Our previous studies demonstrated that 12-O-tetradecanoylphorbol-13-acetate (TPA) had pharmacological activity for the treatment of myeloid leukemia patients. In the present study, we investigated the effects of TPA alone or in combination with capsaicin (8-methyl-N-vanillyl-6-nonenamide) on growth and differentiation in myeloid leukemia HL-60 cells and in a TPA-resistant HL-60 variant cell line termed HL-525. Treatment of HL-60 cells with TPA (0.16-1.6 nM) for 48 h resulted in concentration-dependent growth inhibition and cell differentiation (via the macrophage pathway). Capsaicin (5-50 microM) inhibited the growth of HL-60 cells in a concentration-dependent manner. Treatment of HL-60 cells with capsaicin alone only resulted in a small increase in the number of differentiated cells but treatment of the cells with TPA in combination with capsaicin synergistically increased differentiation. Moreover, inhibitors of protein kinase C (PKC), 7-hydroxystaurosporin (UCN-01; 100 nM) and chelerythrine (0.5 microM), significantly decreased HL-60 cell differentiation induced by the combination of TPA and capsaicin. These results suggest that PKC may be involved in HL-60 cell differentiation induced by TPA in combination with capsaicin. Capsaicin alone caused a very small increase in differentiation in the TPA-resistant HL-525 cells. However, treatment of HL-525 cells with combinations of TPA (0.16 nM) and capsaicin (10-50 microM) caused a strong synergistic increase in differentiation. Results from the present study suggest that a combination of TPA and capsaicin may improve the therapeutic efficacy of TPA and overcome resistance to TPA in some myeloid leukemia patients.