CYP27A1 acts on the pre-vitamin D3 photoproduct, biologically active hydroxy-metabolites lumisterol, producing biologically active hydroxy-metabolites

Prolonged exposure of the skin to UV radiation causes previtamin D3, the initial photoproduct formed by opening of the B ring of 7-dehydrocholesterol, to undergo a second photochemical reaction where the B-ring is reformed giving lumisterol3 (L3), a stereoisomer of 7-dehydrocholesterol. L3 was believed to be an inactive photoproduct of excessive UV radiation whose formation prevents excessive vitamin D production. Recently, we reported that L3 is present in serum and that CYP11A1 can act on L3 producing monohydroxy- and dihydroxy-metabolites which inhibit skin cell proliferation similarly to la,25-dihydroxyvitamin D3. In this study we tested the ability of human CYP27A1 to hydroxylate L3. L3 was metabolized by purified CYP27A1 to 3 major products identified as 25-hydroxyL3, (25R)-27-hydroxyL3 and (25S)-27-hydroxyL3, by NMR. These three products were also seen when mouse liver mitochondria containing CYP27A1 were incubated with L3. The requirement for CYP27A1 for their formation by mitochondria was confirmed by the inhibition of their synthesis by 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol, an intermediate in bile acid synthesis which serves as an efficient competitive substrate for CYP27A1. CYP27A1 displayed a high k(eat) for the metabolism of L3 (76 mol product/min/mol CYP27A1) and a catalytic efficiency (K-cat/K-m) that was 260-fold higher than that for vitamin D3. The CYP27A1-derived hydroxy-derivatives inhibited the proliferation of cultured human melanoma cells and colony formation with IC50 values in the nM range. Thus, L3 is metabolized efficiently by CYP27A1 with hydroxylation at C25 or C27 producing metabolites potent in their ability to inhibit melanoma cell proliferation, supporting that L3 is a prohormone which can be activated by CYP-dependent hydroxylations.