Pancreatic cancer is a highly malignant cancer and lacks effective therapeutic targets. Protein-tyrosine phosphatase 1B (PTP1B), a validated therapeutic target for diabetes and obesity, also plays a critical positive or negative role in tumorigenesis. However, the role of PTP1B in pancreatic cancer remains elusive. Here, we initially demonstrated that PTP1B was highly expressed in pancreatic tumors, and was positively correlated with distant metastasis and tumor staging, and indicated poor survival. Then, inhibition of PTP1B either by shRNA or by a specific small-molecule inhibitor significantly suppressed pancreatic cancer cell growth, migration and colony formation with cell cycle arrest in vitro and inhibited pancreatic cancer progression in vivo. Mechanism studies revealed that PTP1B targeted the PKM2/AMPK/mTOC1 signaling pathway to regulate cell growth. PTP1B inhibition directly increased PKM2 Tyr-105 phosphorylation to further result in significant activation of AMPK, which decreased mTOC1 activity and led to inhibition of p70S6K. Meanwhile, the decreased phosphorylation of PRAS40 caused by decreased PKM2 activity also helped to inhibit mTOC1. Collectively, these findings support the notion of PTP1B as an oncogene and a promising therapeutic target for PDAC.