title: Hydroxylation of CYP11A1-derived products of vitamin D3 metabolism by human and mouse CYP27B1.
abstract: CYP11A1 can hydroxylate vitamin D3 at carbons 17, 20, 22, and 23, producing a range of secosteroids which are biologically active with respect to their ability to inhibit proliferation and stimulate differentiation of various cell types, including cancer cells. As 1α-hydroxylation of the primary metabolite of CYP11A1 action, 20S-hydroxyvitamin D3 [20(OH)D3], greatly influences its properties, we examined the ability of both human and mouse CYP27B1 to 1α-hydroxylate six secosteroids generated by CYP11A1. Based on their kcat/Km values, all CYP11A1-derived metabolites are poor substrates for CYP27B1 from both species compared with 25-hydroxyvitamin D3. No hydroxylation of metabolites with a 17α-hydroxyl group was observed. 17α,20-Dihydroxyvitamin D3 acted as an inhibitor on human CYP27B1 but not the mouse enzyme. We also tested CYP27B1 activity on 20,24-, 20,25-, and 20,26-dihydroxyvitamin D3, which are products of CYP24A1 or CYP27A1 activity on 20(OH)D3. All three compounds were metabolized with higher catalytic efficiency (kcat/Km) by both mouse and human CYP27B1 than 25-hydroxyvitamin D3. CYP27B1 action on these new dihydroxy derivatives was confirmed to be 1α-hydroxylation by mass spectrometry and nuclear magnetic resonance analyses. Both 1,20,25- and 1,20,26- trihydroxyvitamin D3 were tested for their ability to inhibit melanoma (SKMEL-188) colony formation, and were significantly more active than 20(OH)D3. This study shows that CYP11A1-derived secosteroids are 1α-hydroxylated by both human and mouse CYP27B1 with low catalytic efficiency, and that the presence of a 17α-hydroxyl group completely blocks 1α-hydroxylation. In contrast, the secondary metabolites produced by subsequent hydroxylation of 20(OH)D3 at C24, C25, or C26 are very good substrates for CYP27B1.
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Edith K Y
authors: EK
authors: Tang
authors: Jianjun
authors: J
authors: Chen
authors: Zorica
authors: Z
authors: Janjetovic
authors: Elaine W
authors: EW
authors: Tieu
authors: Andrzej T
authors: AT
authors: Slominski
authors: Wei
authors: W
authors: Li
authors: Robert C
authors: RC
authors: Tuckey
mesh_terms: D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D002762:Cholecalciferol / Q000031:analogs & derivatives / Q000378:metabolism*; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006900:Hydroxylation; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D051379:Mice
chemical_list: D004791:Enzyme Inhibitors; D002762:Cholecalciferol; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase; D002786:Cholesterol Side-Chain Cleavage Enzyme