Progesterone Inhibits Endometrial Cancer Invasiveness by Inhibiting the TGFb Pathway

Increased expression of TGFb isoforms in human endometrial cancer correlates with decreased survival and poor prognosis. Progesterone has been shown to exert a chemoprotective effect against endometrial cancer, and previous animal models have suggested that these effects are accompanied by changes in TGFb. The goal of this study was to characterize the effect of progesterone on TGFb signaling pathway components and on TGFb-induced protumorigenic activities in endometrial cancer cell lines. Progesterone significantly decreased expression of three TGFb isoforms at 72 hours after treatment except for TGFb2 in HEC-1B and TGFb3 in Ishikawa cells. Progesterone treatment for 120 hours attenuated expression of the three isoforms in all cell lines. Progesterone exposure for 72 hours reduced expression of TGFb receptors in HEC-1B cells and all but TGFbR1 in Ishikawa cells. Progesterone reduced TGFbR3 expression in RL-95 cells at 72 hours, but TGFbR1 and bR2 expression levels were not affected by progesterone at any time point. SMAD2/3 and pSMAD2/3 were substantially reduced at 72 hours in all cell lines. SMAD4 expression was reduced in RL-95 cells at 24 hours and in HEC-1B and Ishikawa cells at 72 hours following progesterone treatment. Furthermore, progesterone effectively inhibited basal and TGFb1-induced cancer cell viability and invasion, which was accompanied by increased E-cadherin and decreased vimentin expression. An inhibitor of TGFbRI blocked TGFb1induced effects on cell viability and invasion and attenuated antitumor effects of progesterone. These results suggest that downregulation of TGFb signaling is a key mechanism underlying progesterone inhibition of endometrial cancer growth. Cancer Prev Res; 7(10); 1045–55. 2014 AACR. Introduction The TGFb family members are secreted cytokines that regulate a broad array of cellular responses, including proliferation, differentiation, migration, and apoptosis (1, 2). The role of TGFb in tumor biology is complex. It has tumor-suppressing activity in the early stages of carcinogenesis, whereas in the later stages of carcinogenesis, it functions as a tumor promoter (2, 3). The growth-inhibitory function of TGFb is selectively lost in advanced cancers due to mutational inactivation or dysregulated expression of various components of the TGFb signaling pathway resulting in growth, invasion, and metastasis of cancer cells (4). Human endometrial tumors contain the three TGFb isoforms in both the epithelial and stromal counterparts of the tumors, and these proteins are responsible for autocrine as well as paracrine signaling in the microenvironment of these tumors. The different TGFb isoforms can sometimes differentially activate signaling pathways in cancer cells, leading to isoform-specific effects on cellular phenotype (5, 6). Overexpression of TGFb1 has been reported in many (pancreas, breast, prostate, ovary, and endometrium) tumors and is associated with metastatic phenotypes and poor patient outcome (7, 8). TGFb1 signals are largely known as tumor promoters of cellular responses such as proliferation, survival, migration, and invasion (4). Compared with TGFb2 and TGFb3, high levels of TGFb1 are reported in endometrial cancer cell lines and these findings are corroborated with abundant TGFb1 immunoreactivity in human endometrial tumors in vivo (8). TGFb2 is expressed in a variety of cancer cell lines, including glioma, prostate, breast, lung and kidney, and endometrium (5). In endometrial cancer of the three isoforms, TGFb2 is the least expressed. Increase of TGFb2 protein levels was observed in advanced-stage endometrial tumors (9). TGFb2 promotes the survival of tumor cells by Department ofObstetrics andGynecology, UniformedServicesUniversity of the Health Sciences, Bethesda, Maryland. Division of Gynecologic Oncology, and Gynecologic Cancer Translational Research Center of Excellence,Walter ReedNationalMilitaryMedical Center, Bethesda,Maryland. Department of Obstetrics and Gynecology and Women's Health Integrated Research Center, Inova Fairfax Hospital, Falls Church, Virginia. Division of Gynecologic Oncology, North Shore University Health System, University of Chicago, Evanston, Illinois. Department of Molecular and Cell Biology, Uniformed Services University of the Health Sciences, Bethesda, Maryland. Corresponding Author: Viqar Syed, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Road, Bethesda, MD 20814. Phone: 301-295-3128; Fax: 301-295-6774; E-mail: [email protected] doi: 10.1158/1940-6207.CAPR-14-0054 2014 American Association for Cancer Research. Cancer Prevention Research www.aacrjournals.org 1045 Research. on October 18, 2017. © 2014 American Association for Cancer cancerpreventionresearch.aacrjournals.org Downloaded from Published OnlineFirst July 28, 2014; DOI: 10.1158/1940-6207.CAPR-14-0054