12-O-Tetradecanoylphorbol-13-acetate (TPA)-mediated oxidative stress in HeLa cells and its inhibition were studied by fluorometric measurement of H2O2 and by 3H-postlabeling of the oxidized bases 8-hydroxyl-2′-de-oxyguanosine (8-OHdG) and 5-hydroxymethyl-2′-deoxyuridine (HMdU). TPA treatment (10 fmol/cell) caused ≈7-fold increase in H2O2 levels (0.1 nmol TPA/ml), and 5-10-fold increase in 8-OHdG and HMdU (10 nmol TPA/ml). Naturally occurring compounds [caffeic acid phenethyl ester (CAPE), (-)·epigallocatechin gallate (EGCG), penta-O-galloyl-β-d-glu-cose (PGG) and sarcophytol A (Sarp A)] and the anticancer drug tamoxifen (TAM) were tested as potential chemopreventive agents. These agents dose-dependently inhibited TPA-induced H2O2,8-OHdG and HMdU. The doses required for a 50% decrease in H2O2 were ≈2.5 µm for TAM; 5 µm for CAPE, EGCG and PGG; and 75 µm for Sarp A. TAM and PGG (10 µm), EGCG (25 µm), and CAPE (50 µm) abolished TPA-mediated H2O2 production, even below the normal cellular levels. TAM (2.5–20 µm) decreased TPA-mediated HMdU and 8-OHdG formation 2–29 times. Maximum inhibition occurred at 20 µm TAM, which caused an ≈95% decline in HMdU and 8-OHdG. CAPE was effective at 0.5–50 µm. CAPE (25 µm) decreased 8-OHdG 95% and HMdU 58%, while Sarp A (250 µm) reduced 8-OHdG by 93% and HMdU by 78%. EGCG (1–25 µm) and PGG (1–10 µm) inhibited formation of 8-OHdG and HMdU dose-dependently. However, higher doses (50 and 100 µm) decreased the efficacy of that inhibition. Of those agents tested, TAM appears to be the most and Sarp A the least effective. Our results point to these 5 compounds as being potential chemopreventive agents, which at very low doses decrease the tumor promoter-mediated oxidative processes.
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Presented in part at the Annual Meeting of the American Association for Cancer Research (57). This publication was supported by Grant CA 37858 from the National Cancer Institute (its contents are solely the responsibility of the authors and do not necessarily represent the official views of the National Cancer Institute), and by Center Grant ES 00260 from the National Institute of Environmental Health Sciences.