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.