To develop efficient synergistic or additive combinations of chemopreventive and nutritional agents to reduce the risk of colon cancer, experiments were designed to test the application of a selective cyclooxygenase-2 (COX-2) inhibitor together with dietary omega-3 polyunsaturated fatty acids (PUFAs), such as decosahexaenoic acid (DHA). Thus, individual application of celecoxib, a COX-2 inhibitor, DHA, a omega-3 PUFA, and combinations of both were tested for their effectiveness using cell proliferation, apoptosis, and COX-2 expression as markers in the human colon cancer HCA-7 cell line. HCA-7 cells exposed to various subtoxic doses of celecoxib, DHA, or combinations of both were analyzed for inhibition of cell proliferation by trypan blue exclusion and proliferating cell nuclear antigen methods, induction of apoptosis by 4',6-diamidino-2-phenylindole method, and COX-2 by reverse transcription-PCR and Western blot analysis. In addition, we examined the inhibitory potential of celecoxib and DHA on (14)C-arachidonic acid metabolism mediated by COX-2 in the HCA-7 cell line. We found that treatment with celecoxib (50-150 micro M) or DHA (150-225 micro M) individually induces apoptosis and inhibits cell proliferation only at high concentrations in HCA-7 cell lines. A synergistic effect was observed on induction of apoptosis and inhibition of proliferation when cells were exposed to low doses of celecoxib (50-100 micro M) together with DHA (75 micro M). At high concentrations, celecoxib and DHA blocked the increase in COX-2 protein and mRNA expression in HCA-7 cells. Importantly, the inhibition of COX-2 expression was more pronounced in cells treated with low-dose combinations than with individual agents at high concentrations. In addition, celecoxib and DHA at low-dose levels inhibited (14)C-arachidonic acid metabolism (50-85%, P < 0.0001) leading to very low levels of type 2 series prostaglandin formation. These findings provide the basis for the development of combinations of low-dose regimens of a COX-2 inhibitor and omega-3 PUFAs such as DHA for the prevention and treatment of colon cancer. We are currently testing this concept in preclinical models.