Curcumin Suppresses Doxorubicin Resistance In Breast Cancer Cells Via MDR1 And MDR3 Function Inhibition

Objective:Doxorubicin is one of the first-line chemotherapy drugs in the treatment of breast cancer,but most patients use of doxorubicin will appear drug resistance phenomenon,thereby reducing the efficacy.current studies have shown that drug transporter expression and function changes are closely related to tumor cell acquired resistance.recent years,curcumin has been found to have antitumor effects and increase the sensitivity of various chemotherapeutic drugs by regulating drug transporters.This study was to investigate the role of ABC family transporters MDR1 and MDR3 in the acquisition of doxorubicin in breast cancer and to explore the reversal effect of curcumin on these transporter-mediated acquired resistance and its molecular mechanism.Methods:1.Real-time PCR and immunofluorescence were used to screen the drug transporters that are closely related to acquired drug resistance in doxorubicin-sensitive MCF-7 cells and doxorubicin-resistant MCF-7/DOX cells.2.MDCK? monolayer cells Bidirectional transport function experiment were used to observe the effect of MDR1 and MDR3 on the delivery of doxorubicin in MCF-7 and MCF-7/DOX cells.The effects of two drug transporters on the accumulation of doxorubicin in the cells were examined by flow cytometry.3.The effects of DNA methylation and histone acetylation on the expression of MDR1 and MDR3 were observed by bisulfite sequencing(BSP)and chromatin immunoprecipitation(ChIP).4.The effects of curcumin on the sensitivity and intracellular accumulation of doxorubicin in MCF-7 and MCF-7/DOX cells were examined by CCK-8 and flow cytometry.5.The effects of curcumin on the expression of MDR1 and MDR3 in MCF-7 and MCF-7/DOX cells were observed by real-time fluorescence quantitative PCR and immunofluorescence.BSP and CHIP were used to observe the effect of curcumin on MDR1 and MDR3 gene methylation And the effect of histone acetylation modification.6.The effect of curcumin on MDR1 and MDR3 transport of doxorubicin was observed by MDCK? monolayer cells bidirectional transport function experiment.ATPase activity was used to observe the mechanism of curcumin on MDR1 and MDR3 function.Results:1.Compared with doxorubicin-sensitive MCF-7,the expression of MDR1 and MDR3 in the drug-resistant MCF-7/DOX cells significantly increased(4299.64 ± 326.96)and(350.651 ± 42.18)times(P <0.05).Their protein levels were upregulated(4.83±0.37)and(2.46±0.23)times,respectively(P <0.05).2.MDR1 and MDR3 both have the transport function to doxorubicin,the transfer kinetic parameters Km values are(18.74±6.35)and(63.12±20.13)??;Vmax values were(400.78±120.45)and(194.18±89.62)nmol·min-1·mg-1.3.the expression level of MDR1 and MDR3 were inversely proportional to the sensitivity and intracellular doxorubicin concentration in breast cancer cells.3.The up-regulation of the expression of MDR1 and MDR3 in MCF-7/DOX cells is closely related to the low methylation of DNA and the high acetylation level of histone in the promoter region.4.Curcumin can significantly increase the doxorubicin sensitivity and intracellular accumulation of doxorubicin in MCF-7 and MCF-7 / DOX cells in a concentration-and time-dependent manner.5.The effect of curcumin on the expression of MDR1 and MDR3 in MCF-7 and MCF-7/DOX cells was correlated with the methylation level and the level of histone acetylation related in MDR1 and MDR3 promoter.6.Curcumin has a significant inhibitory effect on MDR1 and MDR3 mediated doxorubicin efflux,which is closely related to inhibition of ATPase activity in both transporters.Conclusions:ABC family drug transporter MDR1 and MDR3 have a transport function to doxorubicin,and their expression level in breast cancer MCF-7 cells doxorubicin acquired drug resistance was significantly increased;reduce MDR1 and MDR3 expression Level of MCF-7 and MCF-7/DOX cells can increase the concentration of doxorubicin in MCF-7 and MCF-7/DOX cells,and increase the sensitivity of MDR1 and MDR3.The up-regulation of MDR1 and MDR3 is closely related to methylation of histone and histone acetylation.Curcumin can significantly increase the sensitivity and intracellular accumulation of doxorubicin in breast cancer cells,mainly related to its inhibition of MDR1 and MDR3 ATPase activity and thus affect the efflux of doxorubicin.