The legalization of cannabis in many parts of the United States and other countries has led to a need for a more comprehensive understanding of cannabis constituents and their potential for drug-drug interactions. Although (−)-trans-Δ⁹-tetrahydrocannabinol (THC), cannabidiol (CBD), and cannabinol (CBN) are the most abundant cannabinoids present in cannabis, THC metabolites are found in plasma at higher concentrations and for a longer duration than that of the parent cannabinoids. To understand the potential for drug-drug interactions, the inhibition potential of major cannabinoids and their metabolites on major hepatic cytochrome P450 (P450) enzymes was examined. In vitro assays with P450-overexpressing cell microsomes demonstrated that the major THC metabolites 11-hydroxy-Δ⁹-tetra-hydrocannabinol and 11-nor-9-carboxy-Δ⁹-THC-glucuronide competitively inhibited several major P450 enzymes, including CYP2B6, CYP2C9, and CYP2D6 (apparent K_i,u values = 0.086 ± 0.066 µM and 0.90 ± 0.54 µM, 0.057 ± 0.044 µM and 2.1 ± 0.81 µM, 0.15 ± 0.067 µM and 2.3 ± 0.54 µM, respectively). 11-Nor-9-carboxy-Δ⁹- tetrahydrocannabinol exhibited no inhibitory activity against any CYP450 tested. THC competitively inhibited CYP1A2, CYP2B6, CYP2C9, and CYP2D6; CBD competitively inhibited CYP3A4, CYP2B6, CYP2C9, CYP2D6, and CYP2E1; and CBN competitively inhibited CYP2B6, CYP2C9, and CYP2E1. THC and CBD showed mixed-type inhibition for CYP2C19 and CYP1A2, respectively. These data suggest that cannabinoids and major THC metabolites are able to inhibit the activities of multiple P450 enzymes, and basic static modeling of these data suggest the possibility of pharmacokinetic interactions between these cannabinoids and xenobiotics extensively metabolized by CYP2B6, CYP2C9, and CYP2D6. SIGNIFICANCE STATEMENT Major cannabinoids and their metabolites found in the plasma of cannabis users inhibit several P450 enzymes, including CYP2B6, CYP2C9, and CYP2D6. This study is the first to show the inhibition potential of the most abundant plasma cannabinoid metabolite, THC-COO-Gluc, and suggests that circulating metabolites of cannabinoids play an essential role in CYP450 enzyme inhibition as well as drug-drug interactions.

The UDP-glucuronosyltransferase (UGT) 1A enzymes are involved in the phase II metabolism of many important endogenous and exogenous compounds. The nine UGT1A isoforms exhibit high interindividual differences in expression, but their epigenetic regulation is not well understood. The purpose of the present study was to examine microRNA (miRNA) regulation of hepatic UGT1A enzymes and determine whether or not that regulation impacts enzymatic activity. In silico analysis identified miRNA 491-3p (miR-491-3p) as a potential regulator of the UGT1A gene family via binding to the shared UGT1A 3′-untranslated region common to all UGT1A enzymes. Transfection of miR-491-3p mimic into HuH-7 cells significantly repressed UGT1A1 (P < 0.001), UGT1A3 (P < 0.05), and UGT1A6 (P < 0.05) mRNA levels. For UGT1A1, this repression correlated with significantly reduced metabolism of raloxifene into raloxifene-6-glucuronide (ral-6-gluc; P < 0.01) and raloxifene-4′-glucuronide (ral-4′-gluc; P < 0.01). In HuH-7 cells with repressed miR-491-3p expression, there was a significant increase (∼80%; P < 0.01) in UGT1A1 mRNA and a corresponding increase in glucuronidation of raloxifene into ral-6-gluc (50%; P < 0.05) and ral-4′-gluc (22%; P < 0.01). Knockdown of endogenous miR-491-3p in HepG2 cells did not significantly alter UGT1A1 mRNA levels but did increase the formation of ral-6-gluc (50%; P < 0.05) and ral-4′-gluc (34%; P < 0.001). A significant inverse correlation between miR-491-3p expression and both UGT1A3 (P < 0.05) and UGT1A6 (P < 0.01) mRNA levels was observed in a panel of normal human liver specimens, with a significant (P < 0.05) increase in UGT1A3 and UGT1A6 mRNA levels observed in miR-491-3p nonexpressing versus expressing liver specimens. These results suggest that miR-491-3p is an important factor in regulating the expression of UGT1A enzymes in vivo.

