Long-term effect of Wnt/β-catenin small molecule inhibitor CWP232291 on intestinal carcinogenesis in novel GEM model deficient in Smad4 and p53.

Abstract

594 Background: We have established and characterized a novel genetically-engineered mouse (GEM) model for colorectal cancer by performing intestinal epithelium-specific knockout of Smad4 and p53 under Villin promoter. CWP232291 (JW Pharmaceutical Corporation, Seoul, Korea) is a potent Wnt/ β-catenin small molecule inhibitor currently tested in several Phase I trials. We evaluated chemopreventive efficacy of long-term CWP232291 treatment for intestinal carcinogenesis in our novel GEM model. Methods: Three-week-old GEM mice were intraperitoneally injected BIW for 17 weeks with 100 mg/kg CWP232291 (n = 19) or vehicle normal saline (n = 27) and then sacrificed. Results: All Villin-Cre;Smad4F/F;Trp53F/F mice developed multiple spontaneous intestinal tumors with the high proliferative activity and the activation of Wnt signaling pathway (median adenocarcinoma-free survival, 5.4 months; incidence of adenocarcinoma, 75.9 %). Whereas, Villin-Cre;Smad4F/F or Villin-Cre;Trp53F/F micedidn’t develop any intestinal tumors until 20 weeks. A histological assessment showed that tumor-bearing mice were significantly reduced after CWP232291 treatment (CWP232291, 50.0% vs vehicle, 84.0%; P < 0.01). More importantly, CWP232291 showed significant decrease in the incidence of malignant tumors (CWP232291, 37.5% vs vehicle, 78.3%; P < 0.05). In addition, tumor multiplicity is significantly decreased in CWP232291-treated mice (CWP232291, 1.2 ± 0.4 vs vehicle, 2.8 ± 0.4, P < 0.01). Immunohistochemistry showed that CWP232291 treatment reduced β-catenin, myc and cyclin D1 levels in tumors. In-vivo limiting dilution assay using primary mouse cancer cells showed that the frequency of cancer stem cells (CSCs) was 1/3,223 for the untreated group and 1/48,069 for the CWP291-treated group (p < 0.001). Conclusions: These results showed that CWP232291 inhibited Wnt signaling and reduced the CSCs through down-regulation of β-catenin and Wnt target genes and exerted a chemopreventive effect in our GEM model. This suggests that CWP232291 may be a promising candidate for colorecal cancer chemoprevention and maintenance.