Boswellic acid sensitizes gastric cancer cells to Cisplatin-induced apoptosis via p53-mediated pathway

Abstract

Background: Cisplatin (CDDP) is an effective anticancer drug for Gastric cancer (GC) that induces apoptosis by altering pro- (p53) and anti-apoptotic (Akt and NFkB) proteins; however, chemoresistance remains a big challenge. Additional compounds with promising anticancer effects such as AKBA (Acetyl-keto-beta boswellic acid) may overcome the resistance. However, its role in CDDP-induced apoptosis in GC has not been studied. This study aimed to examine the effectiveness of AKBA on p53-mediated, CDDP-induced apoptosis in GC cells. AGS and NCI-N87 cells were treated with different concentrations (0, 25, 50, 100 μM) of CDDP and/or AKBA.

Methods: P53, Akt and NFkB proteins and apoptosis were assessed by Western blot and flow cytometry. The role of p53 was determined by inhibiting its function via the siRNA approach.

Results: The results revealed that CDDP and AKBA significantly increased p53 content in both cells, while Akt and NFkB were significantly decreased. Both compounds significantly induced apoptosis in a dose-dependent manner. AKBA sensitized GC cells to CDDP-induced apoptosis by altering the protein expression. P53 downregulation affected Akt and NFkB proteins with a slight increase in apoptosis induction in the combination treated groups.

Conclusions: Altogether, our findings suggest that AKBA enhances GC cell sensitivity to CDDP-induced apoptosis via the p53 pathway.

Keywords: AKBA; Apoptosis and gastric cancer; CDDP; P53.

Fig. 1

CDDP had differential effect on proteins expression and induced apoptosis in GC cells. a Western blot shows the effect of CDDP on protein expression in AGS and NCI - N87 cell lines. The expression of tumor suppressor protein 53 increased with CDDP concentration, whilst Akt and NFkB expressions reduced, notice the cleaved PARP which indicates apoptosis b Graphical presentation of apoptosis percentages in AGS and NCI-N87 cell lines treated with different concentrations of CDDP showing significant increase in apoptosis in comparison to control concentration (*P < 0.05, **P < 0.01, ***P < 0.001). The response of AGS cells is more evident than NCI-N87. c Flow cytometry graphs showing the effect of different doses of CDDP on AGS and NCI-N87 cells, each graph shows alive cells, early and late apoptotic cells, and dead cells

Fig. 2

AKBA exhibited similar effect as CDDP on proteins expression and apoptosis in GC cells except for p53. a Western blot shows the effect of AKBA on protein expression in AGS and NCI - N87 cell lines. The expression followed a similar pattern to the cells treated with CDDP b Graphical presentation of apoptosis percentage in AGS and NCI-N87 cell lines treated with different concentrations of AKBA showing significant increase in apoptosis in comparison to control concentration (*P < 0.05, **P < 0.01, ***P < 0.001). The concentration of 100 μM was found to be highly toxic and the apoptosis percentages reached 71.9 and 85.4% in AGS and NCI-N87 cell lines respectively. c Flow cytometry graphs showing the effect of different doses of AKBA on AGS and NCI-N87 cells. Each graph shows alive cells, early and late apoptotic cells, and dead cells

Fig. 3

AKBA sensitized GC cells to CDDP-induced apoptosis via altering proteins expression. a Western blot shows the effect of combined CDDP and AKBA on protein expression in AGS and NCI - N87 cell lines. b Graphical presentation of apoptosis percentage in AGS and NCI-N87 cell lines treated with different concentrations of CDDP and AKBA showing significant increase in apoptosis in comparison to control concentration (*P < 0.05, **P < 0.01, ***P < 0.001)

Fig. 4

P53-dependent pathway mediated CDDP- and AKBA-induced apoptosis in GC a Western blot shows the effect of siRNA, CDDP and AKBA on protein expression in NCI - N87 cell line. P53 siRNA was successful in reducing the increase in p53 expression post treatment of CDDP and AKBA up to approximately 70%. The expression of Akt and NFkB proteins was further reduced in the combined treatment when p53 protein was down-regulated b Graphical presentation of apoptosis percentage in NCI-N87 cell line treated with different treatments showing varying degrees in apoptosis in comparison to control concentration. Apoptosis induction showed a slight trend of increase with no significant difference between treated groups with and without p53 down regulation. (*P < 0.05, **P < 0.01, ***P < 0.001). (c) Flow cytometry graphs showing the effect of different treatments on NCI-N87 cells. Each graph shows alive cells, early and late apoptotic cells, and dead cells