to doses of UV-B radiation lower than the single minimum dose required for inducing cataract leads to cataract in wild-type and Epha2 heterozygous knockout mice. Furthermore, this study indicates, for the first time, a potentially favourable effect of partial Epha2 deficiency against UV radiation-induced damage in the lens., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Fuchs endothelial corneal dystrophy (FECD) is a progressive and potentially a sight threatening disease, and a common indication for corneal grafting in the elderly. Aberrant thickening of Descemet's membrane, formation of microscopic excrescences (guttae) and gradual loss of corneal endothelial cells are the hallmarks of the disease. The aim of this study was to identify differentially abundant proteins between FECD-affected and unaffected Descemet's membrane., Methods: Label-free quantitative proteomics using nanoscale ultra-performance liquid chromatography-mass spectrometry (nUPLC-MS E ) was employed on affected and unaffected Descemet's membrane extracts, and interesting findings were further investigated using quantitative reverse transcription-polymerase chain reaction and immunohistochemical techniques., Results: Quantitative proteomics revealed significantly lower abundance of apolipoprotein E (APOE) and immunoglobulin heavy constant gamma 1 protein (IGHG1) in affected Descemet's membrane. The difference in the distribution of APOE between affected and unaffected Descemet's membrane and of IGHG1 detected by immunohistochemistry support their down-regulation in the disease. Comparative gene expression analysis showed significantly lower APOE mRNA levels in FECD-affected than unaffected corneal endothelium. IGHG1 gene is expressed at extremely low levels in the corneal endothelium, precluding relative expression analysis., Conclusions: This is the first study to report comparative proteomics of Descemet's membrane tissue, and implicates dysregulation of APOE and IGHG1 proteins in the pathogenesis of Fuchs endothelial corneal dystrophy., (© 2019 Royal Australian and New Zealand College of Ophthalmologists.)

Purpose: Few studies have explored the association of genetic variants in microRNA genes and binding sites with diabetic retinopathy (DR) in type 1 diabetes. We conducted a genome-wide scan for single nucleotide polymorphisms (SNPs) in these genes by using data from a genome-wide association study (GWAS)., Methods: All known SNPs were imputed from our GWAS data (n = 325) of DR cases and diabetic controls (no DR). Relevant SNPS were extracted using miRNASNP and PolymiRTS (version 2) databases. χ2 tests and logistic regression (adjusting for age, sex, duration of diabetes, HbA1c, and hypertension) were used to test the association between the imputed SNPs and DR phenotypes (any DR, nonproliferative DR [NPDR], proliferative DR [PDR], diabetic macular edema [DME], and sight-threatening DR defined as PDR, severe NPDR, or clinically significant macula edema [CSME]) compared with diabetic controls. Top-ranking SNPs were genotyped in a larger cohort (N = 560) to confirm their association with DR., Results: Three SNPs (rs10061133, rs1049835, rs9501255) were selected and genotyped in the final cohort. Rs10061133 in MIR449b was protective of sight-threatening DR (odds ratio [OR] = 0.32, P = 3.68 × 10-4) and PDR (OR = 0.30, P = 8.12 × 10-4), and the associations became more significant as the cohort increased in size., Conclusions: Rs10061133 in MIR449b is significantly associated with a decreased risk of PDR and sight-threatening DR in Caucasian patients with type 1 diabetes. This can guide future studies on genetic risk profiling and on developing microRNA-related therapies for sight-threatening DR.

