Products made from the leaves of Camellia sinensis (green tea) are among the most widely consumed natural products worldwide. Claims including cardioprotection, chemoprevention, and weight loss drive green tea beverage and dietary supplement popularity. The increasing popularity of green tea products increases the likelihood of co-consumption with conventional medications resulting in potentially important green tea-drug interactions.
A comprehensive literature search conducted by the Pharmacology Core, combined with compelling preliminary in vitro data, identified intestinal UDP-glucuronosyltransferases (UGTs) as a high priority target for green tea-precipitated natural product-drug interactions. Specifically, methanolic and aqueous extracts prepared from representative green teas (by the Analytical Core) were screened for inhibitory effects on intestinal UGT activity by the Pharmacology Core. All of these extracts showed clear concentration-dependent inhibition. Five abundant green tea catechins [(+)-catechin, (-)-epicatechin, and (-)-epigallocatechin, (-)-epicatechin gallate and (-)-epigallocatechin gallate] were subsequently tested individually as inhibitors of intestinal UGT activity. The inhibitory potency (IC50) of two of these catechins, (-)-epicatechin gallate and (-)-epigallocatechin 3-gallate, were near or below concentrations measured in a cup (250 mL) of brewed tea prepared from a reference standard obtained from the National Institute of Standards and Technology. These data suggest a potential interaction between green tea and drugs metabolized by intestinal UGT. In response, further mechanistic in vitro studies and clinical evaluation were conducted by the NaPDI Center to assess the clinical relevance of a green tea-drug interaction mediated via intestinal UGT inhibition.
Selection of Green Tea as a model precipitant
A comprehensive literature search conducted by the Pharmacology Core, combined with compelling preliminary in vitro data, identified intestinal UDP-glucuronosyltransferases (UGTs) as a high priority target for green tea-precipitated NPDIs. These effects have been attributed to polyphenols known as catechins, which constitute 30-42% of solid green tea extract by weight. As such, the catechins represent the most important and extensively studied green tea constituents in terms of potential precipitants of drug metabolizing enzyme- and transporter-mediated interactions with drugs.
Sourcing appropriate Green Tea products
The Analytical Core used metabolomics and biochemometrics approaches to prioritize candidate green tea products to study as precipitants of an intestinal UGT-mediated NPDI in human subjects. A single tea product was selected from 34 commercially available green tea samples based on the combination of metabolomics and chemometric data within the context of practical considerations including available single lot quantities, shelf-life, and sales. The selected green tea product underwent further quality assurance including key constituent quantification at least in triplicate to ensure reproducibility. A methanolic extract and selected fractions of this product were tested as inhibitors of intestinal UGT activity to confirm bioactivity.
Mechanistic in vitro evaluation
The green tea extract and fractions were prepared by the Analytical Core and provided to the Pharmacology Core for initial high throughput testing using human intestinal microsomes and a fluorescent UGT probe substrate. Raloxifene, an established in vitro and in vivo UGT1A object drug, was selected to complete detailed evaluation of the intestinal UGT inhibitory effects of green tea. The inhibitory potency (Ki) green tea product extract (provided by the Analytical Core), (-)-epicatechin gallate, and (-)-epigallocatechin gallate was determined using human intestinal microsomes. The induction potential of the clinical green tea product extract towards major drug metabolizing enzymes and transporters was also determined using sandwich-cultured human hepatocytes. The recovered inhibition and induction kinetics of green tea extracts/constituents provided the necessary mechanistic information to design a clinical green tea-raloxifene interaction study.
Clinical Study Design
Sixteen healthy adults (8 men, 8 women) participated in a crossover, open-label, three-phase, fixed-sequence study. The three phases included baseline (no green tea), short-term (acute) green tea administration, and long-term (chronic) green tea administration. Each phase consisted of one 12-hour inpatient visit and four 1-hour outpatient visits. Phase I involved administration of a single raloxifene dose (60 mg po) to obtain baseline raloxifene pharmacokinetics. Phase II involved administration of a single raloxifene dose (60 mg po) with brewed green tea (3 cups consumed within the first 6 hours) to determine the effects of acute green tea administration on raloxifene pharmacokinetics. Phase III involved administration of a single raloxifene dose (60 mg po) following a 5-day pre-treatment with green tea (3 cups consumed within the 8-hour work day) to determine the effects of chronic green tea administration on raloxifene pharmacokinetics. Each phase was separated by at least one week to ensure sufficient washout of raloxifene (half-life ~28 hours), raloxifene 4’- and 6-glucuronides (half-lives <20 hours), and green tea catechins (half-lives <2 hours).
