SYNERGISTIC INDUCTION OF HL60 CELL-DIFFERENTIATION BY KETOCONAZOLE AND 1-DEOXY ANALOGS OF VITAMIN-D-3

Background: The goal of differentiation therapy is to induce cancer ce lls to stop proliferating and to express characteristics of normal cel ls. Vitamin D analogues, such as the deltanoids, are being evaluated a s differentiation agents in the treatment of several human cancers (e. g., myeloid leukemias); however, these compounds have a tendency to pr oduce hypercalcemia in patients receiving therapy. A combination of a differentiation-inducing deltanoid with a compound that blocks entry o f calcium into cells (e.g., ketoconazole) may offer a new approach to differentiation therapy and address the problem of hypercalcemia. We i nvestigated whether various ketoconazole-deltanoid combinations would alter cellular differentiation or intracellular calcium homeostasis in comparison with deltanoids used alone. Methods: Cultured human leukem ia HL60 cells were treated with ketoconazole-deltanoid combinations. M arkers of differentiation (expression of CD11b and CD14 antigens and o f non-specific esterase) were measured by flow cytometry and cytochemi stry; cell cycle distribution was measured by flow cytometry of propid ium iodide-stained cells. Expression of differentiation-related genes was assessed by northern blotting and immunoblotting, and changes in i ntracellular calcium homeostasis were monitored by fluorescence analys is of fura-2-containing cells. Results: Ketoconazole strongly potentia ted the differentiating activity of the deltanoids, which exhibited lo w potency when used alone, Ketoconazole-deltanoid combinations had lit tle effect on HL60 cell-cycle distribution, although the cells did sto p proliferating and they differentiated, Ketoconazole-deltanoid combin ations produced only minor changes in intracellular calcium homeostasi s compared with changes produced by 1,25-dihydroxyvitamin D-3, either alone or in combination with ketoconazole. Conclusion: These results s uggest that ketoconazole may be useful in combination with vitamin D a nalogues in the differentiation therapy for myeloid leukemias.