In this study, we detected the cytotoxic activity of gemcitabine-induced autophagy in breast cancer cell lines with different ER status (ER positive MCF-7, ER negative MDA-MB-231 and ER− Bcap 37/ER+ Bcap 37), and investigated the effect of gemcitabine-induced autophagy after inhibition of ERα-mediated signal pathway in MCF-7 cells. This evidence concerns the gene ESR1 and breast cancer.