Formation of urothelial and hepatic DNA adducts from the carcinogen 2-naphthylamine

Abstract

The carcinogen, 2-naphthylamine (2-NA), induces tumor formation in the urinary bladder but not the liver of several species, including humans and dogs. Since its proximate carcinogenic metabolite, N-hydroxy-2-NA, was known to react directly with DNA in vitro to give specific carcinogen-base adducts, we investigated the in vivo formation and persistence of (2-NA)-DNA adducts in the bladder and liver and attempted to determine whether or not these lesions correlated with tissue susceptibility. Male beagle dogs were administered [3H]2-NA and sacrificed after 2 or 7 days. The DNA was isolated from the liver and urothelium and hydrolyzed enzymatically. The (2-NA)-deoxyribonucleoside adducts, which were quantitated by high pressure liquid chromatographic analysis, were the same as those found in vitro, namely 1-(deoxyguanosin-N2-yl)-2-NA, 1-(deoxyadenosin-N6-yl)-2-NA, and an imidazole ring-opened derivative of N-(deoxyguanosin-8-yl)-2-NA. The major difference detected between target and non-target tissues was in the total level of binding to DNA, which was 4-fold higher in the urothelium at 2 days and 8-fold higher at 7 days after 2-NA dosing. Analysis of specific adducts suggested that this difference may be due to the relative persistence of the C-8-guanine adduct in the urothelium as compared to the liver. Similar experiments with the non-carcinogen, 1-naphthylamine, failed to reveal binding in urothelial DNA and indicated a 20-fold lower binding level in hepatic DNA. Evidence for binding of 2-NA to glycogen is also presented and problems associated with measuring total radioactivity in glycogen-contaminated DNA fractions are discussed. The data obtained in this study, through from a necessarily limited number of animals, are consistent with the hypothesis that the formation and persistence of DNA-carcinogen adducts may be important in the initiation of the neoplastic process.

Formation and persistence of arylamine DNA adducts in vivo.

“…This reduced binding was probably responsible for the small tumor yield being recorded with N-hydroxy-2-NA. The DNA adduct derived from 2-NA in target (urothelium) and nontarget (liver) tissues of dogs have also been reported (10,11). Two days following the oral administration of 2-NA, the same three adducts that were found in vitro from N-hydroxy-2-NA were detected in both tissues with 4-fold higher binding levels being observed in the urothelial DNA.…”

“…When the extent ofbinding was measured 7 days after 2-NA administration, the N2-deoxyguanosine adduct appeared to persist in the dog liver, while both this adduct and the C8-deoxyguanosine adduct persisted in the bladder (10,11). The differential loss of adducts indicates that active repair processes are present in both tissues, and the relative persistence of the C8-deoxyguanosine adduct in target but not in the nontarget tissue suggests that this adduct may be a critical lesion for the initiation of urinary bladder tumors.…”

“…Therefore, as part of a study relating the extent of adduct formation to carcinogenic potency, the binding of 1-NA to hepatic and urothelial DNA of dogs was assessed (10,11). Adduct fonnation from 1-NA was not detected in urothelial DNA and only very low levels were found in hepatic DNA.…”

Lifetime carcinogenicity study of 1- and 2-naphthylamine in dogs

“…This quantitative difference in the metabolism of 1-and 2-naphthylamine is reflected in a similar difference in the level of reaction with DNA in the urothelium. DNAnaphthylamine adducts could be detected in the urothelium (18.5 adducts/108 nucleotides) 2 days after administration of 2-naphthylamine to a dog; but none were detected after administration of 1-naphthylamine (Kadlubar et al, 1981). Because the limit of detection of adducts was 1/108 nucleotides, the magnitude of the difference can only be expressed as at least 18-fold.…”

Aminofluorene-DNA adduct formation in Salmonella typhimurium exposed to the carcinogen N-hydroxy-2-acetylaminofluorene.

“…Two other bands of radioactivity were detected at 18.5 and 41.0 min. The identity of the early peak is not known; it coeluted with the ribose analogue N-(guanosin-8-yl)-AF and could not be detected after DNA banding on a cesium chloride gradient (27). The (20) indicated that it contained a small amount of highly labeled DNA (=30 ,g DNA per sample; 22 nCi/mg of DNA).…”