Prostate-Specific Human N-Acetyltransferase 2 (NAT2) Expression in the Mouse 1 MATTHEW A. LEFF, 2 PAUL N. EPSTEIN, MARK A. DOLL, ADRIAN J. FRETLAND, UDAYA-SANKAR DEVANABOYINA, 3 TIMOTHY D. RUSTAN, and DAVID W. HEIN Departments of Pharmacology and Toxicology, University of Louisville School of Medicine, Louisville Kentucky (M.A.L., P.N.E., M.A.D., A.J.F., U.-S.D., D.W.H.); and University of North Dakota School of Medicine and Health Sciences, Grand Forks, North Dakota (T.D.R., P.N.E., D.W.H.) Accepted for publication March 23, 1999 This paper is available online at http://www.jpet.org ABSTRACT 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a heterocyclic amine identified in the human diet and in cigarette smoke that produces prostate tumors in the rat. PhIP is bioac- tivated by cytochrome P-450 enzymes to N-hydroxylated me- tabolites that undergo further activation by conjugation en- zymes, including the N-acetyltransferases, NAT1 and NAT2. To investigate the role of prostate-specific expression of human N-acetyltransferase 2 (NAT2) on PhIP-induced prostate cancer, we constructed a transgenic mouse model that targeted ex- pression of human NAT2 to the prostate. Following construc- tion, prostate, liver, lung, colon, small intestine, urinary bladder, and kidney cytosols were tested for human NAT1- and NAT2- specific N-acetyltransferase activities. Human NAT2-specific N-acetyltransferase activities were 15-fold higher in prostate of transgenic mice versus control mice, but were equivalent be- tween transgenic mice and control mice in all other tissues tested. Human NAT1-specific N-acetyltransferase activities did not differ between transgenic and control mice in any tissue tested. Prostate cytosols from transgenic and control mice did not differ in their capacity to catalyze the N-acetylation of 2-aminofluorene, the O-acetylation of N-hydroxy-2-aminoflu- orene and N-hydroxy-PhIP or the N,O-acetylation of N-hy- droxy-2-acetylaminofluorene. Transgenic and control mice ad- ministered PhIP did not differ in PhIP-DNA adduct levels in the prostate. This study is the first to report transgenic expression of human NAT2 in the mouse. The results do not support a critical role for bioactivation of heterocyclic amine carcinogens by human N-acetyltransferase-2 in the prostate. However, the lack of an effect may relate to the level of overexpression achieved and the presence of endogenous mouse acetyltrans- ferases and/or sulfotransferases. Recent studies have shown that the dietary carcinogen, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in- duces tumors in rat prostate (Shirai et al., 1997). Heterocy- clic amine carcinogens such as PhIP may undergo bioactiva- tion by cytochrome P-450 (CYP) 1A2 in the liver (Butler et al., 1989; Turesky et al., 1998) or by CYP1A1 and/or CYP1B1 in extrahepatic tissues (Crofts et al., 1998). The N-hydroxy me- tabolite(s) formed can be further bioactivated by N-acetyl- transferases (Hein et al., 1993, 1995; Turesky et al., 1991; Minchin et al., 1992; Lin et al., 1995) and sulfotransferases (Chou et al., 1995; Lin et al., 1995) to yield electrophilic arylnitrenium ions that bind covalently to DNA to form ad- ducts that can initiate carcinogenesis. Our laboratory has focused on the role of acetyltrans- ferases in the bioactivation of heterocyclic amine carcino- gens. Two major N-acetyltransferase isozymes [EC 2.3.1.5], NAT1 and NAT2, are expressed in human and other mam- malian species (Vatsis et al., 1995; Grant et al., 1997; Hein et al., 1997). Drugs such as sulfamethazine (SMZ) are selec- tively metabolized by human NAT2, whereas drugs such as p-aminobenzoic acid (PABA) are selectively metabolized by human NAT1 (Grant et al., 1991; Hein et al., 1993). Genetic polymorphisms exist for both human NAT1 and NAT2 (Vat- sis et al., 1995; Grant et al., 1997; Hein et al., 1997). The NAT2 acetylation polymorphism is more completely charac- terized than NAT1. NAT2*4 is the most common allele asso- ciated with rapid acetylator phenotype (Vatsis et al., 1995) and recombinant human NAT2 4 acetyltransferase bioacti- vates N-hydroxy-PhIP at higher rates than recombinant hu- Received for publication December 29, 1998. 1 This work was partially supported by United States Public Health Service Grant CA34627 from the National Cancer Institute. A preliminary report of this work was presented at the 1998 annual meeting of the Society of Toxicol- ogy (Leff et al., 1998; Toxicol Sci 42:318). 2 This work constitutes partial fulfillment by Matthew Leff for the Ph.D. in Pharmacology and Toxicology at the University of Louisville. 3 Present address: Toxicology and Pathology Services, Inc., 10424 Middle Mount Vernon Rd., Mt. Vernon, IN 47620. ABBREVIATIONS: PhIP, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine; NAT2, N-acetyltransferase 2; NAT1, N-acetyltransferase 1; SMZ, sulfamethazine; PABA, p-aminobenzoic acid; PB, probasin; PCR, polymerase chain reaction; bp, base pair; AF, 2-aminofluorene; DTT; dithio- threitol. 0022-3565/99/2901-0182$03.00/0 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 290, No. 1 Copyright © 1999 by The American Society for Pharmacology and Experimental Therapeutics Printed in U.S.A. JPET 290:182–187, 1999 182 at ASPET Journals on December 10, 2016 jpet.aspetjournals.org Downloaded from