Abstract 3847: Lupeol, a novel inhibitor of Wnt/β-catenin signaling: Implications in colon cancer therapy

Abstract

Abstract Introduction: Wnt/β-catenin signaling pathway which regulates a variety of cellular processes has been reported to play a critical role in the pathogenesis of human colon cancer and found in almost 90% of the colon cancer patients. Recent studies from our laboratory and others showed that Lupeol a natural triterpene acts as a potent and selective inhibitor of Wnt signaling in prostate and melanoma tumor cells (Saleem et al, Carcinogenesis, 30: 801-17, 2009). Keeping in view these reports, we hypothesized that Lupeol will inhibit the tumorigenic growth of colon cancer cells (exhibiting aberrant Wnt/α signaling) under in vitro and in vivo conditions. Objectives: We investigated the effect of Lupeol on growth of human normal colon and carcinoma cells (exhibiting varied Wnt profile) and determined if Lupeol could enhances the efficacy of Sulindac, (Wnt signaling inhibitor and chemotherapeutic agent) that is used in clinics to treat human colon cancer. Methods: To test our hypothesis we employed in vitro and in vivo strategies. Firstly, normal human colon epithelial cells (FHC) and colon carcinoma cell lines (SW-480, HCT-116, HT-29) treated with either Lupeol (10-50 μM), or Sulindac (100-200 μM), or combination (Lupeol + Sulindac) were tested for growth, proliferation, clonogenicity and surrogate biomarkers of Wnt signaling by employing 3[H]thymidine uptake, MTT, Soft-agar colony, FACS, immunoblot and luciferase-reporter assays. Secondly, we investigated whether Lupeol is bio-available to mice after oral and intraperitoneal (i.p.) administration by using mass spectrometry. Next, we treated athymic mice bearing colon tumors-derived from HT-29 cells (highly chemoresistant among all colon cells) with either Lupeol (40 mg/kg) or Sulindac (20 mg/kg) or combination and measured tumor growth. In vivo proliferation and biochemical analysis of harvested tumors for surrogate biomarkers of apoptosis, proliferation and Wnt signaling were measured by employing BrdU uptake, immunoblot and immunhistochemical analysis. Results: Lupeol treatment was observed to (i) inhibit the proliferation of colon cancer cells (exhibiting activated Wnt) while sparing normal cells, (ii) induce G0/G1 arrest and apoptosis, (iii) decrease cytoslic as well as nuclear β-catenin levels (iv) increase degradation of cytosolic β-catenin, (iv) decrease TCF-transcriptional activity and (v) decrease the expression of Wnt target genes. Lupeol was observed to bioavailable in mice after oral and i.p. administration. Lupeol significantly inhibited the tumorigenicity of colon tumor cells in a xenograft mouse model. Most importantly, Lupeol treatment significantly sensitized highly chemoresistant colon cancer cells to Sulindac therapy, in vitro and in vivo. Conclusion: Lupeol alone or as an adjuvant to current therapies could be developed as an agent to treat subtype of human colon cancer exhibiting constitutive activation of Wnt signaling. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3847. doi:1538-7445.AM2012-3847