The overall goal of this project is to use immunological and molecular techniques to identify and study the cell surface molecules involved in breast tumor interactions with stromal tissue and to determine the importance of these molecules in tumor metastasis. The cell surface transmembrane hyaluronate receptor CD44 has been shown to be important for lymphocyte adhesion and activation. Recent studies have identified multiple isoforms of CD44, at least one of which confers the ability to metastasize to a normally non-metastatic cell line. In this project, the isoforms of CD44 expressed in human breast cancer cell lines, and in primary and metastatic human breast cancers will be identified. Local expression of CD44 isoforms within cancers will be identified and correlated with patient outcome. The long range goal of this work is to improve prognostic accuracy and to devise novel strategies to inhibit breast cancer metastasis. The individual specific aims in this project are: 1) Oligonucleotide probes specific for each isoform of human CD44 will be developed. A panel of polyclonal antisera and monoclonal antibodies specifically reacting with each isoform will be developed. 2) The expression of CD44 isoforms in normal breast tissue and in primary and metastatic breast carcinoma patient specimens will be characterized using immunohistochemical, in situ hybridization, polymerase chain reaction, and. northern and western blot assays. The relationship between expression of specific isoforms and patient prognosis will be analyzed for a large panel of breast cancer patients followed by the Duke University Cancer Center Tumor Registry. 3) Breast carcinoma cell lines stably expressing specific CD44 isoforms will be developed, and correlations between isoform expression and ability to bind hyaluronate and to interact with lymphocytes will be made. Epithelial cell and lymphocyte interactions will be studied as a function of the CD44 isoform expressed by each cell type. 4) In vitro and in vivo model systems will be used to assess the metastatic behavior of breast cancer cells expressing defined CD44 isoforms. The effects of CD44 peptides and monoclonal antibodies against defined CD44 epitopes on the biologic behavior of the breast cancer cell lines will be determined. These studies should provide much new information on factors affecting the behavior of breast carcinomas in humans.
