Abstract
Acquisition of resistance to docetaxel (Doc) is one of the most important problems in treatment of breast cancer patients, but the underlying mechanisms are still not fully understood. In present study, Doc-resistant MDA-MB-231 and MCF-7 breast cancer cell lines (MDA-MB-231/Doc and MCF-7/Doc) were successfully established in vitro by gradually increasing Doc concentration on the basis of parental MDA-MB-231 and MCF-7 cell lines (MDA-MB-231/S and MCF-7/S). The potential miRNAs relevant to the Doc resistance were screened by miRNA microarray. We selected 5 upregulated miRNAs (has-miR-3646, has-miR-3658, has-miR-4438, has-miR-1246, and has-miR-574-3p) from the results of microarray for RT-qPCR validation. The results showed that expression level of miR-3646 in MDA-MB-231/Doc cells was significantly higher than in MDA-MB-231/S cells. Compared to MCF-7/S cells, miR-3646 expression was up-regulated in MCF-7/Doc cells. Further studies revealed that transfection of miR-3646 mimics into MDA-MB-231/S or MCF-7/S cells remarkably increased their drug resistance, in contrast, transfection of miR-3646 inhibitors into MDA-MB-231/Doc or MCF-7/Doc cells resulted in significant reduction of the drug resistance. By the pathway enrichment analyses for miR-3646, we found that GSK-3β/β-catenin signaling pathway was a significant pathway, in which GSK-3β was an essential member. RT-qPCR and Western blot results demonstrated that miR-3646 could regulate GSK-3β mRNA and protein expressions. Furthermore, a marked increase of both nuclear and cytoplasmic β-catenin expressions (with phosphorylated-β-catenin decrease) was observed in MDA-MB-231/Doc cells compared with MDA-MB-231/S cells, and their expression were positively related to miR-3646 and negatively correlated with GSK-3β. Taken together, our results suggest that miR-3646-mediated Doc resistance of breast cancer cells maybe, at least in part, through suppressing expression of GSK-3β and resultantly activating GSK-3β/β-catenin signaling pathway.
Highlights
Breast cancer is one of the most commonly diagnosed types of malignant tumors among women of all racial and ethnic groups, and is expected to account for 29% of all new cancer cases among American women in 2015 [1]
We selected miR-3646 to investigate its association with the sensitivity of breast cancer cells to Doc and potential mechanism
RTqPCR analysis indicated that miR-3646 was up-regulated by 15.67-fold compared to negative control of mimics (NC) in MDA-MB-231/S cells, and increased by 18.38-fold compared to NC in MCF-7/S cells (Fig 1B and 1E)
Summary
Breast cancer is one of the most commonly diagnosed types of malignant tumors among women of all racial and ethnic groups, and is expected to account for 29% of all new cancer cases among American women in 2015 [1]. Docetaxel (Doc) as a taxane compound by disrupting tumor cell mitosis has been routinely used solely or in combination with other anti-cancer drugs in the treatment of advanced or metastasis breast cancer. The resistance to Doc is often observed during treatment of breast cancer patients [3]. A number of studies have shown several mechanisms involved in acquiring drug resistance in breast cancer, including the alteration of drug transporters effluxed anticancer agents, DNA methylation, histone modification, activation of cell-survival pathway and/or inhibition of apoptotic pathway [4,5,6]. Several studies have demonstrated that deregulation of miRNAs are associated with drug-resistance of cancers including breast cancer [9]
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