Purpose: To investigate which clinical measures influence whether an individual demonstrates earliest glaucomatous structural progression on peripapillary retinal nerve fiber layer (pRNFL) or macular ganglion cell-inner plexiform layer (mGCIPL)., Design: Prospective, longitudinal cohort study., Participants: Two hundred seventy-one eyes from 207 individuals with statistically significant evidence of glaucomatous progression on OCT Guided Progression Analysis (GPA) software were drawn from a total of 1271 eyes from 686 individuals categorized as glaucoma suspect or having early manifest glaucoma undergoing glaucoma surveillance., Methods: Individuals demonstrating earliest evidence of longitudinal progression on mGCIPL GPA event analysis were compared with individuals demonstrating evidence of earliest longitudinal progression on pRNFL GPA event analysis., Main Outcome Measures: Correlation of OCT event change analysis with intraocular pressure (IOP), clinical variables, and baseline thickness of the pRNFL and mGCIPL., Results: Intraocular pressure, baseline pRNFL thickness, baseline mGCIPL thickness, and systemic hypertension were associated with location of first progression. Eyes demonstrating earliest longitudinal progression on mGCIPL had significantly lower maximum-recorded pretreatment IOP (mean difference, 3.90 mmHg; 95% confidence interval [CI], 2.37-5.43 mmHg; P < 0.001). The interval between progression on pRNFL and progression on mGCIPL increased by 12.4 months for every 5-mmHg increase in IOP (95% CI, 10.32-15.72 months). Eyes demonstrating earliest longitudinal progression on mGCIPL showed significantly lower baseline average pRNFL thickness than eyes progressing on pRNFL first (mean difference, 7.07 μm; 95% CI, 4.38-9.77 μm; P < 0.001). Eyes progressing first on mGCIPL parameters were 3.03 times more likely to demonstrate a new paracentral field defect than eyes progressing first on pRNFL parameters (odds ratio, 3.03; 95% CI, 1.26-7.28; P = 0.01)., Conclusions: Clinical features, particularly pretreatment IOP, influence whether structural glaucoma progression is detected earlier with mGCIPL or pRNFL imaging. These data support the usefulness of mGCIPL imaging in addition to pRNFL analysis for detection of glaucoma progression, particularly in patients with normal IOP., (Copyright © 2019 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Importance: Both primary and secondary forms of childhood glaucoma have many distinct causative mechanisms, and in many cases a cause is not immediately clear. The broad phenotypic spectrum of secondary glaucoma, particularly in individuals with variants in FOXC1 or PITX2 genes associated with Axenfeld-Rieger syndrome, makes it more difficult to diagnose patients with milder phenotypes. These cases are occasionally classified and managed as primary congenital glaucoma., Objective: To investigate the prevalence of FOXC1 variants in participants with a suspected diagnosis of primary congenital glaucoma., Design, Setting, and Participants: Australian and Italian cohorts were recruited from January 1, 2007, through March 1, 2016. Australian individuals were recruited through the Australian and New Zealand Registry of Advanced Glaucoma and Italian individuals through the Genetic and Ophthalmology Unit of l'Azienda Socio-Sanitaria Territoriale Grande Ospedale Metropolitano Niguarda in Milan, Italy. We performed exome sequencing, in combination with Sanger sequencing and multiplex ligation-dependent probe amplification, to detect variants of FOXC1 in individuals with a suspected diagnosis of primary congenital glaucoma established by their treating specialist. Data analysis was completed from June 2015 to November 2017., Main Outcome and Measures: Identification of single-nucleotide and copy number variants in FOXC1, along with phenotypic characterization of the individuals who carried them., Results: A total of 131 individuals with a suspected diagnosis of primary congenital glaucoma were included. The mean (SD) age at recruitment in the Australian cohort was 24.3 (18.1) years; 37 of 84 Australian participants (44.0%) were female, and 71 of 84 (84.5%) were of European ancestry. The mean (SD) age at recruitment was 22.5 (18.4) years in the Italian cohort; 21 of 47 Italian participants (44.7%) were female, and 45 of 47 (95.7%) were of European ancestry. We observed rare, predicted deleterious FOXC1 variants in 8 of 131 participants (6.1%), or 8 of 166 participants (4.8%) when including those explained by variants in CYP1B1. On reexamination or reinvestigation, all of these individuals had at least 1 detectable ocular and/or systemic feature associated with Axenfeld-Rieger syndrome., Conclusions and Relevance: These data highlight the genetic and phenotypic heterogeneity of childhood glaucoma and support the use of gene panels incorporating FOXC1 as a diagnostic aid, especially because clinical features of Axenfeld-Rieger syndrome can be subtle. Further replication of these results will be needed to support the future use of such panels.

