Clinical pharmacology of resveratrol and its metabolites in colorectal cancer patients

Abstract

Resveratrol is a phytochemical with chemopreventive activity in preclinical rodent models of colorectal carcinogenesis. Antiproliferation is one of the many chemopreventive modes of action it has been shown to engage in. Concentrations of resveratrol, which can be achieved in human tissues after p.o. administration, have not yet been defined. The purpose of this study was to measure concentrations of resveratrol and its metabolites in the colorectal tissue of humans who ingested resveratrol. Twenty patients with histologically confirmed colorectal cancer consumed eight daily doses of resveratrol at 0.5 or 1.0 g before surgical resection. Resveratrol was found to be well tolerated. Normal and malignant biopsy tissue samples were obtained before dosing. Parent compound plus its metabolites resveratrol-3-O-glucuronide, resveratrol-4'-O-glucuronide, resveratrol-3-O-sulfate, resveratrol-4'-O-sulfate, resveratrol sulfate glucuronide, and resveratrol disulfate were identified by high-performance liquid chromatography (HPLC) with UV or mass spectrometric detection in colorectal resection tissue. Quantitation was achieved by HPLC/UV. Cell proliferation, as reflected by Ki-67 staining, was compared in preintervention and postintervention tissue samples. Resveratrol and resveratrol-3-O-glucuronide were recovered from tissues at maximal mean concentrations of 674 and 86.0 nmol/g, respectively. Levels of resveratrol and its metabolites were consistently higher in tissues originating in the right side of the colon compared with the left. Consumption of resveratrol reduced tumor cell proliferation by 5% (P = 0.05). The results suggest that daily p.o. doses of resveratrol at 0.5 or 1.0 g produce levels in the human gastrointestinal tract of an order of magnitude sufficient to elicit anticarcinogenic effects. Resveratrol merits further clinical evaluation as a potential colorectal cancer chemopreventive agent.

Figure 1

Figure 1 A. HPLC/UV chromatograms of extracts of normal colon tissue resected from a patient who had received resveratrol 1.0g daily for 8 days (solid line) and a patient who had refrained from resveratrol ingestion during the 7 days prior to resection (broken line). Identity of resveratrol-derived species as established by co-chromatography with authentic reference material and LC/MS/MS analysis is indicated above the peaks. Insert shows structures of resveratrol and its metabolites: R₁, R₂=H: resveratrol “7”, R₁=sulfate R₂=H: resveratrol-3-O-sulfate “6”, R₁=H, R₂=sulfate: resveratrol-4′-O-sulfate “5”, R₁=glucuronide, R₂=H: resveratrol-3-O-glucuronide “4”, R₁=H, R₂=glucuronide: resveratrol-4′-O-glucuronide “2”. Positions of the sulfonic and glucuronic acid moieties in resveratrol disulfate “3” and resveratrol sulfate glucuronide “1” are probably 3 and 4′, but this needs confirmation by ¹H-NMR. Naringenin “8” was the internal standard. For details of tissue procurement, extraction and chemical analysis see Materials and Methods. Figure 1 B. LC/MS selected reaction monitoring (SRM) of transitions for the identification of resveratrol metabolites in extracts of colon tissue taken from a patient who had received 0.5g of resveratrol for 8 days. Metabolites identified were resveratrol sulfate glucuronide (m/z 483>227), resveratrol disulfate (m/z 387>227) resveratrol-3-O-glucuronide and resveratrol-4′-O-glucuronide (m/z 403>227), resveratrol-3-O-sulfate and resveratrol-4′-O-sulfate (307>227). Resveratrol was also identified (m/z 227>184). The total ion current from the analysis of a mixture of authentic standards is also shown for comparison. The mixture contained resveratrol-4′-O-glucuronide (1), resveratrol-3-O-glucuronide (2), dehydrated resveratrol glucuronide (exhibits the transition 385>227 and is not present in the patient samples) (3), resveratrol-4′-O-sulfate (4), resveratrol-3-O-sulfate (5) and resveratrol (6).

Figure 2

Concentrations of resveratrol and resveratrol-3-O-sulfate in normal colon and tumor tissue of a patient with a cecal (right-sided) and a sigmoid colonic tumor (left-sided), who had received resveratrol at a dose of 1.0g daily for 8 days. Normal tissue samples were taken at a distance of 5cm proximal or distal from tumor, and from the distal resection margin (RM). Differences in levels between the left and right side, similar to those shown here, were observed for resveratrol disulfate and the two resveratrol monoglucuronides, but not for resveratrol sulfate glucuronide.