2-Nitropropane (2-NP), a secondary nitroalkane, has been extensively used in industry as a solvent for extractions and as a component of paints, inks, and varnishes. The World Health Organization estimates that approximately 4000 U.S. workers may have had significant occupational exposure to this chemical. 2-NP is a bacterial mutagen, produces unscheduled DNA synthesis in rat hepatocytes, and is a strong hepatocarcinogen in male Sprague-Dawley rats. The overall goal of this project is to elucidate the mechanism of the genotoxicity and carcinogenicity of 2-NP, thereby obtaining information for predicting whether or not the compound could also cause neoplasms in man. Previous work by this laboratory has shown that 2-NP produces specific and unique base modifications in rat liver DNA and RNA, namely the appearance of 8- aminoguanine, increases in 8-oxoguanine, and the appearance of thus far unidentified """"""""X1"""""""" adducts in RNA and DNA. The first Specific Aim of this proposal is to structurally identify these X1 adducts. Since the formation of 8-aminoguanine and X1 in DNA may represent tumor initiating events, the second Specific Aim is to determine the mutagenic potency of these lesions in bacterial and mammalian cells using a site-specifically modified single stranded shuttle vector. Metabolic conversion of 2-NP to a species capable of aminating guanine residues at C8 is mediated through aryl sulfotransferase (AST), probably isozyme IV.
Specific Aim 3 is to test the proposed central role of AST in 2-NP-induced hepatocarcinogenesis by a bioassay of 2-NP in control and pentachlorophenol-pretreated 12-day old B6C3F1 mice and in 12-day old normal and brachymorphic mice. For the AST-catalyzed conversion of 2-NP to a species capable of aminating guanine at C8 a mechanism has been proposed involving acetoxime-O-sulfonic acid and hydroxylamine-O-sulfonic acid as intermediates.
In Specific Aim 4, the mutagenicity of these intermediates will be determined and 35S-labeled PAPS will be used in attempts to detect these intermediates in the in vitro AST reactions using 2-NP nitronate as substrate. Since the AST-catalysed activation of 2-NP nitronate is a completely novel biochemical reaction, its kinetic parameters will be determined using purified rat liver ASTIV, (Specific Aim 5). The genotoxicity of secondary nitroalkanes, including 2-NP, depends on the dissociation of the a-hydrogen to yield stable nitronate anions.
In Specific Aim 6, primary nitroalkanes with acidic a-hydrogens will be synthesized and tested for mutagenicity and ability to serve as substrates in the AST-catalysed activation. The 7th Specific Aim is to determine the extents of metabolic activation and detoxication of 2-NP in various rodent species of both sexes and correlate these with 2-NP- induced rat liver nucleic acid modifications and susceptibility to 2-NP hepatocarcinogenesis. Since acute exposure to 2-NP can result in severe hepatotoxicity, it is proposed to establish whether this toxicity is also mediated through the activation of 2-NP by AST, and whether its severity can be reduced by inhibitors of AST (Specific Aim 8). Having obtained information on the extents of the activation and detoxication pathways of 2-NP in rodent species of varying susceptibility to its carcinogenicity, the last Specific Aim (9) is to determine the ability of normal human liver to activate and detoxify 2-NP in order to assess its potential human carcinogenicity.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
5R01ES003257-13
Application #
2634326
Study Section
Chemical Pathology Study Section (CPA)
Project Start
1984-01-01
Project End
2000-12-31
Budget Start
1998-01-01
Budget End
1998-12-31
Support Year
13
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Institute for Cancer Prevention
Department
Type
DUNS #
City
Valhalla
State
NY
Country
United States
Zip Code
10595
Sodum, R S; Fiala, E S (2001) Analysis of peroxynitrite reactions with guanine, xanthine, and adenine nucleosides by high-pressure liquid chromatography with electrochemical detection: C8-nitration and -oxidation. Chem Res Toxicol 14:438-50
Sodum, R S; Akerkar, S A; Fiala, E S (2000) Determination of 3-nitrotyrosine by high-pressure liquid chromatography with a dual-mode electrochemical detector. Anal Biochem 280:278-85
Sohn, O S; Fiala, E S (2000) Analysis of nitrite/nitrate in biological fluids: denitrification of 2-nitropropane in F344 rats. Anal Biochem 279:202-8
Sodum, R S; Fiala, E S (1998) N2-amination of guanine to 2-hydrazinohypoxanthine, a novel in vivo nucleic acid modification produced by the hepatocarcinogen 2-nitropropane. Chem Res Toxicol 11:1453-9
Fiala, E S; Sohn, O S; Li, H et al. (1997) Inhibition of 2-nitropropane-induced rat liver DNA and RNA damage by benzyl selenocyanate. Carcinogenesis 18:1809-15
Sodum, R S; Fiala, E S (1997) Amination of tyrosine in liver cytosol protein of male F344 rats treated with 2-nitropropane, 2-nitrobutane, 3-nitropentane, or acetoxime. Chem Res Toxicol 10:1420-6
Fiala, E S; Sodum, R S; Hussain, N S et al. (1995) Secondary nitroalkanes: induction of DNA repair in rat hepatocytes, activation by aryl sulfotransferase and hepatocarcinogenicity of 2-nitrobutane and 3-nitropentane in male F344 rats. Toxicology 99:89-97
Sodum, R S; Sohn, O S; Nie, G et al. (1994) Activation of the liver carcinogen 2-nitropropane by aryl sulfotransferase. Chem Res Toxicol 7:344-51
Sodum, R S; Nie, G; Fiala, E S (1993) 8-Aminoguanine: a base modification produced in rat liver nucleic acids by the hepatocarcinogen 2-nitropropane. Chem Res Toxicol 6:269-76
Swenson, D H; el-Bayoumy, K; Shuie, G H et al. (1993) Synthesis of N-(purin-8-yl)arylamines. Chem Res Toxicol 6:480-5

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