The anti-oxidant and the anti-tumor-promotion activities of several hydrolyzable tannins (HTs), including a commercial tannic-acid (TA) mixture, were examined in mouse skin treated with 12-O-tetradecanoylphorbol-13-acetate (TPA) in vivo. A single application of TPA gradually increases the hydroperoxide (HPx)-producing activity of the epidermis, which is maximally stimulated at 3 days and returns to control levels at 9 days. Pre-treatments with TA and ellagic acid (EA) strongly inhibit, in a dose-dependent manner, this HPx response to TPA. Total inhibition by TA lasts for about 16 hr, beyond which it is substantially reduced but not completely lost. TA can also reduce the level of epidermal HPx when it is applied 36 hr after the tumor promoter. EA is an antioxidant 10 times more potent than TA and n-propyl gallate (PG), which are equally effective against TPA-induced HPx production. Gallic acid is the least effective of the HTs in inhibiting HPx formation. TA also inhibits the production of HPx induced by several structurally different tumor promoters and the greater HPx responses produced by repeated TPA treatments. When applied 20 min before each promotion treatment, twice a week for 45 weeks, several HTs inhibit the incidence and yield of papillomas and carcinomas promoted by TPA in initiated skin. Overall, TA is more effective than EA and PG in inhibiting skin-tumor promotion by TPA, suggesting that the anti-oxidant effects of HTs are essential but not sufficient for their anti-tumor-promotion activity.

POU domain transcription factors are expressed in the epidermis and are thought to be important regulators of keratinocyte gene expression. In the present article we demonstrate that POU transcription factors suppress transcription of the human involucrin (hINV) promoter. Cotransfection of pINV-2473, a construct containing 2473 base pairs of hINV upstream sequence linked to luciferase, with POU homeodomain transcription factors Oct1, Oct2, Brn4, SCIP, Skn1a or Skn1i, results in a strong suppression of basal promoter activity. The hINV upstream region includes a consensus POU transcription factor binding site, 5'-ATGCAAAT-3', centered around nucleotide -1277. Although this site interacts with POU factors, assays of promoter activity for a series of progressive 5' end truncations demonstrate that this site is not required for POU factor-dependent transcriptional suppression. Suppression is observed with the shortest truncation construct tested, pINV-41, suggesting that this inhibition may be mediated by effects on TATA box proteins. SCIP mutants that lack transactivation or DNA binding domains were shown to suppress transcription, suggesting that the DNA binding and transactivation domains are not required for suppression. Moreover, cotransfection of the pINV-2473 with pKSM13(+)OCT, which contains a single consensus OCT binding site, results in an increase in basal promoter activity, suggesting that endogenous POU factors suppress hINV promoter activity. In addition to inhibiting basal transcription, POU transcription factors also suppress phorbol ester-stimulated hINV promoter activity. These studies suggest that suppression of hINV promoter activity does not require the amino-terminal segment of the POU factor or direct POU factor interaction with DNA and suggest that the suppression may be via indirect interaction with other proteins in the vicinity of the TATA box. Thus, involucrin joins the ranks of a small set of genes that are regulated by POU factors in an octamer binding site-independent manner.

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Several procyanidin dimers and an epicatechin trimer purified from Douglas fir bark tannins were compared with their monomer components (+)-catechin and (-)-epicatechin for their abilities to inhibit the biochemical effects of the potent tumor promoter 12-O-tetradecanoylphorbol 13-acetate (TPA) in mouse epidermis in vivo. Topical applications of the procyanidins, 15 min before the tumor promoter, inhibit TPA-induced ornithine decarboxylase (ODC) activity and this inhibition increases with the degree of polymerization (trimer > dimer > monomer). At a dose of 10 mumol, all procyanidin dimers inhibit the ODC response to TPA to a greater degree than 20 mumol of epicatechin and 10 mumol of epicatechin and/or catechin. Under similar conditions, catechin and epicatechin fail to inhibit the hydroperoxide (HPx) response to TPA whereas the procyanidin dimers inhibit this response by almost 40%. At a dose of 10 mumol, the epicatechin trimer also inhibits TPA-induced ODC activity and HPx production to a greater degree than 10-30 mumol of epicatechin. However, these various treatments with monomeric, dimeric, and trimeric procyanidins do not differ significantly in their abilities to inhibit TPA-stimulated DNA synthesis. These results suggest that some of the antitumor-promoting effects of procyanidins might increase at the biflavanoid and triflavanoid levels.