Chloroquine increases the anti-cancer activity of epirubicin in A549 lung cancer cells.

Abstract

The present study investigated whether the autophagy inhibitor chloroquine (CQ) can improve the sensitivity of the A549 lung cancer cell line to epirubicin (EPI). The Cell Counting Kit 8 (CCK8) assay was used to determine the EPI IC50 in A549 cells treated for 72 h. A549 cells were treated with Western blot analysis was performed to detect the expression level of the autophagy-associated protein, microtubule associated protein 1 light chain 3 β (LC3B), and apoptosis-associated proteins such as cleaved caspase-9 and cleaved caspase-3. CCK8, colony formation, wound healing and Transwell assays were performed to analyze cell proliferation, migration and invasion capacity. Reverse transcription-quantitative PCR (RT-qPCR) was used to analyze the mRNA expression levels of LC3B and beclin-1, and the apoptosis rate was analyzed by flow cytometry. The IC50 of EPI was 0.03 µg/ml. The CCK8 results demonstrated that the cell survival rate was lower in CQ + EPI-treated cells when compared with the individual treatment groups. The colony formation results revealed that the number of clones in the EPI + CQ-treated group was reduced compared with EPI or CQ treatment alone. The wound healing assay revealed that migration was reduced in the EPI + CQ-treated group compared with the other treatment groups, and the Transwell results indicated that the number of cells passing through the Matrigel and membrane was lowest in the CQ + EPI treatment group. The mRNA expression levels of LC3B and beclin-1 were increased in the CQ + EPI group by 51.5 and 61.2%, respectively, when compared with the control group. The results indicated that LC3B protein expression was enhanced by EPI in a concentration-dependent manner, and the protein levels of cleaved caspase-3 and cleaved caspase-9 were higher in the combination group than in the EPI alone group. The flow cytometry results demonstrated that the apoptosis rate was highest in the EPI + CQ group. In conclusion, the autophagy inhibitor CQ increased the sensitivity of A549 cells to EPI, and the underlying mechanism of action may be associated with the activation of apoptosis.