Resistance of HL-60 promyelocytic leukemia cells to induction of differentiation and its reversal by combination treatment

Two sublines of HL-60 cells differing markedly in their ability to undergo differentiation to granulocytes after treatment with retinoic acid (RA), dimethyl sulfoxide (DMSO) and the pyrimidine analog, 5-aza-2′-deoxycytidine (azadCyd) were studied. The sensitive subline (HL-60 S) responded well to 1 μM RA, 1% DMSO and 1 μM azadCyd, showing 89 ± 5%, 46 ± 5% and 29 ± 6% mature nitroblue tetrazolium (NBT)-positive cells, respectively. However, the resistant subline (HL-60 R) showed only modest maturational effects (12 ± 3%, 11 ± 2% and 9 ± 2%, respectively) after treatment with the same agents. Using the HL-60 R as a model for resistance to differentiation induction in the HL-60 cell line, studies were carried out to determine whether the combined used of RA, DMSO and azadCyd could reverse the resistance of these tumor cells to the induction of maturation expressed by the individual agents. When these agents were given in any combination of 2, a minor increase in differentiation induction was detected (13 ± 6% or less NBT-positive cells). However, when all 3 agents were combined (RA + DMSO + azaCyd), resistance was completely reversed (89 ± 7% mature NBT-positive cells). In addition, different degrees of concentration-dependence of each agent in the combination were observed. The RA + DMSO + azadCyd combination caused a maximal accumulation of NBT-positve cells after 72 to 96 hr of incubation. These results show that the lack of competence for induction of differentiation in resistant HL-60 cells can be completely reversed by the above ternary drug combination. However, the mechanism responsible for this synergistic effect must await further elucidation of the molecular mechanisms by which such agents act.