Pancreatic adenocarcinomas are among the most malignant forms of cancer and, therefore, it is of especial interest to set new strategies aimed at improving the prognostic of this deadly disease. The present study was undertaken to investigate the action of cannabinoids, a new family of potential antitumoral agents, in pancreatic cancer. We show that cannabinoid receptors are expressed in human pancreatic tumor cell lines and tumor biopsies at much higher levels than in normal pancreatic tissue. Studies conducted with MiaPaCa2 and Panc1 cell lines showed that cannabinoid administration (a) induced apoptosis, (b) increased ceramide levels, and (c) up-regulated mRNA levels of the stress protein p8. These effects were prevented by blockade of the CB₂ cannabinoid receptor or by pharmacologic inhibition of ceramide synthesis de novo. Knockdown experiments using selective small interfering RNAs showed the involvement of p8 via its downstream endoplasmic reticulum stress–related targets activating transcription factor 4 (ATF-4) and TRB3 in Δ⁹-tetrahydrocannabinol–induced apoptosis. Cannabinoids also reduced the growth of tumor cells in two animal models of pancreatic cancer. In addition, cannabinoid treatment inhibited the spreading of pancreatic tumor cells. Moreover, cannabinoid administration selectively increased apoptosis and TRB3 expression in pancreatic tumor cells but not in normal tissue. In conclusion, results presented here show that cannabinoids lead to apoptosis of pancreatic tumor cells via a CB₂ receptor and de novo synthesized ceramide-dependent up-regulation of p8 and the endoplasmic reticulum stress–related genes ATF-4 and TRB3. These findings may contribute to set the basis for a new therapeutic approach for the treatment of pancreatic cancer. (Cancer Res 2006; 66(13): 6748-55)