(T0930-11-61) Cannabinoids Selectively Alter Cytochrome P450(CYP) Activity in Primary Cultures of Human Hepatocytes

Purpose: Marijuana/cannabis is one of the most widely used illicit drug in the United States. Medical use of marijuana is legalized in several states. The overall objective of the project is to evaluate the effect of cannabinoids on the pharmacokinetics of co-administered drugs using a physiologically based pharmacokinetic (PBPK) model. As a first step, the goal of this study is to evaluate the effect of acute and chronic treatment of cannabinoids on the activity of various CYP enzymes in-vitro.

Methods: Primary cultures of human hepatocytes were obtained from the hepatocyte isolation facility at University of Pittsburgh. The metabolic capacity of the cells used was assessed using testosterone metabolism and by evaluating changes in testosterone metabolism in response to either rifampin (10 µm) or ketoconazole (10 µm) treatment for 3 days. The cells were treated with three major cannabinoids, tetrahydrocannabinol (THC), cannabidiol (CBD) or cannabinol (CBL) or a mixture (MIX) of the three cannabinoids for either 30 minutes to study acute effect or 4 days to study chronic effect with media change every day. Control cells were treated with vehicle simultaneously. The CYP activity was assessed using a cocktail of 5 CYP probes viz. phenacetin for CYP1A2, diclofenac for CYP2C9, mephenytoin for CYP2C19, dextromethorphan for CYP2D6 and midazolam for CYP3A4. CYP activity was evaluated by measuring the metabolites of the respective CYP probes using an UPLC-MS-MS assay and the mRNA expression of major CYP enzymes was studied using qrt-PCR.

Results: The hepatocytes were responsive to drug treatment and showed at least 4-fold increase and at least 30-fold decrease in production of testosterone metabolite due to rifampin and ketoconazole treatment, respectively. Cannabinoids showed no major effect on CYP2D6 and CYP2C9. THC and MIX appear to induce CYP3A4 as seen by 2-fold increase in the production of midazolam metabolite. There was at least a 2-fold increase and 4-fold increase in the production of phenacetin metabolite by CBL and MIX, respectively indicating induction of CYP1A2. CBL and MIX inhibited CYP1A2 in a concentration dependent manner. A similar trend was observed in activity assessed using CYP cocktail and mRNA expression analysis using qrt-PCR.

Conclusion: Cannabinoids selectively alter certain CYP enzymes which may lead to changes in exposure of corresponding substrates that are administered. Future studies will evaluate in vivo predictions of cannabinoid mediated drug interactions based on the in vitro data collected using PBPK modeling.