International Mammalian Genome Society

The 14th International Mouse Genome Conference (2000)


F11. Molecular Profiling of a Mouse Model for Metastatic Breast Carcinoma

K Jessen, S Liu, L Yong, RD Cardiff1, J Gregg
University of California, Davis School of Medicine, Dept. of Pathology, 4645 Second Avenue Research III, Room 3200C Sacramento, CA., jpgregg@ucdavis.edu

Breast cancer is among the most common human cancers, affecting one in every eight women. One of the significant predictors of prognosis is regional and distant metastasis. As cancer is the consequence of a broad dysregulation or alteration of cell signaling pathways, the ability of cells to metastasize may be due to changes in a limited number of pathways related to invasiveness and metastasis. Our study focuses on gaining a better molecular understanding of these cellular pathways using a mouse model for mammary tumor metastasis. We are working with two transplantable mouse mammary tumor lines with significant differences in metastatic potential. Met-1 tumors develop 100% pulmonary metastases while Db-7 tumors exhibit significantly fewer pulmonary metastases (9%).

In order to extract genes differentially expressed in both Met-1 and Db-7, we performed two suppression subtractive hybridization (SSH) experiments. From each of the subtracted libraries, 2600 clones were PCR-amplified and arrayed. By fluorescently labeling the unsubtracted Met-1 and Db-7 libraries and hybridizing them onto the microarray, we were able to demarcate the truly differentially expressed genes. Clones found to be greater than two-fold differentially expressed were then sequenced for identification. Differential expression of each unique gene was verified by RT-PCR, Northern blot, and Western blot. Additionally, RT-PCR was used to determine expression levels of these genes in other mouse tissue (normal mammary, lung, liver, etc.). In order to determine the function of a subset of these genes, anti-sense and sense constructs were introduced into cell lines derived from these tumors. Migration assays were performed and the metastatic potential of each gene was assessed. Using this methodology, we have identified several interesting genes that are both differentially and functionally important for the metastatic phenotype in this model.

 


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