title: Validation of copy number variation analysis for next-generation sequencing diagnostics.
abstract: Although a common cause of disease, copy number variants (CNVs) have not routinely been identified from next-generation sequencing (NGS) data in a clinical context. This study aimed to examine the sensitivity and specificity of a widely used software package, ExomeDepth, to identify CNVs from targeted NGS data sets. We benchmarked the accuracy of CNV detection using ExomeDepth v1.1.6 applied to targeted NGS data sets by comparison to CNV events detected through whole-genome sequencing for 25 individuals and determined the sensitivity and specificity of ExomeDepth applied to these targeted NGS data sets to be 100% and 99.8%, respectively. To define quality assurance metrics for CNV surveillance through ExomeDepth, we undertook simulation of single-exon (n=1000) and multiple-exon heterozygous deletion events (n=1749), determining a sensitivity of 97% (n=2749). We identified that the extent of sequencing coverage, the inter- and intra-sample variability in the depth of sequencing coverage and the composition of analysis regions are all important determinants of successful CNV surveillance through ExomeDepth. We then applied these quality assurance metrics during CNV surveillance for 140 individuals across 12 distinct clinical areas, encompassing over 500 potential rare disease diagnoses. All 140 individuals lacked molecular diagnoses after routine clinical NGS testing, and by application of ExomeDepth, we identified 17 CNVs contributing to the cause of a Mendelian disorder. Our findings support the integration of CNV detection using ExomeDepth v1.1.6 with routine targeted NGS diagnostic services for Mendelian disorders. Implementation of this strategy increases diagnostic yields and enhances clinical care.
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.|Division of Evolution and Genomic Sciences, Neuroscience and Mental Health Domain, School of Health Sciences, Faculty of Biology, Medicines and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
authors.forename: Jamie M
authors.identifier: 0000-0003-1137-9768
authors.initials: JM
authors.lastname: Ellingford
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.
authors.forename: Christopher
authors.initials: C
authors.lastname: Campbell
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Barton
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.
authors.forename: Sanjeev
authors.initials: S
authors.lastname: Bhaskar
authors.affiliation: Ganit Labs, Bio-IT Centre, Institute of Bioinformatics and Applied Biotechnology, Bangalore, India.
authors.forename: Saurabh
authors.initials: S
authors.lastname: Gupta
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.|Division of Evolution and Genomic Sciences, Neuroscience and Mental Health Domain, School of Health Sciences, Faculty of Biology, Medicines and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
authors.forename: Rachel L
authors.initials: RL
authors.lastname: Taylor
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.
authors.forename: Panagiotis I
authors.initials: PI
authors.lastname: Sergouniotis
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.
authors.forename: Bradley
authors.initials: B
authors.lastname: Horn
authors.affiliation: Division of Population Health, Health Services Research and Primary Care, School of Health Sciences, Faculty of Medicine, Biology and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
authors.forename: Janine A
authors.initials: JA
authors.lastname: Lamb
authors.affiliation: Moorfields Eye Hospital NHS Foundation Trust, London, UK.|UCL Institute of Ophthalmology, Department of Genetics, London, UK.
authors.forename: Michel
authors.initials: M
authors.lastname: Michaelides
authors.affiliation: Moorfields Eye Hospital NHS Foundation Trust, London, UK.|UCL Institute of Ophthalmology, Department of Genetics, London, UK.
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Webster
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.|Division of Evolution and Genomic Sciences, Neuroscience and Mental Health Domain, School of Health Sciences, Faculty of Biology, Medicines and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
authors.forename: William G
authors.initials: WG
authors.lastname: Newman
authors.affiliation: Ganit Labs, Bio-IT Centre, Institute of Bioinformatics and Applied Biotechnology, Bangalore, India.
authors.forename: Binay
authors.initials: B
authors.lastname: Panda
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.
authors.forename: Simon C
authors.initials: SC
authors.lastname: Ramsden
authors.affiliation: Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, St Mary's Hospital, Manchester, UK.|Division of Evolution and Genomic Sciences, Neuroscience and Mental Health Domain, School of Health Sciences, Faculty of Biology, Medicines and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
authors.forename: Graeme Cm
authors.initials: GC
authors.lastname: Black
mesh_terms: D056915:DNA Copy Number Variations; D059472:Exome; D005820:Genetic Testing / Q000379:methods* / Q000592:standards; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods* / Q000592:standards; D006801:Humans; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods* / Q000592:standards