Cannabis is an increasingly popular recreational and medicinal drug in the USA. While cannabis is still a Schedule 1 drug federally, many states have lifted the ban on its use. With its increased u...

The UDP-glucuronosyltransferase (UGT) family of enzymes play a central role in the metabolism and detoxification of a wide range of endogenous and exogenous compounds. UGTs exhibit a high degree of structural similarity and display overlapping substrate specificity, often making estimations of potential drug-drug interactions difficult to fully elucidate. One such interaction yet to be examined may be occurring between UGTs and cannabinoids, as the legalization of recreational and medicinal cannabis and subsequent co-usage of cannabis and therapeutic drugs increases in the United States and internationally. In the present study, the inhibition potential of the major cannabinoids Δ⁹-tetrahydrocannabinol (THC), cannabidiol (CBD), and cannabinol (CBN), as well as their major metabolites, was determined in microsomes isolated from HEK293 cells overexpressing individual recombinant UGTs and in microsomes from human liver and kidney specimens. The highest inhibition was seen by CBD against the glucuronidation activity of UGTs 1A9, 2B4, 1A6, and 2B7, with binding-corrected IC₅₀ values of 0.12 ± 0.020 µM, 0.22 ± 0.045 µM, 0.40 ± 0.10 µM, and 0.82 ± 0.15 µM, respectively. Strong inhibition of UGT1A9 was also demonstrated by THC and CBN, with binding-corrected IC₅₀ values of 0.45 ± 0.12 μM and 0.51 ± 0.063 μM, respectively. Strong inhibition of UGT2B7 was also observed for THC and CBN; no or weak inhibition was observed with cannabinoid metabolites. This inhibition of UGT activity suggests that in addition to playing an important role in drug-drug interactions, cannabinoid exposure may have important implications in patients with impaired hepatic or kidney function. SIGNIFICANCE STATEMENT Major cannabinoids found in the plasma of cannabis users inhibit several UDP-glucuronosyltransferase (UGT) enzymes, including UGT1A6, UGT1A9, UGT2B4, and UGT2B7. This study is the first to show the potential of cannabinoids and their metabolites to inhibit all the major kidney UGTs as well as the two most abundant UGTs present in liver. This study suggests that as all three major kidney UGTs are inhibited by cannabinoids, greater drug-drug interaction effects might be observed from co-use of cannabinods and therapeutics that are cleared renally.

The UDP-glucuronosyltransferase (UGT) 2B enzymes are important in the detoxification of a variety of endogenous and exogenous compounds, including many hormones, drugs, and carcinogens. Identifying novel mechanisms governing their expression is important in understanding patient-specific response to drugs and cancer risk factors. In silico prediction algorithm programs were used to screen for microRNAs (miRNAs) as potential regulators of UGT2B enzymes, with miR-216b-5p identified as a potential candidate. Luciferase data suggested the presence of a functional miR-216b-5p binding motif within the 3′ untranslated regions of UGTs 2B7, 2B4, and 2B10. Overexpression of miR-216b-5p mimics significantly repressed UGT2B7 (P < 0.001) and UGT2B10 (P = 0.0018) mRNA levels in HuH-7 cells and UGT2B4 (P < 0.001) and UGT2B10 (P = 0.018) mRNA in Hep3B cells. UGT2B7 protein levels were repressed in both HuH-7 and Hep3B cells in the presence of increasing miR-216b-5p concentrations, corresponding with significant (P < 0.001 and P = 0.011, respectively) decreases in glucuronidation activity against the UGT2B7-specific substrate epirubicin. Inhibition of endogenous miR-216b-5p levels significantly increased UGT2B7 mRNA levels in HuH-7 (P = 0.021) and Hep3B (P = 0.0068) cells, and increased epirubicin glucuronidation by 85% (P = 0.057) and 50% (P = 0.012) for HuH-7 and Hep3B cells, respectively. UGT2B4 activity against codeine and UGT2B10 activity against nicotine were significantly decreased in both HuH-7 and Hep3B cells (P < 0.001 and P = 0.0048, and P = 0.017 and P = 0.043, respectively) after overexpression of miR-216b-5p mimic. This is the first evidence that miRNAs regulate UGT 2B7, 2B4, and 2B10 expression, and that miR-216b-5p regulation of UGT2B proteins may be important in regulating the metabolism of UGT2B substrates.