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Hypermethylation of CpG islands in the promoter regions is an important mechanism to silence the expression of many important genes in cancer. The hypermethylation status is passed to the daughter cells through the methylation of the newly synthesized DNA strand by 5-cytosine DNA methyltransferase (DNMT). We report herein that (-)-epigallocatechin-3-gallate (EGCG), the major polyphenol from green tea, can inhibit DNMT activity and reactivate methylation-silenced genes in cancer cells. With nuclear extracts as the enzyme source and polydeoxyinosine-deoxycytosine as the substrate, EGCG dose-dependently inhibited DNMT activity, showing competitive inhibition with a Ki of 6.89 μM. Studies with structural analogs of EGCG suggest the importance of D and B ring structures in the inhibitory activity. Molecular modeling studies also support this conclusion and suggest that EGCG can form hydrogen bonds with Pro1223, Glu1265, Cys1225, Ser1229, and Arg1309 in the catalytic pocket of DNMT. Treatment of human esophageal cancer KYSE 510 cells with 5 to 50 μM of EGCG for 12 to 144 h caused a concentration- and time-dependent reversal of hypermethylation of p16INK4a, retinoic acid receptor beta (RARβ), O6-methylguanine methyltransferase (MGMT), and human mutL homolog 1 (hMLH1) genes as determined by the appearance of the unmethylation-specific bands in PCR. This was accompanied by the expression of mRNA of these genes as determined by RT-PCR. The re-expression of RARβ and hMLH1 proteins by EGCG was demonstrated by Western blot. Reactivation of some methylation-silenced genes by EGCG was also demonstrated in human colon cancer HT-29 cells, esophageal cancer KYSE 150 cells, and prostate cancer PC3 cells. The results demonstrate for the first time the inhibition of DNA methylation by a commonly consumed dietary constituent and suggest the potential use of EGCG for the prevention or reversal of related gene-silencing in the prevention of carcinogenesis.

[Proc Amer Assoc Cancer Res, Volume 45, 2004]