EFFECTS OF AMINOSALICYLATES AND IMMUNOSUPPRESSIVE AGENTS ON NITRIC OXIDE-DEPENDENT N-NITROSATION REACTIONS

Recent studies have demonstrated that nitric oxide (NO) rapidly and spontaneously decomposes in oxygenated solutions to generate potent N-nitrosating agents. These electrophilic substances have been shown to mediate mutagenesis and carcinogenesis via the formation of aliphatic and aromatic nitrosamines. We have also demonstrated that extravasated neutrophils and macrophages produce significant amounts of N-nitrosating agents derived exclusively from NO. During the course of these studies, we found that certain antioxidants, including 5-aminosalicylic acid (5-ASA), inhibited the leukocyte-mediated N-nitrosation reaction. Because 5-ASA and other anti-inflammatory and immunosuppressive drugs are used to treat inflammatory bowel disease, we wondered if any of these other compounds might also modulate N-nitrosation reactions in vitro. Therefore, the objectives of this study were to assess the ability of aminosalicylates and certain immunosuppressive agents to inhibit NO-dependent N-nitrosation of a model aromatic amine (2,3-diaminonaphthalene) and to determine whether this inhibitory activity correlated with their oxidation potential. We found that the concentrations necessary to inhibit the N-nitrosation reaction by 50% (IC50) were 25, 50 and 100 mu M for 5-ASA, olsalazine (dimeric 5-ASA) and sulfasalazine, respectively. In contrast, sulfapyridine, 4-ASA, N-acetyl-5-ASA, 6-mercaptopurine, azathioprine, and methotrexate were either much less effective or inactive at inhibiting the N-nitrosation reaction. Although 5-ASA was able to fully scavenge the stable free radical 1,1-diphenyl-2-picrylhydrazyl, neither olsalazine nor sulfasalazine was found to be effective at scavenging this weak oxidant. We did find that olsalazine possessed an oxidation potential substantially less than that of sulfasalazine, suggesting that it may, in fact, scavenge more potent oxidizing agents such as the N-nitrosating agent. We conclude that 5-ASA and olsalazine inhibit NO-dependent N-nitrosation reactions by scavenging or decomposing the nitrosating agent(s). We propose that the secondary nitrogen unique to sulfasalazine interacts with the nitrosating agent to yield a secondary nitrosamine, thereby competing for N-nitrosation of our detector.