The Effect Of Epirubicin Sequential NK Cells Against Breast Cancer Cells In Vitro

Purpose:To study the role of Epirubicin (EPI) on cytotoxicity of NK cells against breastcancer cells in vitro and analyze the potential mechanisms.Methods:Three types of breast cancer cell lines (MCF-7, SKBR-3, MDA-MB-231) withdifferent expressions statuses of ER, PR and HER-2were chosen. ER+, ER+, HER-2–breast cancer cell line: MCF-7; ER–, ER–, HER-2+++breast cancer cell line:SKBR-3and ER–, ER–, HER-2–breast cancer cell line: MDA-MB-231. EPI withdifferent concentrations inhibited the proliferation of breast cancer cells and NK cellswere detected by MTT assay. Calcein-AM released assays were applied to detect thechange of cytotoxicity by EPI sequential NK cells against breast cancer cells. Theexpression of NKG2D ligands (ULBP1, ULBP2and MICA) on the surface of breastcancer cells before or after treatment of EPI were analyzed by flow cytometry. Thesecretion of IFN-γ and TNF-were detected by ELISA, the expression of perforin andGranzyme B was analyzed by flow cytometry.Results:EPI significantly inhibit the proliferation of three breast cancer cells and NKcells, the inhibition effects were dose and time dependent. The killing effect of NKcells to the three kinds of breast cancer cells were obviously increased in vitro, whichwas (effector: target ratio) dependent. The cytotoxicity of NK cells against breastcancer cells with12hr EPI (5.0μg/ml) pretreatment was significantly higher thanuntreated and EPI alone. Thus, EPI sequential NK cells show the synergisticcytotoxicity effects against breast cancer cells. Breast cancer cells show increasedexpression of NKG2D ligands (ULBP1, ULBP2and MICA) after treatment of EPI,therefore, increased NKG2D ligands bring out increased NK cells killng effect.Additionally, EPI sequential NK cells secret more IFN-γ and TNF-, amplify the killng effect. EPI sequential NK cells showed decreased expression of perforin andGranzyme B, which need to further study.Conclusion:EPI sequential NK cells show the synergistic cytotoxicity effects against breastcancer cells. These would be explained by up-regulat NKG2D ligands expression onbreast cancer cells, and the increased secretion of IFN-γ and TNF-. EPI sequentialNK cells therapy was suitable for almost all breast cancer types.