Publications by authors named "Lisa S Kearns"

58 Publications

Childhood and Early Onset Glaucoma Classification and Genetic Profile in a Large Australasian Disease Registry.

Ophthalmology 2021 Apr 20. Epub 2021 Apr 20.

Department of Ophthalmology, Flinders University, Flinders Medical Centre, Adelaide, Australia.

Purpose: To report the relative frequencies of childhood and early onset glaucoma subtypes and their genetic findings in a large single cohort.

Design: Retrospective clinical and molecular study.

Participants: All individuals with childhood glaucoma (diagnosed 0 to <18 years) and early onset glaucoma (diagnosed 18 to <40 years) referred to a national disease registry.

Methods: We retrospectively reviewed the referrals of all individuals with glaucoma diagnosed at <40 years of age recruited to the Australian and New Zealand Registry of Advanced Glaucoma (ANZRAG). Subtypes of glaucoma were determined using the Childhood Glaucoma Research Network (CGRN) classification system. DNA extracted from blood or saliva samples underwent sequencing of genes associated with glaucoma.

Main Outcome Measures: The phenotype and genotype distribution of glaucoma diagnosed at <40 years of age.

Results: A total of 290 individuals (533 eyes) with childhood glaucoma and 370 individuals (686 eyes) with early onset glaucoma were referred to the ANZRAG. Primary glaucoma was the most prevalent condition in both cohorts. In the childhood cohort, 57.6% of individuals (167/290, 303 eyes) had primary congenital glaucoma (PCG), and 19.3% (56/290, 109 eyes) had juvenile open-angle glaucoma. Juvenile open-angle glaucoma constituted 73.2% of the early onset glaucoma cohort (271/370, 513 eyes). Genetic testing in probands resulted in a diagnostic yield of 24.7% (125/506) and a reclassification of glaucoma subtype in 10.4% of probands (13/125). The highest molecular diagnostic rate was achieved in probands with glaucoma associated with nonacquired ocular anomalies (56.5%). Biallelic variants in CYP1B1 (n = 29, 23.2%) and heterozygous variants in MYOC (n = 24, 19.2%) and FOXC1 (n = 21, 16.8%) were most commonly reported among probands with a molecular diagnosis. Biallelic CYP1B1 variants were reported in twice as many female individuals as male individuals with PCG (66.7% vs. 33.3%, P = 0.02).

Conclusions: We report on the largest cohort of individuals with childhood and early onset glaucoma from Australasia using the CGRN classification. Primary glaucoma was most prevalent. Genetic diagnoses ascertained in 24.7% of probands supported clinical diagnoses and genetic counseling. International collaborative efforts are required to identify further genes because the majority of individuals still lack a clear molecular diagnosis.
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http://dx.doi.org/10.1016/j.ophtha.2021.04.016DOI Listing
April 2021

Adherence to home-based videogame treatment for amblyopia in children and adults.

Clin Exp Optom 2021 Mar 1:1-7. Epub 2021 Mar 1.

School of Optometry and Vision Science, University of Auckland, Auckland, New Zealand.

: Home-based videogame treatments are increasingly popular for amblyopia treatment. However, at-home treatments tend to be done in short sessions and with frequent disruptions, which may reduce the effectiveness of binocular visual stimulation. These treatment adherence patterns need to be accounted for when considering dose-response relationships and treatment effectiveness.: Home-based videogame treatments are increasingly being used for various sensory conditions, including amblyopia ('lazy eye'), but treatment adherence continues to limit success. To examine detailed behavioural patterns associated with home-based videogame treatment, we analysed in detail the videogame adherence data from the Binocular tReatment of Amblyopia with VideOgames (BRAVO) clinical trial (ACTRN12613001004752).: Children (7-12 years), teenagers (13-17 years) and adults (≥ 18 years) with unilateral amblyopia were loaned iPod Touch devices with either an active treatment or placebo videogame and instructed to play for a total of 1-2 hours/day for six weeks at home. Objectively-recorded adherence data from device software were used to analyse adherence patterns such as session length, daily distribution of gameplay, use of the pause function, and differences between age groups. Objectively-recorded adherence was also compared to subjectively-reported adherence from paper-based diaries.: One hundred and five of the 115 randomised participants completed six weeks of videogame training. Average adherence was 65% (SD 37%) of the minimum hours prescribed. Game training was generally performed in short sessions (mean 21.5, SD 11.2 minutes), mostly in the evening, with frequent pauses (median every 4.1 minutes, IQR 6.1). Children played in significantly shorter sessions and paused more frequently than older age groups (p < 0.0001). Participants tended to over-report adherence in subjective diaries compared to objectively-recorded gameplay time.: Adherence to home-based videogame treatment was characterised by short sessions interspersed with frequent pauses, suggesting regular disengagement. This complicates dose-response calculations and may interfere with the effectiveness of treatments like binocular treatments for amblyopia, which require sustained visual stimulation.
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http://dx.doi.org/10.1080/08164622.2021.1878834DOI Listing
March 2021

