Method and compositions for administering resveratrol and pterostilbene

The present application claims priority from provisional application 61/019,745, filed Jan. 8, 2008.

The present invention relates to methods and compositions for administering resveratrol and pterostilbene.

Resveratrol and its analogs, pterostilbene (3,5-dimethoxy-4′-hydroxy-trans-stilbene), TMS (3,4′,5-reimwrhoxzy-trans-stilbene), 3,4′,4-DH-5-MS (3,4′-dihydroxy5-methoxy-trans-stilbnene) and 3,5-DH-4′MS(3,5-dihydroxy-4′-,ethoxy-trans-stilbene), have been shown to have chemopreventaive activity against cardiovascular disease and a variety of cancers in model systems. However, it is not clear whether the drugs reach the proposed sites of action in vivo after oral administration, especially in humans.

Walle et al., in Drug Metabolism and Disposition 32: 1377-1382, 2004, examined the absorption, bioavailability and metabolism of 14C-resveratrol after oral and intravenous doses in six human volunteers. When healthy men and women took an oral dose of 2 grams of trans-resveratrol, only traces of the unchanged resveratrol were detected in plasma. However, the resveratrol is rapidly transformed in the liver to the glucurunide, which makes the resveratrol not bioavailable.

Resveratrol is found in limited quantities in wine and some foods. The amount of resveratrol in wines ranges from about 0.05 mg/liter to about 7.13 mg/L, with red wines having the most resveratrol. Other sources of resveratrol are peanuts and peanut butter (about 1 mg per cup (about 150 g), and red grapes, which

Resveratrol and pterostilbene have been shown to have chemopreventive activity against cardiovascular disease and a variety of cancers in model systems. However, it is not clear if the drug reaches the proposed sites of action in vivo after oral ingestion, especially in humans. Unfortunately, as reported by Walle et al., Drug Metabolism and Disposition 32:1377-1382, 2004, absorption of a dietary relevant 25-mg oral dose was at least 70% with peak plasma levels of resveratrol and metabolites of 491±90 ng/ml (about 2 microM) and a plasma half-life of 9.2±0.6 hours. However, only trace amounts of unchanged resveratrol (<5 ng/ml) could be detected in plasma. Most of the oral dose was recovered in urine, and liquid chromatography/mass spectrometry analysis identified three metabolic pathways: sulfate and glucuronic acid conjugation of the phenolic groups and hydrogenation of the aliphatic double bond, the latter likely produced by intestinal microflora. Extremely rapid sulfate conjugation by the intestine/liver appears to be the rate-limiting step in the bioavailability of resveratrol and related compounds such as pterostilbene.

In order to avoid this entero-hepatic first exposure, an exposure that deactivates resveratrol, it was discovered that absorption through the buccal mucosa could bypass the enter-hepatic circulation and thus enable the resveratrol to be absorbed unchanged. Indeed, non-conjugated resveratrol levels were substantially increased in the serum in this way. It was also found that ethanol and DMSO in concentrations from 5% to about 50% could be used as vehicles or as transfer agents for resveratrol, which can increase absorption up to ten-fold.

Accordingly, Resveratrol, pterostilbene, and related compounds can be administered transmucosally or buccally by incorporating one or more of these compounds into any foodstuff that remains in the mouth for a period of time of at least five seconds or more, and preferably up to twenty minutes, such as chewing gum, taffy, hard candy, jerky, etc. Detailed Description of the Invention

Significant reductions in cardiovascular disease risk have been associated with moderate consumption of alcoholic beverages (43). This “French Paradox”, the observation that mortality from coronary heart disease is relatively low in France despite relatively high levels of dietary saturated fat and cigarette smoking, suggested that the regular consumption of red wine could provide additional protection from cardiovascular disease (44, 45). Red wine contains resveratrol and even higher levels of flavonoids. These polyphenolic compounds have antioxidant, anti-inflammatory, and other potentially anti-atherogenic effects in the test tube and in some animal modes of atherosclerosis (46). Results of epidemiological studies addressing the value of these polyphenolic compound has been inconsistent. While some large prospective studies found that wine drinkers were at lower risk of cardiovascular disease than been or liquor drinkers (47-49), others fond no difference (50-52). Several studies have discovered that people who drink wine have higher incomes, more education, smoke less, and eat more fruits and vegetables and less saturated fat than people who prefer beer or liquor (53-55). This may explain some of the differences.

