20-Hydroxyvitamin D2is a noncalcemic analog of vitamin D with potent antiproliferative and prodifferentiation activities in normal and malignant cells

Abstract

20-hydroxyvitamin D2[20(OH)D2] inhibits DNA synthesis in epidermal keratinocytes, melanocytes, and melanoma cells in a dose- and time-dependent manner. This inhibition is dependent on cell type, with keratinocytes and melanoma cells being more sensitive than normal melanocytes. The antiproliferative activity of 20(OH)D2is similar to that of 1,25(OH)2D3and of newly synthesized 1,20(OH)2D2but significantly higher than that of 25(OH)D3. 20(OH)D2also displays tumorostatic effects. In keratinocytes 20(OH)D2inhibits expression of cyclins and stimulates involucrin expression. It also stimulates CYP24 expression, however, to a significantly lower degree than that by 1,25(OH)2D3or 25(OH)D3. 20(OH)D2is a poor substrate for CYP27B1 with overall catalytic efficiency being 24- and 41-fold lower than for 25(OH)D3with the mouse and human enzymes, respectively. No conversion of 20(OH)D2to 1,20(OH)2D2was detected in intact HaCaT keratinocytes. 20(OH)D2also demonstrates anti-leukemic activity but with lower potency than 1,25(OH)2D3. The phenotypic effects of 20(OH)D2are mediated through interaction with the vitamin D receptor (VDR) as documented by attenuation of cell proliferation after silencing of VDR, by enhancement of the inhibitory effect through stable overexpression of VDR and by the demonstration that 20(OH)D2induces time-dependent translocation of VDR from the cytoplasm to the nucleus at a comparable rate to that for 1,25(OH)2D3. In vivo tests show that while 1,25(OH)2D3at doses as low as 0.8 μg/kg induces calcium deposits in the kidney and heart, 20(OH)D2is devoid of such activity even at doses as high as 4 μg/kg. Silencing of CY27B1 in human keratinocytes showed that 20(OH)D2does not require its transformation to 1,20(OH)2D2for its biological activity. Thus 20(OH)D2shows cell-type dependent antiproliferative and prodifferentiation activities through activation of VDR, while having no detectable toxic calcemic activity, and is a poor substrate for CYP27B1.