Effects of itraconazole on proliferation and apoptosis of pancreatic cancer cells

Abstract

Objective To investigate the role and mechanism of the activation of B cell lymphoma/leukemia-2-antagonist/killer 1 (bak1) signaling pathway in anticancer effects of itraconazole on pancreatic cancers. Methods The experiment was divided into control group and itraconazole group according to the different detection indicators. Human pancreatic cancer MIA PaCa-2 and CFPAC-1 cells were treated with different concentrations of itraconazole (0, 10, 30, 50, 70, 90 mg/L) bought from Merk’s company in USA. Cell proliferation was evaluated by 3-(4, 5-dimethylhiazol-2-yl)-3, 5-diphenyltetrazolium bromide (MTT) assay. Cell cycle was analyzed by flow cytometry, and cell apoptosis was detected by TdT-mediated dUTP nick end labeling (TUNEL) immunofluorescence assay. Moreover, bak1 small interfering RNA (siRNA) was used to inhibit the expression of bak1, and then the proliferation and apoptosis of pancreatic cancer cells treated with itraconazole were evaluated. The expression of bak1 and its downstream bax and cysteinyl aspartate-specific protease (Caspase)-3 proteins was measured by Western blotting. SPSS 19.0 statistical software was used to analyze the measurement data. The mean soil standard deviation (Mean±SD) was used to express the measurement data. The analysis of variance was used for the comparison between groups. Results The inhibitory rates of 10, 30, 50, 70 and 90 mg/L itraconazole Mia PaCa-2 were 6.211%, 34.741%, 63.226%, 82.531% and 89.112% respectively, and the inhibitory rates of itraconazole on CFPAC-1 were 9.726%, 47.322%, 53.631%, 72.629% and 92.641% respectively. Before and after 50 μmol/L itraconazole intervention, Mia PaCa-2 G0/G1 phase was 43.142% and 57.341% (t=21.762, P<0.05), the difference was statistically significant; CFPAC-1 G0/G1 phase was 40.107% and 63.216% (t=17.186, P<0.05), the difference was statistically significant. 50 mg /L itraconazole could effectively induce the expression of bak1, Bax and cleaved caspase-3 in Mia PaCa-2 cells, the difference was statistically significant (3.406%, 10.712% and 22.626%, t=40.835, P<0.05), the difference was statistically significant (5.041%, 16.135% and 23.701%, t=36.761, P<0.05). After the inhibition of bak1 expression by bak1 siRNA, the inhibition of proliferation and apoptosis induced by itraconazole decreased significantly. Conclusion Itraconazole exerts anti pancreatic cancer effect by regulating the activation of bak1 signaling pathway. Key words: Itraconazole; Pancreatic cancer; Proliferation; Apoptosis; B cell lymphoma/leukemia-2-antagonist/killer 1