OXPHOS bioenergetic compensation does not explain disease penetrance in Leber hereditary optic neuropathy.

Mitochondrion 2020 09 18;54:113-121. Epub 2020 Jul 18.

Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, 32 Gisborne Street, East Melbourne, 3002 Victoria, Australia; Ophthalmology, University of Melbourne, Department of Surgery, Parkville 3010 Victoria, Australia. Electronic address:

Leber hereditary optic neuropathy (LHON) is one of the most common primary mitochondrial diseases. It is caused by point mutations in mitochondrial DNA (mtDNA) genes and in some cases, it can result in irreversible vision loss, primarily in young men. It is currently unknown why LHON mutations affect only some carriers and whether bioenergetic compensation enables unaffected carriers to overcome mitochondrial impairment and preserve cellular function. Here, we conducted bioenergetic metabolic assays and RNA sequencing to address this question using male-only, age-matched, m.11778G > A lymphoblasts and primary fibroblasts from both unaffected carriers and affected individuals. Our work indicates that OXPHOS bioenergetic compensation in LHON peripheral cells does not explain disease phenotype. We show that complex I impairment is similar in cells from unaffected carrier and affected patients, despite a transcriptional downregulation of metabolic pathways including glycolysis in affected cells relative to carriers detected by RNA sequencing. Although we did not detect OXPHOS bioenergetic compensation in carrier cells under basal conditions, our results indicate that cells from affected patients suffer a growth impairment under metabolic challenge compared to carrier cells, which were unaffected by metabolic challenge. If recapitulated in retinal ganglion cells, decreased susceptibility to metabolic challenge in unaffected carriers may help preserve metabolic homeostasis in the face of the mitochondrial complex I bioenergetic defect.
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http://dx.doi.org/10.1016/j.mito.2020.07.003DOI Listing
September 2020

A novel AFG3L2 mutation close to AAA domain leads to aberrant OMA1 and OPA1 processing in a family with optic atrophy.

Acta Neuropathol Commun 2020 06 29;8(1):93. Epub 2020 Jun 29.

Mitochondrial dysfunctions in neurodegeneration Unit, Division of Neuroscience, Ospedale San Raffaele, Milan, Italy.

Autosomal dominant optic atrophy (ADOA) is a neuro-ophthalmic condition characterized by bilateral degeneration of the optic nerves. Although heterozygous mutations in OPA1 represent the most common genetic cause of ADOA, a significant number of cases remain undiagnosed.Here, we describe a family with a strong ADOA history with most family members spanning three generation having childhood onset of visual symptoms. The proband, in addition to optic atrophy, had neurological symptoms consistent with relapsing remitting multiple sclerosis. Clinical exome analysis detected a novel mutation in the AFG3L2 gene (NM_006796.2:c.1010G > A; p.G337E), which segregated with optic atrophy in family members. AFG3L2 is a metalloprotease of the AAA subfamily which exerts quality control in the inner mitochondrial membrane. Interestingly, the identified mutation localizes close to the AAA domain of AFG3L2, while those localized in the proteolytic domain cause dominant spinocerebellar ataxia type 28 (SCA28) or recessive spastic ataxia with epilepsy (SPAX5). Functional studies in patient fibroblasts demonstrate that the p.G337E AFG3L2 mutation strongly destabilizes the long isoforms of OPA1 via OMA hyper-activation and leads to mitochondrial fragmentation, thus explaining the family phenotype. This study widens the clinical spectrum of neurodegenerative diseases caused by AFG3L2 mutations, which shall be considered as genetic cause of ADOA.
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http://dx.doi.org/10.1186/s40478-020-00975-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7325028PMC
June 2020

Biallelic CPAMD8 Variants Are a Frequent Cause of Childhood and Juvenile Open-Angle Glaucoma.

