Oxidation mechanism of 7-hydroxy-δ ⁸-tetrahydrocannabinol and 8-hydroxy-δ9-tetrahydrocannabinol to the corresponding ketones by CYP3A11

A cDNA isolated from a C57BL/6 mouse liver cDNA library had the identical nucleotide sequence in coding region with the mouse CYP3A11, and the NH ₂-terminal sequence was also identical to that of cytochrome P450 (P450) MDX-B, a microsomal alcohol oxygenase. The COS-7 cells transfected with the CYP3A11 expression vector formed 7-oxo-Δ ⁸-tetrahydrocannabinol (7-oxo-Δ ⁸-THC) from 7α- and 7β-hydro-Δ ⁸-THC. An immunologically related protein with P450 MDX-B was expressed in the COS-7 cell microsomes. The cell microsomes expressed CYP3A11; COS-3A11 catalyzed the oxidation of 7-hydroxy-Δ ⁸-THC and 8-hydroxy-Δ ⁹-THC to 7-oxo-Δ ⁸-THC and 8-oxo-Δ ⁹-THC, respectively, in a reconstituted system. ¹⁸O derived from atmospheric oxygen was incorporated into about 30% of the corresponding ketones formed from 7α-hydroxy-Δ ⁸-THC and 8β-hydroxy-Δ ⁹-THC by mouse hepatic microsomes, P450 MDX-B, and COS-3A11, although incorporation of the stable isotope into the oxidized metabolites from 7β-hydroxy-Δ ⁸-THC and 8α-hydroxy-Δ ⁹ -THC was negligible. ¹⁸O, however, was not incorporated into 7-oxo-Δ ⁸ -THC formed from 7α-hydroxy-Δ ⁸THC by using cumene hydroperoxide instead of NADPH under ¹⁸O ₂. When ¹⁸O-labeled 7α-hydroxy-Δ ⁸ -THC and 8β-hydroxy-Δ ⁹-THC were incubated with above enzymes under air, about 30% of the ketones formed released ¹⁸O from a hydroxy group at the 7 and 8 positions in the course of the oxidation. These results suggest that 7α-hydroxy-Δ ⁸ -THC and 8β-hydroxy-Δ ⁹-THC may be oxidized to the corresponding ketones by CYP3A11 via a gem-diol pathway. 7β-Hydroxy-Δ ⁸-THC and 8α-hydroxy-Δ ⁹-THC may be also converted to the ketones through a stereoselective dehydration of an enzyme-bound gem-diol rather than through a direct hydrogen extraction as a peroxy form of the enzyme.