Cancer Control (Mar 2023)

Detection of Nine Oncogenes Amplification in Lung and Colorectal Cancer Formalin-Fixed Paraffin-Embedded Tissue Samples using Combined Next-Generation Sequencing-Based Script and Digital Droplet Polymerase Chain Reaction

  • Christophe Bontoux MD,
  • Alice Guyard MD,
  • Audrey Lupo MD, PhD,
  • Karim Diallo MSc,
  • Khedidja Rebah MD,
  • Agathe Hercent MD,
  • Edouard Guenzi MD,
  • Jérome Cros MD, PhD,
  • Jérome Lamoril MD,
  • Karen Leroy PharmD, PhD,
  • Nathalie Theou-Anton PharmD, PhD

DOI
https://doi.org/10.1177/10732748231167257
Journal volume & issue
Vol. 30

Abstract

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Introduction Gene copy number variations have theranostic impact and require reliable methods for their identification. We aimed to evaluate the reliability of combined next-generation sequencing (NGS) and digital droplet PCR (ddPCR) method for gene amplification evaluation. Methods We conducted a retrospective multicentric observational study. MET/ERBB2 amplifications were assessed in patients with lung or colorectal carcinoma (cohort A), from 2016 to 2020, by fluorescence in situ hybridization (FISH)/immunohistochemistry (IHC), NGS and ddPCR. NGS-based script and ddPCR were then used to detect amplifications of 7 additional oncogenes ( EGFR, KRAS, BRAF, FGFR1, FGFR2, FGFR3, PIK3CA) in a cohort of patients (cohort B). Results 55 patients (9 control, 25 ERBB2- amplified and 21 MET- amplified) out of 3779 patients tested were included in cohort A. Correlation coefficient between NGS-based script and FISH/IHC results were .88 for MET (P < .001) and .89 (P < .001) for ERBB2 . Using a threshold ratio of 1.56 with the NGS-based script, the sensitivity was 100% for both genes and the specificity 69% for MET and 90% for ERBB2 , respectively . With an alternative 1.76 threshold, sensitivity was 94% for MET and 96% for ERBB2 , while specificity was 85% for MET and 90% for ERBB2 . Correlation coefficient between FISH and ddPCR ratio was .90 for MET and .88 for ERBB2 . In both cohorts, NGS-based script and ddPCR results were significantly correlated regarding all genes (P < .001). Conclusion Combined NGS-based script and ddPCR method is reliable and easily feasible for the detection of gene amplifications, providing useful data for guided therapy in cancer.