Ophthalmology 2020 06 7;127(6):758-766. Epub 2020 Jan 7.

Department of Ophthalmology, Flinders University, Adelaide, Australia.

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) 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.
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http://dx.doi.org/10.1016/j.ophtha.2019.12.024DOI Listing
June 2020

The genetic and clinical landscape of nanophthalmos and posterior microphthalmos in an Australian cohort.

Clin Genet 2020 05 5;97(5):764-769. Epub 2020 Mar 5.

Department of Ophthalmology, Flinders University, Adelaide, Australia.

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.
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http://dx.doi.org/10.1111/cge.13722DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7811993PMC
May 2020

The challenge of an adequate outcome in trials for genetic eye disease such as Leber hereditary optic neuropathy.

Clin Exp Ophthalmol 2019 08;47(6):704-705

Centre for Eye Research Australia, University of Melbourne, Royal Victorian Eye and Ear Hospital, Melbourne, Victoria, Australia.

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http://dx.doi.org/10.1111/ceo.13586DOI Listing
August 2019

Prevalence of FOXC1 Variants in Individuals With a Suspected Diagnosis of Primary Congenital Glaucoma.

JAMA Ophthalmol 2019 04;137(4):348-355

Department of Ophthalmology, Flinders University, Flinders Medical Centre, Adelaide, Australia.

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.
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http://dx.doi.org/10.1001/jamaophthalmol.2018.5646DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6459212PMC
April 2019

Mitochondrial DNA Variation and Disease Susceptibility in Primary Open-Angle Glaucoma.

Invest Ophthalmol Vis Sci 2018 09;59(11):4598-4602

Center for Eye Research Australia, Ophthalmology, University of Melbourne Department of Surgery, Melbourne, Australia.

Purpose: To determine whether mitochondrial DNA haplogroups or rare variants associate with primary open-angle glaucoma in subjects of European descent.

Methods: A case-control comparison of age- and sex-matched cohorts of 90 primary open-angle glaucoma patients and 95 population controls. Full mitochondrial DNA sequences from peripheral blood were generated by next-generation sequencing and compared to the revised Cambridge Reference Sequence to define mitochondrial haplogroups and variants.

Results: Most subjects were of the major European haplogroups H, J, K, U, and T. Logistic regression analysis showed haplogroup U to be significantly underrepresented in male primary open-angle glaucoma subjects (odds ratio 0.25; 95% confidence interval [CI] 0.09-0.67; P = 0.007; Bonferroni multiple testing P = 0.022). Variants in the mitochondrial DNA gene MT-ND2 were overrepresented in the control group (P = 0.005; Bonferroni multiple testing correction P = 0.015).

Conclusions: Mitochondrial DNA ancestral lineages modulate the risk for primary open-angle glaucoma in populations of European descent. Haplogroup U and rare variants in the mitochondrial DNA-encoded MT-ND2 gene may be protective against primary open-angle glaucoma. Larger studies are warranted to explore haplogroup associations with disease risk in different ethnic groups and define biomarkers of primary open-angle glaucoma endophenotypes to target therapeutic strategies.
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http://dx.doi.org/10.1167/iovs.18-25085DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6138263PMC
September 2018

Cross-ancestry genome-wide association analysis of corneal thickness strengthens link between complex and Mendelian eye diseases.

Nat Commun 2018 05 14;9(1):1864. Epub 2018 May 14.

Department of Ophthalmology, Flinders University, SA 5042, Adelaide, Australia.

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) 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.
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http://dx.doi.org/10.1038/s41467-018-03646-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5951816PMC
May 2018

Crowd-sourced Ontology for Photoleukocoria: Identifying Common Internet Search Terms for a Potentially Important Pediatric Ophthalmic Sign.

Transl Vis Sci Technol 2018 Feb 15;7(1):18. Epub 2018 Feb 15.

Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, Melbourne, VIC, Australia.

