MiR-375 is epigenetically downregulated due to promoter methylation and modulates multi-drug resistance in breast cancer cells via targeting YBX1

S.-L. Liu, Y.-F. SUI, M.-Z. Lin

Clinical Laboratory, the People’s Hospital of Changle, Weifang, Shandong, China. mingzhenlin1@outlook.com

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• Oncology

OBJECTIVE: In this study, we firstly verified how miR-375 is downregulated in breast cancer cells with multi-drug resistance (MDR) and further investigated the regulative effect of miR-375 on Ybx1 expression.

MATERIALS AND METHODS: MiR-375 expression and promoter methylation status were studied by retrieving data in NCBI GEO Datasets, qRT-PCR and Methylation-Specific PCR (MSP) assay. Drug sensitivity of the cancer cells was assessed using MTT assay. The binding between miR-375 and YBX1 gene was predicted using Targetscan 7.1 and verified using western blot and dual luciferase assay.

RESULTS: MiR-375 is significantly downregulated in both MCF-7/ADM and MCF-7/PTX cells than in MCF-7 cells. MCF-7/ADM and MCF-7/PTX cells had significantly higher level of promoter methylation than MCF-7 cells. 5-AZA-dC treatment significantly reduced the methylation in MCF-7/ADM and MCF-7/PTX cells and increased miR-375 expression. MiR-375 can directly target 3’UTR of YBX1 and thereby decrease its expression in MCF-7/ADM and MCF-7/PTX cells. Both miR-375 overexpression and YBX1 knockdown significantly decreased P-gp expression and increased chemosensitivity of the cancer cells.

CONCLUSIONS: MiR-375 is downregulated in MCF-7/ADM and MCF-7/PTX cells, and its downregulation is a result of promoter methylation. MiR-375 can directly target 3’UTR of YBX1 and thereby decrease its expression, which might be an important mechanism of MDR in breast cancer cells.