Abstract
Abstract Background: Metastatic disease significantly contributes to mortality in ovarian cancer. Unfortunately, the source of metastatic cells and the processes involved in the generation of metastatic ovarian cancer is not well characterized. Epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial (MET) transitions have key roles in the process of tumor formation and metastasis. In this study we tested the hypothesis that epithelial ovarian cancer stem cells (EOC stem cells) are the source of metastasis. We demonstrate that the EOC stem cells can undergo both EMT and MET to yield cells with metastatic potential in vitro and create metastatic ovarian cancer in vivo. Methods: EOC stem cells were cultured in very confluent conditions for 30d. Changes in cellular morphology were monitored using the IncuCyte video imaging system. Levels of epithelial, mesenchymal, and stem cell markers were determined using RT-qPCR, Western blot analyses, and Flow cytometry. Gene expression profile was determined using EMT gene array. Results: I.p. injection of a pure culture of EOC stem cells suspended in matrigel in nude mice generated a solid tumor mass, whereas, injection of EOC stem cells without matrigel created carcinomatosis. Molecular characterization showed that the cells forming the solid tumor maintain an epithelial phenotype whereas cells forming the carcinomatosis acquire the mesenchymal markers Vimentin and Twist-1. In vitro, EOC stem cells grown in very confluent cultures formed viable mesenchymal-like spheroid cells (mspheroid cells) by day 30. EMT array results showed that the newly formed spheroids lost epithelial markers Ck18 and Ck19 but gained mesenchymal markers, Foxc2, Slug, Twist-1 and Vimentin. Moreover, the mspheroid cells exhibited a 4-fold increase in invasion capacity and significantly higher levels of MMP2 and MMP9 compared to the EOC stem cells. Finally, mspheroid cells plated in tissue culture flask re-attached and formed a monolayer of epithelial cells loosing expression of Foxc2 and Slug, suggesting MET. These results were not observed on parallel experiments performed on mature epithelial ovarian cancer cells (mOCCs), which do not possess stemness properties. Conclusion: We showed that the in vitro generation of mspheroid cells with highly metastatic potential and the creation of carcinomatosis in vivo occur only from a pure culture of EOC stem cells and not from cultures of mOCCs. Furthermore, we demonstrate the involvement of EMT and MET in this process. These results suggest that the EOC stem cells represent an early progenitor stage in the primary tumor, which have the capacity to differentiate, acquire the ability to detach and metastasize, and revert back to an epithelial phenotype at distant sites, thereby establishing metastatic disease. Acknowledgements: This study is supported in part by the Sands foundation, Brozman foundation, and grant from the NCI (RO1CA127913) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3405. doi:10.1158/1538-7445.AM2011-3405
Published Version
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