Interferons (IFN)-alpha, -beta, and -gamma enhance the activity of 5-fluorouracil (5-FU) in vitro and in vivo. Various mechanisms have been identified to account for this modulation. First, IFN induces the enzyme thymidine phosphorylase, thereby enhancing the conversion of 5-FU to its active metabolite, 5-fluorodeoxyuridine monophosphate (FdUMP), leading to increased depletion of thymidine triphosphate pools and increased DNA fragmentation. Second, IFN treatment leads to abrogation of an 5-FU-associated increase in the enzyme thymidylate synthase (TS), thus increasing tumor sensitivity to 5-FU. Finally, IFN augments plasma 5-FU levels. Single-institution studies of 5-FU in combination with IFN-alpha showed high response rates; however, randomized trials demonstrated equivalent survival to 5-FU alone or in combination with leucovorin (LV). Randomized trials of 5-FU double-modulated by both IFN-alpha and LV showed no response or survival advantage compared with 5-FU/LV, and greater toxicity. The randomized trials are all limited by inconsistent schedules of administration of IFN-alpha. The combination of 5-FU and IFN-beta has shown promising results in single-arm and small randomized trials. A large randomized trial, requiring a standardized schedule of administration of IFN-beta, has been initiated.