Literature data describing green tea metabolism and disposition and comprehensive data generated by the NaPDI Center interaction project to describe the green tea-raloxifene interaction have been uploaded into the data repository using standard operating procedures developed by the Informatics Core.
Summary of Data
- Methanolic and aqueous extracts prepared from representative green teas (by the Analytical Core) were screened for inhibitory effects on intestinal UGT activity (4-methylumbelliferone glucuronidation by human intestinal microsomes) by the Pharmacology Core. All extracts showed clear concentration-dependent inhibition. Five abundant green tea catechins [(+)-catechin, (-)-epicatechin, and (-)-epigallocatechin, (-)-epicatechin gallate and (-)-epigallocatechin gallate] were subsequently tested as inhibitors of intestinal UGT activity using raloxifene as a clinically relevant in vitro object drug.
- (-)-Epicatechin gallate and (-)-epigallocatechin gallate competitively inhibited raloxifene-4’-glucuronide formation with Ki values (mean ± S.D.) of 0.8 ± 0.2 and 2.0 ± 0.3 µM, respectively. (-)-Epicatechin gallate and (-)-epigallocatechin gallate competitively inhibited raloxifene-6-glucuronide formation with Ki values (mean ± S.D.) of 1.0 ± 0.1 and 2.0 ± 0.3 µM, respectively. (+)-Catechin, (-)-epicatechin, and (-)-epigallocatechin demonstratedrelatively weak inhibition (20-50%) of raloxifene glucuronidation when tested at 100 μM.
- Cytochrome P450 induction potential of a methanolic extract of the selected green tea product was evaluated in cryopreserved sandwich-cultured human hepatocytes. No significant (≥ 20% of adjusted positive control response) induction of CYP1A2, CYP2B6, and CYP3A4 mRNA content or activity were observed.
- Transporter induction by a methanolic extract of the selected green tea extract was evaluated in cryopreserved sandwich-cultured human hepatocytes. No significant (≥ 20% of adjusted positive control response) induction of SLCO1B3 (OATP1B3) and ABCB1 (Pgp, MDR1) mRNA content were observed.
- A mechanistic model was applied to predict raloxifene AUC change in the presence of (-)-epicatechin gallate or (-)-epigallocatechin 3-gallate. Intestinal lumen concentrations of ECG and EGCG were predicted to be 4.4 and 15.2 μM, respectively, after administration a bag of the selected green tea product. Using Ki values of 1 and 2 µM for (-)-epicatechin gallate or (-)-epigallocatechin 3-gallate, respectively, the AUCi/AUC ratio was 4.4 and 6.1, respectively. The average maximum enterocyte concentrations of ECG and EGCG were estimated to be 0.18 and 0.54 µM, respectively, after administration a bag of the selected green tea product, producing an AUCi/AUC ratio of 1.2 and 1.3, respectively.
Healthy volunteers (n=58) aged 18-65 years were pre-screened to assess eligibility and gauge interest in clinical study participation. Forty-three individuals were eligible based upon pre-screening and twenty eligible individuals were consented, screened, and met enrollment criteria. Four subjects were withdrawn during the study, resulting in sixteen individuals that completed all study phases. A pharmacokinetic interaction was observed between a well-characterized green tea and the intestinal UGT substrate raloxifene, as reflected by the geometric mean ratio raloxifene AUC0-96h and Cmax decreasing to below the pre-defined no effect range (0.75-1.33). The unaltered raloxifene and glucuronide terminal half-lives, combined with unaltered ratios of glucuronide-to-raloxifene AUC0-96h, in the presence of green tea, suggested inhibition of intestinal UGT activity was not responsible for the observed interaction, different from the IVIVE prediction. The greater decrease in raloxifene geometric mean Cmax relative to AUC0-96h further suggested that green tea alters primarily processes in the intestine, which could include permeability, transport and/or physicochemical processes involved in raloxifene absorption.
Green tea catechins potently inhibit intestinal UGT activity in vitro. However, inhibition of raloxifene intestinal glucuronidation does not appear to be the primary mechanism underlying clinical green tea-raloxifene interactions.