Purpose: Leukocoria is the most common presenting sign for pediatric eye disease including retinoblastoma and cataract, with worse outcomes if diagnosis is delayed. We investigated whether individuals could identify leukocoria in photographs (photoleukocoria) and examined their subsequent Internet search behavior.

Methods: Using a web-based questionnaire, in this cross-sectional study we invited adults aged over 18 years to view two photographs of a child with photoleukocoria, and then search the Internet to determine a possible diagnosis and action plan. The most commonly used search terms and websites accessed were recorded.

Results: The questionnaire was completed by 1639 individuals. Facebook advertisement was the most effective recruitment strategy. The mean age of all respondents was 38.95 ± 14.59 years (range, 18-83), 94% were female, and 59.3% had children. An abnormality in the images presented was identified by 1613 (98.4%) participants. The most commonly used search terms were: "white," "pupil," "photo," and "eye" reaching a variety of appropriate websites or links to print or social media articles.

Conclusions: Different words or phrases were used to describe the same observation of photoleukocoria leading to a range of websites. Variations in the description of observed signs and search words influenced the sites reached, information obtained, and subsequent help-seeking intentions.

Translational Relevance: Identifying the most commonly used search terms for photoleukocoria is an important step for search engine optimization. Being directed to the most appropriate websites informing of the significance of photoleukocoria and the appropriate actions to take could improve delays in diagnosis of important pediatric eye disease such as retinoblastoma or cataract.
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http://dx.doi.org/10.1167/tvst.7.1.18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5815559PMC
February 2018

Effectiveness of a Binocular Video Game vs Placebo Video Game for Improving Visual Functions in Older Children, Teenagers, and Adults With Amblyopia: A Randomized Clinical Trial.

JAMA Ophthalmol 2018 02;136(2):172-181

School of Optometry and Vision Science, The University of Auckland, Auckland, New Zealand.

Importance: Binocular amblyopia treatment using contrast-rebalanced stimuli showed promise in laboratory studies and requires clinical trial investigation in a home-based setting.

Objective: To compare the effectiveness of a binocular video game with a placebo video game for improving visual functions in older children and adults.

Design, Setting, And Participants: The Binocular Treatment of Amblyopia Using Videogames clinical trial was a multicenter, double-masked, randomized clinical trial. Between March 2014 and June 2016, 115 participants 7 years and older with unilateral amblyopia (amblyopic eye visual acuity, 0.30-1.00 logMAR; Snellen equivalent, 20/40-20/200) due to anisometropia, strabismus, or both were recruited. Eligible participants were allocated with equal chance to receive either the active or the placebo video game, with minimization stratified by age group (child, age 7 to 12 years; teenager, age 13 to 17 years; and adult, 18 years and older).

Interventions: Falling-blocks video games played at home on an iPod Touch for 1 hour per day for 6 weeks. The active video game had game elements split between eyes with a dichoptic contrast offset (mean [SD] initial fellow eye contrast, 0.23 [0.14]). The placebo video game presented identical images to both eyes.

Main Outcomes And Measures: Change in amblyopic eye visual acuity at 6 weeks. Secondary outcomes included compliance, stereoacuity, and interocular suppression. Participants and clinicians who measured outcomes were masked to treatment allocation.

Results: Of the 115 included participants, 65 (56.5%) were male and 83 (72.2%) were white, and the mean (SD) age at randomization was 21.5 (13.6) years. There were 89 participants (77.4%) who had prior occlusion. The mean (SD) amblyopic eye visual acuity improved 0.06 (0.12) logMAR from baseline in the active group (n = 56) and 0.07 (0.10) logMAR in the placebo group (n = 59). The mean treatment difference between groups, adjusted for baseline visual acuity and age group, was -0.02 logMAR (95% CI, -0.06 to 0.02; P = .25). Compliance with more than 25% of prescribed game play was achieved by 36 participants (64%) in the active group and by 49 (83%) in the placebo group. At 6 weeks, 36 participants (64%) in the active group achieved fellow eye contrast greater than 0.9 in the binocular video game. No group differences were observed for any secondary outcomes. Adverse effects included 3 reports of transient asthenopia.

Conclusions And Relevance: The specific home-based binocular falling-blocks video game used in this clinical trial did not improve visual outcomes more than the placebo video game despite increases in fellow eye contrast during game play. More engaging video games with considerations for compliance may improve effectiveness.

