Abstract 1969: Cancer exosomes as a new mediator to spread drug resistance to EGFR tyrosine kinase inhibitors

Abstract

Abstract Aim: The study aims to identify new therapeutic targets from cancer exosomes, which may spread drug resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in the treatment of non-small-cell lung cancer (NSCLC). Background: EGFR TKIs have exhibited extraordinary clinical efficacy in advanced lung cancer. However, their usefulness is compromised by drug resistance mediated by various mechanisms, including the emergence of a secondary EGFR T790M mutation. Exosomes are small membrane vesicles derived from the cellular phenomenon of membrane blebbing under both normal and pathological conditions. They mediate intercellular cross-talk by transferring receptors, antigens and cytokines from donor to recipient cells. It has been reported that EGFR TKI resistance mutation exists before drug exposure. Selective pressure from EGFR TKI therapy may thus cause the expansion of the latent resistant subclones, resulting in the emergence of a resistant phenotype in the entire tumor population. We hypothesize that exosomes may mediate the transfer of the secondary EGFR mutant or other mediators from the resistant subclones to neighboring sensitive cells. Method: The possible exosome-mediated transfer of secondary EGFR mutant (or other protein mediators) from EGFR T790M-bearing NSCLC (H1975) cells to a sensitive NSCLC cell line (HCC827) was evaluated. Exosomes were isolated from H1975 cells-conditioned medium by the standard differential centrifugation method. The secretion of protein mediators into exosomes by resistant H1975 cells was studied by protein array. The EGFR kinase activity of the exosomes was evaluated by biochemical assay. The uptake of fluorescence-labeled exosome from the resistant cells by the sensitive cells was visualized by fluorescence microscopy. The activation of the EGFR signaling pathway in the recipient sensitive cells after incubation with exosomes derived from the resistant cells was examined by Western blot analysis. Result: Exosomes were successfully isolated from H1975-conditioned medium and confirmed by the expression of exosomal markers CD63 and HSP70. Compared with exosomes derived from sensitive HCC827 cells, exosomes secreted from H1975 cells were enriched with apoptosis-regulating proteins including Src. H1975-derived exosomes increased the viability of HCC827 cells in the presence of gefitinib and inhibited the apoptosis induced by gefitinib. Western blot analysis showed that the exosomes can block the reduction of BCL-2 and full-length PARP, while abolishing the increase of BAX and cleaved PARP induced by gefitinib in HCC827 cells. Conclusion: H9175-derived exosomes may be involved in the spreading of drug resistance to EGFR TKIs by regulating cell apoptosis. New treatment strategies may be designed to target and inhibit this exosome-mediated transfer of mediators implicated in the acquired resistance to EGFR TKIs. Citation Format: Kenneth KW To, Christy WS Tong, Mia MX Wu. Cancer exosomes as a new mediator to spread drug resistance to EGFR tyrosine kinase inhibitors [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 1969.