Relationship between mechanisms, bioavailibility, and preclinical chemopreventive efficacy of resveratrol: a conundrum.

Experience has taught us that the rational clinical development of novel putative cancer chemopreventive agents needs to be based on robust preclinical information. Such information should relate to the mechanism of action, pharmacodynamics, and pharmacokinetics of the agent under study. Deficiencies in knowledge of any of these areas confound the understanding of the reasons for failure, when in a costly intervention trial, an agent is found to lack activity. The ramifications of such deficiencies are exemplified by the discussions engendered by the -carotene intervention trials in the early 1990s (1). In this commentary, we scrutinize recent preclinical information on the promising and novel diet-derived putative cancer chemopreventive agent resveratrol (3,5,4 -trihydroxy-trans-stilbene), with the aim of preparing the ground for its clinical evaluation. Resveratrol is a phytoalexin generated in response to environmental stress or pathogenic attack in grapes, mulberries, cranberries, peanuts, and plants of the Cassia quinquangulata family. Activity of Resveratrol in Cells and Biosystems in Vitro. In 1996, Jang et al. (2) published a paper in which the ability of resveratrol to inhibit diverse cellular events associated with the three major stages of carcinogenesis (initiation, promotion, and progression) was described. This seminal paper undoubtedly fired the imagination of the cancer chemoprevention research community. During the 6 years since its publication, 100 reports have appeared in the literature in which cellular and biochemical mechanisms of this agent have been elucidated, and their potential role in the suggested cancer chemopreventive activity of this agent has been discussed. These mechanistic studies have been comprehensively summarized in several reviews (3–7), and Table 1 illustrates some of the mechanistic properties of resveratrol in vitro, but the list is certainly not exhaustive. Collectively, the investigations of biochemical and cellular mechanisms of activity of resveratrol in vitro (Table 1) suggest that it engenders changes relevant to cancer chemoprevention at the level of the cell or the isolated subcellular target system, when used at concentrations between 10 and 100 M. Only on very few occasions have concentrations 10 M been shown to elicit bioactivity, e.g., IC50s for cellular growth inhibition by resveratrol tend to span the 5–10 M range (3); it inhibited recombinant cytochrome P450 CYP1B1 enzyme activity at 1 M (8), and it exerted antiestrogenicity in cells at submicromolar concentrations (9). There are two pivotal questions relating to the appropriateness of translating these in vitro results to animals and humans in vivo: (a) can 10 -10 4 M concentrations of resveratrol be achieved in vivo in the intact mammalian target organ in which malignancies are to be prevented; and (b) are doses of resveratrol, which have been shown to delay or prevent malignancies in rodent models, consistent with the attainment of such target organ levels? It is perhaps surprising that, to our knowledge, there has been no study on resveratrol published thus far probing the link between target organ levels, efficacy in vivo, and activity observed in cells in vitro. Levels of Resveratrol and its Metabolites in Humans and Rodents. How much resveratrol can be recovered from the human or rodent organism after resveratrol consumption? The resveratrol content of wine is 5 mg/liter. Assuming moderate wine consumption (250 ml in a 70-kg person), the intake of resveratrol with wine in humans is 18 g/kg/day. In a recent study in healthy volunteers, resveratrol was administered at a dose of 360 g/kg either dissolved in grape juice, vegetable juice, or white wine, i.e., at a dose which was 20 times that associated with “normal” wine intake (10). The authors used a very sensitive gas chromatography-mass spectrometry method and found plasma peak levels of 20 nM authentic resveratrol and 2 M “total” resveratrol (i.e., genuine resveratrol plus resveratrol generated by hydrolysis of its conjugates) 30 min after ingestion, irrespective of dietary matrix. Results from preclinical studies in rats, using exclusively high-performance liquid chromatography methods, suggest consistent attainment of plasma peak levels 5–10 min post-oral administration and a rapid plasma elimination half-life of 12–15 min. However, these studies differ as to the actual peak level values: doses of 2 mg/kg (11), 20 mg/kg (12), and 50 mg/kg resveratrol (13), each given via the i.g. route, generated peak values of 2, 1.2, and 6.6 M, respectively. In the latter study, the peak level of resveratrol glucuronide was as high as 105 M, and the authors present convincing evidence for extensive enterohepatic circulation (14). Using radiolabelled resveratrol administered by the oral route, an appreciable fraction, 50–75% of the dose, was absorbed in rats (14), and radioactivity could be recovered from the stomach, liver, kidney, intestine, bile, and urine in mice (15). Studies in mice, rats, and dogs suggest consistently that resveratrol is well absorbed and rapidly glucuronidated and sulfated both in the liver and intestinal epithelial cells (14–18). In investigations using a perfused rat small intestine model, ample uptake and metabolism of resveratrol occurred in the gut ex vivo (17, 19). Furthermore, resveratrol underwent glucuronidation and sulfation readily in liver cells and human and rodent liver and gut subcellular fractions (18, 20, 21). Taken together, all of these metabolism and pharmacokinetic investigations suggest convincingly that resveratrol is satisfactorily absorbed from the rodent gastrointestinal tract and efficiently Received 6/27/03; accepted 7/1/03. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 To whom requests for reprints should be addressed, at Department of Oncology, RKCSB, Leicester Royal Infirmary, University of Leicester, Leicester, LE2 7LX, United Kingdom. Phone: 0044 116 2231856; Fax: 0044 116 2231855; E-mail: [email protected]. 953 Vol. 12, 953–957, October 2003 Cancer Epidemiology, Biomarkers & Prevention