Proteinase‐activated receptor‐2 (PAR‐2), which is a G protein‐coupled receptor, is activated in inflammatory processes and cell proliferation. We previously demonstrated that an anti‐PAR‐2 antibody suppresses proliferation of human pancreatic cells in vitro. However, there have been no studies of PAR‐2 signaling pathways in vivo. The aim of this study was to determine whether blockade of PAR‐2 by RNA interference influences pancreatic tumor growth. We originally constructed small interfering RNAs (siRNAs) targeting human PAR‐2, and performed cell proliferation assays of Panc1 human pancreatic cancer cell line with these siRNAs. Intratumoral treatment with these PAR‐2 siRNAs and atelocollagen was also performed in a xenograft model with nude mice and Panc1 cells. siRNAs against human PAR‐2 inhibited proliferation of Panc1 cells, whereas control scramble siRNAs had no effect on proliferation. The PAR‐2 siRNAs dramatically suppressed tumor growth in the xenograft model. PAR‐2‐specific siRNA inhibited growth of human pancreatic cancer cells both in vitro and in vivo. Blockade of PAR‐2 signaling by siRNA may be a novel strategy to treat pancreatic cancer. © 2007 Wiley‐Liss, Inc.