Tiger Zhou

Common variants near ABCA1 , AFAP1 and GMDS confer risk of primary open-angle glaucoma

Primary open-angle glaucoma (POAG) is a major cause of irreversible blindness worldwide. We performed a genome-wide association study in an Australian discovery cohort comprising 1,155 cases with advanced POAG and 1,992 controls. We investigated the association of the top SNPs from the discovery stage in two Australian replication cohorts (932 cases and 6,862 controls total) and two US replication cohorts (2,616 cases and 2,634 controls total). Meta-analysis of all cohorts identified three loci newly associated with development of POAG. These loci are located upstream of ABCA1 (rs2472493[G], odds ratio (OR) = 1.31, P = 2.1 × 10−19), within AFAP1 (rs4619890[G], OR = 1.20, P = 7.0 × 10−10) and within GMDS (rs11969985[G], OR = 1.31, P = 7.7 × 10−10). Using RT-PCR and immunolabeling, we show that these genes are expressed within human retina, optic nerve and trabecular meshwork and that ABCA1 and AFAP1 are also expressed in retinal ganglion cells.

Angiopoietin receptor TEK mutations underlie primary congenital glaucoma with variable expressivity

Primary congenital glaucoma (PCG) is a devastating eye disease and an important cause of childhood blindness worldwide. In PCG, defects in the anterior chamber aqueous humor outflow structures of the eye result in elevated intraocular pressure (IOP); however, the genes and molecular mechanisms involved in the etiology of these defects have not been fully characterized. Previously, we observed PCG-like phenotypes in transgenic mice that lack functional angiopoietin-TEK signaling. Herein, we identified rare TEK variants in 10 of 189 unrelated PCG families and demonstrated that each mutation results in haploinsufficiency due to protein loss of function. Multiple cellular mechanisms were responsible for the loss of protein function resulting from individual TEK variants, including an absence of normal protein production, protein aggregate formation, enhanced proteasomal degradation, altered subcellular localization, and reduced responsiveness to ligand stimulation. Further, in mice, hemizygosity for Tek led to the formation of severely hypomorphic Schlemms canal and trabecular meshwork, as well as elevated IOP, demonstrating that anterior chamber vascular development is sensitive to Tek gene dosage and the resulting decrease in angiopoietin-TEK signaling. Collectively, these results identify TEK mutations in patients with PCG that likely underlie disease and are transmitted in an autosomal dominant pattern with variable expressivity.

Meta-analysis of Genome-Wide Association Studies Identifies Novel Loci Associated With Optic Disc Morphology

Primary open‐angle glaucoma is the most common optic neuropathy and an important cause of irreversible blindness worldwide. The optic nerve head or optic disc is divided in two parts: a central cup (without nerve fibers) surrounded by the neuroretinal rim (containing axons of the retinal ganglion cells). The International Glaucoma Genetics Consortium conducted a meta‐analysis of genome‐wide association studies consisting of 17,248 individuals of European ancestry and 6,841 individuals of Asian ancestry. The outcomes of the genome‐wide association studies were disc area and cup area. These specific measurements describe optic nerve morphology in another way than the vertical cup‐disc ratio, which is a clinically used measurement, and may shed light on new glaucoma mechanisms. We identified 10 new loci associated with disc area (CDC42BPA, F5, DIRC3, RARB, ABI3BP, DCAF4L2, ELP4, TMTC2, NR2F2, and HORMAD2) and another 10 new loci associated with cup area (DHRS3, TRIB2, EFEMP1, FLNB, FAM101, DDHD1, ASB7, KPNB1, BCAS3, and TRIOBP). The new genes participate in a number of pathways and future work is likely to identify more functions related to the pathogenesis of glaucoma.

