Ethacrynic acid and 1α,25-dihydroxyvitamin D 3 cooperatively inhibit proliferation and induce differentiation of human myeloid leukemia cells

Abstract

The active form of vitamin D, 1α,25-dihydroxyvitamin D 3 (VD 3), inhibits proliferation and induces differentiation of leukemia cells, but its clinical use is limited by the adverse effect of hypercalcemia. In this study we found that the loop diuretic ethacrynic acid, which is used to treat hypercalcemia, enhanced the differentiation of human leukemia cells induced by VD 3. Ethacrynic acid alone inhibited the proliferation of human promyelocytic HL-60 cells while only slightly increasing differentiation markers such as nitroblue tetrazolium (NBT)-reducing and lysozyme activities. Ethacrynic acid effectively enhanced the growth-inhibiting action of VD 3. In the presence of ethacrynic acid, VD 3 increased the NBT-reducing and lysozyme activities and the CD11b expression of HL-60 cells more effectively than VD 3 alone. Other loop diuretics, furosemide and bumetanide, also enhanced the differentiation of HL-60 cells induced by VD 3, but to a lesser extent than ethacrynic acid. The differentiation of HL-60 cells induced by all- trans retinoic acid, dimethyl sulfoxide or phorbol-12-myristate 13-acetate was also enhanced by ethacrynic acid with increasing NBT-reducing and lysozyme activities and the expression of CD11b or CD14 surface antigen. Morphologically, ethacrynic acid enhanced the monocytic differentiation of HL-60 cells induced by VD 3 and phorbol ester and the granulocytic differentiation by retinoic acid and dimethyl sulfoxide. Other human myelomonocytic leukemia ML-1, U937, P39/TSU and P31/FUJ cells were induced to differentiate by VD 3 and this was also enhanced by ethacrynic acid. The long-term culture of HL-60 cells showed that ethacrynic acid plus VD 3 induced the complete growth arrest of HL-60 cells. Therefore ethacrynic acid, which is used to treat hypercalcemia, enhanced the proliferation-inhibiting and differentiation-inducing activities of VD 3 and the combination of ethacrynic acid and VD 3 may be useful in therapy for myeloid leukemia.