ANTIPROLIFERATIVE ACTION OF ESTROGEN RECEPTOR β AND HES-1 IN BREAST CANCER

Breast cancer is the most common malignancy among Swedish women. Although the mechanism behind the tumorigenesis remains unclear, estrogen receptor α (ERα) plays an important role in the progression of breast cancer and is regarded as a target for endocrine therapy. In this thesis, focus is on the second estrogen receptor, ERβ and its function in breast cancer. In addition, the significance of the transcription factors Hes-1 and Hes-6 in breast cancer and their relation to ERα has been studied. By using T47D breast cancer cells with inducible ERβ expression, the role of ERβ has been characterised with respect to proliferation and cell-cycle regulation. In contrast to ERα, expression of ERβ inhibited the proliferation of 17β-estradiol (E2) treated breast cancer cells and caused significantly changed levels of cell-cycle regulators. In response to ERβ expression, the levels of the Cdk2-activating phosphatase Cdc25A as well as cyclin E and E2F1 were reduced with a subsequent decrease of the Cdk2activity. Moreover, expression of ERβ reduced the number of tumor associated blood vessels as well as tumor volume in a mouse xenograft model. In addition to the anti-tumorigenic effects, expression of ERβ reduced the levels of secreted growth factors in vitro as well as in vivo. The transcriptional repressor Hes-1 has been described as an essential factor during embryonic development. However, Hes-1 is also important in breast cancer cells where it inhibits proliferation. Downregulation of Hes-1 by ERα is a crucial step in E2 stimulated proliferation of breast cancer cells. We expressed Hes-1 in breast cancer cells to study the mechanism behind its antiproliferative properties upon E2 treatment. Hes-1 expression induced a G1 cell-cycle phase arrest and a concomitant reduction of the E2F1-level. By real-time quantitative PCR and electrophilic mobility shift assay, we can conclude that Hes-1 inhibits E2F1 at the promoter level. Hes-6 is an inhibitor of Hes-1 and has been associated with tumorigenesis and metastasis. When Hes-6 was expressed in T47D breast cancer cells, proliferation was increased as well as tumor growth in immunodeficient mice. Furthermore, E2F1 was identified as an important target gene, induced by Hes-6 in breast cancer cells. In conclusion, these studies have significantly contributed to the knowledge of estrogen receptor function in breast cancer as well as to elucidate important roles of Hes-1 and Hes-6 in estrogen signalling. LIST OF PUBLICATIONS I. Anders Ström, Johan Hartman, James S Foster, Jay Wimalasena and JanÅke Gustafsson. Estrogen receptor β inhibits 17β-estradiol-stimulated proliferation of the breast cancer cell line T47D. Proc Natl Acad Sci U S A. 2004 Feb 10;101(6):1566-71 II. Johan Hartman, Karolina Lindberg, Andrea Morani, José Inzunza, Anders Ström and Jan-Åke Gustafsson. Estrogen receptor β inhibits angiogenesis and growth of T47D breast cancer xenografts. Cancer Res. 2006 Dec 1;66(23):11207-13. III. Johan Hartman, Patrick Müller, James S Foster, Jay Wimalasena, JanÅke Gustafsson and Anders Ström. HES-1 inhibits 17β-estradiol and heregulin-β1-mediated upregulation of E2F-1. Oncogene. 2004 Nov 18;23(54):8826-33 IV. Johan Hartman, Eric W. -F. Lam, Jan-Åke Gustafsson and Anders Ström. Hes-6, an inhibitor of Hes-1, is regulated by 17β-estradiol and increases the malignant behaviour of breast cancer cells. Submitted manuscript.