Elsevier

Life Sciences

Volume 80, Issue 15, 20 March 2007, Pages 1415-1419
Life Sciences

Cytochrome P450 enzymes involved in the metabolism of tetrahydrocannabinols and cannabinol by human hepatic microsomes

https://doi.org/10.1016/j.lfs.2006.12.032Get rights and content

Abstract

In this study, tetrahydrocannabinols (THCs) were mainly oxidized at the 11-position and allylic sites at the 7α-position for Δ8-THC and the 8β-position for Δ9-THC by human hepatic microsomes. Cannabinol (CBN) was also mainly metabolized to 11-hydroxy-CBN and 8-hydroxy-CBN by the microsomes. The 11-hydroxylation of three cannabinoids by the microsomes was markedly inhibited by sulfaphenazole, a selective inhibitor of CYP2C enzymes, while the hydroxylations at the 7α-(Δ8-THC), 8β-(Δ9-THC) and 8-positions (CBN) of the corresponding cannabinoids were highly inhibited by ketoconazole, a selective inhibitor of CYP3A enzymes. Human CYP2C9-Arg expressed in the microsomes of human B lymphoblastoid cells efficiently catalyzed the 11-hydroxylation of Δ8-THC (7.60 nmol/min/nmol CYP), Δ9-THC (19.2 nmol/min/nmol CYP) and CBN (6.62 nmol/min/nmol CYP). Human CYP3A4 expressed in the cells catalyzed the 7α-(5.34 nmol/min/nmol CYP) and 7β-hydroxylation (1.39 nmol/min/nmol CYP) of Δ8-THC, the 8β-hydroxylation (6.10 nmol/min/nmol CYP) and 9α,10α-epoxidation (1.71 nmol/min/nmol CYP) of Δ9-THC, and the 8-hydroxylation of CBN (1.45 nmol/min/nmol CYP). These results indicate that CYP2C9 and CYP3A4 are major enzymes involved in the 11-hydroxylation and the 8-(or the 7-) hydroxylation, respectively, of the cannabinoids by human hepatic microsomes. In addition, CYP3A4 is a major enzyme responsible for the 7α- and 7β-hydroxylation of Δ8-THC, and the 9α,10α-epoxidation of Δ9-THC.

Introduction

Tetrahydrocannabinols (THCs) and cannabinol (CBN), which are the major constituents of marijuana, are known to be extensively metabolized by experimental animals and humans (Harvey, 1984). Cytochrome P450 (CYP) is mainly involved in the primary metabolism of the cannabinoids in hepatic microsomes (Yamamoto et al., 1995). Our previous studies demonstrated that the major CYP enzymes involved in the hepatic metabolism of THCs in mice and male rats are CYP2C29 (Watanabe et al., 1993) and CYP2C11 (Narimatsu et al., 1990), respectively. The metabolic reaction of THCs is complicated and over 40 metabolites have been identified in humans in vivo [Agurell et al., 1986]. Many metabolites of CBN have also been identified in humans in vivo (Agurell et al., 1986). However, relatively limited information is available for in vitro metabolism of these cannabinoids in humans (Halldin et al., 1982, Yamamoto et al., 1983), in particular CYP enzymes involved in the metabolism of THCs and CBN are not fully elucidated. Bornheim et al. (1992) suggested in a study with immunoinhibition and purified enzyme that CYP2C9 was mainly responsible for the 11-hydroxylation of Δ9-THC in human hepatic microsomes. Our previous study suggested that a member of the CYP2C subfamily is primarily responsible for the 11-hydroxylation of Δ8-THC, Δ9-THC, and CBN in human hepatic microsomes from an elderly woman, although the enzyme involved in the reaction was not entirely specified (Watanabe et al., 1995). The 11-hydroxylation of THCs and CBN has been shown to be the metabolic activation pathway of both cannabinoids (Watanabe et al., 1980; Yamamoto et al., 1987, Yamamoto et al., 2003). Recently, Bland et al. (2005) reported the kinetic nature and pharmacogenetics of CYP2C9 enzymes for the metabolic interaction of Δ9-THC and phenytoin. The present paper describes the CYP enzymes responsible for the major metabolism of THCs and CBN in human hepatic microsomes.

Section snippets

Chemicals

Δ9-THC and CBN were isolated from cannabis leaves by the methods of Aramaki et al. (1968). Δ8-THC was prepared by acidic isomerization of Δ9-THC as described by Gaoni and Mechoulam (1966). 7α-Hydroxy-Δ8-THC (Mechoulam et al., 1972), 7β-hydroxy-Δ8-THC (Mechoulam et al., 1972), 11-hydroxy-Δ8-THC (Inayama et al., 1974), 8β-hydroxy-Δ9-THC (Pitt et al., 1975), 9α,10α-epoxyhexahydrocannabinol (9α,10α-EHHC) (Narimatsu et al., 1983), 11-hydroxy-Δ9-THC (Pitt et al., 1975), 8-hydroxy-CBN (Inayama et al.,

Metabolism of cannabinoids by human hepatic microsomes

The most predominant metabolites of THCs and CBN formed by pooled human hepatic microsomes were 11-hydroxy metabolites. The catalytic activities (nmol/min/mg protein) for the 11-hydroxylation of pooled microsomes for Δ8-THC, Δ9-THC, and CBN were 0.492, 0.515, and 0.448, respectively. Δ8-THC, Δ9-THC, and CBN were also biotransformed to 7α-hydroxy-Δ8-THC (0.355 nmol/min/mg protein), 8β-hydroxy-Δ9-THC (0.344 nmol/min/mg protein), and 8-hydroxy-CBN (0.039 nmol/min/mg protein), respectively, as the

Discussion

Δ8-THC was mainly converted to 7α-hydroxy-Δ8-THC and 11-hydroxy-Δ8-THC by human hepatic microsomes. Δ9-THC and CBN were also primarily oxidized at the 8- and 11-positions by human hepatic microsomes. The 11-hydroxylation of three cannabinoids was highly inhibited by sulfaphenazole, which has been confirmed as a potent inhibitor of CYP2C enzymes, suggesting that CYP2C enzymes are involved in the 11-hydroxylation of the cannabinoids in human hepatic microsomes as indicated in the case of Δ9-THC

Acknowledgement

A part of this work was supported by the Academic Frontier Project for Private Universities from the Ministry of Education, Culture, Sports, Science and Technology of Japan (2005–2009).

References (30)

  • L.M. Bornheim et al.

    Human hepatic microsomal metabolism of Δ1-tetrahydrocannabinol

    Drug Metabolism and Disposition

    (1992)
  • C. Emoto et al.

    Cooperativity of α-naphthoflavone in cytochrome P4503A-dependent drug oxidation activities in hepatic microsomes from mouse and human

    Xenobiotica

    (2001)
  • M. Halldin et al.

    Identification of in vitro metabolites of Δ1-tetrahydrocannabinol formed by human liver

    Drug Metabolism and Disposition

    (1982)
  • D.J. Harvey

    Chemistry, metabolism, and pharmacokinetics of the cannabinoids

  • S. Inayama et al.

    The oxidation of Δ1- and Δ6-tetrahydrocannabinol with selenium dioxide

    Chemical and Pharmaceutical Bulletin

    (1974)
  • Cited by (189)

    • Cannabis, cannabinoids, and receptor responses

      2023, Cannabis Use, Neurobiology, Psychology, and Treatment
    View all citing articles on Scopus
    View full text