Fig. 7
Inhibiting CDK1-STAT3 signaling by fisetin enhanced effect of gemcitabine through suppressing cancer stemness. a, b Real-time proliferation analysis of human PANC-1 cells and mice KPC203 cells. Cells were treated with DMSO, fisetin (100 µM), gemcitabine (10 µM) or combination therapy of fisetin (100 µM) and gemcitabine (10 µM) for three days. Cell growth was monitored using the Incucyte SX5 system, as area of cells was recorded every hour. The times on the x- axis indicated the times after treatment. The y- axis reflected phase area of cells per well normalized to 0 h. (n = 6, mean ± SD). Fis, fisetin, Gem, gemcitabine. c, d Sphere formation assay of human PANC-1 cells and mice KPC203 cells with DMSO, fisetin alone, gemcitabine alone or combination therapy. Cells were pretreated with fisetin, gemcitabine or combination therapy for 48 h, then 1 × 103 cells were seed in ultra low ultra-low cluster plates to perform sphere forming analysis. Scale bars 100 μm. Data are presented as mean ± SD (n = 3). *P < 0.05, ***P < 0.001. e Representative flow cytometry plots for CD44 and CD24 expression in human pancreatic cancer PANC-1 cells with DMSO, fisetin alone, gemcitabine alone or combination therapy treatment as above. Proportion of CD44+/CD24+ positive cells were increased by gemcitabine, which can be attenuated by fisetin in combination group. f Protein levels of HDAC3, p-STAT3, STAT3, p-CDK1, CDK1, CD44 and Sox2 in PANC-1 cells treated with the indicated treatment, was detected by western blot. g Photographs and h weights of allograft tumors of KPC (KrasLSL−G12D p53LSL−R172H Pdx-1-Cre) mice at day 21 after first treatment. Data are presented as mean ± SD (n = 4); **P < 0.01, ***P < 0.001