Ewans, LJ, Colley, A, Gaston-Massuet, C, Gualtieri, A, Cowley, MJ, McCabe, MJ, Anand, D, Lachke, SA, Scietti, L, Forneris, F, Zhu, Y, Ying, K, Walsh, C, Kirk, EP, Miller, D, Giunta, C, Sillence, D, Dinger, M, Buckley, M & Roscioli, T 2019, 'Pathogenic variants in PLOD3 result in a Stickler syndrome-like connective tissue disorder with vascular complications.', Journal of medical genetics, vol. 56, no. 9, pp. 629-638.View/Download from: Publisher's site
BACKGROUND:Pathogenic PLOD3 variants cause a connective tissue disorder (CTD) that has been described rarely. We further characterise this CTD and propose a clinical diagnostic label to improve recognition and diagnosis of PLOD3-related disease. METHODS:Reported PLOD3 phenotypes were compared with known CTDs utilising data from three further individuals from a consanguineous family with a homozygous PLOD3 c.809C>T; p.(Pro270Leu) variant. PLOD3 mRNA expression in the developing embryo was analysed for tissue-specific localisation. Mouse microarray expression data were assessed for phylogenetic gene expression similarities across CTDs with overlapping clinical features. RESULTS:Key clinical features included ocular abnormalities with risk for retinal detachment, sensorineural hearing loss, reduced palmar creases, finger contractures, prominent knees, scoliosis, low bone mineral density, recognisable craniofacial dysmorphisms, developmental delay and risk for vascular dissection. Collated clinical features showed most overlap with Stickler syndrome with variable features of Ehlers-Danlos syndrome (EDS) and epidermolysis bullosa (EB). Human lysyl hydroxylase 3/PLOD3 expression was localised to the developing cochlea, eyes, skin, forelimbs, heart and cartilage, mirroring the clinical phenotype of this disorder. CONCLUSION:These data are consistent with pathogenic variants in PLOD3 resulting in a clinically distinct Stickler-like syndrome with vascular complications and variable features of EDS and EB. Early identification of PLOD3 variants would improve monitoring for comorbidities and may avoid serious adverse ocular and vascular outcomes.
McCabe, MJ, Gauthier, M-EA, Chan, C-L, Thompson, TJ, De Sousa, SMC, Puttick, C, Grady, JP, Gayevskiy, V, Tao, J, Ying, K, Cipponi, A, Deng, N, Swarbrick, A, Thomas, ML, Lord, RV, Johns, AL, Kohonen-Corish, M, O'Toole, SA, Clark, J, Mueller, SA, Gupta, R, McCormack, AI, Dinger, ME, Cowley, MJ & kConFab 2019, 'Development and validation of a targeted gene sequencing panel for application to disparate cancers.', Scientific reports, vol. 9, no. 1.View/Download from: Publisher's site
Next generation sequencing has revolutionised genomic studies of cancer, having facilitated the development of precision oncology treatments based on a tumour's molecular profile. We aimed to develop a targeted gene sequencing panel for application to disparate cancer types with particular focus on tumours of the head and neck, plus test for utility in liquid biopsy. The final panel designed through Roche/Nimblegen combined 451 cancer-associated genes (2.01 Mb target region). 136 patient DNA samples were collected for performance and application testing. Panel sensitivity and precision were measured using well-characterised DNA controls (n = 47), and specificity by Sanger sequencing of the Aryl Hydrocarbon Receptor Interacting Protein (AIP) gene in 89 patients. Assessment of liquid biopsy application employed a pool of synthetic circulating tumour DNA (ctDNA). Library preparation and sequencing were conducted on Illumina-based platforms prior to analysis with our accredited (ISO15189) bioinformatics pipeline. We achieved a mean coverage of 395x, with sensitivity and specificity of >99% and precision of >97%. Liquid biopsy revealed detection to 1.25% variant allele frequency. Application to head and neck tumours/cancers resulted in detection of mutations aligned to published databases. In conclusion, we have developed an analytically-validated panel for application to cancers of disparate types with utility in liquid biopsy.
