2-NITROSOAMINO-3-METHYLIMIDAZO[4,5-F]QUINOLINE STABILITY AND REACTIVITY

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. N-Nitrosamines and nitrosamides can initiate cancer. These studies evaluated the stability and reactivity of 2-nitrosoamino-3-methylimidazo[4,5-f]quinoline (N-NO-IQ) to assess its possible role in the initiation of colon cancer by 2-amino-3-methylimidazo[4,5-f]quinoline (IQ). 14C-N-NO-IQ was incubated with different solvents and pHs in the presence and in the absence of nucleophiles and analyzed by HPLC. The products identified by electrospray ionization mass spectrometry include 2-chloro-3-methylimidazo[4,5-f]quinoline (2-Cl-IQ), 2,2'-azo-3,3'-dimethylimidazo[4,5-f]quinoline (AZO-IQ), 2-azido-IQ (2-N3-IQ), 3-methylimidazo[4,5-f]quinoline (deamino-IQ), and IQ. A variety of organic solvents were tested with 0.1 N HCl. 2-Cl-IQ and IQ were formed following acidification of all solvents. AZO-IQ was only formed in methanol. Deamino-IQ was the major product formed in all of the alcohols tested, except for methanol. Under acidic conditions that completely convert N-NO-IQ in 5 min (acetonitrile with 0.1 N HCl), 62% of N-NO-IQ remains after 30 min if dimethyl sulfoxide is substituted for acetonitrile. N-NO-IQ was stable in the physiologic pH range of 5.5-9.0 and did not react with nucleophiles over a 4 h period at pH 7.4 and 37 C. At acidic pH (pH 2.0) for 30 min and 37 C, N-NO-IQ becomes labile forming electrophile(s), which combine with biologically relevant nucleophiles. The reaction of N-NO-IQ at pH 2.0 followed first-order kinetics (t1/2 = 10 ¿ 2 min) and was significantly increased in 10 mM NaN3 (t1/2 = 2 ¿ 0.1 min). 2-N3-IQ was the major product observed in the latter incubation. N-NO-IQ binding to DNA at pH 2.0 is 100-fold more than that at pH 7.4. At pH 2.0, greater than 90% of the binding was inhibited by 10 mM NaN3. Thus, N-NO-IQ forms a reactive electrophile(s) at acidic pH, which binds DNA. N-NO-IQ reaction products may depend on the pH and the hydrophobic milieu of cells or tissues.