Calycosin-induced Inhibitory Effects On Migration And Invasion Of Human Breast Cancer Cells Via Rab27b-β-catenin-VEGF Signaling Pathway In Vitro And In Vivo

Objective: To explore the effect of the phytoestrogen calycosin on invasion and migration of human breast cancer cells MCF-7 and MDA-MB-231,and investigate the role of Rab27B-dependent signaling pathway in this effect.Methods: In in vitro experiment,MCF-7 and MDA-MB-231 cells were treated with different concentrations of calycosin（0,50,100,150,200 μM）respectively.The changes in cell migration and invasion were detected by wound healing and transwell assays,while the expression levels of Rab27 B by q RT-PCR and Western blot.Next,MCF-7 and MDA-MB-231 cells were transfected with Lv-Rab27B-GFP or Lv-sh Rab27B-GFP,and then subjected to incubation with calycosin.Finally,the migration and invasiveness of the transfected cells were assessed,as well as the Rab27 B,β-catenin and VEGF expression.In in vivo experiment,MDA-MB-231 cells were injected into tail vein of nude mice to produce metastatic tumors.The tumor-bearing mice were treated with intraperitoneal administration of calycosin（0,60,90 mg/kg）.Subsequently,the therapeutic efficacy of calycosin on tumor metastasis were evaluated,and the levels of Rab27 B,β-catenin and VEGF in metastatic tumors were determined.Result: The migration and invasion of MCF-7 and MDA-MB-231 cells were attenuated by calycosin in a dose-dependent manner in vitro,accompanied with decreased Rab27 B expression（P<0.05 or P<0.01）.In addition,Rab27 B overexpression stimulated expression of β-catenin and VEGF in the two breast cancer cell lines,resulting in promotion of cell migration and invasion（P<0.05）.On the contrary,down-regulated Rab27 B expression exerted inhibitory effects against the cancer cell metastasis,as well as β-catenin and VEGF expression（P<0.05）.Therefore,it was found that calycosin-induced inhibition on breast cancer cell metastasis could be further enhanced by Rab27 B down-regulation,whereas blocked by Rab27 B up-regulation（P<0.05）.Similarly,in vivo,calycosin significantly suppressed tumor metastasis of MDA-MB-231 cells to lung,as evidenced by the decreased number of tumor nodules（P<0.05）.Also,calycosin treatment reduced the expression levels of Rab27 B,β-catenin and VEGF in tumor tissue（P<0.05）.Conclusion: The metastatic activity of human breast cancer cells could be inhibited by calycosin,which may be mediated by inactivation of Rab27B-β-catenin-VEGF signaling pathway.This will provide us with new therapeutic approaches and targets for clinical treatment of breast cancer.