Trial Registration: anzctr.org.au Identifier: ACTRN12613001004752.
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http://dx.doi.org/10.1001/jamaophthalmol.2017.6090DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6584719PMC
February 2018

Generation of a human induced pluripotent stem cell line CERAi001-A-6 using episomal vectors.

Stem Cell Res 2017 07 19;22:13-15. Epub 2017 May 19.

Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, Australia; Ophthalmology, Department of Surgery, The University of Melbourne, Australia. Electronic address:

We report the generation of the hiPSC line CERAi001-A-6 from primary human dermal fibroblasts. Reprogramming was performed using episomal vector delivery of OCT4, SOX2, KLF4, L-MYC, LIN28 and shRNA for p53.
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http://dx.doi.org/10.1016/j.scr.2017.05.007DOI Listing
July 2017

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

Eur J Hum Genet 2017 06 17;25(7):839-847. Epub 2017 May 17.

Department of Ophthalmology, Flinders University, Flinders Medical Centre, Adelaide, SA, Australia.

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.
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http://dx.doi.org/10.1038/ejhg.2017.59DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5520071PMC
June 2017

Mitochondrial replacement in an iPSC model of Leber's hereditary optic neuropathy.

Aging (Albany NY) 2017 04;9(4):1341-1350

Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, East Melbourne, Australia.

Cybrid technology was used to replace Leber hereditary optic neuropathy (LHON) causing mitochondrial DNA (mtDNA) mutations from patient-specific fibroblasts with wildtype mtDNA, and mutation-free induced pluripotent stem cells (iPSCs) were generated subsequently. Retinal ganglion cell (RGC) differentiation demonstrates increased cell death in LHON-RGCs and can be rescued in cybrid corrected RGCs.
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http://dx.doi.org/10.18632/aging.101231DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5425131PMC
April 2017

Development of a Modular Automated System for Maintenance and Differentiation of Adherent Human Pluripotent Stem Cells.

SLAS Discov 2017 09 13;22(8):1016-1025. Epub 2017 Mar 13.

1 Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital & Department of Ophthalmology, the University of Melbourne, East Melbourne, Victoria, Australia.

Patient-specific induced pluripotent stem cells (iPSCs) have tremendous potential for development of regenerative medicine, disease modeling, and drug discovery. However, the processes of reprogramming, maintenance, and differentiation are labor intensive and subject to intertechnician variability. To address these issues, we established and optimized protocols to allow for the automated maintenance of reprogrammed somatic cells into iPSCs to enable the large-scale culture and passaging of human pluripotent stem cells (PSCs) using a customized TECAN Freedom EVO. Generation of iPSCs was performed offline by nucleofection followed by selection of TRA-1-60-positive cells using a Miltenyi MultiMACS24 Separator. Pluripotency markers were assessed to confirm pluripotency of the generated iPSCs. Passaging was performed using an enzyme-free dissociation method. Proof of concept of differentiation was obtained by differentiating human PSCs into cells of the retinal lineage. Key advantages of this automated approach are the ability to increase sample size, reduce variability during reprogramming or differentiation, and enable medium- to high-throughput analysis of human PSCs and derivatives. These techniques will become increasingly important with the emergence of clinical trials using stem cells.
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http://dx.doi.org/10.1177/2472555217696797DOI Listing
September 2017

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

Hum Mol Genet 2017 01;26(2):438-453

Centre for Eye Research Australia (CERA), University of Melbourne, Royal Victorian Eye and Ear Hospital, Melbourne, Victoria, Australia.

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.
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http://dx.doi.org/10.1093/hmg/ddw399DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5968632PMC
January 2017

Gene-Based Therapies for Leber Hereditary Optic Neuropathy. Hype or Hope?

Asia Pac J Ophthalmol (Phila) 2016 Jul-Aug;5(4):253-5

From the *Lions Eye Institute, Centre for Ophthalmology and Visual Science, University of Western Australia, Perth; †Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, University of Melbourne, Melbourne; and ‡School of Medicine, Menzies Institute for Medical Research, University of Tasmania, Hobart, Australia.

