Synergistic effect of simvastatin plus NS398 on inhibition of proliferation and survival in hepatocellular carcinoma cell line

Background and Aims: NS398, a selective cyclooxygenase-2 inhibitor, and simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor, both exert an anticancer effect on hepatocellular carcinoma cells, but the effect of co-administration of the two drugs remains unknown. We aimed to investigate the synergistic in vitro anticancer effect of co-administration of NS398 and simvastatin and its mechanism. Methods: The Hep3B and Huh-7 cell lines were cultured. Cells were treated with simvastatin, NS398, or a combination. 5-bromo-2'-deoxyuridine ELISA assay, flow cytometry, Western blot analyses, and immunofluorescence assay were performed. Results: In both cell lines, co-administration of simvastatin and NS398 resulted in a greater effect on proliferation and apoptosis. In Hep3B cells, co-administration of the two drugs resulted in a greater decrease in procaspase 3 and Bcl-2 and an increase in cleaved caspase 9 than that noted with monotherapy. In Huh-7 cells, co-administration of the two drugs resulted in a greater decrease in procaspase 3 and cyclin D1 and an increase in cleaved caspase 9. Expression of NF-κB and Akt were also decreased to a greater extent when the two drugs were co-administered in both cell lines. Immunofluorescence assay showed suppression of the nuclear localization of NF-κB by simvastatin or NS398. The effect was greater by co-administration. Conclusions: The co-administration of NS398 and simvastatin produced greater antiproliferative and proapoptotic effects against Hep3B cells and Huh-7 cells. Inhibition of the NF-κB and Akt pathway and activation of caspase cascade, which are considered as the major mechanism of synergistic anticancer properties, were observed in both cell lines.