Synergistic effects of clinically achievable concentrations of 12-O-tetradecanoylphorbol-13-acetate in combination with all-trans retinoic acid, 1 alpha,25-dihydroxyvitamin D-3, and sodium butyrate on differentiation in HL-60 cells

Our recent studies demonstrated that 12-O-tetradecanoylphorbol-13-acetate ( TPA) has pharmacological activity for the treatment of acute myelocytic leu kemia patients. In the present study, we investigated the potential synergi stic effect of all-trans retinoic acid (RA), 1 alpha ,25-dihydroxyvitamin D -3 (VD3), and sodium butyrate (NaB) on TPA-induced differentiation in HL-60 human promyelocytic leukemia cells. The cells were treated once with these agents for 48 h or treated every 24 h for 96 h. Treatment of HL-60 cells o nce with TPA, RA, VD3, or NaB for 48 h resulted in concentration-dependent growth inhibition and cell differentiation. At clinically achievable concen trations, TPA (0.16 nM) increased the number of adherent cells and RA (0.1- 1 muM) increased the number of nitroblue tetrazolium (NDT)-positive cells. The combinations of TPA (0.16 nM) with RA (0.1-1 muM), VD3 (1 nM), or NaB ( 100 muM) for 48 h synergistically increased differentiation as measured by the formation of adherent cells (P less than or equal to 0.01). Moreover, c ells treated with various combinations of low concentrations of TPA, RA, VD 3, and NaB every 24 h for 96 h resulted in a further decrease in cell growt h and an increase in differentiation. At clinically achievable concentratio ns, the strongest stimulation of differentiation was achieved in cells trea ted with a "cocktail" that combined TPA, RA, VD3, and NaB. The synergistic effect of combinations of TPA with RA or NaB at clinically effective concen trations on HL-60 cell differentiation suggests that the combination of the se agents may improve the therapeutic efficacy of TPA for the treatment of acute promyelocytic leukemia (APL) patients. A differentiation "cocktail" t hat combines TPA, RA, VD3, and NaB may provide an even more effective strat egy for improving the therapeutic efficacy of TPA and RA.