Transformation of Human Endometrial Epithelial Cells

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DESCRIPTION (provided by applicant): Endometrial cancer is the most abundant cancer of the genital tract in American women and ranks fourth in overall incidence among cancers in women. Therefore, discoveries that contribute significantly to our understanding of this disease and hat provide new methods with which novel therapies may be discovered, would have a high impact on cancer in women. In this application we propose a pilot study to culture normal endometrial epithelial cells, which have very limited longevity in culture, and derive cultures from them that have a greatly increased longevity and capacity for proliferation in vitro. In the past we did with transfers of SV40 large T antigen but the cells that result are notably abnormal, behaving like severely hyperplastic to early malignant endometrial cells. Furthermore, we used an agent that appears to have no role in endometrial cancer development. In this pilot study we seek to develop propagable human endometrial epithelial cells that have been genetically modified in a manner that better approximates changes occurring during endometrial carcinogenesis. We seek to develop cells with an enhanced capacity for proliferation. To accomplish this we propose the following three specific aims 1) to determine the best genetic transfer agents and techniques applicable to primary cultures of normal epithelial endometrial cells. 2) to test the best way to impair PTEN function in these cells, recreating a recognized early event in endometrial carcinogenesis, and determine whether this change results in cells with increased capacity for cell proliferation. 3) to discover further mutations specifically relevant to development of endometrial cancer using various genetic libraries as "virtual mutagens." These studies will assess increased capacity for cell proliferation and will employ novel selection protocols that will mimic the selection pressures exerted by basement membrane and stromal cells in the uterine tissue. By identifying the alterations in cell physiology produced by this process, we hope to clarify the mechanisms that allow carcinogenic endometrial epithelial cells to escape the homeostatic controls on cell proliferation that normally operate in within endometrial tissue.