The role of epithelial-mesenchymal transition in the acquired resistance to epidermal growth factor receptor-tyrosine kinase inhibitors in non-small cell lung cancer

ZENG Yunyun; ZHANG Weimin; ZHANG Xi

Abstract：Objective To explore the role of epithelial?mesenchymal transition( EMT) in the acquired resistance to epidermal growth factor receptor?tyrosine kinase inhibitors ( EGFR?TKIs ) in non?small cell lung cancer ( NSCLC ) . Methods The EGFR del E746?A750?mutated human lung adenocarcinoma PC9/AB cell line and the EGFR wild?type H460/ER cell line were used in this study. The role of EMT in the acquired resistance to EGFR?TKIs was investigated by establishing stable E?cadherin over?expression cell lines( PC9/AB?CDH1 and H460/ER?CDH1) by transforming gene?CDH1 with lentivirus. MTT assay was used to measure the cell pro?liferation. Wound?healing assay and Transwell assay were adopted to determine the migration and invasion ability of the cells. The mR?NA and protein expressions of E?cadherin, Vimentin, Snail,β?catenin and EGFR were determined by the real?time fluorescence quan?titative PCR( qRT?PCR) and Western blotting, respectively. Results EMT( low E?cadherin and high Vimentin) was found in H460/ER and PC9/AB cells, neither T790M mutation nor c?Met amplification were detected. Over?expression of E?cadherin in both PC9/AB?CDH1 and H460/ER?CDH1 cells reversed morphological signature of EMT. Reversing of EMT remarkably increased the sensitivity to EGFR?TKIs in PC9/AB?CDH1and H460/ER?CDH1 cells. Compared with PC9/AB cells, the sensitivity to gefitinib in PC9/AB?CDH1 cells increased 11?4 folds. The half?inhibition concentration(IC50)of PC9/AB?CDH1 and PC9/AB cells was (0?70±0?22)μmol/L and (8?68±0?44)μmol/L with statistical significance(P<0?05). Compared with H460/ER cells, the sensitivity to erlotinib in H460/ER?CDH1 cells increased 6?1 folds. The IC50 of H460/ER?CDH1 and H460/ER cells was ( 7?51 ± 1?12) μmol/L and ( 53?72 ± 12?95) μmol/L with statistical significance(P<0?05). The expressions of EGFR and its phosphorylation form in both PC9/AB?CDH1and H460/ER?CDH1 cells were significantly increased( P<0?05) . Conclusion It demonstrates that reversing of EMT can re?verse the acquired gefitinib/erlotinib?resistance in NSCLC, which suggests that EMT plays an important role in the acquired resistance to EGFR?TKIs in NSCLC, possibly through down?regulating the phosphorylation of EGFR.