Abstract 4809: Bromo-ormeloxifene inhibits epithelial mesenchymal transition via targeting β-catenin signaling pathways in cervical cancer cells

Abstract

Abstract Background: Cervical cancer (CxC) is a leading cause of mortality and morbidity among women worldwide. Current chemotherapeutic agents for CxC have shown systemic toxicity in CxC patients. Ormeloxifene (ORM) is a non-toxic and non-steroidal drug with well-defined pharmacokinetic and pharmacodynamic properties in humans. Studies have shown its anti-cancer potential in various pre-clinical mouse models. Here, we have synthesized and characterized a novel analogue of ormeloxifene, Bromo-ormeloxifene (Br-ORM), which showed more therapeutic efficacy against CxC in vitro and in vivo model systems. Methodology: The effect of Br-ORM on CxC cells (CaSki and SiHa) growth and proliferation was determined by colony formation and MTS assays. Molecular docking of Br-ORM with β-catenin was done by AutoDock4 software. Effect of Br-ORM on the expression of epithelial-to mesenchymal (EMT) markers (N-cadherin, slug, snail), MMPs (MMP2 and MMP3) and miR-200a was analyzed by Western blot and qPCR analyses respectively. Apoptosis analysis was done by Annexin-V staining kit. Effect of Br-ORM on β-catenin cellular localization in CxC cells was analyzed by immunofluorescence analysis. The anti-tumor efficacy of Br-ORM was investigated in orthotopic xenograft mouse model of CxC. Results: Br-ORM (10-20 µM) effectively inhibited growth and proliferation of CxC cells in a dose and time-dependent manner as compared to ORM. Br-ORM efficiently suppressed metastatic phenotypes of CxC cells as determined by significant (P<0.05) decrease in invasion and migration potential of CxC cells. Moreover, Br-ORM showed increased apoptosis, which was observed by enhanced Annexin-V staining and PARP protein cleavage. Br-ORM markedly reduced the EMT process as evident by repression of N-cadherin, slug, snail, MMPs (MMP2 and MMP3) and β-catenin/TCF-4 transcriptional activity. Br-ORM potently reduced the translocation of β-catenin in the nucleus. Bioinformatic analysis revealed that Br-ORM proficiently binds into active site of β-catenin with a minimum energy -7.6 kcal/mol. Br-ORM treatment replenished the expression of miR-200a, which directly targets β-catenin in CxC cells. Br-ORM treatment (250 µg/mouse, three times a week) significantly (P<0.01) regressed the cervical tumor growth in orthotopic xenograft mouse model. Similar molecular effects of Br-ORM were observed in excised tumor tissues. Conclusion: These results suggest that Br-ORM inhibits the metastatic phenotypes of CxC cells via targeting β-catenin signaling pathway. Br-ORM could be used as a novel therapeutic modality for the treatment of CxC. Citation Format: Mohammed Sikander, Shabnam Malik, Bilal B. Hafeez, Sonam Kumari, Sheema Khan, John Apraku, Hassan Mandil, Andrew E. Massey, Aditya Ganju, Parvez Khan, Fathi T. Halaweish, Subhash C. Chauhan, Meena Jaggi. Bromo-ormeloxifene inhibits epithelial mesenchymal transition via targeting β-catenin signaling pathways in cervical cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4809.