Copy number variations of TBK1 in Australian patients with primary open-angle glaucoma

PURPOSE To investigate the presence of TBK1 copy number variations in a large, well-characterized Australian cohort of patients with glaucoma comprising both normal-tension glaucoma and high-tension glaucoma cases. DESIGN A retrospective cohort study. METHODS DNA samples from patients with normal-tension glaucoma and high-tension glaucoma and unaffected controls were screened for TBK1 copy number variations using real-time quantitative polymerase chain reaction. Samples with additional copies of the TBK1 gene were further tested using custom comparative genomic hybridization arrays. RESULTS Four out of 334 normal-tension glaucoma cases (1.2%) were found to carry TBK1 copy number variations using quantitative polymerase chain reaction. One extra dose of the TBK1 gene (duplication) was detected in 3 normal-tension glaucoma patients, while 2 extra doses of the gene (triplication) were detected in a fourth normal-tension glaucoma patient. The results were further confirmed by custom comparative genomic hybridization arrays. Further, the TBK1 copy number variation segregated with normal-tension glaucoma in the family members of the probands, showing an autosomal dominant pattern of inheritance. No TBK1 copy number variations were detected in 1045 Australian patients with high-tension glaucoma or in 254 unaffected controls. CONCLUSION We report the presence of TBK1 copy number variations in our Australian normal-tension glaucoma cohort, including the first example of more than 1 extra copy of this gene in glaucoma patients (gene triplication). These results confirm TBK1 to be an important cause of normal-tension glaucoma, but do not suggest common involvement in high-tension glaucoma.

New insights into the genetics of primary open-angle glaucoma based on meta-analyses of intraocular pressure and optic disc characteristics

Primary open-angle glaucoma (POAG), the most common optic neuropathy, is a heritable disease. Siblings of POAG cases have a ten-fold increased risk of developing the disease. Intraocular pressure (IOP) and optic nerve head characteristics are used clinically to predict POAG risk. We conducted a genome-wide association meta-analysis of IOP and optic disc parameters and validated our findings in multiple sets of POAG cases and controls. Using imputation to the 1000 genomes (1000G) reference set, we identified 9 new genomic regions associated with vertical cup-disc ratio (VCDR) and 1 new region associated with IOP. Additionally, we found 5 novel loci for optic nerve cup area and 6 for disc area. Previously it was assumed that genetic variation influenced POAG either through IOP or via changes to the optic nerve head; here we present evidence that some genomic regions affect both IOP and the disc parameters. We characterized the effect of the novel loci through pathway analysis and found that pathways involved are not entirely distinct as assumed so far. Further, we identified a novel association between CDKN1A and POAG. Using a zebrafish model we show that six6b (associated with POAG and optic nerve head variation) alters the expression of cdkn1a. In summary, we have identified several novel genes influencing the major clinical risk predictors of POAG and showed that genetic variation in CDKN1A is important in POAG risk.

Genetic Association at the 9p21 Glaucoma Locus Contributes to Sex Bias in Normal-Tension Glaucoma

PURPOSE Many genome-wide association studies have identified common single nucleotide polymorphisms (SNPs) at the 9p21 glaucoma locus (CDKN2B/CDKN2B-AS1) to be significantly associated with primary open-angle glaucoma (POAG), with association being stronger in normal tension glaucoma (NTG) and advanced glaucoma. We aimed to determine whether any observed differences in genetic association at the 9p21 locus are influenced by sex. METHODS Sex was assessed as a risk factor for POAG for 2241 glaucoma participants from the Australian and New Zealand Registry of Advanced Glaucoma, the Glaucoma Inheritance Study in Tasmania, and the Flinders Medical Centre. A total of 3176 controls were drawn from the Blue Mountains Eye Study and South Australia: 1523 advanced POAG and 718 nonadvanced POAG cases were genotyped along with 3176 controls. We selected 13 SNPs at the 9p21 locus, and association results were subanalyszd by sex for high-tension glaucoma (HTG) and NTG. Odds ratios (ORs) between sexes were compared. RESULTS A sex bias was present within advanced NTG cases (57.1% female versus 42.9% male, P = 0.0026). In all POAG cases, the strongest associated SNP at 9p21 was rs1063192 (OR, 1.43; P = 4 × 10-18). This association was stronger in females (OR, 1.5; P = 5 × 10-13) than in males (OR, 1.35; P = 7 × 10-7), with a statistically significant difference in female to male OR comparison (P = 1.0 × 10-2). An NTG to HTG subanalysis yielded statistically significant results only in females (OR, 1.63; P = 1.5 × 10-4) but not in males (OR, 1.15; P = 2.8 × 10-1), with a statistically significant difference in female to male OR comparison (P = 1.4 × 10-4). CONCLUSIONS This study demonstrated that female sex is a risk factor for developing advanced NTG. The stronger genetic signals at the 9p21 locus among females may contribute at least in part to the observed sex bias for NTG.

