Studies on DNA adducts and carcinogenesis with 2-nitrofluorene as a model substance

Abstract: Studies on DNA Adducts and Carcinogenesis with 2-Nitrofluorene as a Model Substance Xian-Shu Cui Center for Nutrition and Toxicology Department of Biosciences at Novum, Karolinska institute Novum, S-141 57 Huddinge, Sweden A number of carcinogens or their electrophilic intermediates are able to bind covalently to DNA, forming DNA adducts. DNA adducts may cause gene mutations if they are misrepaired or remain unrepaired before DNA replication occurs. DNA adduct formation is believed to be the initial step of chemical carcinogenesis. 2-Nitrofluorene (NF) is an air pollutant and a carcinogen. Metabolism of NF is generally via nitro-reductive and oxidative pathways. Nitro-reductive metabolism of NF results in formation of 2- aminofluorene and 2-acetylaminofluorene, two well known carcinogens. The present study focused on investigating DNA adducts and carcinogenesis using NF and some of its derivatives as model substances. The nitro function is supposed to be crucial for the genotoxic effects of nitroarenes. However, the great difference between NF and 2,7-dinitrofluorene (2,7-dNF) in the Salmonella mutagenicity test was not seen in the in vivo genotoxic assays. After intraperitoneal or oral administration, 2,7-dNF induced a lower amount of DNA adducts and a similar or higher amount of preneoplastic lesions in the liver compared to NF. Oral administration of NF or 2,7-dNF induced a higher amount of DNA adducts and preneoplastic liver lesions than intraperitoneal injection. The genotoxicity of oxidized metabolites of NF, 5-, 7- and 9-OH-NF, was studied in vitro and in vivo. Generally, capacity of these substances to initiate preneoplastic liver lesions was associated with their potential to form DNA adducts. 5-OH-NF showed low or no potential to induce DNA adduct formation and preneoplastic liver lesions. 7- and 9-OH-NF induced less DNA adducts and preneoplastic liver lesions than NF. The long-term tumor study showed that NF was a potent carcinogen, primarily targeting the liver, forestomach and kidney. NF induced tumor development and DNA adduct formation in a dose-dependent manner. The levels of DNA adducts in different organs during NF administration were associated with the location of tumors. Four major NF-DNA adducts were found. One DNA adduct co-migrated with C3- (deoxyguanosine-N2-yl)-2-acetylaminofluorene and one co-migrated with N-(deoxyguanosine-C8-yl)-2- aminofluorene by 32P-HPLC co-chromatography. The other two, which constituted the major part (>80%) of NF-DNA adducts after a long period of NF administration, were not characterized. The four NF-DNA adducts showed different recovery after nuclease Pl or butanol enrichment. Three out of the four NF-DNA adducts were still detectable in rat liver after 11 months on basal diet. Cell proliferation is a crucial factor for transforming DNA lesions to mutations. In the present study, the effects of NF on cell proliferation of the tumor-target tissues were investigated. Cell proliferation was induced by a short-term NF administration (4 days) in the forestomach mucosa. As a non-tumor target tissue, the glandular stomach mucosa were inhibited regarding the cell proliferation. Proximal straight tubules of kidney was the only section in kidney showing increased rate of cell proliferation after NF treatment, indicating the possible site of origin for the resulting renal tumors. However, NF inhibited hepatocyte proliferation, suggesting a different mechanism of carcinogenesis in hepatocytes compared to others. In conclusion, NF and most of its derivatives studied are capable of binding to DNA and capable of initiating preneoplastic lesions. The levels of NF-DNA adducts in the tested tissues are associated with the location of NF-induced tumors. Cell proliferation may play a role in the process of NF-induced tumor formation. Key words: 2-nitrofluorene, carcinogens, DNA adducts, preneoplastic lesion, tumor