We recently identified 1alpha,25-dihydroxy 3 epi-vitamin D3 (1alpha,25(OH)2-3-epi-D3) as a major metabolite of 1alpha,25- dihydroxy vitamin D3 (1alpha,25(OH)2D3) in the primary cultures of human keratinocytes. It is noted that the production of 1alpha,25(OH)2-3-epi-D3 is tissue specific. Also, 1alpha,25(OH)2-3-epi-D3 when compared to 1alph,25(OH)2D3 was found to be less active in generating some of the classical 1alpha,25(OH)2D3 mediated genomic biological responses. These biological activity findings indicated that the metabolism of 1alpha,25(OH)2D3 into 1alpha,25(OH)2-3-epi-D3 in target tissues leads to significant reduction in the activity of 1alpha,25(OH)2D3. Furthermore, we also identified 1alpha,25(OH)2-3-epi-D3 as a circulating metabolite in rats treated with pharmacological amounts of either 1alpha(OH)D3 or 1alpha,25(OH)2D3. Thus, this newly discovered metabolic pathway of 1alpha,25(OH)2D3 like the previously well established C-24 oxidation pathway, appears to play a significant role in the target tissue inactivation of 1alpha,25(OH)2D3. In the present grant proposal we coined the term """"""""3beta-hydroxy epimerase"""""""", for the enzyme(s) responsible for epimerization of 3beta-hydroxyl of 1alha,25(OH)2D3. The following studies are proposed to elucidate the overall functional significance of our novel observation of the tissue specific metabolism of 1alpha,25(OH)2D3 into 1alpha,25(OH)2-3-epi-D3 in the vitamin D-endocrine system. 1) further investigations towards establishing the target tissue specificity for the production of 1alpha,25(OH)2-3-epi-D3 in normal tissues from human, rat and chick species; 2) subcellular localization and enzyme kinetic studies to characterize the enzyme(s) responsible for the production of 1alpha,25(OH)2-3- epi-D3; 3) studies of further target tissue metabolism of 1alpha,25(OH)2-3-epi-D3; 4) comparative pharmacokinetic and biological activity studies between 1alpha,25(OH)2D3 and the newly discovered metabolite 1alpha,25(O2H)-3-epi-3-D. 5) studies of 1alpha,25(OH)2-3-epi-D3 production in mice that lack 24-hydroxylase to identify the 3beta-hydroxy epimerization as the possible alternate pathway for the inactivation of 1alpha,25(OH)2D3. It is anticipated that knowledge of target tissue specific metabolism of 1alpha,25(OH)2D3 will provide important information for our improved understanding of how 1alpha,25(OH)2D3 is inactivated in various different target tissue.
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