Chemoattractants expressed at bony sites and pelvic lymph nodes are thought to promote the preferential metastasis of human prostate tumor cells to these organs. Epidermal growth factor (EGF) is a potent chemoattractant for several human metastatic prostate tumor cell lines, including the TSU-prl cell line, and EGF has been localized to the stroma of both bony sites and pelvic lymph nodes in humans. Hence, we investigated whether the TSU-prl cell line expresses a functional EGF receptor (EGFR), which when antagonized reduces EGF-mediated chemomigration of this cell line. In this context, the EGFR immunoprecipitated from cell lysates of TSU-prl cells comigrated with the EGFR from A431 cells at a molecular weight of 170 kD. Addition of human EGF (hEGF) to the TSU-prl cells for 5 min stimulated the dose-dependent biphasic phosphorylation of the EGFR, with maximal stimulation of EGFR phosphorylation occurring at 2 ng/ml hEGF. In addition, treatment of hEGF-stimulated (2 ng/ml) TSU-prl cells with 0.5 kg/ml anti-hEGFR monoclonal antibody or 100 nM staurosporine inhibited EGFR phosphorylation. Conversely, as negative controls, treatment of hEGF-stimulated (2 ng/ml) TSU-prl cells with K252a or dimethyl sulfoxide (DMSO) vehicle did not inhibit EGFR phosphorylation. TSU-prl cells were stimulated to migration in 4 hr across Boyden chambers in response to 10 ng/ml hEGF. Treatment of the TSU-prl cells with anti-hEGFR monoclonal antibody inhibited in a dosedependent manner the chemomigration of the TSU-prl cells across Boyden chambers. Similarly, treatment of the TSU-prl cells with staurosporine inhibited in a dose-dependent manner the chemomigration of the TSU-prl cells across Boyden chambers. These results demonstrate that antagonists of hEGF-mediated hEGFR phosphorylation also antagonize chemomigration of the TSU-prl cells across Boyden chambers, suggesting that antagonists of the EGFR in prostate cancer may be useful in the treatment of metastatic disease.