Studies on the carcinogenic mechanisms of histamine N-methyltransferase-mediated breast cancer formation

Previous results from our laboratory have demonstrated that mRNA expression of the histamine N-methyltransferase (HNMT) in tumor tissues (n = 240) from breast cancer patients is higher than in normal tissues （ tumor/normal > 16 fold ） . However, the role HNMT-mediated breast cancer formation is still uncertain. Here, we outline research goals that we hope to achieve within the next three years to help answer these questions. Year-1 study: Demographic study on the expression level of HNMT in breast tumor tissues In the year-1 proposal, we first want to investigate whether the higher expression level of HNMT was occurred in advanced stage breast cancer cells. Immunohistochemical staining and confocal microscopic observations for the HNMT expressions in the tumor tissue will be performed. Laser capture microscopic (LCM) dissected human breast cancer cells will be isolated for detection of the expression levels of HNMT mRNA. We will identify the regions regulated by transcription factors on the HNMT gene using the luciferase promoter reporter system and identification of the transcription factors involved in regulating HNMT gene expression using chromatin immunoprecipitation (CHIP). Year-2 study: In vitro studies of the HNMT’s role in regulating oncogenic signals in breast cancer tumorigenesis. In the year-2 proposal, the HNMT SiRNA knock-down and adenoviral Tet-off overexpression cells lines will further be established in our lab and adapted as a research model cells for ask on the synergistic effects of AKT-mediated HNMT activity in breast cancer carcinogenesis. Our preliminary data further demonstrated that HNMT protein will be translocated to cell nuclear and binding to Her-2 gene promoter through EGF stimulation. Such results pinpoint the specific role of HNMT on the breast cancer formation. Year-3 study: In vivo studies of the HNMT's role in regulating the oncogenic signals in breast cancer tumorigenesis. In the year-3 proposal, several specific aims will be completed. 1.Establish tumor models of HNMT knock-down and HNMT over-expression in vivo. 2. Study HNMT oncogenic signal transduction in xenograft models using either siRNA mediated repression of HNMT expression, or Tet-off mediated HNMT over-expression. 3. Characterize the in vivo oncogenic signal that stimulates the nuclear translocation of both estrogen and nicotine, which, leads to cancer cell proliferation. We further want to 1.Study the anti-tumor effects of natural HNMT antagonists in vivo. 2. Search for HNMT selective inhibitors with anti-tumor effects. 3. Assess whether the anti-tumor effects of these natural antagonists are mediated by their ability to modify expression levels of HNMT in a SCID mouse model. These results should assist the development of new therapeutic strategies for breast cancer patients.