A super-enhancer (SE) is a cluster of enhancers with a relatively high density of particular chromatin features. SEs typically regulate key genes that can determine cell identity and differentiation. Identifying SEs and their effects may be critical in predicting key regulatory genes, such as master transcription factor genes or oncogenes. Signal inducible SEs are dense stretches of signal terminal transcription factor (TF) binding regions, and may modulate the interaction between environmental factors (e.g., Vitamin D) and genetic factors (i.e., risk variants) in complex diseases such as multiple sclerosis (MS). As a complex autoimmune disease, the etiology and progression of MS, including the interaction between Vitamin D and MS risk variants, is still unclear and can be explored from the aspect of signal SEs. Vitamin D [with its active form: 1,25(OH) 2 D 3 ], is an environmental risk factor for MS. It binds the Vitamin D receptor (VDR) and regulates gene expression. This study explores the association between VDR super-enhancers (VSEs) and MS risk variants. Firstly, we reanalyse public ChIP-seq and RNA-seq data to classify VSEs into three categories according to their combinations of persistent and secondary VDR binding. Secondly, we indicate the genes with VSE regions that are near MS risk variants. Furthermore, we find that MS risk variants are enriched in VSE regions, and we indicate some genes with a VSE overlapping MS risk variant for further exploration. We also find two clusters of genes from the set of genes showing correlation of expression patterns with the MS risk gene ZMIZ1 that appear to be regulated by VSEs in THP-1 cells. It is the first time that VSEs have been analyzed, and we directly connect the genetic risk factors for MS risk with Vitamin D based on VSEs.

To identify genetic variants associated with diabetic retinopathy (DR), we performed a large multiethnic genome-wide association study. Discovery included eight European cohorts ( n = 3,246) and seven African American cohorts ( n = 2,611). We meta-analyzed across cohorts using inverse-variance weighting, with and without liability threshold modeling of glycemic control and duration of diabetes. Variants with a P value <1 × 10 -5 were investigated in replication cohorts that included 18,545 European, 16,453 Asian, and 2,710 Hispanic subjects. After correction for multiple testing, the C allele of rs142293996 in an intron of nuclear VCP-like ( NVL ) was associated with DR in European discovery cohorts ( P = 2.1 × 10 -9 ), but did not reach genome-wide significance after meta-analysis with replication cohorts. We applied the Disease Association Protein-Protein Link Evaluator (DAPPLE) to our discovery results to test for evidence of risk being spread across underlying molecular pathways. One protein-protein interaction network built from genes in regions associated with proliferative DR was found to have significant connectivity ( P = 0.0009) and corroborated with gene set enrichment analyses. These findings suggest that genetic variation in NVL, as well as variation within a protein-protein interaction network that includes genes implicated in inflammation, may influence risk for DR., (© 2018 by the American Diabetes Association.)

Mitochondrial haplogroups H1, H2 and UK have previously been reported to be associated with proliferative diabetic retinopathy (PDR) in Caucasian patients with diabetes. We aimed to replicate this finding with a larger sample and expand the analysis to include different severities of DR, and diabetic macular edema (DME). Caucasian participants (n = 2935) with either type 1 or type 2 diabetes from the Australian Registry of Advanced Diabetic Retinopathy were enrolled in this study. Twenty-two mitochondrial single nucleotide polymorphisms were genotyped by MassArray and haplogroups reconstructed using Haplogrep. Chi square tests and logistic regressions were used to test associations between haplogroup and DR phenotypes including any DR, non-proliferative DR (NPDR), proliferative DR (PDR) and DME. After stratifying the samples in type 1 and type 2 diabetes groups, and adjusting for sex, age, diabetes duration, concurrent HbA1c and hypertension, neither haplogroups H1, H2, UK, K or JT were associated with any DR, NPDR, PDR or DME.

Emmanuelle Souzeau, who contributed to analysis of data, was inadvertently omitted from the author list in the originally published version of this Article. This has now been corrected in both the PDF and HTML versions of the Article.

