To clarify the potential involvement of plasmin(ogen) cascade proteins in the cell dissociation and subsequent invasion of pancreatic cancer cells, western blot analysis, immunocytochemistry, immunohistochemistry, and in vitro invasion assay were performed in the cell lines or tissue of pancreatic cancer. The strong expression of plasmin(ogen), urokinase type plasminogen activator (uPA) and uPA receptor (uPAR), and apparently weak expression of the relevant proteins were found in the conditioned medium of dissociated (PC-1.0) and non-dissociated (PC-1) pancreatic cancer cells, respectively. Furthermore, uPA-treatment significantly induced the expression of plasmin(ogen) and uPAR in the conditioned medium of non-dissociated (PC-1) pancreatic cancer cells. Moreover, the expression of plasmin(ogen) and uPAR was stronger at the invasive front than at the center of human pancreatic cancer tissue. On the other hand, plasmin-treatment induced the expression of matrix metalloproteinase-2 (MMP-2), MMP-7 and MMP-9 in PC-1 cells. Simultaneously, plasmin- or uPA-treatments obviously induced the dissociation of cell colonies and in vitro invasiveness in PC-1 cells. The plasmin(ogen) cascade is closely involved in the invasion of pancreatic cancer cells and, especially in its early stage, cell dissociation. Targeting the plasmin(ogen) cascade may provide a new insight into molecular target therapy based on anti-invasion and anti-metastasis for pancreatic cancer.