Leber hereditary optic neuropathy has now joined Leber congenital amaurosis in the list of genetic eye diseases undergoing gene therapy clinical trials. Although a dramatic response to treatment would be welcome, a minor improvement in vision is a major challenge in efficacy assessment, given this may occur spontaneously as part of the natural history of minor recovery in some patients. Thus, we must await the outcome of adequately powered clinical trials to know if the treatment is effective, particularly given the likely high cost of such therapeutic interventions in the future. We need global cooperation to ensure that the most suitable patients are enrolled in these trials and that support is provided for participants who need to travel from the Asia-Pacific region to Europe or North America if there are no local arms of these trials.
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http://dx.doi.org/10.1097/APO.0000000000000220DOI Listing
January 2017

Participant understanding and recall of informed consent for induced pluripotent stem cell biobanking.

Cell Tissue Bank 2016 Sep 14;17(3):449-56. Epub 2016 Jun 14.

Department of Surgery, Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, University of Melbourne, 32 Gisborne Street, East Melbourne, VIC, Australia.

The ability to generate human induced pluripotent stem cells (iPSCs) has opened new avenues for human disease modelling and therapy. The aim of our study was to determine research participants' understanding of the information given when donating skin biopsies for the generation of patient-specific iPSCs. A customised 35-item questionnaire based on previous iPSC consent guidelines was sent to participants who had previously donated samples for iPSC research. The questionnaire asked pertinent demographic details, participants' motivation to take part in iPSC research and their attitudes towards related ethical issues. 234 participants were contacted with 141 (60.3 %) complete responses received. The median duration between recruitment and follow-up questioning was 313 days (range 10-573 days). The majority of participants (n = 129, 91.5 %) believed they understood what a stem cell was; however, only 22 (16.1 %) correctly answered questions related to basic stem cell properties. We found no statistically significant difference in responses from participants with different levels of education, or those with a health sciences background. The poor understanding amongst participants of iPSC research is unlikely to be unique to our study and may impact future research if not improved. As such, there is a need to develop an easily understood yet comprehensive consent process to ensure ongoing ethical progress of iPSC biobanking.
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http://dx.doi.org/10.1007/s10561-016-9563-8DOI Listing
September 2016

Response: Cycloplegia in refraction: age and cycloplegics.

Acta Ophthalmol 2016 Aug 11;94(5):e373. Epub 2016 May 11.

Centre for Ophthalmology and Visual Science, Lions Eye Institute, University of Western Australia, Perth, Western Australia, Australia.

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http://dx.doi.org/10.1111/aos.13082DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6519986PMC
August 2016

Heterogeneity of Human Research Ethics Committees and Research Governance Offices across Australia: An observational study.

Australas Med J 2016 29;9(2):33-9. Epub 2016 Feb 29.

Menzies Institute for Medical Research, School of Medicine, University of Tasmania, TAS, Australia.

Background: Conducting ethically grounded research is a fundamental facet of all investigations. Nevertheless, the administrative burdens of current ethics review are substantial, and calls have been made for a reduction in research waste.

Aims: To describe the heterogeneity in administration and documentation required by Human Research Ethics Committees (HRECs) and Research Governance Offices (RGOs) across Australia.

Methods: In establishing a nationwide study to investigate the molecular aetiology of Giant Cell Arteritis (GCA), for which archived pathological specimens from around Australia are being recruited, we identified variation across separate HREC and RGO requirements. Submission paperwork and correspondence from each collaborating site and its representative office for research were reviewed. This data was interrogated to evaluate differences in current guidelines.

Results: Twenty-five pathology departments across seven Australian States collaborated in this study. All states, except Victoria, employed a single ethics review model. There was discrepancy amongst HRECs as to which application process applied to our study: seven requested completion of a "National Ethics Application Form" and three a "Low Negligible Risk" form. Noticeable differences in guidelines included whether electronic submission was sufficient. There was variability in the total number of documents submitted (range five to 22) and panel review turnaround time (range nine to 136 days).

Conclusion: We demonstrate the challenges and illustrate the heavy workload involved in receiving widespread ethics and governance approval across Australia. We highlight the need to simplify, homogenise, and nationalise human ethics for non-clinical trial studies. Reducing unnecessary administration will enable investigators to achieve research aims more efficiently.
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http://dx.doi.org/10.4066/AMJ.2015.2587DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4780210PMC
March 2016

Measurement of Systemic Mitochondrial Function in Advanced Primary Open-Angle Glaucoma and Leber Hereditary Optic Neuropathy.

