The Cripto-1 (CR-1) derived EGF-CFC family was overexpressed in tumor development enhancing proliferation, epithelial-mesenchymal transition (EMT) and migration of tumor cells. However, correlation between CR-1 and prostate cancer (PCa) remains still unclear. In the present study, we proved that CR-1 was expressed in PCa and its function was in the progression of PCa. Compared with benign prostatic hyperplasia (BPH) tissues, we confirmed that PCa tissues had high expression of CR-1 by immunohistochemistry and statistical data showed that CR-1 promoted properties of EMT in PCa tissues, including the downregulation of the cell adhesion molecules β-catenin (membrane) and E-cadherin while upregulating transcription factors β-catenin. Overexpression of CR-1 had close relationship with PSA, Gleason, clinical staging and lymph node metastasis in PCa patients. Then, we found that PC-3 cells transfected with CR-1-shRNA inhibited EMT using RT-PCR, RT-qPCR, western blotting and immunofluorescence. Also, we evaluated cell invasive ability in vitro by transwell and wound-healing assay. Our data showed that transfected CR-1-shRNA altered EMT including β-catenin, E-cadherin, c-myc, GSK-3, p-GSK and Wnt/β-catenin pathway in PC-3. It also suppressed PC-3 cell migration. Additionally, our results displayed that Licl had antitumor activity against PC-3 through the inhibition of Wnt/β-catenin pathway. Inhibition of cell viability was dose-time dependent. The present study proved that CR-1 regulates EMT of PCa by Wnt/β-catenin pathway. Hence, CR-1 may provide a new biological marker, and possibly contributes to clinical treatment against PCa.