The aim of this study was to investigate the expression of the protein tyrosine phosphatases (PTP) PRL-1, PRL-2, and PRL-3 in human breast cancer and to evaluate its clinical and prognostic significance. PRL-PTP mRNA expression was examined in malignant (n = 7) and nonmalignant (n = 7) cryoconserved breast tissue samples as well as in eight breast cancer cell lines by RT-PCR. Furthermore, protein expression of PRL-3 was analysed semiquantitatively by immunohistochemistry in ductal breast carcinoma in situ (n = 135) and invasive breast cancer (n = 147) by use of tissue microarray technology (TMA). In 24 lymph node-positive patients we selected the corresponding lymph node metastases for analysis of PRL-3 expression, and a validation set (n = 99) of invasive breast cancer samples was examined. Staining results were correlated with clinicopathological parameters and long-term follow-up. PRL-3 mRNA expression was significantly higher in malignant compared to benign breast tissue. For PRL-1 and PRL-2 expression no significant differences were observed. Staining of TMAs showed PRL-3 expression in 85.9% ductal carcinoma in situ and 75.5% invasive breast carcinomas. Analysis of survival parameters revealed a shorter disease-free survival (DFS) in patients with PRL-3-positive carcinomas, and in particular a significantly shorter DFS in nodal-positive patients with PRL-3 overexpressing tumours as compared to PRL-3-negative breast carcinomas (66+/-7 months (95% CI, 52-80) vs 97+/-9 months (95% CI, 79-115); P = 0.032). Moreover, we found a more frequent expression of PRL-3 in lymph node metastases as compared to the primary tumours (91.7 vs 66.7%; P = 0.033). Our results suggest that PRL-3 might serve as a novel prognostic factor in breast cancer, which may help to predict an adverse disease outcome.