1,25-dihydroxyvitamin D_3 induces cell cycle arrest of MDA-MB-231 cells transfected with exogenous ER? expression vector containing vitamin D response elements / 第三军医大学学报

ABSTRACT

Objective To investigate the effects and molecular mechanism of tamoxifen on cell cycle distribution of ER-negative MDA-MB-231 cells re-expressing exogenous ER? gene under induction of Vit D3. Methods ER?/pcDNA3.1+ eukaryotic plasmid containing 4 copies of vitamin D response elements (VDRE) and Tk promoter was transfected into MDA-MB-231 cells. MTT assay and flow cytometry were used to analyze the inhibitory effects of Vit D3 and Tamoxifen on cell proliferation and cell cycle distribution. Then RT-PCR and Western blot assay were employed to detect the expressions of Cyclin D1, CDK4, Rb and phosphorylated Rb. The binding affinity of Cyclin D1/CDK4 complex was also analyzed by using immunoprecipitation assay. Results After concomitant treatment with Vit D3 and Tamoxifen for 72 h, Vit D3 and Tamoxifen synergetically inhibited proliferation of MDA-MB-231VDRE-ER? cells, and the majority of cells were arrested in G0/G1 phase. Cyclin D1 expression and the binding affinity of Cyclin D1/CDK4 complex were markedly down-regulated after 24 h of the concomitant treatment. Concurrently, the phosphorylation of Rb protein was also effectively inhibited while its expression exhibited no evident changes. Conclusion Vit D3 could effectively restore the sensitivity of ER-negative breast cancer cells transfected with exogenous ER? vector containing VDREs-Tk promoter to Tamoxifen by regulating the expression and activity of G1/S phase-related molecules.