Introduction: Tissue rebiopsy has limited value for detecting T790M mutation to prescribe the third-generation EGFR-TKIs. Liquid biopsy using cell-free DNA is being adopted but it plays a complementary role because of low sensitivity. Recently, extracellular vesicles (EV) are proven to contain double-stranded DNA and liquid biopsy using EV DNA is highly promising owing to its intact stability. We aimed to investigate the feasibility of liquid biopsy using EV DNA isolated from bronchoalveolar lavage fluid (BALF) and plasma in addition to tissue rebiopsy to detect T790M mutation in NSCLC patients who developed acquired resistance to EGFR-TKIs

Methods: From October 2014 to December 2016, we enrolled 30 patients who had disease progression after previous treatment with EGFR-TKIs. Rebiopsy was performed in 23 patients and liquid biopsy was done in 16 BALF and 6 plasma samples. Sixteen BALF samples were obtained in 9 T790M-negative cases by rebiopsy and 7 patients ineligible for rebiopsy. Plasma testing was done in 6 BALF T790M-negative cases. EV were isolated from BALF and plasma by ultracentrifuge method. After DNA extraction from the isolated EV, EGFR genotyping to detect T790M mutation was done by peptide nucleic acid (PNA)-clamping method. We retrospectively reviewed patient characteristics, types of tumor rebiopsy procedure, and responses to third generation EGFR-TKIs.

Results: Thirty patients who experienced progression to EGFR-TKIs were evaluated. Tumor rebiopsy was not possible in 7 patients. Out of 23 patients who performed tumor rebiopsy, 22 patients achieved adequate tissue specimen for evaluation, among whom 5 patients conducted rebiopsy twice (4 patients) or three times (1 patient), but 1 patient did not get evaluable tumor cells. Twenty two evaluable tumor rebiopsy revealed 54.5% (12/22) of T790M positivity. Among 17 patients who showed T790M negativity for tumor rebiopsy (n=10) or were ineligible for tumor rebiopsy (n=7), BAL was done in 16 patients and EGFR genotyping using BALF EV DNA showed 31.3% (5/16) of T790M positivity. In 11 patients with T790M negativity from EGFR genotyping using BALF EV DNA, 6 available plasma samples were tested for EGFR genotyping using plasma EV DNA and 3 patients revealed T790M positivity (3/6 = 50%). Finally, we found 8 more T790M positive patients by liquid biopsy (BALF EV DNA = 5, plasma EV DNA = 3) in addition to standard tissue rebiopsy (n=12).

Conclusion: Liquid biopsy using EV DNA isolated from BALF and plasma could be an alternative and promising method to detect T790M mutation in NSCLC patients with acquired resistance to EGFR-TKIs.

Citation Format: Jong Sik Lee, Hee Joung Kim, In Ae Kim, Jae Young Hur, Kye Young Lee. Feasibility of liquid biopsy using extracellular vesicle-derived DNA isolated from bronchoalveolar lavage fluid and plasma in addition to tissue rebiopsy to detect T790M mutation in NSCLC patients with acquired resistance to EGFR-TKIs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1596.