Ewans, LJ, Schofield, D, Shrestha, R, Zhu, Y, Gayevskiy, V, Ying, K, Walsh, C, Lee, E, Kirk, EP, Colley, A, Ellaway, C, Turner, A, Mowat, D, Worgan, L, Freckmann, M-L, Lipke, M, Sachdev, R, Miller, D, Field, M, Dinger, ME, Buckley, MF, Cowley, MJ & Roscioli, T 2018, 'Whole-exome sequencing reanalysis at 12 months boosts diagnosis and is cost-effective when applied early in Mendelian disorders.', Genetics in medicine : official journal of the American College of Medical Genetics, vol. 20, no. 12, pp. 1564-1574.View/Download from: Publisher's site
PURPOSE:Whole-exome sequencing (WES) has revolutionized Mendelian diagnostics, however, there is no consensus on the timing of data review in undiagnosed individuals and only preliminary data on the cost-effectiveness of this technology. We aimed to assess the utility of WES data reanalysis for diagnosis in Mendelian disorders and to analyze the cost-effectiveness of this technology compared with a traditional diagnostic pathway. METHODS:WES was applied to a cohort of 54 patients from 37 families with a variety of Mendelian disorders to identify the genetic etiology. Reanalysis was performed after 12 months with an improved WES diagnostic pipeline. A comparison was made between costs of a modeled WES pathway and a traditional diagnostic pathway in a cohort with intellectual disability (ID). RESULTS:Reanalysis of WES data at 12 months improved diagnostic success from 30 to 41% due to interim publication of disease genes, expanded phenotype data from referrer, and an improved bioinformatics pipeline. Cost analysis on the ID cohort showed average cost savings of US$586 (AU$782) for each additional diagnosis. CONCLUSION:Early application of WES in Mendelian disorders is cost-effective and reanalysis of an undiagnosed individual at a 12-month time point increases total diagnoses by 11%.
Gennarino, VA, Palmer, EE, McDonell, LM, Wang, L, Adamski, CJ, Koire, A, See, L, Chen, CA, Schaaf, CP, Rosenfeld, JA, Panzer, JA, Moog, U, Hao, S, Bye, A, Kirk, EP, Stankiewicz, P, Breman, AM, McBride, A, Kandula, T, Dubbs, HA, Macintosh, R, Cardamone, M, Zhu, Y, Ying, K, Dias, KR, Cho, MT, Henderson, LB, Baskin, B, Morris, P, Tao, J, Cowley, MJ, Dinger, ME, Roscioli, T, Caluseriu, O, Suchowersky, O, Sachdev, RK, Lichtarge, O, Tang, J, Boycott, KM, Holder, JL & Zoghbi, HY 2018, 'A Mild PUM1 Mutation Is Associated with Adult-Onset Ataxia, whereas Haploinsufficiency Causes Developmental Delay and Seizures', Cell, vol. 172, no. 5, pp. 924-932.e11.View/Download from: Publisher's site
© 2018 Elsevier Inc. Certain mutations can cause proteins to accumulate in neurons, leading to neurodegeneration. We recently showed, however, that upregulation of a wild-type protein, Ataxin1, caused by haploinsufficiency of its repressor, the RNA-binding protein Pumilio1 (PUM1), also causes neurodegeneration in mice. We therefore searched for human patients with PUM1 mutations. We identified eleven individuals with either PUM1 deletions or de novo missense variants who suffer a developmental syndrome (Pumilio1-associated developmental disability, ataxia, and seizure; PADDAS). We also identified a milder missense mutation in a family with adult-onset ataxia with incomplete penetrance (Pumilio1-related cerebellar ataxia, PRCA). Studies in patient-derived cells revealed that the missense mutations reduced PUM1 protein levels by ∼25% in the adult-onset cases and by ∼50% in the infantile-onset cases; levels of known PUM1 targets increased accordingly. Changes in protein levels thus track with phenotypic severity, and identifying posttranscriptional modulators of protein expression should identify new candidate disease genes. Different dosages of an RNA-binding protein result in human neurological diseases of corresponding severities.
