Activation of trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene to genotoxic metabolites by rat and human cytochromes P450

Hiroshi Yamazaki, Mayumi Mimura, Yoshimitsu Oda, Frank J. Gonzalez, Karam El-bayoumy, Young Heum Chae, F. Peter Guengerich, Tsutomu Shimada

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

In order to address the hypothesis that 6-aminochrysene (6-AC) is converted to genotoxic products by cytochrome P450 enzymes via two activation pathways (N-hydroxylation and epoxidation), the activation of 6-AC and trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene (6-AC-diol) to genotoxic metabolites was examined in rat and human liver microsomal cytochrome P450 enzymes using Salmonella typhimurium TA1535/pSK1002 and TA1535/pSK1002/pNM12 (NM2009) as tester strains. The latter bacteria, an O-acetyl-transferase-overexpressing strain, was highly sensitive to metabolites derived from activation of 6-AC, but not those from 6-AC-diol, using liver microsomes from phenobarbital-treated rats or a reconstituted monooxygenase system containing P4502B1 or -2B2, thus suggesting the roles of P450 and acetyltransferase systems in the activation process. 6-AC-diol, on the other hand, was activated very efficiently by liver microsomes prepared from β-naphthoflavone-treated rats or a reconstituted system containing P4501A1 or -1A2; the activation reaction is considered to proceed through diol-epoxide formation. The contribution of rat P4501A enzymes towards activation of 6-AC-diol was confirmed by the inhibitory effects on the activation process of α-naphthoflavone, a specific inhibitor of P4501 A-related activities, and antibodies raised against purified P4501A1 and -1A2. In humans, P4501A2 was found to be the major enzyme involved in the activation of 6-AC-diol to genotoxic metabolites while the parent compound 6-AC was activated mainly by P4503A4. Experiments using recombinant P450 proteins expressed in human lymphoblastoid cells lines showed that human P4501A1 could also activate 6-AC-diol to reactive metabolites at almost the same rate measured with P4501A2. In addition, P4502B6 was found to efficiently catalyze the activation of 6-AC to genotoxic metabolites, and P4503A4 was active in the activation of 6-AC-diol as well as 6-AC. Addition of purified rat epoxide hydrolase to the incubation mixture containing purified rat P4501A1 or microsomes expressing human P4501A1 caused inhibition of activation of 6-AC-diol. These results suggest the existence of different enzymatic activation pathways for 6-AC and 6-AC-diol. The former carcinogen may be N-hydroxylated principally by P4502B enzymes in rats and P4503A4 and -2B6 in humans and activation to its ultimate metabolites may proceed through esterification of the N-hydroxy metabolites by an N-acetyltransferase. The 6-AC-diol is metabolized to its ultimate diolepoxide product by P4501A enzymes in rat and human liver microsomes. P4503A4 (humans) and P4503A2 (rats) may also contribute to some extent in the activation of 6-AC-diol, albeit at lower rates than those of P4501A enzymes.

Original languageEnglish (US)
Pages (from-to)465-470
Number of pages6
JournalCarcinogenesis
Volume15
Issue number3
DOIs
StatePublished - Mar 1 1994

Fingerprint

Cytochrome P-450 Enzyme System
Liver Microsomes
6-chrysenamine
Acetyltransferases
Enzymes
Epoxide Hydrolases
Enzyme Activation
Esterification
Epoxy Compounds
Salmonella typhimurium
Hydroxylation
Phenobarbital
Transferases
Microsomes
Mixed Function Oxygenases
Recombinant Proteins
Carcinogens

