The synthetic retinoid N-(4-hydroxphenyl) retinamide (4HPR) has manifold actions, which may contribute to its chemopreventive effects on breast cancer cell growth and progression. A role for ceramide as a stress-response signal is investigated here during the cytotoxic action of 4HPR in MCF-7 cells. N-(4-hydroxphenyl) retinamide induced a dose-dependent decline in cell growth and survival associated with a maximal 10-fold increase in ceramide production at 10 microM. N-(4-hydroxphenyl) retinamide exhibited a greater potency than all-trans retinoic acid (ATRA) on growth inhibition and ceramide production. The synthetic peroxisome proliferator-activated receptors agonist troglitazone (TGZ), but not the native ligand 15-deoxy-delta 12,14-prostaglandin J2, abrogated both these actions of 4HPR but not that of ATRA. The antioxidant N-acetylcysteine mimicked the abrogative effect of TGZ on 4HPR action, while the exogenous oxidant H2O2 also stimulated ceramide production. The inhibitors of de novo ceramide synthesis, fumonisin B1 and myriocin, blocked the ceramide response to 4HPR and partially reversed the apoptotic response, but did not prevent the overall decline in cell survival. The pancaspase inhibitor Z-VAD fmk reduced the decrease in cell survival caused by 4HPR, but did not affect the ceramide response. These findings describe a novel redox-sensitive elevation of ceramide levels associated with the cytotoxic response of breast cancer cells to 4HPR. However, a major mediatory role for this sphingolipid in this context remains equivocal.