This study investigated the influence of cathepsin B (CATB), a downstream target of Hedgehog (Hh) signaling, in pancreatic cancer. Cyclopamine (Hh signal inhibitor) suppressed expression of Shh, as well as Hh-induced transcription factor Gli1, and induced apoptosis in Shh-positive pancreatic cancer cell line (PANC-1). Microarray analysis revealed CATB as a gene downregulated by Hh. Cyclopamine reduced CATB protein and mRNA levels. Cyclopamine or CATB inhibitor reduced PANC-1 cell invasiveness (P<0.05). CATB expression in human pancreatic cancer tissues tended to correlate with Shh expression (P=0.053). Conclusively, Hh targets CATB and Hh signaling through CATB might influence pancreatic cancer cell invasiveness.