RF15 | PMON293 Metabolic activation of tachysterol to biologically active hydroxyderivatives that act on VDR, AhR, LXRs and PPARγ receptors

Absorption of ultraviolet B (UVB) radiation by the B ring of 7-dehydrocholesterol (7-DHC) leads to the transformation of 7-DHC to previtamin D(3), which after absorption of additional UVB isomerizes to tachysterol(3) (T(3)) and lumisterol3 (L(3)). Previously we demonstrated that CYP11A1 can hydroxylate the side chain of vitamin D(3) (D(3)), 7-DHC and L(3). Similarly CYP27A1 can hydroxylate the side chain of L(3) to biologically active hydroxyderivatives. In a continuation of these studies, we report that CYP11A1 and CYP27A1 hydroxylate T(3) to 20S(OH)T(3) and 25(OH)T(3), respectively, plus minor unidentified hydroxyderivatives. Both 20S(OH)T(3) and 25(OH)T(3)were detected in the human epidermis and serum. T(3) was also detected in human serum and was present at a concentration of 7.3±2.5 ng/ml. 20S(OH)T(3) and 25(OH)T(3) inhibited the proliferation of epidermal keratinocytes and dermal fibroblasts and stimulated the expression of differentiation and anti-oxidative genes in keratinocytes in a similar manner to 1,25-dihydroxyvitamin D3 . They acted on the vitamin D receptor (VDR) as demonstrated by image flow cytometry and the translocation of VDR-coupled GFP (VDR-GFP) from the cytoplasm to the nucleus of melanoma cells, as well as by the stimulation of CYP24A1 expression. Functional studies using a human aryl hydrocarbon receptor (AhR) reporter assay revealed marked activation of AhR by 20S(OH)T(3), a smaller effect by 25(OH)T3 and only minimal activation by T(3). The T(3) hydroxyderivatives showed high affinity binding to the ligand binding domain (LBD) of the liver X receptor (LXR) α and β, and to the peroxisome proliferator-activated receptor γ (PPARg) in LanthaScreen TR-FRET coactivator assays. Molecular docking using crystal structures of the LBDs of VDR, AhR, LXRs and PPARγ revealed high docking scores for 20S(OH)T(3) and 25(OH)T(3), comparable to their previously characterized ligands. The scores for binding to the non-genomic site of the VDR were very low indicating a lack of interaction with hydroxy-T(3) ligands. In conclusion, we have identified 20S(OH)T(3) and 25(OH)T(3) in the human epidermis and serum, identified CYP enzymes responsible for their production, demonstrated phenotypic effects on skin cells, and identified VDR, AhR, LXRs and PPARγ, and possibly RORs, as their genomic receptor targets. Thus CYP11A1, aside from its role in steroidogenesis, not only metabolizes vitamin D(3), 7-DHC and L(3) to biologically active metabolites, but also activates T(3) to biologically active 20S(OH)T(3). Similarly, CYP27A1, previously reported to act on L(3), D(3) and 7DHC, also activates T(3) via hydroxylation at C25. We believe that these novel findings open new areas for future studies on the role of active forms of T(3), not only in the skin, but also in systemic physiology and pathology. Presentation: Sunday, June 12, 2022 12:54 p.m. - 12:59 p.m., Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.