Caspase-mediated pro-apoptotic interaction of panaxadiol and irinotecan in human colorectal cancer cells

Abstract

Objectives: Panaxadiol is a purified sapogenin of ginseng saponins that exhibits anticancer activity. Irinotecan is a second-line anticancer drug, but clinical treatment with irinotecan is limited due to its side effects. In this study, we have investigated the possible synergistic anticancer effects of panaxadiol and irinotecan on human colorectal cancer cells and explored the potential role of apoptosis in their synergistic activity.

Key findings: The combination of panaxadiol and irinotecan significantly enhanced antiproliferative effects in HCT-116 cells (P< 0.05). Cell cycle analysis demonstrated that combining irinotecan treatment with panaxadiol significantly increased the G1-phase fractions of cells, compared with irinotecan treatment alone. In apoptotic assays, the combination of panaxadiol and irinotecan significantly increased the percentage of apoptotic cells compared with irinotecan alone (P<0.01). Increased activity of caspase-3 and caspase-9 was observed after treating with panaxadiol and irinotecan. The synergistic apoptotic effects were supported by docking analysis, which demonstrated that panaxadiol and irinotecan bound two different chains of the caspase-3 protein.

Conclusions: Data from this study suggested that caspase-3- and caspase-9-mediated apoptosis may play an important role in the panaxadiol enhanced antiproliferative effects of irinotecan on human colorectal cancer cells.

Fig. 1

Chemical structures of panaxadiol (PD) and irinotecan (IRN).

Fig. 2

Antiproliferative effects of irinotecan and panaxadiol on human colorectal cancer cells. HCT-116 (a) and SW-480 (b) cells were treated with panaxadiol (10 μM) and/or irinotecan (1, 3, 5, 10 or 20 μM) for 48 hours. Data are presented as mean ± SE. *P < 0.05, **P < 0.01, compared to corresponding groups of IRN only.

Fig. 3

Effects of irinotecan and panaxadiol on HCT-116 cell cycle. HCT-116 cells were treated with panaxadiol (10 μM) and/or irinotecan (1, 3, 5, 10 or 20 μM) for 48 h. Cell cycle profile was determined using flow cytometry after staining with PI/RNase. (a) Representative histograms of cell cycle distribution. (b) Percentage of each cell cycle phase with various treatments or with control. Data are presented as the mean ± SE of triplicate experiments.

Fig. 4

Effects of irinotecan and panaxadiol on HCT-116 cell apoptosis. HCT-116 cells were treated with panaxadiol (10 μM) and/or IRN (1, 3, 5, 10 or 20 μM) for 48 h. Apoptosis was quantified using flow cytometry after staining with annexin V/PI. (a) Representative scatter plots of PI (y-axis) vs annexin V (x-axis). (b) Percentage of viable, early apoptotic and late apoptotic cells. Data are presented as the mean ± SE of triplicate experiments.

Fig. 5

Effect of PD and IRN on Caspase 3/9 activity in HCT-116 cells. HCT-116 cells were treated with panaxadiol (PD; 10 μM) and/or irinotecan (IRN; 10 μM) for 6, 10 or 24 h. Caspase activities were measured by caspase 3, 9/CPP 32 Colorimetric Assay kits. Data are presented as mean ± SE of mean. *P < 0.05, **P < 0.01, versus corresponding control groups.

Fig. 6

Docking simulation of irinotecan and panaxadiol on the caspase-3 protein. Panaxadiol (PD) and irinotecan (IRN) bind to distinct binding sites on the protein. A surface and ribbon view is shown above (a), while an insight ligand-site model is shown below (b).