Critical tumor suppressor function mediated by epithelial Mig-6 in endometrial 1 cancer. 2

26 Endometrial cancer is preceded by endometrial hyperplasia, unopposed estrogen 27 exposure and genetic alterations, but the precise causes of endometrial cancer remain 28 uncertain. Mig-6 , mainly known as a negative regulator of the EGF receptor, is an 29 important mediator of progesterone signaling in the uterus, where it mediates tumor 30 suppression by modulating endometrial stromal-epithelial communications. In this 31 study, we investigated the function of Mig-6 in the uterine epithelium using a tissue- 32 specific gene knockout strategy, in which floxed Mig-6 ( Mig-6 f/f ) mice were crossed to 33 Wnt7a-Cre mice ( Wnt7a cre+ Mig-6 f/f ). Wnt7a cre+ Mig-6 f/f mice developed endometrial 34 hyperplasia and estrogen-dependent endometrial cancer, exhibiting increased 35 proliferation in epithelial cells as well as apoptosis in sub-epithelial stromal cells. We 36 documented increased expression of NOTCH1 and BIRC3 in epithelial cells of 37 Wnt7a cre+ Mig-6 f/f mice and decreased expression of the progesterone receptor (PR) in 38 stromal cells. Progesterone therapy controls endometrial growth and prevents 39 endometrial cancer, but the effectiveness of progesterone as a treatment for women with 40 endometrial cancer is less clear. We noted that the hyperplasic phenotype of Wnt7a cre+ 41 Mig-6 f/f mice was prevented by progesterone treatment, whereas this treatment had no 42 effect in PR cre/+ Mig-6 f/f mice where Mig-6 was deleted in both the epithelial and Mig-6 the the the presence of progesterone and that Mig-6 regulates PTEN/PI3K/AKT signaling In searching for relevance of the findings from the animal study to human endometrial cancer, we found a significantly lower MIG-6 expression in human endometrioid carcinoma cells compared to normal uterine cells, suggesting a similar Mig-6 regulated system present in the human uterus (20). Tumor suppressor function of Mig-6 90 coordinates endometrial stromal-epithelial communication. The expression of Mig-6 in these cellular compartments is under tight temporal and endocrine control. However, the 92 function of Mig-6 in endometrial cancer has remained elusive. In this study, we used endometrial epithelial cell specific Mig-6 knockout ( Wnt7a cre+ Mig-6 f/f ) mice to assess the role of epithelial Mig-6 in steroid hormone 95 responsiveness and tumorigenesis (18, 22). We observed that Wnt7a cre+ Mig-6 f/f mice 96 display normal P4 attenuation of E2-mediated uterine hypertrophy. However, Wnt7a cre+ 97 Mig-6 f/f mice develop endometrial hyperplasia as well as estrogen-induced endometrial cancer. The development of endometrial hyperplasia in Wnt7a cre+ Mig-6 f/f mice is prevented by P4 treatment. These data suggest that P4-induced stromal Mig-6 prevents 100 hyperplasia seen in Wnt7a cre+ Mig-6 f/f mice by regulating estrogen signaling. mRNA (ER target Mig-6 , and 18S (for primers cDNA RNA Transcriptase PCR All real-time PCR was by using five independent RNA sets. The relative expression of each transcript was normalized to 18S rRNA using ABI rRNA control reagents. Statistical analyses were performed using Student’s t test and one-way ANOVA. Differences between multiple groups were determined by Tukey’s post hoc multiple comparisons test. cre/+ These results therapy. It is sufficient to play as a tumor suppressor and/or mediator of PR-P4 393 signaling, even though expression of Mig-6 in Wnt7a cre+ Mig-6 f/f mice is lower than control mice after P4 treatment. Our results suggest that activated stromal Mig-6 can regulate proliferation and apoptosis via regulating ER α activity in epithelium, can contribute to the prevention of endometrial hyperplasia and that epithelial Mig-6 is a critical tumor suppressor involved in P4 mediated protection against the development endometrial cancer. These results suggest that epithelial Mig-6 is critical for a tumor suppressor function in endometrial cancer. this hyperplastic phenotype ER α The Wnt7a cre+ Mig-6 f/f model is useful for studying new targets during cancer progression and can be exploited therapeutically in order to identify new therapies for the prevention and treatment of endometrial cancer. Determining the role of Mig-6 in stromal-epithelial cross talk will be critical in understanding the role of steroid hormone signaling in endometrial function and dysfunction associated with infertility and endometriosis and in developing therapy for both of these common uterine diseases. analysis of α target genes ( Muc-1 , Clca3 , Ltf, Birc1a and Birc1b ) was performed on uteri of control mice, PR mice and after vehicle or treatment. The results represent the relative expression of transcripts