Comparison of next-generation sequencing methodologies and direct sequencing for the detection of EGFR mutations in non-small-cell lung carcinoma: Implications for optimal clinical testing.

Abstract

e22025 Background: The efficacy of next-generation sequencing (NGS) for the prediction of clinical benefits from EGFR TKIs in non small cell lung cancer(NSCLC) has yet to be fully established. The purpose of the present study was to evaluate the predictive effect of EGFR-evaluating methodologies on the use of EGFR TKIs in patients with NSCLC. Methods: cTumor samples were obtained from patients with NSCLC at 5 institutions in Korea. Study cohort A consisted of surgically resected or biopsy NSCLC samples enriched with adenocarcinoma histology. Study cohort B consisted of patients who received EGFR TKI during the treatment period. EGFR status was tested with direct sequencing, Peptide nucleic acid-locked nucleic acid PCR clamp (PNA-LNA PCR Clamp) method, and Personal genome machine (PGM) sequencing using the Ion AmpliSeq Cancer Panel on the Ion Torrent. Results: EGFR mutations in cohort A were identified in 16 (28.1%), 27 (47.4%), and 29 (50.9%) of 57 patients by direct sequencing, PNA-LNA clamping, and the Ion Torrent PGM, respectively. In 2 cases of EGFR mutant detected by Ion Torrento PGM in contrast to EGFR wild type by PNA-LNA clamping, EGFR mutation was found on novel sequence other than classical EGFR mutations. In total, 33 out of 37 patients in cohort B whose frozen NSCLC tissue was available were examined. Patients with EGFR mutations tested by PNA-LNA clamping did not show a significantly higher response rate than did patients with the wild-type EGFR (P=0.607). Mutational status detected by 2 methods did not significantly predict overall survival and progression free survival. However, curves showed a greater differentiation when the mutation was detected by PNA-LNA clamp as compared to mutations detected by direct sequencing. Conclusions: Though this study was conducted in a small number of cases, the results suggest that both PNA-LNA clamping and the Ion Torrent PGM are highly sensitive procedures relative to direct sequencing, and are useful screening tools for the detection of EGFR mutations in clinical practice.