Validation of the Oncomine™ focus panel for next-generation sequencing of clinical tumour samples.
Virchows Arch
; 473(4): 489-503, 2018 Oct.
Article
in En
| MEDLINE
| ID: mdl-30105577
ABSTRACT
The clinical utility of next-generation sequencing (NGS) for a diverse range of targets is expanding, increasing the need for multiplexed analysis of both DNA and RNA. However, translation into daily use requires a rigorous and comprehensive validation strategy. The aim of this clinical validation was to assess the performance of the Ion Torrent Personal Genome Machine (IonPGM™) and validate the Oncomine™ Focus DNA and RNA Fusion panels for clinical application in solid tumour testing of formalin-fixed, paraffin-embedded (FFPE) tissue. Using a mixture of routine FFPE and reference material across a variety of tissue and specimen types, we sequenced 86 and 31 samples on the Oncomine™ Focus DNA and RNA Fusion assays, respectively. This validation considered a number of parameters including the clinical robustness of the bioinformatics pipeline for variant detection and interpretation. The Oncomine™ Focus DNA assay had a sample and variant-based sensitivity of 99.1 and 97.1%, respectively, and an assay specificity of 100%. The Oncomine™ Focus Fusion panel had a good sensitivity and specificity based upon the samples assessed, however requires further validation to confirm findings due to limited sample numbers. We observed a good sequencing performance based upon amplicon, gene (hotspot variants within gene) and sample specific analysis with 92% of clinical samples obtaining an average amplicon coverage above 500X. Detection of some indels was challenging for the routine IonReporter™ workflow; however, the addition of NextGENe® software improved indel identification demonstrating the importance of both bench and bioinformatic validation. With an increasing number of clinically actionable targets requiring a variety of methodologies, NGS provides a cost-effective and time-saving methodology to assess multiple targets across different modalities. We suggest the use of multiple analysis software to ensure identification of clinically applicable variants.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Biomarkers, Tumor
/
Sequence Analysis, RNA
/
Sequence Analysis, DNA
/
Molecular Diagnostic Techniques
/
High-Throughput Nucleotide Sequencing
/
Neoplasms
Type of study:
Diagnostic_studies
/
Prognostic_studies
Limits:
Humans
Language:
En
Journal:
Virchows Arch
Journal subject:
BIOLOGIA MOLECULAR
/
PATOLOGIA
Year:
2018
Document type:
Article
Affiliation country:
United kingdom