Clinical Validation of Targeted Next Generation Sequencing for Colon and Lung Cancers

PLoS One. 2015 Sep 14;10(9):e0138245. doi: 10.1371/journal.pone.0138245. eCollection 2015.

Abstract

Objective: Recently, Next Generation Sequencing (NGS) has begun to supplant other technologies for gene mutation testing that is now required for targeted therapies. However, transfer of NGS technology to clinical daily practice requires validation.

Methods: We validated the Ion Torrent AmpliSeq Colon and Lung cancer panel interrogating 1850 hotspots in 22 genes using the Ion Torrent Personal Genome Machine. First, we used commercial reference standards that carry mutations at defined allelic frequency (AF). Then, 51 colorectal adenocarcinomas (CRC) and 39 non small cell lung carcinomas (NSCLC) were retrospectively analyzed.

Results: Sensitivity and accuracy for detecting variants at an AF >4% was 100% for commercial reference standards. Among the 90 cases, 89 (98.9%) were successfully sequenced. Among the 86 samples for which NGS and the reference test were both informative, 83 showed concordant results between NGS and the reference test; i.e. KRAS and BRAF for CRC and EGFR for NSCLC, with the 3 discordant cases each characterized by an AF <10%.

Conclusions: Overall, the AmpliSeq colon/lung cancer panel was specific and sensitive for mutation analysis of gene panels and can be incorporated into clinical daily practice.

Publication types

  • Clinical Trial
  • Research Support, Non-U.S. Gov't
  • Validation Study

MeSH terms

  • Carcinoma, Non-Small-Cell Lung / genetics*
  • Colonic Neoplasms / genetics*
  • DNA Mutational Analysis / methods
  • DNA, Neoplasm / genetics
  • Female
  • Gene Frequency*
  • High-Throughput Nucleotide Sequencing / methods*
  • Humans
  • Lung Neoplasms / genetics*
  • Male
  • Neoplasm Proteins / genetics*
  • Sensitivity and Specificity

Substances

  • DNA, Neoplasm
  • Neoplasm Proteins

Grants and funding

This work was supported by funding from the “Fonds Yvonne Boël” (Brussels, Belgium). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.