All Science Journal Classification (ASJC) codes

  • Cancer Research

Cite this

Yamazaki, Hiroshi ; Mimura, Mayumi ; Oda, Yoshimitsu ; Gonzalez, Frank J. ; El-bayoumy, Karam ; Chae, Young Heum ; Guengerich, F. Peter ; Shimada, Tsutomu. / Activation of trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene to genotoxic metabolites by rat and human cytochromes P450. In: Carcinogenesis. 1994 ; Vol. 15, No. 3. pp. 465-470.
@article{f61ecf39fef349f993e005df6ba29303,
title = "Activation of trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene to genotoxic metabolites by rat and human cytochromes P450",
abstract = "In order to address the hypothesis that 6-aminochrysene (6-AC) is converted to genotoxic products by cytochrome P450 enzymes via two activation pathways (N-hydroxylation and epoxidation), the activation of 6-AC and trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene (6-AC-diol) to genotoxic metabolites was examined in rat and human liver microsomal cytochrome P450 enzymes using Salmonella typhimurium TA1535/pSK1002 and TA1535/pSK1002/pNM12 (NM2009) as tester strains. The latter bacteria, an O-acetyl-transferase-overexpressing strain, was highly sensitive to metabolites derived from activation of 6-AC, but not those from 6-AC-diol, using liver microsomes from phenobarbital-treated rats or a reconstituted monooxygenase system containing P4502B1 or -2B2, thus suggesting the roles of P450 and acetyltransferase systems in the activation process. 6-AC-diol, on the other hand, was activated very efficiently by liver microsomes prepared from β-naphthoflavone-treated rats or a reconstituted system containing P4501A1 or -1A2; the activation reaction is considered to proceed through diol-epoxide formation. The contribution of rat P4501A enzymes towards activation of 6-AC-diol was confirmed by the inhibitory effects on the activation process of α-naphthoflavone, a specific inhibitor of P4501 A-related activities, and antibodies raised against purified P4501A1 and -1A2. In humans, P4501A2 was found to be the major enzyme involved in the activation of 6-AC-diol to genotoxic metabolites while the parent compound 6-AC was activated mainly by P4503A4. Experiments using recombinant P450 proteins expressed in human lymphoblastoid cells lines showed that human P4501A1 could also activate 6-AC-diol to reactive metabolites at almost the same rate measured with P4501A2. In addition, P4502B6 was found to efficiently catalyze the activation of 6-AC to genotoxic metabolites, and P4503A4 was active in the activation of 6-AC-diol as well as 6-AC. Addition of purified rat epoxide hydrolase to the incubation mixture containing purified rat P4501A1 or microsomes expressing human P4501A1 caused inhibition of activation of 6-AC-diol. These results suggest the existence of different enzymatic activation pathways for 6-AC and 6-AC-diol. The former carcinogen may be N-hydroxylated principally by P4502B enzymes in rats and P4503A4 and -2B6 in humans and activation to its ultimate metabolites may proceed through esterification of the N-hydroxy metabolites by an N-acetyltransferase. The 6-AC-diol is metabolized to its ultimate diolepoxide product by P4501A enzymes in rat and human liver microsomes. P4503A4 (humans) and P4503A2 (rats) may also contribute to some extent in the activation of 6-AC-diol, albeit at lower rates than those of P4501A enzymes.",
author = "Hiroshi Yamazaki and Mayumi Mimura and Yoshimitsu Oda and Gonzalez, {Frank J.} and Karam El-bayoumy and Chae, {Young Heum} and Guengerich, {F. Peter} and Tsutomu Shimada",
year = "1994",
month = "3",
day = "1",
doi = "10.1093/carcin/15.3.465",
language = "English (US)",
volume = "15",
pages = "465--470",
journal = "Carcinogenesis",
issn = "0143-3334",
publisher = "Oxford University Press",
number = "3",

}

Activation of trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene to genotoxic metabolites by rat and human cytochromes P450. / Yamazaki, Hiroshi; Mimura, Mayumi; Oda, Yoshimitsu; Gonzalez, Frank J.; El-bayoumy, Karam; Chae, Young Heum; Guengerich, F. Peter; Shimada, Tsutomu.

In: Carcinogenesis, Vol. 15, No. 3, 01.03.1994, p. 465-470.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Activation of trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene to genotoxic metabolites by rat and human cytochromes P450

AU - Yamazaki, Hiroshi

AU - Mimura, Mayumi

AU - Oda, Yoshimitsu

AU - Gonzalez, Frank J.

