Cystatin SN (cystatin 1, CST1) is a member of the cystatin superfamily that inhibits the proteolytic activity of cysteine proteases. CST1 is a tumor biomarker that provides useful information for the diagnosis of esophageal, gastric, and colorectal carcinomas. However, the significance of CST1 in pancreatic cancer is unknown. The aim of this study was to assess whether CST1 is a potential biomarker for early diagnosis of malignant pancreatic neoplasms. Microarray analysis of mRNA extracted from pancreatic cancer and pancreatic normal tissues was performed. Bioinformatics revealed that CST1 was one of the highest expressed genes on the array in pancreatic cancer, compared with normal tissue. In addition, the upregulation of CST1 in pancreatic cancer and several pancreatic cancer cell lines was confirmed using real-time PCR (RT-PCR), immunohistochemistry, and Western blotting. Next, CST1 knockdown using siRNA reduced the expression of the proliferation-related proteins p-AKT and PCNA significantly, as well as colony formation and xenograft development in vitro. Consistent with this, CST1 mRNA overexpression was correlated closely with malignancy-associated proteins such as PCNA, cyclin D1, cyclin A2, and cyclin E in pancreatic cancer cell lines. In conclusion, our data suggest that CST1 might contribute to the proliferation of pancreatic cancer cells and could be a potential biomarker for the early detection of pancreatic cancer.