Effect of tamoxifen on ceramide cytotoxicity in colon cancer cells.

Abstract

e14112 Background: Poor prognosis in patients with colorectal cancer necessitates the search for new treatment strategies. Ceramide, because of its role in induction of apoptosis in cancer cells, is a promising strategy. Ceramide can be generated in situ by exposure to chemotherapy and ionizing radiation, or it can be administered in the form of short-chain analogs, including nanoliposomal formulations that can be selectively apoptotic for cancer cells. However, conversion of ceramide to inactive metabolites greatly diminishes apoptotic potential. The aim of our investigation was to determine whether cytotoxicity of C6-ceramide could be increased by tamoxifen, which has been shown to be a potent regulator of ceramide metabolism via it’s interaction with Golgi-localized P-glycoprotein (P-gp). Methods: Human colon cancer cell lines HCT-15, HT-29, and LoVo were used. Cell viability (24-72 hr) was assessed spectrophotometrically (n = 6 ± S.D.). Apoptosis was gauged by caspase activation. Metabolic studies utilized [14C]C6-ceramide. Results: C6-ceramide (5.0 µM) and tamoxifen (10 µM) decreased HCT-15 cell viability to 79 ± 6 and 54 ± 3% of control, respectively; the combination decreased viability to 16 ± 4%. This was accompanied by a synergistic 7-fold increase in caspase 3/7 activation. HT-29 cells responded to combination C6-ceramide/tamoxifen in a similar fashion. In LoVo cells, C6-ceramide, tamoxifen, and the combination decreased viability to 84 ± 4, 87 ± 6, and 17 ± 1% of control, respectively, and the combination was synergistic for caspase-3 cleavage. Nanoliposomal formulations of C6-ceramide, tamoxifen, and composites were also effective, yielding viabilities of 76 ± 4, 96 ± 4, and 36 ± 3% of ghost control, respectively. LoVo cells converted C6-ceramide chiefly to C6-glucosylceramide and C6-sphingomyelin; tamoxifen blocked synthesis of the former. Cyclosporin A, another P-gp antagonist, also enhanced C6-ceramide cytotoxicity. Conclusions: Tamoxifen is a favorable partnering agent for enhancing C6-ceramide cytotoxicity in colon cancer cells. The high-frequency expression of P-gp in colorectal cancer presents an adventitious target for augmenting ceramide-based therapies, a strategy that could hold promise for treatment.