Abstract 5156: Wnt5A enhances the malignant phenotype of lung cancer cells via non-canonical signaling

Abstract

Abstract Objective: Previously, we reported that cigarette smoke condensate dramatically increases tumorigenicity of lung cancer cells via epigenetic silencing of Dickkopf-1(Dkk-1), which encodes an inhibitor of Wnt stem cell signaling. Furthermore, we demonstrated that knock-down of Dkk-1 up-regulates Wnt5A in lung cancer cells. Expression of this non-canonical Wnt ligand correlates with adverse outcome in lung cancer patients. The present study was undertaken to ascertain if Wnt5A directly modulates the malignant phenotype of lung cancer cells. Methods: Calu-6, H358, A549, and H841 lung cancer lines were transduced with lentiviral vectors expressing Wnt5A or control sequences. Cell count, scratch, and matrigel assays were used to evaluate in-vitro proliferation, migration, and invasion of transduced cells. Similar experiments were performed with exogenous recombinant Wnt5A (rWnt5A). Nude mouse xenograft experiments were used to assess tumorigenicity of lung cancer cells transduced with Wnt5A or vector controls. Canonical and non-canonical Wnt signal pathways were evaluated using western blot, promoter-reporter, and focused quantitative RT-PCR array techniques. Results: Wnt5A transduced lung cancer cells exhibited significantly increased proliferation, migration, and invasion relative to vector controls (p<0.05; 2-tailed t-test); this phenomenon was pronounced under low serum conditions, and was observed following exposure of parental cells to rWnt5A. Relative to vector controls, Calu-6 and H358 cells constitutively expressing Wnt5A exhibited increased tumor take (p<0.05) and size of subcutaneous xenografts (p<0.05). Whereas microarray and Wnt Super Array experiments demonstrated upregulation of genes implicated in canonical as well as non-canonical Wnt signaling, western blot and promoter-reporter assays revealed that Wnt5A predominantly mediated activation of non-canonical signaling. Calu-6 and H358 cells constitutively expressing Wnt5A exhibited upregulation of the nuclear orphan receptor, ROR2, as well as activation of c-jun terminal N-kinase (Jnk) and c-jun kinase, which mediate non-canonical Wnt signaling. Consistent with these findings, rWnt5A abrogated rWnt3A-mediated activation of β-catenin signaling in lung cancer cells. Conclusion: Wnt5A enhances proliferation, migration, invasion, and tumorigenicity of cultured lung cancer cells via non-canonical pathways. Collectively, these findings warrant evaluation of inhibitors of Wnt5A-mediated signaling for lung cancer therapy. 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 5156.