Simple method to detect important epidermal growth factor receptor gene mutations with bronchoscopic specimens of lung cancer patients for gefitinib treatment

Recent studies have demonstrated that epidermal growth factor receptor (EGFR) mutations are a predictive molecular marker of tumor response to gefitinib treatment in non-small-cell lung cancer (NSCLC). Early studies of EGFR mutations have been analyzed by direct sequencing with surgically resected specimens. However, it is often difficult to obtain sufficient and tumor-enriched tissue specimens from inoperative NSCLC patients for mutation analysis. Therefore, we set out to develop a simple, sensitive and reliable method based on polymerase chain reaction (PCR) and restriction endonuclease treatment to detect important EGFR mutations for gefitinib treatment, including G719S, deletion or insertion in exons 19 and 20, T790M, L858R and L861Q using bronchoscopic specimens. Two hundred and eleven bronchoscopic samples cytologically diagnosed as malignant were analyzed and mutations detected from 60 patients (28.4%). The present assay could detect deletion in exon 19 and L858R mutations in the presence of at least 1% and 5% mutant cells, respectively, from a background of normal cells. Comparison of mutation detection between bronchoscopic and surgical specimens from 42 patients indicated that the sensitivity of this assay using bronchoscopic specimens was 95% (18/19) with a specificity of 100% (23/23). Furthermore, deletion variants in exon 19 could be distinguished by native polyacrylamide gel electrophoresis. Consequently, this simple, sensitive and reliable assay can provide important information pertaining to optimal therapeutic strategies.