Fibroblast growth factors (FGF), and in particular FGF8, have been strongly implicated in prostate carcinogenesis. This study investigated the expression of Sef, a key inhibitory regulator of FGF signalling, in prostate cancer. In a panel of cell lines, hSef was detected in both androgen-dependent and independent cells but was significantly reduced in highly metastatic derivative clones. hSef expression was not influenced by androgenic stimulation. Forced downregulation of hSef by siRNA increased FGF8b induced cell migration (P=0.02) and invasion (P=0.007). Reduced hSef levels also enhanced FGF8b stimulated expression of MMP9 (P=0.005). mRNA in situ hybridization revealed hSef expression in 80% (8/10) of benign biopsies but in only 69% (23/33) of Gleason sum 4-7 and 35% (10/28) of Gleason sum 8-10 cancer biopsies (P=0.004). Quantitative PCR of microdissected glands confirmed this trend (P=0.001). hSef was expressed in 69% (27/39) of non-metastatic tumours but in only 18% (2/11) of metastatic tumours (P=0.004, n=50). hSef expression was next correlated with earlier data on FGF8b expression in a subgroup of cancers. In this cohort, 86% (19/22) of high-grade cancers expressed FGF8 but only 31% (7/22) expressed hSef. Positive FGF8 expression but a loss of hSef was observed in 88% (7/8) of metastatic tumours. In contrast, metastasis was evident in only 10% (1/10) of tumours, which co-expressed both FGF8 and hSef (P<0.001). These results suggest evidence that hSef is downregulated in advanced prostate cancer and might facilitate an enhanced tumorigenic response to FGFs. Further research into the role of hSef in cancer cell signalling and the mechanism of its downregulation may contribute to more effective targeting of growth factors in prostate cancer.