Response of Apc^min and A33^ΔNβ-cat mutant mice to treatment with tea, sulindac, and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)

Abstract

There is growing interest in the potential health benefits of tea, and a recent report described the potent antimutagenic activity of white tea in comparison with green tea against several heterocyclic amines, including 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) [Mutat. Res. 495 (2001) 61]. We compared the inhibitory effects of white and green teas with sulindac, a nonsteroidal anti-inflammatory agent, in two different mouse models of intestinal tumorigenesis. In the Apc^min mouse, white and green teas given at human-relevant concentrations (1.5% w/v, 2-min brew), and sulindac (80 ppm in the drinking water), each suppressed polyp formation by ∼50%, and the combination of white tea plus sulindac was more effective than either treatment alone (P=0.05). Mice expressing an N-terminally truncated, oncogenic version of β-catenin (A33^ΔNβ-cat mutant mice) developed colonic aberrant crypt foci (ACF) spontaneously, but PhIP treatment increased the incidence and number of ACF per colon. In the normal-looking intestinal mucosa of Apc^min and A33^ΔNβ-cat mice, white tea plus sulindac treatment markedly attenuated the expression of β-catenin protein, and this was recapitulated in vitro in cells transiently transfected with β-catenin plus Tcf-4 and treated with tea or the major tea polyphenol epigallocatechin-3-gallate (EGCG). Expression of a β-catenin/Tcf reporter was inhibited by EGCG in the transfected cells, and the β-catenin/Tcf target genes cyclin D1 and c-jun were downregulated in vivo by tea plus sulindac treatment. Collectively, the data support a chemopreventive role for tea and sulindac against intermediate and late stages of colon cancer, via effects on the β-catenin/Tcf signaling pathway.

Introduction

The genetic model of colorectal cancer proposes that an accumulation of “genetic hits” in such genes as K-RAS, DCC, and P53 is required in order to produce late stage tumors [1]. According to this model, mutations occur at an early stage in APC or CTNNB1 genes, causing accumulation of β-catenin protein and activation of β-catenin/Tcf target genes, such as cyclin D1 and c-jun (see [2] for a review). Because diet is an important risk factor for colorectal cancer development [3], we have examined the role of dietary constituents in modulating the pathways regulated by Apc and Ctnnb1 genes in animal models. In this report, we describe preliminary findings using two different mouse models of intestinal neoplasia after treatment with tea, sulindac, or 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP).

The first study used the Apc^min mouse. Apc^min mice are genetically predisposed to intestinal polyp formation due to impaired function of the Apc protein resulting from a mutation that introduces a premature translational stop codon at amino acid 850, and these mice respond well to tumor suppression by nonsteroidal anti-inflammatory drugs (NSAIDs), such as sulindac [4]. As part of our continuing effort to characterize the possible role of white and green teas in preventing or reducing the risk of cancers in the GI tract [5], [6], [7], we examined the suppressing effects of tea and sulindac, alone and in combination, in the Apc^min mouse.

We also describe here the preliminary characterization of a new mouse model, designated A33^ΔNβ-cat, which expresses in the intestinal epithelium an oncogenic form of β-catenin as a “knock-in” transgene which lacks the amino-terminal amino acids that constitute the glycogen synthase kinase-3β (GSK-3β) regulatory region. We found that in Apc^min and A33^ΔNβ-cat mice, tea and sulindac caused marked reductions in β-catenin expression in the intestinal mucosa and altered the expression of β-catenin/Tcf target genes, leading to suppression of tumorigenesis.