Palmer, EE, Schofield, D, Shrestha, R, Kandula, T, Macintosh, R, Lawson, JA, Andrews, I, Sampaio, H, Johnson, AM, Farrar, MA, Cardamone, M, Mowat, D, Elakis, G, Lo, W, Zhu, Y, Ying, K, Morris, P, Tao, J, Dias, K-R, Buckley, M, Dinger, ME, Cowley, MJ, Roscioli, T, Kirk, EP, Bye, A & Sachdev, RK 2018, 'Integrating exome sequencing into a diagnostic pathway for epileptic encephalopathy: Evidence of clinical utility and cost effectiveness.', Molecular genetics & genomic medicine, vol. 6, no. 2, pp. 186-199.View/Download from: Publisher's site
Epileptic encephalopathies are a devastating group of neurological conditions in which etiological diagnosis can alter management and clinical outcome. Exome sequencing and gene panel testing can improve diagnostic yield but there is no cost-effectiveness analysis of their use or consensus on how to best integrate these tests into clinical diagnostic pathways.We conducted a retrospective cost-effectiveness study comparing trio exome sequencing with a standard diagnostic approach, for a well-phenotyped cohort of 32 patients with epileptic encephalopathy, who remained undiagnosed after "first-tier" testing. Sensitivity analysis was included with a range of commercial exome and multigene panels.The diagnostic yield was higher for the exome sequencing (16/32; 50%) than the standard arm (2/32; 6.2%). The trio exome sequencing pathway was cost-effective compared to the standard diagnostic pathway with a cost saving of AU$5,236 (95% confidence intervals $2,482; $9,784) per additional diagnosis; the standard pathway cost approximately 10 times more per diagnosis. Sensitivity analysis demonstrated that the majority of commercial exome sequencing and multigene panels studied were also cost-effective. The clinical utility of all diagnoses was reported.Our study supports the integration of exome sequencing and gene panel testing into the diagnostic pathway for epileptic encephalopathy, both in terms of cost effectiveness and clinical utility. We propose a diagnostic pathway that integrates initial rapid screening for treatable causes and comprehensive genomic screening. This study has important implications for health policy and public funding for epileptic encephalopathy and other neurological conditions.
Gururaj, S, Palmer, EE, Sheehan, GD, Kandula, T, Macintosh, R, Ying, K, Morris, P, Tao, J, Dias, KR, Zhu, Y, Dinger, ME, Cowley, MJ, Kirk, EP, Roscioli, T, Sachdev, R, Duffey, ME, Bye, A & Bhattacharjee, A 2017, 'A De Novo Mutation in the Sodium-Activated Potassium Channel KCNT2 Alters Ion Selectivity and Causes Epileptic Encephalopathy', Cell Reports, vol. 21, no. 4, pp. 926-933.View/Download from: Publisher's site
© 2017 The Author(s) Early infantile epileptic encephalopathies (EOEE) are a debilitating spectrum of disorders associated with cognitive impairments. We present a clinical report of a KCNT2 mutation in an EOEE patient. The de novo heterozygous variant Phe240Leu SLICK was identified by exome sequencing and confirmed by Sanger sequencing. Phe240Leu rSlick and hSLICK channels were electrophysiologically, heterologously characterized to reveal three significant alterations to channel function. First, [Cl−]i sensitivity was reversed in Phe240Leu channels. Second, predominantly K+-selective WT channels were made to favor Na+ over K+ by Phe240Leu. Third, and consequent to altered ion selectivity, Phe240Leu channels had larger inward conductance. Further, rSlick channels induced membrane hyperexcitability when expressed in primary neurons, resembling the cellular seizure phenotype. Taken together, our results confirm that Phe240Leu is a "change-of-function" KCNT2 mutation, demonstrating unusual altered selectivity in KNa channels. These findings establish pathogenicity of the Phe240Leu KCNT2 mutation in the reported EOEE patient. Gururaj et al. report a KCNT2 mutation in a patient with epileptic encephalopathy and employ electrophysiological analyses to establish channel properties that could underlie epileptogenesis: namely, inhibition by high [Cl−]i and loss of exclusive selectivity to K+. Furthermore, primary neurons expressing Ph240Leu display a hyperexcitable phenotype.
Warton, K, Lin, V, Navin, T, Armstrong, NJ, Kaplan, W, Ying, K, Gloss, B, Mangs, H, Nair, SS, Hacker, NF, Sutherland, RL, Clark, SJ & Samimi, G 2014, 'Methylation-capture and Next-Generation Sequencing of free circulating DNA from human plasma', BMC GENOMICS, vol. 15.View/Download from: Publisher's site
Constantinescu, L, Cowley, M, Ying, K, Budd, P, Lin, D, Kaplan, W & Dinger, M 2014, 'Implementing a clinical genomics infrastructure to sequence 18,000 human genomes per year', CEUR Workshop Proceedings, pp. 26-27.
Copyright © 2014 for the individual papers by the papers' authors. Clinical genomics is a rapidly evolving field focused on the use of genome sequencing information to guide patient diagnosis and treatment. Whole genome sequencing has been dubbed "the test to replace all genetic tests", since one sequencing run can identify all genetic variants present in a patient's genome. Implementing clinical-grade, whole genome sequencing across large patient cohorts represents a substantial big data challenge. We will present our "Sabretooth" plan for scaling operations in our centre from an estimated 800 to 18,000 genomes per year.