Importance: The p.Gln368Ter (rs74315329) risk allele in the myocilin gene (MYOC) was initially reported to have high penetrance in glaucoma registry-based studies, but much lower estimates were recently obtained from population-based studies. We investigated this disparity using data from Australia and the United Kingdom., Objectives: To examine the penetrance and effect size of the MYOC p.Gln368Ter variant with glaucoma and ocular hypertension (OHT)., Design, Setting, and Participants: This cross-sectional study within the UK Biobank (UKBB) included participants of white British ancestry. Glaucoma cases were defined by International Classification of Diseases, Ninth Revision (ICD-9) and Tenth Revision (ICD-10) diagnoses and self-reported questionnaires. Carriers of the MYOC p.Gln368Ter variant were identified using genotype imputation from arrays. In contrast, 2 Australian registry-based studies, the Australian and New Zealand Registry of Advanced Glaucoma and the Glaucoma Inheritance Study in Tasmania, ascertained glaucoma cases referred by eye care clinicians, with historic control participants recruited from other Australian studies. Samples were either directly sequenced or had genotypes determined by imputation (for the Australian registry and historic control participants). Recruitment to the UKBB occurred between 2006 and 2010, and data analysis occurred from September 2017 to July 2018., Main Outcomes and Measures: The penetrance and odds ratio (OR) were estimated for the MYOC p.Gln368Ter variants in participants with glaucoma and OHT., Results: A total of 411 337 UKBB participants of white British ancestry (mean [SD] age, 56.6 [8.0] years) were included, plus 3071 Australian registry and 6750 historic control participants. In the UKBB, the minor allele frequency of the MYOC p.Gln368Ter variant was 1 in 786 individuals (0.13%). The odds ratio of p.Gln368Ter in patients with primary open-angle glaucoma (POAG) was 6.76 (95% CI, 4.05-11.29); glaucoma (POAG, self-reported glaucoma, and unspecified glaucoma), 4.40 (95% CI, 3.38-5.71); OHT, 3.56 (95% CI, 2.53-4.92); and OHT and glaucoma combined, 4.18 (95% CI, 3.05-5.67). The penetrance of the MYOC p.Gln368Ter variant was 7.6% in patients with glaucoma, 24.3% in patients with OHT, and 30.8% in patients with OHT and glaucoma combined. In the Australian registry studies, the odds of MYOC p.Gln368Ter variant were 12.16 (95% CI, 6.34-24.97) in patients with advanced glaucoma and 3.97 (95% CI, 1.55-9.75) in those with nonadvanced glaucoma; the penetrance of glaucoma was 56.1%, and penetrance in those considered to have glaucoma or be glaucoma suspects was 69.5%., Conclusions and Relevance: The MYOC p.Gln368Ter variant confers a very high-risk effect size for advanced glaucoma; the risk is lower in nonadvanced glaucoma and OHT. In the general population sample, approximately 50% of MYOC p.Gln368Ter carriers 65 years and older had glaucoma or OHT, with higher prevalence in the Australian registry studies.

Purpose: Diabetic retinopathy is the most common eye complication in patients with diabetes. The purpose of this study is to identify genetic factors contributing to severe diabetic retinopathy., Methods: A genome-wide association approach was applied. In the Genetics of Diabetes Audit and Research in Tayside Scotland (GoDARTS) datasets, cases of severe diabetic retinopathy were defined as type 2 diabetic patients who were ever graded as having severe background retinopathy (Level R3) or proliferative retinopathy (Level R4) in at least one eye according to the Scottish Diabetic Retinopathy Grading Scheme or who were once treated by laser photocoagulation. Controls were diabetic individuals whose longitudinal retinopathy screening records were either normal (Level R0) or only with mild background retinopathy (Level R1) in both eyes. Significant Single Nucleotide Polymorphisms (SNPs) were taken forward for meta-analysis using multiple Caucasian cohorts., Results: Five hundred and sixty cases of type 2 diabetes with severe diabetic retinopathy and 4,106 controls were identified in the GoDARTS cohort. We revealed that rs3913535 in the NADPH Oxidase 4 (NOX4) gene reached a p value of 4.05 × 10 -9 . Two nearby SNPs, rs10765219 and rs11018670 also showed promising p values (p values = 7.41 × 10 -8 and 1.23 × 10 -8 , respectively). In the meta-analysis using multiple Caucasian cohorts (excluding GoDARTS), rs10765219 and rs11018670 showed associations for diabetic retinopathy (p = 0.003 and 0.007, respectively), while the p value of rs3913535 was not significant (p = 0.429)., Conclusion: This genome-wide association study of severe diabetic retinopathy suggests new evidence for the involvement of the NOX4 gene., (© 2018 The Authors. Acta Ophthalmologica published by John Wiley & Sons Ltd on behalf of Acta Ophthalmologica Scandinavica Foundation.)

Intraocular pressure (IOP) is currently the sole modifiable risk factor for primary open-angle glaucoma (POAG), one of the leading causes of blindness worldwide 1 . Both IOP and POAG are highly heritable 2 . We report a combined analysis of participants from the UK Biobank (n = 103,914) and previously published data from the International Glaucoma Genetic Consortium (n = 29,578) 3,4 that identified 101 statistically independent genome-wide-significant SNPs for IOP, 85 of which have not been previously reported 4-12 . We examined these SNPs in 11,018 glaucoma cases and 126,069 controls, and 53 SNPs showed evidence of association. Gene-based tests implicated an additional 22 independent genes associated with IOP. We derived an allele score based on the IOP loci and loci influencing optic nerve head morphology. In 1,734 people with advanced glaucoma and 2,938 controls, participants in the top decile of the allele score were at increased risk (odds ratio (OR) = 5.6; 95% confidence interval (CI): 4.1-7.6) of glaucoma relative to the bottom decile.

