Abstract 2072: Ormeloxifene attenuates wnt/β-catenin signaling in colon cancer cells by modulation of PKD1 and glycolytic pathways

Abstract

Abstract Background: Colorectal cancer is the third most commonly diagnosed cancer and third leading cause of cancer-related deaths. Aberrant Wnt/β-catenin signaling is implicated in development and progression of colon cancers. Therefore, modulation of this master regulatory signaling is essential for the prevention and treatment of colon cancer. Protein Kinase D1 (PKD1) which is downregulated in colon cancer, regulates β-catenin transcriptional activity by its phosphorylation. Therefore, drug mediated upregulation of PKD1 may have high clinical significance for the prevention/treatment of Wnt/β-catenin signaling induced cancers. We have identified, ormeloxifene, a synthetic non-steroidal selective estrogen receptor modulator (SERM), as a novel PKD1 modulator. Herein, we have investigated the effect of ormeloxifene on tumorigenic phenotypes of colon cancer cells and its effect on PKD1 and glycolytic pathways mediated oncogenic β-catenin signaling. Materials and Methods: We have used SW480-PKD1-GFP and SW480-GFP colon cancer cells (SW480 has a mutated APC gene leading to stabilization of β-catenin due to lack of degradation) to investigate the effect of PKD1 on β-catenin activity and xenograft mouse model. SW480, SW48, SW620 and Lovo cell lines have been used to observe the effect of ormeloxifene (5-20 μM) on various cellular characteristics, including cell proliferation, clonogenic potential, motility and aggregation. β-catenin/T-cell factor 4 (TCF4) transcription activity was measured by Dual-Glo luciferase reporter assays. The expression and subcellular localization of proteins was determined by immunoblotting and confocal microscopy. Chemokine (C-C) motif ligand 2 (CCL2) and glucose assay were performed using ELISA technique. Results: Ormeloxifene treatment restores PKD1 expression, thereby, attenuates β-catenin transcriptional activity. We have also observed that ormeloxifene treatment effectively inhibits CCL2 expression, glucose uptake, glucose metabolism and Glut1 receptor expression in cancer cells. Ormeloxifene (5-20μM) inhibits cell proliferation and motility of colon cancer cells. Additionally, ormeloxifene effectively inhibited CCL2 mediated cellular migration of colon cancer cells. Furthermore, Western blot results depict that PKD1 inhibits TCF4 activity in colon cancer cells. SW480 PKD1 overexpressing cells clearly delayed tumor formation in nude mice, compared to control cells. In addition, SW480-PKD1-GFP cells showed less hypoxia (Glut1) and increased Vasculature (CD31) in tumors. Conclusion: Our findings demonstrate that ormeloxifene treatment effectively attenuates β-catenin mediated oncogenic signaling and tumorigenic phenotypes of colon cancer cells via upregulation of PKD1 and suppression of glycolytic pathways. Thus it can be used as a potential new preventive/treatment modality for colon cancer. Citation Format: Aditya Ganju, Rishi Gara, Sonam Kumari, Man Mohan Singh, Subhash Chauhan, Meena Jaggi. Ormeloxifene attenuates wnt/β-catenin signaling in colon cancer cells by modulation of PKD1 and glycolytic pathways. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2072. doi:10.1158/1538-7445.AM2015-2072