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Background  Circulating cell-free DNA (cfDNA) is emerging as a surrogate sample type for mutation analyses. To improve the clinical utility of cfDNA, we developed a sensitive peptide nucleic acid (PNA)-based method for analyzing  EGFR  and  KRAS  mutations in the plasma cfDNA of patients with advanced non-small cell lung cancer (NSCLC).    Methods  Baseline tissue and plasma samples were collected from treatment-naïve advanced NSCLC patients participated in a randomized phase II study, which was registered with ClinicalTrials.gov at Feb. 2009 (NCT01003964).  EGFR  and  KRAS  mutations in the plasma cfDNA were analyzed retrospectively using a PNA clamping-assisted fluorescence melting curve analysis. The results were compared with those obtained from tissue analysis performed using the direct sequencing. Exploratory analyses were performed to determine survival predicted by the plasma and tissue mutation status.    Results  Mutation analyses in matched tissue and plasma samples were available for 194 patients for  EGFR  and 135 patients for  KRAS . The mutation concordance rates were 82.0 % (95 % confidence interval [CI], 76.5–87.4) for  EGFR  and 85.9 % (95 % CI, 80.1–91.8) for  KRAS . The plasma  EGFR  mutation test sensitivity and specificity were 66.7 % (95 % CI, 60.0–73.3) and 87.4 % (95 % CI, 82.7–92.1), respectively, and the plasma  KRAS  mutation test sensitivity and specificity were 50.0 % (95 % CI, 41.6–58.4) and 89.4 % (95 % CI, 84.2–94.6), respectively. The predictive value of the plasma EGFR and KRAS mutation status with respect to survival was comparable with that of the tissue mutation status.    Conclusions  These data suggest that plasma  EGFR  and  KRAS  mutations can be analyzed using PNA-based real-time PCR methods and used as an alternative to tumor genotyping for NSCLC patients when tumor tissue is not available.    Electronic supplementary material  The online version of this article (doi:10.1186/s12885-016-2678-2) contains supplementary material, which is available to authorized users.

bmc cancer

PNA clamping-assisted fluorescence melting curve analysis for detecting EGFR and KRAS mutations in the circulating tumor DNA of patients with advanced non-small cell lung cancer