Sphingolipids, which include ceramides and sphingosine, are essential structural components of cell membranes that also have messenger functions that regulate the proliferation, survival, and death of cells. Exogenous application of ceramide is cytotoxic, and exposure of cells to radiation or chemotherapy is associated with increased ceramide levels due to enhanced de novo synthesis, catabolism of sphingomyelin, or both. Ceramide can be metabolized to less toxic forms by glycosylation, acylation, or by catabolism to sphingosine, which is then phosphorylated to the anti-apoptotic sphingosine 1-phosphate. Glucosylceramide synthase overexpression has been shown to enhance resistance to doxorubicin, suggesting that inhibition of ceramide metabolism or catabolism might enhance cancer chemotherapy. Several anticancer agents, including the cytotoxic retinoid, fenretinide (4-HPR), have been shown to act, at least in part, by increasing tumor cell ceramide via de novo synthesis. Combinations of 4-HPR and modulators of ceramide action and/or metabolism demonstrated increased anti-tumor activity in pre-clinical models with minimal toxicity for non-malignant cells, and were effective in a p53-independent manner against tumor cell lines resistant to standard cytotoxic agents. Phase I trials of ceramide metabolism inhibitors in combination with 4-HPR and with other cytotoxic agents are in development. Thus, pharmacological manipulation of sphingolipid metabolism to enhance tumor cell ceramide is being realized and offers a novel approach to cancer chemotherapy.