Effects of 5-fluorouracil and 6-azauridine on interferon action.

During the past few years, several laboratories reported that actinomycin D blocked the antiviral activity of interferon (E. Heller, Virology 21:652, 1963; J. Taylor, Biochem. Biophys. Res. Commun. 14:447, 1964). This finding suggests that de novo synthesis of cellular ribonucleic acid (RNA) is required for the functioning of interferon, because actinomycin D is known to block the synthesis of cellular RNA (E. R. Reich et al., Science 134:556, 1961). However, several instances can be cited where actinomycin D had a mode of action unrelated to the inhibition of transcription (D. Kom et al., Biochem. Biophys. Res. Commun. 19:473, 1965; M. Revel et al., Proc. Natl. Acad. Sci. U.S. 51:810, 1964; G. R. Honig and M. Rabinovitz, Science 149:1504, 1965). In the present report, experiments were performed with interferon and antimetabolites which affect either the synthesis or activity of RNA. The effects of these antimetabolites, 5fluorouracil and 6-azauridine, on the viral inhibitory activity of interferon in chick embryo fibroblast cells should provide information on the role of RNA during interferon action. The methods for preparing chick embryo fibroblast cultures and interferon have been described by M. Ho and M. K. Breinig (J. Immunol. 89:177, 1962). Hanks medium containing 4% bovine serum and Sindbis virus (AR 339) were employed. 6-Azauridine (6-AU) was purchased from Schwarz Bio Research Inc., Orangeburg, N.Y., and 5-fluorouracil (5-FU) was a gift of Paul Calabresi, Yale Medical School. The results of one study are summarized in Table 1 and confirm the results obtained by others with actinomycin D. At a concentration of 1 Ag/ml, actinomycin D completely counteracted the inhibitor activity of interferon on Sindbis virus. 5-FU (10-3 M) and 6-AU (1 mg/ml) did not inhibit virus development when present in the medium before infection, although both drugs boosted the virus titer about threeto fourfold in the presence of interferon. In seven separate studies, this value ranged from threeto ninefold for both drugs. Since 6-AU is known to block RNA synthesis through the formation of 6-azauridine 5'-phosphate, a compound which inhibits the enzymatic decarboxylation of orotidylic acid (R. E. Handschumacher, J. Biol. Chem. 235:2917, 1960), and since 5-FU is incorporated into RNA causing miscoding during protein synthesis (F. Gros and S. Naono, p. 195, in J. C. Harris [ed.], Protein Biosynthesis, Academic Press, Inc., New York, 1961), the results suggest that interferon action requires the syn-