Major in vitro phase I and phase II metabolites of cannabichromene

Abstract Background Following the passage of the Agricultural Improvement Act of 2018, the public of the United States has largely unobstructed access to minor cannabinoids. Cannabichromene (CBC) is a prevalent minor cannabinoid which has grown in popularity due to anecdotal claims comprising anti-inflammatory, anti-depressant, and anti-tumor properties. However, there is limited data regarding pharmacological activity, drug metabolism and pharmacokinetics. Given the expected growth in the minor cannabinoid market, it becomes increasingly imperative to investigate the fundamental characteristics of CBC exposure, including metabolism and identification of key CYP450s and UGTs involved in the generation of major metabolites. Methods HepaRG cells and human liver microsomes were used to generate CBC metabolites in vitro at 2- and 24-h timepoints; cell extracts were analyzed via high-resolution mass spectrometry. Various dosages in HepaRG cells were analyzed for potential cytotoxicity. Human liver microsomes were then used to characterize the phenotypic generation of major CBC metabolites. A one-way ANOVA was used to compare the statistical difference between control groups and treatment groups. Results The major phase I and phase II metabolites were detected in CBC-exposed HepaRG cells; the major oxidative metabolite was 2’-hydroxycannabicitran while the major conjugated metabolite was putatively identified as CBC-glucuronide. Enzyme inhibition studies identified CYP3A, UGT1A9, and UGT1A1 to be involved in the generation of major metabolites. CBC and CBC metabolites were well tolerated in HepaRG cells. Conclusions This is the first study to profile CBC metabolism in HepaRG cells and human liver microsomes in an orthogonal manner. 2’-hydroxycannabicitran and a proposed CBC-glucuronide were generated and detected as major metabolites in both systems.2’-hydroxycannabicitran was generated by CYP3A, while UGT1A9 and UGT1A1 contributed to the generation of CBC-glucuronide. Ultimately, the identification of CBC metabolites and contributing metabolic isoforms will support future clinical research on CBC pharmacokinetics and potentially contribute to the development of future safety and toxicological guidelines.