Gui-Li Huang1, Dong-Yan Shen1, Cheng-Fu Cai1, Qiu-Yan Zhang1, Hong-Yue Ren1, Qing-Xi Chen1. 1. Gui-Li Huang, Qiu-Yan Zhang, Qing-Xi Chen, State Key Laboratory of Cellular Stress Biology, Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystems, School of Life Sciences, Xiamen University, Xiamen 361005, Fujian Province, China.
Abstract
AIM: To develop a safe and effective agent for cholangiocarcinoma (CCA) chemotherapy. METHODS: A drug combination experiment was conducted to determine the effects of β-escin in combination with chemotherapy on CCA cells. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was performed to determine the effects of β-escin and common chemotherapeutics on the proliferation of human CCA cells (QBC939, Sk-ChA-1, and MZ-ChA-1). Immunocytochemistry was used to detect the expression of P-glycoprotein (P-gp) protein. Luciferase reporter assay was used to detect the activation of the Wnt/β-catenin pathway. The protein levels of P-gp, pS9-GSK3β, pT216-GSK3β, GSK3β, β-catenin, and p-β-catenin were further confirmed by western blotting. RESULTS: The drug sensitivity of QBC939 and QBC939/5-fluorouracil (5-FU) cells to 5-FU, vincristine sulfate (VCR), or mitomycin C was significantly enhanced by β-escin compared with either agent alone (P<0.05). In addition, the combination of β-escin (20 μmol/L) with 5-FU and VCR was synergic with a combination index<1. Further investigation found that the mRNA and protein expression of P-gp was down-regulated by β-escin. Moreover, β-escin induced GSK3β phosphorylation at Tyr-216 and dephosphorylation at Ser-9, resulting in phosphorylation and degradation of β-catenin. Interestingly, activation of the GSK3β/β-catenin pathway induced by Wnt3a resulted in up-regulation of P-gp, which was effectively abolished by β-escin, indicating that β-escin down-regulated P-gp expression in a GSK3β-dependent manner. CONCLUSION: β-escin was a potent reverser of P-gp-dependent multidrug resistance, with said effect likely being achieved via inhibition of the GSK3β/β-catenin pathway and thus suggesting a promising strategy of developing combination drugs for CCA.
AIM: To develop a safe and effective agent for cholangiocarcinoma (CCA) chemotherapy. METHODS: A drug combination experiment was conducted to determine the effects of β-escin in combination with chemotherapy on CCA cells. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was performed to determine the effects of β-escin and common chemotherapeutics on the proliferation of human CCA cells (QBC939, Sk-ChA-1, and MZ-ChA-1). Immunocytochemistry was used to detect the expression of P-glycoprotein (P-gp) protein. Luciferase reporter assay was used to detect the activation of the Wnt/β-catenin pathway. The protein levels of P-gp, pS9-GSK3β, pT216-GSK3β, GSK3β, β-catenin, and p-β-catenin were further confirmed by western blotting. RESULTS: The drug sensitivity of QBC939 and QBC939/5-fluorouracil (5-FU) cells to 5-FU, vincristine sulfate (VCR), or mitomycin C was significantly enhanced by β-escin compared with either agent alone (P<0.05). In addition, the combination of β-escin (20 μmol/L) with 5-FU and VCR was synergic with a combination index<1. Further investigation found that the mRNA and protein expression of P-gp was down-regulated by β-escin. Moreover, β-escin induced GSK3β phosphorylation at Tyr-216 and dephosphorylation at Ser-9, resulting in phosphorylation and degradation of β-catenin. Interestingly, activation of the GSK3β/β-catenin pathway induced by Wnt3a resulted in up-regulation of P-gp, which was effectively abolished by β-escin, indicating that β-escin down-regulated P-gp expression in a GSK3β-dependent manner. CONCLUSION: β-escin was a potent reverser of P-gp-dependent multidrug resistance, with said effect likely being achieved via inhibition of the GSK3β/β-catenin pathway and thus suggesting a promising strategy of developing combination drugs for CCA.