1 alpha 25-dihydroxy-3-epi-vitamin D-3 a physiological metabolite of 1 alpha,25-dihydroxyvitamin D-3: Its production and metabolism in primary human keratinocytes

Recent studies of metabolism using pharmacological substrate concentrations of 1 alpha ,25-dihydroxyvitamin D-3 [1 alpha ,25(OH)(2)D-3] in several tis sues including primary cultures of human keratinocytes, bovine parathyroid cells and bone cells led to the identification of 1 alpha ,25-dihydroxy-3-e pi-vitamin D-3 [1 alpha ,25(OH)(2)-3-epi-D-3] as a major natural metabolite of 1 alpha ,25(OH)(2)D-3. In the present study, we demonstrate that human keratinocytes incubated with 25-hydroxy[26,27-H-3] vitamin D-3 produce 1 al pha ,25(OH)(2)-3-epi-D-3 along with 1 alpha ,25(OH)(2)D-3. The production o f 1 alpha ,25(OH)(2)-3-epi-D-3 is also identified in human keratinocytes in cubated with physiological substrate concentrations of 1 alpha ,25(OH)(2)D- 3. Unlike 24-hydroxylase, the major enzyme involved in the further metaboli sm of 1 alpha ,25(OH)(2)D-3 in human keratinocytes, the enzyme(s) responsib le for the production of 1 alpha ,25(OH)(2)-3-epi-D-3 is constitutive and i s not inhibited by ketoconazole. It is also noted that 1 alpha ,25(OH)(2)-3 -epi-D-3 is further metabolised in human keratinocytes into several as yet unidentified metabolites, the production of which is inhibited to a great e xtent by SDZ 89-443, an inhibitor of 24-hydroxylase. This finding indicates that the 24-hydroxylase like in the case of 1 alpha ,25(OH)(2)D-3, also pl ays a major role in the metabolism of 1 alpha ,25(OH)(2)-3-epi-D-3. The res ults obtained from the metabolism studies performed in parallel among 25OHD (3), 1 alpha ,25(OH)(2)D-3 and 1 alpha ,25(OH)(2)-3-epi-D-3 indicate that 1 alpha ,25(OH)(2)-3-epi-D-3 and its metabolites exhibit higher metabolic st ability. In summary, we demonstrate for the first time that 1 alpha ,25(OH) (2)-3-epi-D-3 is a physiological metabolite of 1 alpha ,25(OH)(2)D-3 in hum an keratinocytes. Also, 1 alpha ,25(OH)(2)-3-epi-D-3 is further metabolised in human keratinocytes mainly through the activity of 24-hydroxylase. Furt hermore, our finding of the relative metabolic stability of 1 alpha ,25(OH) (2)-3-epi-D-3 and especially its metabolites when compared to 1 alpha ,25(O H)(2)D-3 and its metabolites provides an important explanation for its prev iously observed potent inhibitory effect on keratinocyte growth in spite of its low affinity to vitamin D receptor. (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.