Abstract 1485: Inhibition of nuclear β-catenin translocation by a chemopreventive dietary triterpene lupeol

Abstract

Abstract Aberrant Wnt/β-catenin signaling is observed in several malignancies and these subsets are mostly associated with very poor prognosis. Thus, agents that attenuate the translocation of β-catenin from the cytoplasm to the nucleus could be extremely valuable for tumors that exhibit constitutive Wnt/β-catenin signaling. We previously demonstrated that lupeol, a dietary triterpene, preferentially inhibits viability and clonogenic potential of human melanoma Mel 928 and Mel 1241 cells that exhibit constitutive Wnt/β-catenin signaling compared to Mel 1011 cells which lack constitutive Wnt activation. Here, we hypothesized that lupeol inhibits nuclear-cytoplasmic localization of β-catenin thereby suppressing Wnt signaling. To test our hypothesis, first we probed the cytosolic and nuclear fractions of lupeol treated melanoma cells using enzyme based assays and observed an increase in cytosolic with a concomitant increase in nuclear β-catenin levels indicating that it inhibits nuclear β-catenin localization; thereby modulating Wnt signaling and inhibiting downstream Wnt targets. We then probed lupeol treated melanoma cells for β-catenin using immunofluorescence techniques and observed that it decreases the translocation of β-catenin into the nucleus in Mel 928 and Mel 1241 cells that harbor constitutive Wnt/β-catenin signaling. No change in the rate of translocation was observed in Mel 1011 cells that do not exhibit constitutive Wnt activation. Next, we wanted to demonstrate if these findings could be extended in the subset of colorectal cancer cells of varied Wnt/β-catenin signaling status. We treated colorectal cancer cells with lupeol (20-40 µm) and employed immunofluorescence imaging to study the localization of β-catenin. Lupeol was found to inhibit the nuclear localization of β-catenin in DLD 1 and HCT 116 cells which harbor constitutive Wnt/β-catenin signaling while producing no such effects in RKO cells that lack constitutive Wnt/β-catenin signaling. We then determined the transcriptional activity and expression of Wnt target genes in colorectal cancer cells (DLD 1, HCT 116) treated with lupeol. We observed a decrease in β-catenin/TCF4 transcriptional activity as indicated by TOPFlash luciferase activity and inhibition of downstream Wnt targets viz. Axin2 and TCF1 as assessed by immunoblotting. Next, using dominant-negative TCF4, we inhibited Wnt signaling in HCT 116 colorectal cancer cells harboring constitutive Wnt activation and found that in these cells the effects of lupeol on cytoplasmic-nuclear localization of β-catenin were lost. Taking together, our data strongly advocates the efficacy of lupeol against melanoma and colorectal cancer cells that harbor constitutive Wnt signaling. Validation of these cell culture data to animal model systems could pave the way for developing new strategies for chemoprevention of cancers harboring constitutive Wnt/β-catenin signaling. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1485.