The Role Of C-Myc-mediated Warburg Effect In The Inhibition Of Invasion And Metastasis Of Breast Cancer Cells Via Calycosin

Objective: To explore the effect of the phytoestrogen calycosin on migration and invasion ability of ER-positive breast cancer cells MCF-7and ER-negative breast cancer cells MDA-MB-231,and investigate the role of c-Myc-mediated Warburg effect in this anti-metastastic effects.Methods: MCF-7 and MDA-MB-231 of breast cancer cells were respectively treated with different concentrations of calycosin(0,25,50,100,150 ?M).The effects of calycosin on cell migration and invasion were detected by wound-healing and transwell assays.Glucose intake,lactate dehydrogenase(LDH)activity and concentrations of pyruvate and lactate in cells were respectively determined.The expression levels of c-Myc and glycolytic enzymes were assessed by q RT-PCR and Western blot.Next,MCF-7 and MDA-MB-231 cells were respectively transfected with pc DNA-c-Myc and pc DNA-si RNA-c-Myc to up-regulate or down-regulate c-Myc gene expression,followed by treatment with 150?M calycosin.The alteration of cell migration and invasion was detected by wound-healing and transwell assays,while the alteration of glycolysis metabolism by glucose,LDH,pyruvate,lactate assays and Western blot.Result: Calycosin inhibited the migration and invasion of both ER-positive MCF-7 cells and ER-negative MDA-MB-231 cells in aconcentration-dependent manner.It was found that calycosin decreased glucose uptake,LDH activity,cytoplasmic lactate and pyruvate content,accompanied with inhibition of c-myc expression and glycolytic enzymes activities in cells(P<0.05 or P<0.01).Meanwhile,c-Myc overexpression significantly accelerated cell migration and invasion,and increased the Warburg effects(P<0.05 or P<0.01)in MCF-7 and MDA-MB-231 cells.On the contrary,the down-regulation of c-Myc had the opposite effect on cell metastasis and Warburg effect in the two cell lines.Notably,compared with calycosin alone,the down-regulated c-Myc expression enhanced the inhibitory effect of calycosin on MCF-7 and MDA-MB-231 cells.Cell migration and invasion was further inhibited,as well as the glucose uptake,LDH activity,cytoplasmic lactate and pyruvate content,and glycolytic enzymes activities(P<0.05 or P<0.01).On the other hand,calycosin-induced inhibitory effects on cell metastasis and Warburg effect was partially blocked by c-Myc overexpression(P<0.05 or P<0.01).Conclusion: By targeting c-Myc-mediated signaling,calycosin could reverse the Warburg effect in both ER-positive and ER-negative breast cancer cells,and then inhibit cell migration and invasion.This may provide new therapeutic options and targets for treatment of breast cancer.