Predictive genetic testing experience for myocilin primary open-angle glaucoma using the Australian and New Zealand Registry of Advanced Glaucoma

Purpose:Predictive genetic testing of relatives of known myocilin (MYOC) gene mutation carriers is an appropriate strategy to identify individuals at risk for glaucoma. It is likely to prevent irreversible blindness in this high-risk group because this treatable condition might otherwise be diagnosed late. The Australian and New Zealand Registry of Advanced Glaucoma has established genetic testing protocols for known glaucoma genes, including MYOC.Methods:Through the Australian and New Zealand Registry of Advanced Glaucoma, we investigated the experience of 40 unaffected individuals who had undergone predictive genetic testing for MYOC mutations through questionnaires.Results:The main motivations for being tested were (i) to make appropriate interventions and (ii) to reduce uncertainty. All our respondents perceived strong benefits, either medical or emotional, in being tested. However, different concerns were raised by the respondents that need to be addressed during counseling. Greater family awareness was reported by the majority of the respondents, and the ability to provide information to children was a strong motivation for being tested.Conclusion:This study provides valuable information on the personal and familial impacts of having predictive genetic testing for glaucoma, which will help health professionals to better address the issues faced by patients and provide them adequate support.Genet Med 16 7, 558–563.

Glaucoma spectrum and age-related prevalence of individuals with FOXC1 and PITX2 variants

Variation in FOXC1 and PITX2 is associated with Axenfeld-Rieger syndrome, characterised by structural defects of the anterior chamber of the eye and a range of systemic features. Approximately half of all affected individuals will develop glaucoma, but the age at diagnosis and the phenotypic spectrum have not been well defined. As phenotypic heterogeneity is common, we aimed to delineate the age-related penetrance and the full phenotypic spectrum of glaucoma in FOXC1 or PITX2 carriers recruited through a national disease registry. All coding exons of FOXC1 and PITX2 were directly sequenced and multiplex ligation-dependent probe amplification was performed to detect copy number variation. The cohort included 53 individuals from 24 families with disease-associated FOXC1 or PITX2 variants, including one individual diagnosed with primary congenital glaucoma and five with primary open-angle glaucoma. The overall prevalence of glaucoma was 58.5% and was similar for both genes (53.3% for FOXC1 vs 60.9% for PITX2, P=0.59), however, the median age at glaucoma diagnosis was significantly lower in FOXC1 (6.0±13.0 years) compared with PITX2 carriers (18.0±10.6 years, P=0.04). The penetrance at 10 years old was significantly lower in PITX2 than FOXC1 carriers (13.0% vs 42.9%, P=0.03) but became comparable at 25 years old (71.4% vs 57.7%, P=0.38). These findings have important implications for the genetic counselling of families affected by Axenfeld-Rieger syndrome, and also suggest that FOXC1 and PITX2 contribute to the genetic architecture of primary glaucoma subtypes.

