Abstract LB-14: Raf-1 in epithelial-mesenchymal transition: a novel therapeutic target for ovarian and endometrial cancers

Abstract

Abstract Ovarian and endometrial cancers are the 5th and 8th overall leading causes of female cancer related deaths in the US, respectively. According to the American Cancer Society, 21,990 new ovarian cancer cases and 46,470 new endometrial cancer cases were diagnosed in 2011. These cancers share a common mechanism of pathogenesis, epithelial-mesenchymal transition (EMT). EMT increases the invasiveness and progression of epithelial tumor cells into more aggressive and metastatic disease. Amplification of growth factor signaling is also common in malignancy. Epidermal growth factor receptor (EGFR) is amplified in 60-80% of ovarian and endometrial cancer and its overexpression is associated with increased invasiveness and poor prognosis. Studies have shown that specific cytokines, including transforming growth factor-β (TGFβ) and epidermal growth factor (EGF), interact with the intracellular MEK/ERK signaling cascade, a pathway involved in several malignant processes including EMT. Raf-1 is an intracellular MAPK signaling molecule that is recruited to membrane bound Ras upon activation by growth factors. Activation of Raf-1 includes phosphorylation at ser 338, which in turn activates the downstream MEK/ERK signaling cascade. Our data showed that EGF phosphorylates Raf-1 at ser 338 and ERK in endometrial cancer cells, which is inhibited by the specific Raf-1 antagonist GW5074. Little is known on the involvement of Raf-1 in the pathogenesis of ovarian or endometrial cancer. Our studies show that inhibition of Raf-1 by GW5074 significantly decreases TGFβ and EGF induced invasiveness of immortalized endometrial epithelial cells (EECs). EGF and to a lesser degree TGFβ, enhanced the survival of EECs under serum starvation. GW5074 inhibited this effect as evidenced by increased apoptosis and caspase 3/7 activity, suggesting that Raf-1 mediates EEC survival. While EGF had minimal effects on inducing ovarian cancer cell line SKOV3, the addition of GW5074 significantly reduced SKOV3 cell growth by half. Our data shows that the addition of EGF, but not TGF-β, to endometrial and ovarian epithelial cells induced EMT as evidenced by the disruption of cell-cell contact, loss of the characteristic “cobblestone” appearance of epithelial cells, increased motility and induction of the mesenchymal marker vimentin. Treatment with GW5074 of cells exposed to EGF reversed these changes. Our FACS analysis showed that EGF decreases the cell surface detection of EPCAM, an epithelial cell surface adhesion molecule, by more than half, further suggesting that EGF is involved in EMT in endometrial cancer cells. Our data suggests that multiple steps in the pathogenesis of both ovarian and endometrial cancers are mediated by Raf-1. Raf-1's involvement in inducing EMT suggests that Raf-1 may be a novel therapeutic target for ovarian and endometrial cancers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-14. doi:1538-7445.AM2012-LB-14