It is now widely recognised that the cross-talk between cancer and stromal cells may play a crucial role in cancer progression. However little is known about the complex underlying molecular mechanisms that occur within the tumor microenvironment. Fibroblasts are the major stromal cells with multiple roles, especially towards both the extracellular matrix and the neighbouring cell population, including neoplastic cells. Consequently, proteomic analyses would provide a wider resource for a better understanding of the potential modulating effects exerted by fibroblasts on cancer cells. In this report we describe the effects of fibroblast stimulation on the breast cancer cell line (8701-BC) proteomics, using a trans-well co-culture system. Our results clearly indicate that fibroblasts induce considerable proteomic modulations on 8701-BC, mainly in the cytoskeleton proteins and glycolytic enzymes. Additionally, fibroblast-conditioned medium increased neoplastic cell proliferation and invasion with a concurrent up-regulation of the c-myc oncogene. Collectively these results suggest that fibroblast stimulation may enhance the malignant potential of breast cancer cells in vitro.