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Recommendations for the clinical interpretation and reporting of copy number gains using gene panel NGS analysis in routine diagnostics.

ABSTRACT: Next-generation sequencing (NGS) panel analysis on DNA from formalin-fixed paraffin-embedded (FFPE) tissue is increasingly used to also identify actionable copy number gains (gene amplifications) in addition to sequence variants. While guidelines for the reporting of sequence variants are available, guidance with respect to reporting copy number gains from gene-panel NGS data is limited. Here, we report on Dutch consensus recommendations obtained in the context of the national Predictive Analysis for THerapy (PATH) project, which aims to optimize and harmonize routine diagnostics in molecular pathology. We briefly discuss two common approaches to detect gene copy number gains from NGS data, i.e., the relative coverage and B-allele frequencies. In addition, we provide recommendations for reporting gene copy gains for clinical purposes. In addition to general QC metrics associated with NGS in routine diagnostics, it is recommended to include clinically relevant quantitative parameters of copy number gains in the clinical report, such as (i) relative coverage and estimated copy numbers in neoplastic cells, (ii) statistical scores to show significance (e.g., z-scores), and (iii) the sensitivity of the assay and restrictions of NGS-based detection of copy number gains. Collectively, this information can guide clinical and analytical decisions such as the reliable detection of high-level gene amplifications and the requirement for additional in situ assays in case of borderline results or limited sensitivity.

SUBMITTER: Eijkelenboom A

PROVIDER: S-EPMC6581937 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Recommendations for the clinical interpretation and reporting of copy number gains using gene panel NGS analysis in routine diagnostics.

Eijkelenboom Astrid A Tops Bastiaan B J BBJ Tops Bastiaan B J BBJ van den Berg Anke A van den Brule Adrianus J C AJC Dinjens Winand N M WNM Dubbink Hendrikus J HJ Ter Elst Arja A Geurts-Giele Willemina R R WRR Groenen Patricia J T A PJTA Groenendijk Floris H FH Heideman Daniëlle A M DAM Huibers Manon M H MMH Huijsmans Cornelis J J CJJ Jeuken Judith W M JWM van Kempen Léon C LC Korpershoek Esther E Kroeze Leonie I LI de Leng Wendy W J WWJ van Noesel Carel J M CJM Speel Ernst-Jan M EM Vogel Maartje J MJ van Wezel Tom T Nederlof Petra M PM Schuuring Ed E Ligtenberg Marjolijn J L MJL

Virchows Archiv : an international journal of pathology 20190319 6

Next-generation sequencing (NGS) panel analysis on DNA from formalin-fixed paraffin-embedded (FFPE) tissue is increasingly used to also identify actionable copy number gains (gene amplifications) in addition to sequence variants. While guidelines for the reporting of sequence variants are available, guidance with respect to reporting copy number gains from gene-panel NGS data is limited. Here, we report on Dutch consensus recommendations obtained in the context of the national Predictive Analysi ...[more]

PMID: 30888490

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Project description:Background: Targeted next generation sequencing (NGS) panels are increasingly being used in clinical genomics to increase capacity, throughput and affordability of gene testing. Identifying whole exon deletions or duplications (termed exon copy number variants, 'exon CNVs') in exon-targeted NGS panels has proved challenging, particularly for single exon CNVs. Methods: We developed a tool for the Detection of Exon Copy Number variants (DECoN), which is optimised for analysis of exon-targeted NGS panels in the clinical setting. We evaluated DECoN performance using 96 samples with independently validated exon CNV data. We performed simulations to evaluate DECoN detection performance of single exon CNVs and to evaluate performance using different coverage levels and sample numbers. Finally, we implemented DECoN in a clinical laboratory that tests BRCA1 and BRCA2 with the TruSight Cancer Panel (TSCP). We used DECoN to analyse 1,919 samples, validating exon CNV detections by multiplex ligation-dependent probe amplification (MLPA). Results: In the evaluation set, DECoN achieved 100% sensitivity and 99% specificity for BRCA exon CNVs, including identification of 8 single exon CNVs. DECoN also identified 14/15 exon CNVs in 8 other genes. Simulations of all possible BRCA single exon CNVs gave a mean sensitivity of 98% for deletions and 95% for duplications. DECoN performance remained excellent with different levels of coverage and sample numbers; sensitivity and specificity was >98% with the typical NGS run parameters. In the clinical pipeline, DECoN automatically analyses pools of 48 samples at a time, taking 24 minutes per pool, on average. DECoN detected 24 BRCA exon CNVs, of which 23 were confirmed by MLPA, giving a false discovery rate of 4%. Specificity was 99.7%. Conclusions: DECoN is a fast, accurate, exon CNV detection tool readily implementable in research and clinical NGS pipelines. It has high sensitivity and specificity and acceptable false discovery rate. DECoN is freely available at www.icr.ac.uk/decon.

Dataset Information

Recommendations for the clinical interpretation and reporting of copy number gains using gene panel NGS analysis in routine diagnostics.

Publications

Recommendations for the clinical interpretation and reporting of copy number gains using gene panel NGS analysis in routine diagnostics.

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