Objective To investigate whether transforming growth factor beta (TGFβ) signaling is disrupted in human pancreatic cancer cells, and to study the role of TGFβ receptors and Smad genes. Summary Background Data TGFβ is a known inhibitor of pancreatic growth. Disruption of the TGFβ signaling pathway may play a role in pancreatic cancer development. Methods The effect of TGFβ on the BxPC-3, MiaPaCa-2, and PANC-1 pancreatic cancer cell lines was evaluated by [3H]thymidine incorporation and a TGFβ-responsive reporter assay. Expression of TGFβ receptors and Smads 2 and 3 was assessed by cross-linking assays and reverse transcriptase–polymerase chain reaction (RT-PCR). The ability to restore TGFβ responsiveness was evaluated by transfection of TGFβ signaling components. Results TGFβ produced little inhibition of DNA synthesis and did not activate a TGFβ-responsive reporter in pancreatic cancer cell lines. 125TGFβ cross-linking and RT-PCR confirmed the presence of TGFβ receptors and Smad2 and Smad3 transcripts. Transfection of TGFβ receptors or Smads 2 and 3 did not restore responsiveness. However, transfection of Smad4 into the BxPC-3 pancreatic cancer cell line restored TGFβ responsiveness. Conclusions Pancreatic cancer cells show loss of TGFβ responsiveness. Smads 2 and 3 and TGFβ receptors are not defective in the cell lines studied. Transfection of Smad4 into one of the cell lines restored TGFβ responsiveness, suggesting an important role for Smad4 in pancreatic cancer. It is likely that other, as yet unidentified genes are important in TGFβ resistance in pancreatic cancer cells.