AU - El-bayoumy, Karam

AU - Chae, Young Heum

AU - Guengerich, F. Peter

AU - Shimada, Tsutomu

PY - 1994/3/1

Y1 - 1994/3/1

N2 - In order to address the hypothesis that 6-aminochrysene (6-AC) is converted to genotoxic products by cytochrome P450 enzymes via two activation pathways (N-hydroxylation and epoxidation), the activation of 6-AC and trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene (6-AC-diol) to genotoxic metabolites was examined in rat and human liver microsomal cytochrome P450 enzymes using Salmonella typhimurium TA1535/pSK1002 and TA1535/pSK1002/pNM12 (NM2009) as tester strains. The latter bacteria, an O-acetyl-transferase-overexpressing strain, was highly sensitive to metabolites derived from activation of 6-AC, but not those from 6-AC-diol, using liver microsomes from phenobarbital-treated rats or a reconstituted monooxygenase system containing P4502B1 or -2B2, thus suggesting the roles of P450 and acetyltransferase systems in the activation process. 6-AC-diol, on the other hand, was activated very efficiently by liver microsomes prepared from β-naphthoflavone-treated rats or a reconstituted system containing P4501A1 or -1A2; the activation reaction is considered to proceed through diol-epoxide formation. The contribution of rat P4501A enzymes towards activation of 6-AC-diol was confirmed by the inhibitory effects on the activation process of α-naphthoflavone, a specific inhibitor of P4501 A-related activities, and antibodies raised against purified P4501A1 and -1A2. In humans, P4501A2 was found to be the major enzyme involved in the activation of 6-AC-diol to genotoxic metabolites while the parent compound 6-AC was activated mainly by P4503A4. Experiments using recombinant P450 proteins expressed in human lymphoblastoid cells lines showed that human P4501A1 could also activate 6-AC-diol to reactive metabolites at almost the same rate measured with P4501A2. In addition, P4502B6 was found to efficiently catalyze the activation of 6-AC to genotoxic metabolites, and P4503A4 was active in the activation of 6-AC-diol as well as 6-AC. Addition of purified rat epoxide hydrolase to the incubation mixture containing purified rat P4501A1 or microsomes expressing human P4501A1 caused inhibition of activation of 6-AC-diol. These results suggest the existence of different enzymatic activation pathways for 6-AC and 6-AC-diol. The former carcinogen may be N-hydroxylated principally by P4502B enzymes in rats and P4503A4 and -2B6 in humans and activation to its ultimate metabolites may proceed through esterification of the N-hydroxy metabolites by an N-acetyltransferase. The 6-AC-diol is metabolized to its ultimate diolepoxide product by P4501A enzymes in rat and human liver microsomes. P4503A4 (humans) and P4503A2 (rats) may also contribute to some extent in the activation of 6-AC-diol, albeit at lower rates than those of P4501A enzymes.

AB - In order to address the hypothesis that 6-aminochrysene (6-AC) is converted to genotoxic products by cytochrome P450 enzymes via two activation pathways (N-hydroxylation and epoxidation), the activation of 6-AC and trans-l,2-dihydro-l,2-dihydroxy-6-aminochrysene (6-AC-diol) to genotoxic metabolites was examined in rat and human liver microsomal cytochrome P450 enzymes using Salmonella typhimurium TA1535/pSK1002 and TA1535/pSK1002/pNM12 (NM2009) as tester strains. The latter bacteria, an O-acetyl-transferase-overexpressing strain, was highly sensitive to metabolites derived from activation of 6-AC, but not those from 6-AC-diol, using liver microsomes from phenobarbital-treated rats or a reconstituted monooxygenase system containing P4502B1 or -2B2, thus suggesting the roles of P450 and acetyltransferase systems in the activation process. 6-AC-diol, on the other hand, was activated very efficiently by liver microsomes prepared from β-naphthoflavone-treated rats or a reconstituted system containing P4501A1 or -1A2; the activation reaction is considered to proceed through diol-epoxide formation. The contribution of rat P4501A enzymes towards activation of 6-AC-diol was confirmed by the inhibitory effects on the activation process of α-naphthoflavone, a specific inhibitor of P4501 A-related activities, and antibodies raised against purified P4501A1 and -1A2. In humans, P4501A2 was found to be the major enzyme involved in the activation of 6-AC-diol to genotoxic metabolites while the parent compound 6-AC was activated mainly by P4503A4. Experiments using recombinant P450 proteins expressed in human lymphoblastoid cells lines showed that human P4501A1 could also activate 6-AC-diol to reactive metabolites at almost the same rate measured with P4501A2. In addition, P4502B6 was found to efficiently catalyze the activation of 6-AC to genotoxic metabolites, and P4503A4 was active in the activation of 6-AC-diol as well as 6-AC. Addition of purified rat epoxide hydrolase to the incubation mixture containing purified rat P4501A1 or microsomes expressing human P4501A1 caused inhibition of activation of 6-AC-diol. These results suggest the existence of different enzymatic activation pathways for 6-AC and 6-AC-diol. The former carcinogen may be N-hydroxylated principally by P4502B enzymes in rats and P4503A4 and -2B6 in humans and activation to its ultimate metabolites may proceed through esterification of the N-hydroxy metabolites by an N-acetyltransferase. The 6-AC-diol is metabolized to its ultimate diolepoxide product by P4501A enzymes in rat and human liver microsomes. P4503A4 (humans) and P4503A2 (rats) may also contribute to some extent in the activation of 6-AC-diol, albeit at lower rates than those of P4501A enzymes.

UR - http://www.scopus.com/inward/record.url?scp=0028314292&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0028314292&partnerID=8YFLogxK

U2 - 10.1093/carcin/15.3.465

DO - 10.1093/carcin/15.3.465

M3 - Article

C2 - 8118930

AN - SCOPUS:0028314292

VL - 15

SP - 465

EP - 470

JO - Carcinogenesis

JF - Carcinogenesis

SN - 0143-3334

IS - 3

ER -