Abstract: OBJECTIVE to investigate the role and mechanism of miR-363 on cisplatin-resistant breast cancer cells. METHODS collected the serum from the advanced breast cancer patients who were treated with cisplatin-based chemotherapy. Then the expression of miR-363 in breast cancer patients’ serum was detected before or after chemotherapy using qPCR analysis. Constructed the cisplatin-resistant MCF-7 cell line, the cytotoxicity of cisplatin to MCF-7 cell line and MCF-7-R cell line was measured by MTT assay. Transfected the MCF-7-R cells with miR-363 to determine whether the transfection of miR-363 enhanced cytotoxicity of cisplatin to MCF-7-R cells. Confirmed whether the expression of Mcl-1 was regulated by miR-363 using bioinformatics, qPCR and western blot. Constructed the Mcl-1 expression vector, and detected the cytotoxicity of cisplatin combing with the cotransfection of miR-363 and Mcl-1 expression vector by MTT assay. RESULTS miR-363 levels were significantly decreased in advanced breast cancer patients treated with cisplatin-based chemotherapy. MiR-363 levels were also lower in MCF-7-R cells than in MCF-7 cells. Exogenous miR-363 significantly overcame cisplatin resistance in MCF-7-R cells, whereas miR-363 knockdown increased the cell viability during cisplatin treatment. It was demonstrated that miR-363 directly targeted to Mcl-1, and the downregulation of miR-363 resulted in upregulation of Mcl-1. miR-363 decreased cisplatin resistance of MCF-7-R cells, partly by targeting Mcl-1. CONCLUSION downregulation of miR-363 increases drug resistance in cisplatin-treated MCF-7 by downregulating Mcl-1.