TISSUE FACTOR MESSENGER RNA, ANTIGEN, AND ACTIVITY

点击翻译按钮获取中文摘要

The major objectives of this application are to evaluate the synthesis of tissue factor (TF) and the expression of TF procoagulant activity in cancer cells in order to determine the role of TF in malignancy-associated coagulapathies and the role of TF in the proliferative capacity of tumor cells. The potential benefit of inhibition of TF or the TF-dependent coagulation system for therapeutic benefit and diagnosis of early metastatic foci will be evaluated. The specific aims and methods we will use to achieve these objectives are as follows: 1) TF messenger RNA (mRNA), TF antigen, and TF activity will be determined in various human tumor cell lines in vitro. This will provide baseline information regarding TF gene expression and biological activity. This goal will be accomplished by RNA isolation and Northern blot analysis using cDNA probes which have previously been developed; monoclonal antibody will be used in an ELISA assay to detect TF antigen; TF procoagulant activity will be evaluated by use of a two-stage coagulation assay which will be calibrated with a sample of pure human brain TF. 2) Various agents, such as growth factors, chemotherapeutic agents, or mediators of inflammation, which are known to stimulate or inhibit TF activity in normal cells (fibroblasts, endothelial cells, monocytes, etc.), will be evaluated for effect on TF mRNA, TF antigen, and TF activity in tumor cell lines. This information will serve as a baseline for TF response in cancer cells to various agents. 3) TF mRNA, TF antigen, and TF activity and alteration of these variables in response to various agents will be evaluated in malignant cells originating from the same cell type as the cell lines, but obtained from patients. 4) TF mRNA, TF antigen, and TF activity of tumor cells will be evaluated in vivo following injection and tumor formation in nude mice. 5) The effect on TF mRNA, TF antigen, and TF activity of various agents that inhibit or stimulate TF will be evaluated in vivo in nude mice as well as effect of various agents on tumorigenicity and on survival. Effect on tumorigenicity will be evaluated by tumor size and by histopathologic analysis and will be correlated with diagnostic imaging of labelled TF monoclonal antibody. This project will provide correlation of TF gene expression, both transcription and translation with the appearance of biological activity in malignancies known to be either associated or not associated with increased thrombosis and correlation of TF in malignancies with increased tumorigenic potential versus those with less tumorigenic potential.