Many genes have been suggested as candidate genes for keratoconus based on their function, their proximity to associated polymorphisms or due to the identification of putative causative variants within the gene. However, very few of these genes have been assessed for rare variation in keratoconus more broadly. In contrast, VSX1 and SOD1 have been widely assessed, however, the vast majority of studies have been small and the findings conflicting. In a cohort of Australians of European descent, consisting of 385 keratoconus cases and 396 controls, we screened 21 keratoconus candidate genes: BANP, CAST, COL4A3, COL4A4, COL5A1, FOXO1, FNDC3B, HGF, IL1A, IL1B, ILRN, IMMP2L, MPDZ, NFIB, RAB3GAP1, RAD51, RXRA, SLC4A11, SOD1, TF and VSX1. The candidate genes were sequenced in these individuals by either whole exome sequencing or targeted gene sequencing. Variants were filtered to identify rare (minor allele frequency <1%), potentially pathogenic variants. A total of 164 such variants were identified across the two groups with no variants fulfilling these criteria in cases in IL1RN, BANP, IL1B, RAD51 or SOD1. The frequency of variants was compared between cases and controls using chi-square or Fishers' Exact tests for each gene with at least one rare potentially pathogenic variant identified in the case cohort. The number of rare potentially pathogenic variants per gene ranged from three (RXRA) to 102 (MPDZ), however for all genes, there was no difference in the frequency between the cases and controls. We conclude that rare potentially pathogenic variation in the 21 candidate genes assessed do not play a major role in keratoconus susceptibility and pathogenesis., Competing Interests: Author Dr. Richard Lindsay is employed by Richard Lindsay and Associates. This does not alter our adherence to PLOS ONE policies on sharing data and materials. There are no patents, products in development or marketed products to declare.

Background: Pediatric cataract is an important cause of blindness and visual impairment in children. A large proportion of pediatric cataracts are inherited, and many genes have been described for this heterogeneous Mendelian disease. Surveys of schools for the blind in Bhutan, Cambodia, and Sri Lanka have identified many children with this condition and we aimed to identify the genetic causes of inherited cataract in these populations., Methods: We screened, in parallel, 51 causative genes for inherited cataracts in 33 probands by Ampliseq enrichment and sequencing on an Ion Torrent PGM. Rare novel protein coding variants were assessed for segregation in family members, where possible, by Sanger sequencing., Results: We identified 24 rare (frequency <1% in public databases) or novel protein coding variants in 12 probands and confirmed segregation of variants with disease in the extended family where possible. Of these, six are predicted to be the cause of disease in the patient, with four other variants also highly likely to be pathogenic., Conclusion: This study found that 20%-30% of patients in these countries have a mutation in a known cataract causing gene, which is considerably lower than the 60%-70% reported in Caucasian cohorts. This suggests that additional cataract genes remain to be discovered in this cohort of Asian pediatric cataract patients., (© 2018 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.)

Central corneal thickness (CCT) is a highly heritable trait associated with complex eye diseases such as keratoconus and glaucoma. We perform a genome-wide association meta-analysis of CCT and identify 19 novel regions. In addition to adding support for known connective tissue-related pathways, pathway analyses uncover previously unreported gene sets. Remarkably, >20% of the CCT-loci are near or within Mendelian disorder genes. These included FBN1, ADAMTS2 and TGFB2 which associate with connective tissue disorders (Marfan, Ehlers-Danlos and Loeys-Dietz syndromes), and the LUM-DCN-KERA gene complex involved in myopia, corneal dystrophies and cornea plana. Using index CCT-increasing variants, we find a significant inverse correlation in effect sizes between CCT and keratoconus (r = -0.62, P = 5.30 × 10 -5 ) but not between CCT and primary open-angle glaucoma (r = -0.17, P = 0.2). Our findings provide evidence for shared genetic influences between CCT and keratoconus, and implicate candidate genes acting in collagen and extracellular matrix regulation.