The Role of Genetic Testing in Children Requiring Surgery for Ectopia Lentis.

Non-traumatic ectopia lentis can be isolated or herald an underlying multisystemic disorder. Technological advances have revolutionized genetic testing for many ophthalmic disorders, and this study aims to provide insights into the clinical utility of genetic analysis in paediatric ectopia lentis. Children that underwent lens extraction for ectopia lentis between 2013 and 2017 were identified, and gene panel testing findings and surgical outcomes were collected. Overall, 10/11 cases received a probable molecular diagnosis. Genetic variants were identified in four genes: FBN1 (associated with Marfan syndrome and cardiovascular complications; n = 6), ADAMTSL4 (associated with non-syndromic ectopia lentis; n = 2), LTBP2 (n = 1) and ASPH (n = 1). Parents appeared unaffected in 6/11 cases; the initial presentation of all six of these children was to an ophthalmologist, and only 2/6 had FBN1 variants. Notably, 4/11 cases required surgery before the age of 4 years, and only one of these children carried an FBN1 variant. In summary, in this retrospective cohort study, panel-based genetic testing pointed to a molecular diagnosis in >90% of paediatric ectopia lentis cases requiring surgery. In a subset of study participants, genetic analysis revealed changes in genes that have not been linked to extraocular manifestations and highlighted that extensive systemic investigations were not required in these individuals. We propose the introduction of genetic testing early in the diagnostic pathway in children with ectopia lentis.

EyeG2P: an automated variant filtering approach improves efficiency of diagnostic genomic testing for inherited ophthalmic disorders.

BACKGROUND: Genomic variant prioritisation is one of the most significant bottlenecks to mainstream genomic testing in healthcare. Tools to improve precision while ensuring high recall are critical to successful mainstream clinical genomic testing, in particular for whole genome sequencing where millions of variants must be considered for each patient. METHODS: We developed EyeG2P, a publicly available database and web application using the Ensembl Variant Effect Predictor. EyeG2P is tailored for efficient variant prioritisation for individuals with inherited ophthalmic conditions. We assessed the sensitivity of EyeG2P in 1234 individuals with a broad range of eye conditions who had previously received a confirmed molecular diagnosis through routine genomic diagnostic approaches. For a prospective cohort of 83 individuals, we assessed the precision of EyeG2P in comparison with routine diagnostic approaches. For 10 additional individuals, we assessed the utility of EyeG2P for whole genome analysis. RESULTS: EyeG2P had 99.5% sensitivity for genomic variants previously identified as clinically relevant through routine diagnostic analysis (n=1234 individuals). Prospectively, EyeG2P enabled a significant increase in precision (35% on average) in comparison with routine testing strategies (p<0.001). We demonstrate that incorporation of EyeG2P into whole genome sequencing analysis strategies can reduce the number of variants for analysis to six variants, on average, while maintaining high diagnostic yield. CONCLUSION: Automated filtering of genomic variants through EyeG2P can increase the efficiency of diagnostic testing for individuals with a broad range of inherited ophthalmic disorders.

Assessing the Pathogenicity of In-Frame CACNA1F Indel Variants Using Structural Modeling.

Small in-frame insertion-deletion (indel) variants are a common form of genomic variation whose impact on rare disease phenotypes has been understudied. The prediction of the pathogenicity of such variants remains challenging. X-linked incomplete congenital stationary night blindness type 2 (CSNB2) is a nonprogressive, inherited retinal disorder caused by variants in CACNA1F, encoding the Cav1.4α1 channel protein. Here, structural analysis was used through homology modeling to interpret 10 disease-correlated and 10 putatively benign CACNA1F in-frame indel variants. CSNB2-correlated changes were found to be more highly conserved compared with putative benign variants. Notably, all 10 disease-correlated variants but none of the benign changes were within modeled regions of the protein. Structural analysis revealed that disease-correlated variants are predicted to destabilize the structure and function of the Cav1.4α1 channel protein. Overall, the use of structural information to interpret the consequences of in-frame indel variants provides an important adjunct that can improve the diagnosis for individuals with CSNB2.

Embryonal sarcoma of the liver in a girl with Cockayne syndrome.

The first reported malignancy associated with Cockayne syndrome.

Comparison of in silico strategies to prioritize rare genomic variants impacting RNA splicing for the diagnosis of genomic disorders.

The development of computational methods to assess pathogenicity of pre-messenger RNA splicing variants is critical for diagnosis of human disease. We assessed the capability of eight algorithms, and a consensus approach, to prioritize 249 variants of uncertain significance (VUSs) that underwent splicing functional analyses. The capability of algorithms to differentiate VUSs away from the immediate splice site as being 'pathogenic' or 'benign' is likely to have substantial impact on diagnostic testing. We show that SpliceAI is the best single strategy in this regard, but that combined usage of tools using a weighted approach can increase accuracy further. We incorporated prioritization strategies alongside diagnostic testing for rare disorders. We show that 15% of 2783 referred individuals carry rare variants expected to impact splicing that were not initially identified as 'pathogenic' or 'likely pathogenic'; one in five of these cases could lead to new or refined diagnoses.

Clinical utility of genetic testing in 201 preschool children with inherited eye disorders.

