Pancreatic cancer is usually diagnosed in the advanced stages and is sensitive to only few therapies. The forkhead box L1 (FOXL1) and protein phosphatase 2A (PP2A) have been recognized to be tumor suppressive in human pancreatic cancers. In the present study, we co-expressed the two tumor suppressive molecules with a '2A peptide' linker, which guaranteed the two molecules were transcribed into one mRNA, whereas they were translated into two separate proteins, in pancreatic cancer Panc‑1 cells, and investigated the inhibition of the two molecules on the proliferation and migration of Panc‑1 cells. Results demonstrated that, either overexpression of FOXL1 or PP2A via adenovirus significantly inhibited the proliferation of Panc‑1 cells, whereas promoted apoptosis in such cells. Moreover, the co-expression of both FOXL1 and PP2A exerted synergistic antitumor effect in Panc‑1 cells, with significantly higher proliferation inhibition and tumor induction. In addition, we found that the overexpressed FOXL1 promoted the TNF‑related apoptosis‑inducing ligand (TRAIL), whereas the overexpressed PP2A downregulated the phosphorylation of c‑MYC. The co-expression of FOXL1 and PP2A presented both functions in Panc‑1 cells. In conclusion, the adenovirus‑mediated co‑expression of FOXL1 and PP2A with the 2A peptide linker exterts synergistic suppression of pancreatic cancer cells via inhibiting the growth and promoting apoptosis of cancer cells, probably via upregulating TRAIL and reducing the phosphorylation of MYC.