Based on the reviewers'comments and new findings we revised the study design, dose, timing, tissue processing and statistical evaluation. The objective of the proposed human intervention study remains to determine whether a combined administration of quercetin and green tea (GT) will increase the bioavailability and decrease the metabolism of GT polyphenols (GTPs) such as epigallocatechin gallate (EGCG). Factors limiting the biological activity of GTPs are the reduced bioavailability and extensive metabolism converting EGCG into methylated metabolites with decreased activity. In our previous human intervention study in men drinking 6 cups of green tea daily prior to prostatectomy we demonstrated that GTPs are present in human prostate tissue. However 50 percent of EGCG was found in methylated form. Quercetin, a flavonoid found in onions, apples, broccoli, berries and teas is a natural inhibitor of catechol-O-methyltransferase (COMT) and multidrug resistance transport proteins (MRP1 and MRP2). Our cell culture studies demonstrated that the combined treatment with EGCG and quercetin led to a 4 to 10-fold increase in intracellular concentration of EGCG and significant decrease in EGCG methylation, depending on the cell line used. We demonstrated in mouse prostate cancer xenograft studies that the administration of the combination of GT with quercetin significantly decreased tumor volume, increased EGCG tissue concentration and decreased the methylation of EGCG significantly compared to treatment with GT or quercetin alone. The combined administration also significantly decreased the protein expression of methyltransferases and MRP1. We therefore propose a placebo- controlled, double-blinded, randomized, parallel two arm human pilot intervention study to determine whether the consumption of GT extract and quercetin will increase the bioavailability of GTPs in blood and prostate tissue and decrease the methylation of EGCG. We will administer two capsules of GT extract combined with one capsule of quercetin (N=25) or with one placebo capsule (N=15) twice daily for 3 weeks to men scheduled for prostatectomy. After 10 participants will complete the intervention, liver toxicity will be evaluated. In the absence of lier toxicity the quercetin dose will be doubled (800 mg daily) (N=15). GTPs, quercetin and methylated metabolites will be analyzed in blood and prostate tissue using high performance liquid chromatography (HPLC) with coularray electrochemical detection (ECD). To evaluate the mechanism involved, COMT and DNA methyltransferase enzyme activities and gene and protein expression as well as MRP1 gene and protein expression and global DNA methylation will be determined in tumor and normal prostate tissue and COMT activity in erythrocytes using HPLC-ECD, real time qPCR and Western blot analyses. The results will lay the foundation for future clinical trials to evaluate the interaction of quercetin and GT in chemoprevention of early stages of prostate cancer.
TO PUBLIC HEALTH The potential of green tea to prevent or treat prostate cancer is limited due to restricted uptake and fast metabolism and excretion of the green tea chemicals. We propose a human intervention study to test the potential of another natural chemical from onions and apples called quercetin to increase the tissue concentration and decrease the metabolism/excretion of green tea chemicals.