PURPOSE: A key property to consider in all genetic tests is clinical utility, the ability of the test to influence patient management and health outcomes. Here we assess the current clinical utility of genetic testing in diverse pediatric inherited eye disorders (IEDs). METHODS: Two hundred one unrelated children (0-5 years old) with IEDs were ascertained through the database of the North West Genomic Laboratory Hub, Manchester, UK. The cohort was collected over a 7-year period (2011-2018) and included 74 children with bilateral cataracts, 8 with bilateral ectopia lentis, 28 with bilateral anterior segment dysgenesis, 32 with albinism, and 59 with inherited retinal disorders. All participants underwent panel-based genetic testing. RESULTS: The diagnostic yield of genetic testing for the cohort was 64% (ranging from 39% to 91% depending on the condition). The test result led to altered management (including preventing additional investigations or resulting in the introduction of personalized surveillance measures) in 33% of probands (75% for ectopia lentis, 50% for cataracts, 33% for inherited retinal disorders, 7% for anterior segment dysgenesis, 3% for albinism). CONCLUSION: Genetic testing helped identify an etiological diagnosis in the majority of preschool children with IEDs. This prevented additional unnecessary testing and provided the opportunity for anticipatory guidance in significant subsets of patients.

Diagnostic yield of panel-based genetic testing in syndromic inherited retinal disease.

Thirty percent of all inherited retinal disease (IRD) is accounted for by conditions with extra-ocular features. This study aimed to establish the genetic diagnostic pick-up rate for IRD patients with one or more extra-ocular features undergoing panel-based screening in a clinical setting. One hundred and six participants, tested on a gene panel which contained both isolated and syndromic IRD genes, were retrospectively ascertained from the Manchester Genomic Diagnostics Laboratory database spanning 6 years (2012-2017). Phenotypic features were extracted from the clinical notes and classified according to Human Phenotype Ontology; all identified genetic variants were interpreted in accordance to the American College of Medical Genetics and Genomics guidelines. Overall, 49% (n = 52) of patients received a probable genetic diagnosis. A further 6% (n = 6) had a single disease-associated variant in an autosomal recessive disease-relevant gene. Fifty-two percent (n = 55) of patients had a clinical diagnosis at the time of testing. Of these, 71% (n = 39) received a probable genetic diagnosis. By contrast, for those without a provisional clinical diagnosis (n = 51), only 25% (n = 13) received a probable genetic diagnosis. The clinical diagnosis of Usher (n = 33) and Bardet-Biedl syndrome (n = 10) was confirmed in 67% (n = 22) and 80% (n = 8), respectively. The testing diagnostic rate in patients with clinically diagnosed multisystemic IRD conditions was significantly higher than those without one (71% versus 25%; p value < 0.001). The lower pick-up rate in patients without a clinical diagnosis suggests that panel-based approaches are unlikely to be the most effective means of achieving a molecular diagnosis for this group. Here, we suggest that genome-wide approaches (whole exome or genome) are more appropriate.

Panel-Based Clinical Genetic Testing in 85 Children with Inherited Retinal Disease.

PURPOSE: To assess the clinical usefulness of genetic testing in a pediatric population with inherited retinal disease (IRD). DESIGN: Single-center retrospective case series. PARTICIPANTS: Eighty-five unrelated children with a diagnosis of isolated or syndromic IRD who were referred for clinical genetic testing between January 2014 and July 2016. METHODS: Participants underwent a detailed ophthalmic examination, accompanied by electrodiagnostic testing (EDT) and dysmorphologic assessment where appropriate. Ocular and extraocular features were recorded using Human Phenotype Ontology terms. Subsequently, multigene panel testing (105 or 177 IRD-associated genes) was performed in an accredited diagnostic laboratory, followed by clinical variant interpretation. MAIN OUTCOME MEASURES: Diagnostic yield and clinical usefulness of genetic testing. RESULTS: Overall, 78.8% of patients (n = 67) received a probable molecular diagnosis; 7.5% (n = 5) of these had autosomal dominant disease, 25.4% (n = 17) had X-linked disease, and 67.2% (n = 45) had autosomal recessive disease. In a further 5.9% of patients (n = 5), a single heterozygous ABCA4 variant was identified; all these participants had a spectrum of clinical features consistent with ABCA4 retinopathy. Most participants (84.7%; n = 72) had undergone EDT and 81.9% (n = 59) of these patients received a probable molecular diagnosis. The genes most frequently mutated in the present cohort were CACNA1F and ABCA4, accounting for 14.9% (n = 10) and 11.9% (n = 8) of diagnoses respectively. Notably, in many cases, genetic testing helped to distinguish stationary from progressive IRD subtypes and to establish a precise diagnosis in a timely fashion. CONCLUSIONS: Multigene panel testing pointed to a molecular diagnosis in 84.7% of children with IRD. The diagnostic yield in the study population was significantly higher compared with that in previously reported unselected IRD cohorts. Approaches similar to the one described herein are expected to become a standard component of care in pediatric ophthalmology. We propose the introduction of genetic testing early in the diagnostic pathway in children with clinical and/or electrophysiologic findings, suggestive of IRD.