Occurrence of CYP1B1 Mutations in Juvenile Open-Angle Glaucoma With Advanced Visual Field Loss

IMPORTANCE Juvenile open-angle glaucoma (JOAG) is a severe neurodegenerative eye disorder in which most of the genetic contribution remains unexplained. OBJECTIVE To assess the prevalence of pathogenic CYP1B1 sequence variants in an Australian cohort of patients with JOAG and severe visual field loss. DESIGN, SETTING, AND PARTICIPANTS For this cohort study, we recruited 160 patients with JOAG classified as advanced (n = 118) and nonadvanced (n = 42) through the Australian and New Zealand Registry of Advanced Glaucoma from January 1, 2007, through April 1, 2014. Eighty individuals with no evidence of glaucoma served as a control group. We defined JOAG as diagnosis before age 40 years and advanced JOAG as visual field loss in 2 of the 4 central fixation squares on a reliable visual field test result. We performed direct sequencing of the entire coding region of CYP1B1. Data analysis was performed in October 2014. MAIN OUTCOMES AND MEASURES Identification and characterization of CYP1B1 sequence variants. RESULTS We identified 7 different pathogenic variants among 8 of 118 patients with advanced JOAG (6.8%) but none among the patients with nonadvanced JOAG. Three patients were homozygous or compound heterozygous for CYP1B1 pathogenic variants, which provided a likely basis for their disease. Five patients were heterozygous. The allele frequency among the patients with advanced JOAG (11 in 236 [4.7%]) was higher than among our controls (1 in 160 [0.6%]; P = .02; odds ratio, 7.8 [95% CI, 0.02-1.0]) or among the control population from the Exome Aggregation Consortium database (2946 of 122 960 [2.4%]; P = .02; odds ratio, 2.0 [95% CI, 0.3-0.9]). Individuals with CYP1B1 pathogenic variants, whether heterozygous or homozygous, had worse mean (SD) deviation on visual fields (-24.5 [5.1] [95% CI, -31.8 to -17.2] vs -15.6 [10.0] [95% CI, -17.1 to -13.6] dB; F1,126 = 5.90; P = .02; partial ηp2 = 0.05) and were younger at diagnosis (mean [SD] age, 23.1 [8.4] [95% CI, 17.2-29.1] vs 31.5 [8.0] [95% CI, 30.1-33.0] years; F1,122 = 7.18; P = .008; ηp2 = 0.06) than patients without CYP1B1 pathogenic variants. CONCLUSIONS AND RELEVANCE Patients with advanced JOAG based on visual field loss had enrichment of CYP1B1 pathogenic variants and a more severe phenotype compared with unaffected controls and patients with nonadvanced JOAG.

Rare, potentially pathogenic variants in ZNF469 are not enriched in keratoconus in a large Australian cohort of European descent

Purpose The Zinc Finger Protein 469 (ZNF469) gene has been proposed as a candidate gene for keratoconus due to the association of an upstream polymorphism (rs9938149) with the disease in two independent studies, and the role of the gene in the autosomal recessive disease Brittle Cornea Syndrome. Coding variants in ZNF469 have been assessed for association with keratoconus in several small studies, with conflicting results. We assessed rare, potentially pathogenic variants in ZNF469 for enrichment in keratoconus patients in a cohort larger than all previous studies combined. Methods ZNF469 was sequenced in 385 Australian keratoconus patients of European descent, 346 population controls, and 230 ethnically matched screened controls by either whole exome sequencing or targeted gene sequencing. The frequency of rare and very rare potentially pathogenic variants was compared between cases and controls using χ2 or Fishers exact tests and further explored using a gene based test (Sequence Kernel Association Test [SKAT]), weighting on the rarity of variants. Results A total of 49 rare, including 33 very rare, potentially pathogenic variants were identified across all groups. No enrichment of rare or very rare potentially pathogenic variants in ZNF469 was observed in our cases compared to the control groups following analysis using χ2 or Fishers exact tests. This finding was further supported by the SKAT results, which found no significant difference in the frequency of variants predicted to be damaging between cases and either control group (P = 0.06). Conclusions Rare variants in ZNF469 do not contribute to keratoconus susceptibility and do not account for the association at rs9938149.