ROLE OF METASTASIS GENES AT SPECIFIC STEPS OF METASTASIS

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In order for a cancer cell in a solid tumor to complete the process of establishing a metastatic lesion it must complete a number of steps whose success is likely to depend on quite different phenotypic characteristics (invasion, intravasation, transport, extravasation and growth). Although a variety of genes have been implicated in either enhancement or retardation of the formation of metastatic lesions after implantation of experimental tumors or intravenous injection of tumor cells, there has been no experimental dissection of the stages within the metastatic cascade affected by expression of individual "metastasis" genes. We have available a series of mouse mammary tumor cell lines of varying metastatic potential which were derived from a single spontaneous mouse mammary tumor by in vivo selection. We have developed techniques for determining which step in the metastatic cascade can no longer be efficiently completed by each of the cells lines. Our goal in this study is to determine whether this model system for studies of metastatic behavior can be utilized to define the specific process(es) affected by genes that enhance or inhibit metastasis. We have chosen two prototype genes, activated c-Ha-ras and NM-23. Expression of these genes has been found to correlate with high [ras] and low [NM-23] metastatic potential in a variety of tumor models. There is also good documentation indicating that introduction of either of these genes into tumor cells by transfection affects the metastatic behavior of recipient cells. Thus, our specific aims in this proposal are: 1) To transfect full-length cDNA clones of NM23 or activated Ha-ras in appropriate expression vectors into cells of high or low metastatic potential, 2) To determine the level of expression of each gene in progeny clones, 3) To assess the effect of high or low level expression of these genes on the ability of the transfected cells to complete successive steps in the metastatic cascade, and 4) To reisolate cells from different metastatic sites to confirm that any observed metastatic alterations are associated with levels of expression of these genes. Completion of these studies will both validate a test system for determining where a variety genes already known to affect metastatic behavior act and open the possibility for development of a system wherein genes not previously known to affect steps in the metastatic cascade can be identified.