Epidemiological evidence suggests tea (Camellia sinensis L.) has chemopreventive effects against various tumors. Green tea contains many polyphenols, including epigallocatechin-3 gallate (EGCG), which possess anti-oxidant qualities. Reduction of chemically induced mammary gland carcinogenesis by green tea in a carcinogen-induced rat model has been suggested previously, but the results reported were not statistically significant. Here we have tested the effects of green tea on mammary tumorigenesis using the 7,12-dimethylbenz(a)anthracene (DMBA) Sprague-Dawley (S-D) rat model. We report that green tea significantly increased mean latency to first tumor, and reduced tumor burden and number of invasive tumors per tumor-bearing animal; although, it did not affect tumor number in the female rats. Furthermore, we show that proliferation and/or viability of cultured Hs578T and MDA-MB-231 estrogen receptor-negative breast cancer cell lines was reduced by EGCG treatment. Similar negative effects on proliferation were observed with the DMBA-transformed D3-1 cell line. Growth inhibition of Hs578T cells correlated with induction of p27(Kip1) cyclin-dependent kinase inhibitor (CKI) expression. Hs578T cells expressing elevated levels of p27(Kip1) protein due to stable ectopic expression displayed increased G1 arrest. Thus, green tea had significant chemopreventive effects on carcinogen-induced mammary tumorigenesis in female S-D rats. In culture, inhibition of human breast cancer cell proliferation by EGCG was mediated in part via induction of the p27(Kip1) CKI.

NF-B/Rel is a structurally and evolutionary conserved family of transcription factors distinguished by the presence of an N-terminal 300-amino-acid region, termed the Rel homology domain. The Rel homology domain is responsible for DNA binding, dimerization, nuclear translocation, and binding of Rel factors to the IB inhibitory proteins (reviewed in reference 25). Mammals express five NF-B members, of which RelB, c-Rel, and p65 (RelA) are synthesized as mature products and contain a C-terminal transactivation domain. In contrast, p50 and p52 are synthesized as longer precursors that have C-terminal ankyrin repeats and can serve as inhibitory molecules. The p105 and p100 precursor proteins require proteolytic processing to produce the mature p50 and p52 subunits, respectively, which lack a transactivation domain (46). In most untransformed cells other than B lymphocytes, NF-B complexes are sequestered in the cytoplasm bound to specific inhibitory proteins, termed IBs (2), which have been found to display specificity of interaction (reviewed in references 54 and 59). For example, IB-␣ most strongly interacts with p65 and c-Rel and only weakly with RelB, p50, and p52. Activation of NF-B can occur via multiple pathways. Stimuli such as tumor necrosis factor (TNF) and interleukin-1 lead to induction of p65/p50 complexes with IB-␣ via the canonical pathway involving activation of the IKK complex, consisting of the kinases IKK␣ and IKK␤ (33). In particular, activation of the IKK␤ kinase leads to phosphorylation of IB-␣ on S32/36 and its rapid proteasome-mediated degradation (7, 12), allowing for translocation of the free NF-B to the nucleus (reviewed in reference 59). Thus, an IB-␣ protein with the S32/36A mutation, which is resistant to phosphorylation and subsequent degradation, is termed superrepressor (SR)-IB-␣. A recently reported alternative pathway involves induction of RelB/p52 complexes upon stimulation by such TNF family receptors as lymphotoxin ␤ and CD40 (15,34,62). This signaling leads to activation of the NF-B-inducing kinase (NIK) and subsequent IKK␣-mediated phosphorylation of the p100 component of RelB/p100 cytoplasmic complexes (44, 61), resulting in its processing to p52. Studies by Bravo and coworkers (18) indicate that the resulting RelB/p52 complexes do not interact well with IB-␣ and, thus, are largely free to migrate to the nucleus.NF-B factors have been implicated in the pathogenesis of breast cancer and in development of the normal mammary gland. We and others have demonstrated aberrant constitutive activation of NF-B factors in human and rodent breast cancers (14,29,35,41,51). High levels of nuclear NF-B were found in the majority of primary human and rodent breast tumor tissue samples, breast cancer cell lines, and carcinogentransformed mammary epithelial cells (14, 29). Inhibition of