BACKGROUND: Much attention has been focused on epidermal growth factor receptor (EGFR) mutation testing since the introduction of EGFR-tyrosine kinase inhibitors have improved survival in EGFR-positive lung cancer patients. Liquid biopsy using circulating tumor cells or cell-free DNA (cfDNA) has enabled less invasive testing, but requires a highly sensitive method. To date, liquid biopsy using bronchoalveolar lavage (BAL) fluid has rarely been used. METHODS: From 20 patients with lung adenocarcinoma, we isolated cfDNA from 20 samples of cell-free BAL fluid and 19 cell-free bronchial washing samples. cfDNA was examined for EGFR mutations using peptide nucleic acid (PNA)-mediated PCR clamping method. In cases where the results from the tumor biopsy and BAL-derived cfDNA test were not consistent, PANAMutyper™ R EGFR kit was used along with PNA clamping-assisted fluorescence melting curve analysis. RESULTS: We included 17 patients with advanced stage disease and three with non-advanced stage disease. Tumor biopsy detected EGFR mutations in 12 of the patients. One patient had a p.L858R mutation and a de novo p.T790M mutation. The results from PNA-mediated PCR clamping were 75.0% (9/12) concordant with the tumor biopsy results for EGFR mutation status. PANAMutyper with fluorescence melting curve analysis was performed in three cases, which detected EGFR mutations in two more patients (11/12, 91.7%). EGFR mutations were detected in the cfDNA extracted from two bronchial washing samples. CONCLUSIONS: cfDNA from BAL fluid could be used for molecular testing of EGFR mutations and identification of p.T790M mutations, with an easily applicable method.
BACKGROUND: Much attention has been focused on epidermal growth factor receptor (EGFR) mutation testing since the introduction of EGFR-tyrosine kinase inhibitors have improved survival in EGFR-positive lung cancerpatients. Liquid biopsy using circulating tumor cells or cell-free DNA (cfDNA) has enabled less invasive testing, but requires a highly sensitive method. To date, liquid biopsy using bronchoalveolar lavage (BAL) fluid has rarely been used. METHODS: From 20 patients with lung adenocarcinoma, we isolated cfDNA from 20 samples of cell-free BAL fluid and 19 cell-free bronchial washing samples. cfDNA was examined for EGFR mutations using peptide nucleic acid (PNA)-mediated PCR clamping method. In cases where the results from the tumor biopsy and BAL-derived cfDNA test were not consistent, PANAMutyper™ R EGFR kit was used along with PNA clamping-assisted fluorescence melting curve analysis. RESULTS: We included 17 patients with advanced stage disease and three with non-advanced stage disease. Tumor biopsy detected EGFR mutations in 12 of the patients. One patient had a p.L858R mutation and a de novo p.T790M mutation. The results from PNA-mediated PCR clamping were 75.0% (9/12) concordant with the tumor biopsy results for EGFR mutation status. PANAMutyper with fluorescence melting curve analysis was performed in three cases, which detected EGFR mutations in two more patients (11/12, 91.7%). EGFR mutations were detected in the cfDNA extracted from two bronchial washing samples. CONCLUSIONS: cfDNA from BAL fluid could be used for molecular testing of EGFR mutations and identification of p.T790M mutations, with an easily applicable method.
Authors: Roberta Roncarati; Laura Lupini; Elena Miotto; Elena Saccenti; Susanna Mascetti; Luca Morandi; Cristian Bassi; Debora Rasio; Elisa Callegari; Valentina Conti; Rosa Rinaldi; Giovanni Lanza; Roberta Gafà; Alberto Papi; Antonio Frassoldati; Silvia Sabbioni; Franco Ravenna; Gian L Casoni; Massimo Negrini Journal: Mol Oncol Date: 2020-06-24 Impact factor: 6.603
Authors: Zuzana Pös; Ondrej Pös; Jakub Styk; Angelika Mocova; Lucia Strieskova; Jaroslav Budis; Ludevit Kadasi; Jan Radvanszky; Tomas Szemes Journal: Int J Mol Sci Date: 2020-11-16 Impact factor: 5.923