SYNERGISTIC EFFECTS OF CEPHARANTHINE AND CELECOXIB IN HUMAN COLORECTAL CANCER CELLS

Abstract

Celecoxib, a specific cyclooxygenase 2 (COX-2) inhibitor, is clinically suggested as adjunctive therapy for preventing the progression of colon polyps to colorectal cancer. However, this drug is used at high dose in a long period of time for this indication. This leads to the cardiovascular toxicity concern. Combination of celecoxib with other anticancer agents is a strategy for improving the efficacy of celecoxib. This study aimed to investigate anticancer effect of celecoxib – cepharanthine combination on human colorectal cancer HT-29 cells. Effects of the drugs on cell viability were determined by resazurin assay. Celecoxib and cepharanthine decreased viability of HT-29 cells with IC50 > 50 µM and at 5.22 ± 0.28 µM, respectively after 48 h of exposure. Celecoxib at 5, 10, 20, and 40 µM were combined with cepharanthine at 1.25, 2.5 and 5 µM for HT-29 viability test. All combinations had synergistic effects on the cell viability with combination indices less than one (CI < 1). The combinations of celecoxib at 20 and 40 µM and cepharanthine at 1.25 and 2.5 µM were investigated further. When compared to each drug, these combinations did not have additive/ synergistic effects on COX-2 and NADPH oxidases (NOX1 and NOX2) expression determined by real time RT- PCR, PGE2 production determined by ELISA, and ROS generation determined by DCFH-DA assay. The combinations significantly increased HT-29 cell accumulation at the G1 phase of the cell cycle when compared to the effects of each drug. The cell cycle was determined by propidium iodide (PI) staining with fluorescence flow cytometer. The effects on cell cycle arrest of these combinations may come from the increase in cyclin-dependent kinase (CDK) inhibitor p21 and the decreased in cyclin A expression. The combination of 40 µM celecoxib – 2.5 µM cepharanthine significantly increased p21 but decreased cyclin A expression when compared to each drug. The combinations significantly increased HT-29 apoptosis determined by annexin V/FITC and PI staining with flow cytometer when compared to each drug. Similarly, their apoptotic induction effects correlated with the increase in pro-apoptotic BAX and the decrease in anti-apoptotic Bcl-XL expression induced by these combinations. The results from this study reveal the synergistic anticancer effect of celecoxib – cepharanthine combinations at sub-IC50 concentrations. This finding may be useful for reducing the dose and toxicities of celecoxib in colorectal cancer prevention. Further investigations are needed to confirm these results.