In this study we sought to clarify the role of the pro-apoptotic potential of mitochondria in the death pathway emanating from the TRAIL (APO-2L) and CD95 receptors in pancreatic carcinoma cells. We focused on the role of the Bcl-2 family member Bcl-X L, using three pancreatic carcinoma cell lines as a model system, two of which have high (Panc-1, PancTuI) and one has low (Colo357) Bcl-X L expression. In these cell lines, the expression of Bcl-X L correlated with sensitivity to apoptosis induced by TRAIL or anti-CD95. Flow cytometric analysis revealed cell surface expression of TRAIL-R1 and TRAIL-R2 on PancTuI and Colo357, and TRAIL-R2 on Panc-1 cells. In Colo357 cells retrovirally transduced with Bcl-X L, caspase-8 activation in response to treatment with TRAIL or anti-CD95 antibody was not different from parental cells and EGFP-transfected controls, however, apoptosis was completely suppressed as measured by the mitochondrial transmembrane potential Δψm, caspase-3 activity (PARP cleavage) and DNA-fragmentation. Inhibition of Bcl-X L function by overexpression of Bax or administration of antisense oligonucleotides against Bcl-X L mRNA resulted in sensitization of Panc-1 cells to TRAIL and PancTuI cells to anti-CD95 antibody-induced cell death. The results show that Bcl-X L can protect pancreatic cancer cells from CD95-and TRAIL-mediated apoptosis. Thus, in these epithelial tumour cells the mitochondrially mediated ‘type II’pathway of apoptosis induction is not only operative regarding the CD95 system but also regarding the TRAIL system.