Abstract
Botanical and other natural products (NPs) are often coconsumed with prescription medications, presenting a risk for cytochrome P450 (P450)-mediated NP-drug interactions. The NP goldenseal (Hydrastis canadensis) has exhibited antimicrobial activities in vitro attributed to isoquinoline alkaloids contained in the plant, primarily berberine, (−)-β-hydrastine, and to a lesser extent, hydrastinine. These alkaloids contain methylenedioxyphenyl rings, structural alerts with potential to inactivate P450s through formation of metabolic intermediate complexes. Time-dependent inhibition experiments were conducted to evaluate their ability to inhibit major P450 activities in human liver microsomes by using a cocktail of isozyme-specific substrate probes. Berberine inhibited CYP2D6 (dextromethorphan O-demethylation; KI = 2.7 μM, kinact = 0.065 minute−1) and CYP3A4/5 (midazolam 1′-hydroxylation; KI = 14.8 μM, kinact = 0.019 minute−1); (−)-β-hydrastine inhibited CYP2C9 (diclofenac 4′-hydroxylation; KI = 49 μM, kinact = 0.036 minute−1), CYP2D6 (KI > 250 μM, kinact > 0.06 minute−1), and CYP3A4/5 (KI = 28 μM, kinact = 0.056 minute−1); and hydrastinine inhibited CYP2D6 (KI = 37 μM, kinact = 0.049 minute−1) activity. Berberine additionally exhibited allosteric effects on midazolam hydroxylation, showing both positive and negative heterotropic cooperativity. Experiments with recombinant isozymes showed that berberine activated midazolam 1′-hydroxylation by CYP3A5, lowering Km(app), but showed mixed inhibition and negative cooperativity toward this reaction when catalyzed by CYP3A4. Berberine inactivated CYP3A4 at a much faster rate than CYP3A5 and was a noncompetitive inhibitor of midazolam 4-hydroxylation by CYP3A4 but a strong mixed inhibitor of the CYP3A5 catalyzed reaction. These complex kinetics should be considered when extrapolating the risk for NP-drug interactions involving goldenseal.
SIGNIFICANCE STATEMENT Robust kinetic parameters were determined for the reversible and time-dependent inhibition of CYP2C9, CYP2D6, and CYP3A4/5 activities in human liver microsomes by major component isoquinoline alkaloids contained in the botanical natural product goldenseal. The alkaloid berberine also exhibited opposing, isozyme-specific allosteric effects on midazolam hydroxylation mediated by recombinant CYP3A4 (inhibition) and CYP3A5 (activation). These data will inform the development of a physiologically based pharmacokinetic model that can be used to predict potential clinically relevant goldenseal-drug interactions.
Footnotes
- Received February 27, 2020.
- Accepted June 15, 2020.
↵1 Current affiliation: Drug Disposition, Eli Lilly and Company, Indianapolis, Indiana.
↵2 Current affiliation: Pfizer Inc., Boulder, Colorado.
This work was supported by National Institutes of Health National Center for Complementary and Integrative Health [Grant U54 AT008909]. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
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- Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics
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