In vitro, resveratrol effectively scavenges (neutralizes) free radicals an other oxidants (11) and inhibits oxidation of low density lipoprotein (12, 13). However, there is little evidence that resveratrol is an important antioxidant in vivo (14) because resveratrol is very quickly conjugated in the gastrointestinal/liver system, and intracellular concentrations of resveratrol in humans are likely to be much lower than that of other important antioxidants, such as vitamin C, vitamin E and glutathione. It is believed that the antioxidant activity of resveratrol metabolites, which comprise most of the circulating resveratrol, may be lower than that of resveratrol.

The chemical structure of resveratrol is very similar to that of the synthetic estrogen agonist, diethylstilbestrol, suggesting that resveratrol might also function as an estrogen agonist. However, in cell culture experiments resveratrol acts as an estrogen agonist under some condition, and an estrogen antagonist under other conditions (16, 17). In estrogen receptor-positive breast cancer cells, resveratrol acted as an estrogen agonist in the absence of the endogenous estrogen 17-beta-estradiol, but acted as an estrogen antagonist in the presence of 17-beta estradiol (18, 19). At present it appears that resveratrol has the potential to act as an estrogen agonist or antagonist, depending upon factors such as cell type, estrogen receptor isoform (ER alpha or ER beta), and the presence of endogenous estrogens (15).

Resveratrol, 3,4′-dihydroxystilbene, also known as 3,4′5-stilbenetriol, is a phytoalexin produced naturally by several plants when under attack by bacteria or fungi. Resveratrol has also been produced synthetically [see Farina et al., Nat. Prod. Res. 20(3): 247-252 2006]. A number of beneficial health effects of resveratrol have been reported, including anti-cancer, antiviral, neuroprotective, anti-aging, anti-inflammatory, and life-prolonging effects, although most of the studies used animal subjects rather than humans.

Resveratrol is found in widely varying amounts in grapes (primarily the skins of red grapes), raspberries, mulberries, plums, peanuts, berries of the Vaccinium species, including blueberries, bilberries, cranberries, some pines such as Scotch pine and eastern white pine, and the roots and stalks of giant knotweed and Japanese knotweed. Resveratrol was first isolated from an extract of the Peruvian legume Cassia quinquangulata in 1974.

In grapes, resveratrol is found primarily in the skin and seeds. The amount found in grape skins also varies with the grape cultivar, its geographic origin, and exposure to fungal infection. The amount of fermentation time a wine spends in contact with grape skins is an important determinant of its resveratrol content.

Pterostilbene is a stilbenoid compound chemically related to resveratrol. Other names for pterostilbene are 4-[(E)-2-(3,5-dimethoxyphenyl)ethenyl]phenol; 3′,5′-dimethoxy-4-stilbenol; and 3,5-dimethoxy-4′-hydroxy-trans-stilbene.

Pterostilbene has been found to be equal in potency to resveratrol as a cancer-preventive agent (Rimando et al., Cancer Chemopreventive and Antioxidant Activities of Pterostilbene, a Naturally Occurring Analogue of Resveratrol, Ars.usda.gov). Antioxidants destroy free radicals, highly reactive molecules whose excess has been linked to cancer, which may account for the effectiveness of resveratrol and pterostilbene against cancer. Animal studies have also found that pterostilbene can lower blood glucose and may be a potent antidiabetic agent.

Quantitative studies have shown that for every ten parts resveratrol, there are only one to two parts pterostilbene. The relationship between the two compounds and their unequal content in plants is unclear, but remains the subject of ongoing studies. Dark-skinned grapes are likely to contain the most pterostilbene. For reasons that are not clear, pterostilbene is normally not found in wine. This may be because it is unstable in light and air, which makes it less likely to survive the wine-making process.