PLoS One 2015 23;10(10):e0140919. Epub 2015 Oct 23.

Centre for Eye Research Australia, University of Melbourne, Royal Victorian Eye and Ear Hospital, Melbourne, Australia.

Primary Open Angle Glaucoma (POAG) is a common neurodegenerative disease characterized by the selective and gradual loss of retinal ganglion cells (RGCs). Aging and increased intraocular pressure (IOP) are glaucoma risk factors; nevertheless patients deteriorate at all levels of IOP, implying other causative factors. Recent evidence presents mitochondrial oxidative phosphorylation (OXPHOS) complex-I impairments in POAG. Leber Hereditary Optic Neuropathy (LHON) patients suffer specific and rapid loss of RGCs, predominantly in young adult males, due to complex-I mutations in the mitochondrial genome. This study directly compares the degree of OXPHOS impairment in POAG and LHON patients, testing the hypothesis that the milder clinical disease in POAG is due to a milder complex-I impairment. To assess overall mitochondrial capacity, cells can be forced to produce ATP primarily from mitochondrial OXPHOS by switching the media carbon source to galactose. Under these conditions POAG lymphoblasts grew 1.47 times slower than controls, whilst LHON lymphoblasts demonstrated a greater degree of growth impairment (2.35 times slower). Complex-I enzyme specific activity was reduced by 18% in POAG lymphoblasts and by 29% in LHON lymphoblasts. We also assessed complex-I ATP synthesis, which was 19% decreased in POAG patients and 17% decreased in LHON patients. This study demonstrates both POAG and LHON lymphoblasts have impaired complex-I, and in the majority of aspects the functional defects in POAG were milder than LHON, which could reflect the milder disease development of POAG. This new evidence places POAG in the spectrum of mitochondrial optic neuropathies and raises the possibility for new therapeutic targets aimed at improving mitochondrial function.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0140919PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4619697PMC
June 2016

CYP1B1 copy number variation is not a major contributor to primary congenital glaucoma.

Mol Vis 2015 11;21:160-4. Epub 2015 Feb 11.

Department of Ophthalmology, Flinders University, Flinders Medical Centre, Adelaide, Australia.

Purpose: To evaluate the prevalence and the diagnostic utility of testing for CYP1B1 copy number variation (CNV) in primary congenital glaucoma (PCG) cases unexplained by CYP1B1 point mutations in The Australian and New Zealand Registry of Advanced Glaucoma.

Methods: In total, 50 PCG cases either heterozygous for disease-causing variants or with no CYP1B1 sequence variants were included in the study. CYP1B1 CNV was analyzed by Multiplex Ligation-dependent Probe Amplification (MLPA).

Results: No deletions or duplications were found in any of the cases.

Conclusion: This is the first study to report on CYP1B1 CNV in PCG cases. Our findings show that this mechanism is not a major contributor to the phenotype and is of limited diagnostic utility.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4333725PMC
September 2015

Current landscape of direct-to-consumer genetic testing and its role in ophthalmology: a review.

Clin Exp Ophthalmol 2015 Aug 27;43(6):578-90. Epub 2015 Apr 27.

Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, University of Melbourne, Melbourne, Victoria, Australia.

The sequencing of the human genome has seen the emergence of the direct-to-consumer (DTC) genetic-testing market, which allows individuals to obtain information about their genetic profile and its many health and lifestyle implications. Genetics play an important role in the development of many eye diseases, however, little information is available describing the influence of the DTC industry in ophthalmology. In this review, we examined DTC companies providing genetic test products for eye disease. Of all eye conditions, the majority of DTC companies provided susceptibility testing or risk assessment for age-related macular degeneration (AMD). For the 15 companies noted to offer products, we found considerable variation in the cost, scope and clarity of informational content of DTC genetic testing for ophthalmic conditions. The clinical utility of these tests remains in question, and the American Academy of Ophthalmology recommendations against routine testing for many conditions probably still apply.
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http://dx.doi.org/10.1111/ceo.12508DOI Listing
August 2015

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

Genet Epidemiol 2015 Mar 28;39(3):207-16. Epub 2015 Jan 28.

