Cell lines 4A4 and 2C5 are the respective metastatic and non-metastatic variants (nude mouse system) derived from the human mammary carcinoma cell line MDA-MB-435. In order to identify genes associated with or functionally involved in metastasis, we have extended our previous transcriptional profile from 5000 to 12,000 genes using the Affymetrix Gene Chip array technology. Based on a threshold level of a change factor of > or = 2.5, we found that the steady-state level of 40 genes (0.3%) was up-regulated and conversely 89 genes (0.7%) were down-regulated in the metastatic cell line 4A4. The de-regulated genes were classified into categories such as tumor antigens/transmembrane receptors, enzymes, mediators of signaling, cell migration and angiogenesis, cell-cycle-, apoptosis-, differentiation- and growth-factor related genes, tumor suppression, transcription factors and genes encoding components of the extracellular matrix and the cytoskeleton. As possible mediators of invasion we identified DGCR6, osteopontin, autotaxin and the 65 kD phosphoprotein p65. In addition, three sugar-modifying enzymes were up-regulated in cell line 4A4. Profound differences in G-protein-mediated signaling and down-regulation of the tumor-suppressor genes DPC4, BARDI and DLC-1 were noted in the metastatic cell line 4A4.