Vol. 29, Issue 11, 1485-1491, November 2001

Oxidation Mechanism of 7-Hydroxy-⁸-tetrahydrocannabinol and 8-Hydroxy-⁹-tetrahydrocannabinol to the Corresponding Ketones by CYP3A11

Tamihide Matsunaga, Hiroyuki Tanaka, Shinsuke Higuchi, Kinya Shibayama, Nobuyuki Kishi, Kazuhito Watanabe, and Ikuo Yamamoto

Department of Hygienic Chemistry, Faculty of Pharmaceutical Sciences, Hokuriku University

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 NH₂-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-⁸-tetrahydrocannabinol (7-oxo-⁸-THC) from 7- and 7-hydroxy-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-⁸-THC and 8-hydroxy-⁹-THC to 7-oxo-⁸-THC and 8-oxo-⁹-THC, respectively, in a reconstituted system. ¹⁸O derived from atmospheric oxygen was incorporated into about 30% of the corresponding ketones formed from 7-hydroxy-⁸-THC and 8-hydroxy-⁹-THC by mouse hepatic microsomes, P450 MDX-B, and COS-3A11, although incorporation of the stable isotope into the oxidized metabolites from 7-hydroxy-⁸-THC and 8-hydroxy-⁹-THC was negligible. ¹⁸O, however, was not incorporated into 7-oxo-⁸-THC formed from 7-hydroxy-⁸-THC by using cumene hydroperoxide instead of NADPH under ¹⁸O₂. When ¹⁸O-labeled 7-hydroxy-⁸-THC and 8-hydroxy-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 7-hydroxy-⁸-THC and 8-hydroxy-⁹-THC may be oxidized to the corresponding ketones by CYP3A11 via a gem-diol pathway. 7-Hydroxy-⁸-THC and 8-hydroxy-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.