Department of Epidemiology, Erasmus Medical Center, Rotterdam, the Netherlands; Department of Ophthalmology, Erasmus Medical Center, Rotterdam, the Netherlands.

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.
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http://dx.doi.org/10.1002/gepi.21886DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4480365PMC
March 2015

What is the appropriate age cut-off for cycloplegia in refraction?

Acta Ophthalmol 2014 Sep 19;92(6):e458-62. Epub 2014 Mar 19.

Department of Ophthalmology, Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital, University of Melbourne, Melbourne, Vic., Australia.

Purpose: To investigate the age range for which cycloplegia provides additional information compared with non-cycloplegic refraction in teenagers and young adults.

Methods: Data for 1295 subjects (704 female; 591 male) from the Twins Eye Study in Tasmania (TEST) and the Brisbane Adolescent Twin Study (mean age: 19.65 ± 3.56, range: 13-26 years) were included. For all participants, cycloplegia was induced by instillation of either one drop of 1% cyclopentolate (13-14 years) or one drop of 1% tropicamide (15-26 years). Pre- and postcycloplegic refractive errors for both eyes were measured using a Humphrey-598 automated refractor and spherical equivalents of refractive error were calculated. Generalized Estimating Equations (GEE) were used to model the spherical equivalent refraction (SER) for each eye against age (by year) and axial length (in the given eye).

Results: The mean group difference between pre- and postcycloplegic SER (post minus pre) was 0.17 ± 0.52 D and 0.12 ± 0.51 D for the right and left eyes, respectively, indicating that postcycloplegic refraction was generally more hyperopic/less myopic. The mean difference between pre- and postcycloplegic SER decreased from 0.36 ± 0.41 D in the 13-year-olds to 0.06 ± 0.50 D in people aged 25 years. After adjusting for family-relatedness, the difference between pre- and postcycloplegia SER was significant in all age groups up until the age of 20 years.

Conclusions: Non-cycloplegic autorefraction can result in group mean SER differences of greater myopia than cycloplegic autorefraction and occurs in teenagers (13-19 years of age), but not in adults 20-26 years. These data suggest that cycloplegia is not required in population estimates of refractive error for young adults once they reach approximately 20 years of age.
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http://dx.doi.org/10.1111/aos.12388DOI Listing
September 2014

Interrogation of the platelet-derived growth factor receptor alpha locus and corneal astigmatism in Australians of Northern European ancestry: results of a genome-wide association study.

Mol Vis 2013 6;19:1238-46. Epub 2013 Jun 6.

Centre for Ophthalmology and Visual Science, University of Western Australia, Lions Eye Institute, Perth, Australia.

Purpose: Corneal astigmatism is a common eye disorder characterized by irregularities in corneal curvature. Recently, the rs7677751 single nucleotide polymorphism (SNP) at the platelet-derived growth factor receptor alpha (PDGFRA) locus was found to be associated with corneal astigmatism in people of Asian ancestry. In the present study, we sought to replicate this finding and identify other genetic markers of corneal astigmatism in an Australian population of Northern European ancestry.

Methods: Data from two cohorts were included in this study. The first cohort consisted of 1,013 individuals who were part of the Western Australian Pregnancy Cohort (Raine) Study: 20-year follow-up Eye Study. The second cohort comprised 1,788 individuals of 857 twin families who were recruited through the Twins Eye Study in Tasmania and the Brisbane Adolescent Twin Study. Corneal astigmatism was calculated as the absolute difference between the keratometry readings in two meridians, and genotype data were extracted from genome-wide arrays. Initially, each cohort was analyzed separately, before being combined for meta- and subsequent genome-wide pathway analysis.

Results: Following meta-analysis, SNP rs7677751 at the PDGFRA locus had a combined p=0.32. No variant was found to be statistically significantly associated with corneal astigmatism at the genome-wide level (p<5.0×10(-8)). The SNP with strongest association was rs1164064 (p=1.86×10(-6)) on chromosome 3q13. Gene-based pathway analysis identified a significant association between the Gene Ontology "segmentation" (GO:0035282) pathway, corrected p=0.009.

Conclusions: Our data suggest that the PDGFRA locus does not transfer a major risk of corneal astigmatism in people of Northern European ancestry. Better-powered studies are required to validate the novel putative findings of our study.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3675057PMC
September 2013