title: Effect of ovariectomy on the in vitro and in vivo activation of carcinogenic N-2-fluorenylhydroxamic acids by rat mammary gland and liver.
abstract: N-Hydroxy-N-2-fluorenylacetamide (N-OH-2-FAA) and its benzamide analogue N-OH-2-FBA are mammary gland carcinogens in the female Sprague-Dawley rat. Ovariectomy inhibits tumorigenicity of topically applied N-OH-2-FAA suggesting modulation of carcinogen-activating enzymes in the gland. This study concerned the activation of N-OH-2-FAA and N-OH-2-FBA by the mammary gland and liver, a chief site of metabolism, from 50-day-old female rats and effects on the activation of ovariectomy performed at 22 days of age. The levels of N-debenzolyation of N-OH-2-FBA to N-hydroxy-N-2-fluorenamine (N-OH-2-FA), catalyzed by microsomal carboxylesterases in mammary gland and liver were similar and increased 1.5- and 1.7-fold, respectively, by ovariectomy. N-Debenzoylating activity in cytosols of both tissues appeared to be partially of microsomal origin. Mammary gland cytosol contained N-, O- and N,O-acyltransferase activities at levels 40-50% those of liver. N-Acyltransferase activity was determined via acetyl coenzyme A (AcCoA)-dependent acetylation of 2-FA and a new assay, N-OH-2-FAA-dependent acetylation of 9-oxo-2-FA. The latter activity was decreased in mammary gland by ovariectomy. Microsomal N-acyltransferase activities were <36% those of cytosols. AcCoA-dependent binding of N-OH-2-[ring-[3H]FBA to DNA, catalyzed by cytosol, was consistent with a two-step activation of N-OH-2-FBA involving esterase-catalyzed N-debenzoylation to N-OH-2-FA and its O-acyltransferase-catalyzed acetylation to the electrophilic N-acetoxy-2-FA. O-Acetyltransfer by mammary gland appeared to be rate-limiting since ovariectomy-dependent increases in N-debenzoylation did not increase binding with S9 fraction. Little or no sulfotransferase-catalyzed binding of N-OH-2-[ring-3H]FBA-derived N-OH-2-[ring-3H]FA was detected in the liver or mammary gland cytosol, respectively. The level of binding of N-OH-2-[ring-3H]FAA to DNA catalyzed by cytosolic N,O-acyltransferase was decreased approximately 23% in mammary gland and increased 1.2-fold in liver by ovariectomy. 32P-Postlabeling analyses indicated a single adduct N-(deoxyguanosin-8-yl)-2-fluorenamine in DNA of both tissues 24 h after one intraperitoneal injection of N-OH-2-FBA or N-OH-2-FAA. Respective levels were 3.6- and 5.5-fold greater in liver than mammary gland. After ovariectomy, the adduct levels from N-OH-2-FBA increased 1.8-fold in mammary gland and from N-OH-2-FAA decreased approximately 50% in both tissues. Thus, the ovariectomy-dependent changes in levels of enzymes activating N-OH-2-FBA and N-OH-2-FAA were consistent with in vivo DNA adduct levels in the target mammary gland, but not in the liver.
authors.affiliation: Department of Laboratory Medicine and Pathology, University of Minnesota, Minneapolis 55455, USA.
authors.forename: C L
authors.initials: CL
authors.lastname: Ritter
authors.forename: K K
authors.initials: KK
authors.lastname: Bennett
authors.forename: N F
authors.initials: NF
authors.lastname: Fullerton
authors.forename: F A
authors.initials: FA
authors.lastname: Beland
authors.forename: D
authors.initials: D
authors.lastname: Malejka-Giganti
mesh_terms: D000107:Acetylation; D000818:Animals; D001711:Biotransformation; D001835:Body Weight / Q000187:drug effects / Q000502:physiology; D002273:Carcinogens / Q000378:metabolism / Q000493:pharmacokinetics*; D004247:DNA / Q000187:drug effects / Q000378:metabolism; D018736:DNA Adducts / Q000378:metabolism; D005260:Female; D005449:Fluorenes / Q000378:metabolism / Q000493:pharmacokinetics*; D008099:Liver / Q000201:enzymology / Q000378:metabolism*; D008321:Mammary Glands, Animal / Q000201:enzymology / Q000378:metabolism*; D010052:Ovariectomy; D010053:Ovary / Q000502:physiology*; D010761:Phosphorus Radioisotopes; D011506:Proteins / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014316:Tritium
chemical_list: D002273:Carcinogens; D018736:DNA Adducts; D005449:Fluorenes; D010761:Phosphorus Radioisotopes; D011506:Proteins; C007739:fluorenylhydroxamic acid; D014316:Tritium; D004247:DNA