Metabolic activation and genotoxicity of N-hydroxy-2-acetylaminofluorene and N-hydroxyphenacetin derivatives in Reuber (H4-II-E) hepatoma cells.
Derivatives of both N-hydroxy-2-acetylaminofluorene (N-OH-AAF) and N-hydroxyphenacetin (N-OH-P) were tested for their ability to cause DNA damage in Reuber (H4-II-E) cells using the alkaline elution technique. Reuber cells are devoid of N-OH-AAF deacylase, N,O-acyltransferase, and sulfotransferase activities. The hydroxamic acids themselves caused very little DNA damage, while N-hydroxy-2-aminofluorine (20 to 100 microM), N-hydroxyphenetidine (20 to 200 microM), and p-nitrosophenetole (10 to 100 microM) all caused dose-dependent damage. The dose-dependent DNA damage caused by N-acetoxy-2-acetylaminofluorene (5 to 25 microM) was completely inhibited by the deacylase inhibitor paraoxon (100 microM). In the presence of both partially purified rabbit liver cytosolic N,O-acyltransferase and guinea pig liver microsomal deacylase, N-OH-AAF was genotoxic. Neither paraoxon nor tRNA had any effect on the DNA damage induced by N-OH-AAF in the presence of N,O-acyltransferase, while paraoxon completely inhibited the damage when N-OH-AAF was incubated in the presence of guinea pig deacylase, and N-OH-P only caused slight DNA damage at higher concentrations of enzyme. In addition, partially purified guinea pig liver deacylase and N-OH-AAF (25 microM) caused 2600 revertants in the Salmonella test system, while only 380 revertants were seen with a 40-fold greater concentration of N-OH-P (1000 microM). The mutagenicity of both N-OH-AAF and N-OH-P was completely inhibited by paraoxon. Thus, it is clear that metabolites of N-OH-AAF formed outside the cell are capable of passing both the cellular and nuclear membranes to cause genotoxicity. Metabolic activation of N-OH-AAF by either the membrane-bound deacylase or the cytosolic N,O-acyltransferase caused genotoxicity via a deacetylation process. Metabolic activation of N-OH-P by guinea pig deacylase caused low levels of DNA damage, whereas activation by N,O-acyltransferase was not sufficient to cause genotoxicity.
D015073:2-Acetylaminofluorene / Q000031:analogs & derivatives*; D000217:Acyltransferases / Q000378:metabolism; D000818:Animals; D001711:Biotransformation; D002460:Cell Line; D004261:DNA Replication / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D006168:Guinea Pigs; D006881:Hydroxyacetylaminofluorene / Q000378:metabolism / Q000633:toxicity*; D007700:Kinetics; D008114:Liver Neoplasms, Experimental / Q000378:metabolism / Q000503:physiopathology*; D008862:Microsomes, Liver / Q000201:enzymology; D010615:Phenacetin / Q000031:analogs & derivatives* / Q000378:metabolism / Q000633:toxicity; D011817:Rabbits; D051381:Rats; D011919:Rats, Inbred Strains; D013329:Structure-Activity Relationship
C008068:N-hydroxyphenacetin; D006881:Hydroxyacetylaminofluorene; D015073:2-Acetylaminofluorene; D000217:Acyltransferases; D010615:Phenacetin