Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0596263 (carcinogenesis)
 64,820 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bacterial assays were used to examine the activation of 14 known procarcinogens by cytochrome P450 (P450) enzymes. Human P450s 1A1, 1A2 and 3A4 were expressed in Escherichia coli with slight modification of their N-terminal sequences. Genotoxicity was measured by the induction of the SOS response in Salmonella typhimurium NM2009 (TA1535/pSK1002/pNM12), which contains a umuC regulatory sequence attached to the lacZ reporter gene. Conditions for analysis were examined using E. coli membranes and purified enzymes. Membrane fractions, fortified with NADPH-P450 reductase, were found to be useful preparations for measuring activation of the procarcinogens. Conditions of linearity were established for these assays and the systems were applied to several particular problems related to bioactivation of procarcinogens by P450s. The patterns of activation of the 14 individual chemicals were consistent with the literature developed using human liver microsomes, purified liver P450s and other approaches. The P450s expressed in bacterial membranes could be inhibited by antibodies. 7,8-Benzoflavone inhibited P450s 1A1 and 1A2 and stimulated P450 3A4 in the membranes. The contributions of P450s 1A1 and 1A2 were distinguished with some of the arylamines and 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene. Recombinant P450 3A4 was found to be more active than P450 1A2 in the activation of aflatoxin B1 at all substrate concentrations examined.
Carcinogenesis 1994 Nov
PMID:Activation of procarcinogens by human cytochrome P450 enzymes expressed in Escherichia coli. Simplified bacterial systems for genotoxicity assays. 795 1

Tamoxifen is the major therapeutic agent for the treatment of hormone-dependent breast cancer. Tamoxifen treatment appears to be associated with an increased incidence of endometrial carcinoma in humans and hepatocellular carcinoma in rats. These carcinogenic effects of tamoxifen might be induced by the formation of a tamoxifen reactive intermediate that binds covalently to macromolecules. Liver microsomal cytochrome P450s (CYPs) catalyze the metabolism of tamoxifen, forming a reactive intermediate that binds irreversibly to microsomal proteins, primarily to a 54 kDa protein (Mani, C. and Kupfer, D., Cancer Res., 51, 6052-6058, 1991). The current study identifies the P450 enzymes that catalyze the activation of tamoxifen to a reactive intermediate in rats and humans. Among the species examined, rats, chickens and humans demonstrate low tamoxifen binding activity, ranging from 0.1 to 0.4 nmol bound/mg protein/h. In contrast, hamsters and mice exhibit high binding, 1.2 and 1.6 nmol/mg protein/h respectively. Treatment of male rats with phenobarbital or pregnenolone-16 alpha-carbonitrile (PCN) markedly elevated the binding of tamoxifen to liver microsomal proteins. Methylcholanthrene treatment had no effect on binding. These findings suggested the involvement of CYP3A in catalysis of the covalent binding. Alternate substrates of CYP3A, cortisol and erythromycin, inhibited tamoxifen binding in liver microsomes from PCN- and phenobarbital-treated rats. Treatment of rats with troleandomycin (TAO), an inducer of CYP3A, followed by the dissociation of the TAO-CYP3A complex, elevated the covalent binding to liver microsomes approximately 3-fold. Antibodies against rat CYP3A1 strongly inhibited tamoxifen binding to liver microsomes from PCN- and phenobarbital-treated rats, whereas the antibodies anti-CYP2B1/2B2 did not inhibit binding. In humans, tamoxifen binding was inhibited by the anti-rat CYP3A1 IgG and also by alternate substrates of CYP3A. These results indicate that the activation of tamoxifen to a reactive intermediate by rat and human liver microsomes is principally catalyzed by CYP3A enzymes.
Carcinogenesis 1994 Dec
PMID:Involvement of cytochrome P4503A in catalysis of tamoxifen activation and covalent binding to rat and human liver microsomes. 800 Dec 26

Tamoxifen and its metabolite 4-hydroxytamoxifen can both exist as geometrical isomers. Trans-tamoxifen is an oestrogen receptor antagonist and is used for the treatment of breast cancer. Trans-4-hydroxytamoxifen is 100 times more anti-oestrogenic than trans-tamoxifen. The cis isomers of tamoxifen and 4-hydroxytamoxifen are oestrogenic and weakly anti-oestrogenic or oestrogenic respectively. Both isomers of 4-hydroxytamoxifen have been detected in breast tumours of patients treated with trans-tamoxifen and it has been proposed that enzymatic isomerization of 4-hydroxytamoxifen occurs in vivo, resulting in resistance to tamoxifen therapy. We have investigated the isomerization of 4-hydroxytamoxifen by human liver microsomes and whether it is mediated by cytochromes P450. Microsomes from five of the 12 livers examined catalysed the interconversion of trans- and cis-4-hydroxytamoxifen (0.52 microM) when incubated for 40 min with an NADPH-generating system. Between 51 and 64% conversion of trans-4-hydroxytamoxifen was observed. Cis-4-hydroxytamoxifen was also converted to trans-4-hydroxytamoxifen (range 22-27%). Incubations with control, heat-treated microsomes resulted in approximately 1% isomerization of trans-4-hydroxytamoxifen. The extent of isomerization of trans- to cis-4-hydroxytamoxifen observed in microsomes from the other seven livers (range 2-8%) did not greatly exceed that seen in heat-inactivated microsomes. Enzymatic isomerization required NADPH and was inhibited by SKF 525A and ketoconazole, indicating the involvement of cytochromes P450. Enzymatic isomerization of trans-tamoxifen and trans-droloxifene (the 3-hydroxy synthetic analogue of tamoxifen) was not observed. These findings may have implications for the safe and effective use of tamoxifen.
Carcinogenesis 1994 Dec
PMID:Interindividual variation in the isomerization of 4-hydroxytamoxifen by human liver microsomes: involvement of cytochromes P450. 800 Dec 29

The tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1- butanone (NNK) is a potential human carcinogen that is known to be metabolized to DNA-reactive intermediates by the cytochromes P450. We have examined the nature of NNK's DNA damaging effects in a mammalian cell system expressing a specific human cytochrome P450 (2A6) and containing a target gene for mutagenesis. Human CYP2A6, which is known to activate NNK to a mutagen, was lipofected via a retroviral vector into the Chinese hamster ovary AS52 cell line, which contains the bacterial gpt gene and can be mutated to 6-thioguanine resistance. AS52 cells expressed negligible CYP2A6-specific coumarin 7-hydroxylase activity (0.7 pmol/mg protein/min), while a CYP2A6 transfected clone (AS52-E8) expressed 30 pmol/mg protein/min. Both cell lines were equally sensitive to the cytotoxic and mutagenic effects of the direct-acting mutagen ethylmethanesulfonate; however, only the AS52-E8 cells exhibited a dose-dependent increase in cytotoxicity and mutant frequency upon treatment with NNK. At the highest NNK dose (1200 micrograms/ml), the mutant frequency in AS52-E8 cells was 14-fold (339 x 10(-6)) greater than the spontaneous frequency of 24 x 10(-6). Ninty-eight mutant clones were isolated following NNK treatment. Based on PCR analysis, 21 clones contained deletions/rearrangements and 77 were putative point mutants. Sequencing potential point mutants showed that 81% contained G:C to A:T transitions. Four of six G:C to A:T hotspots were at the second G of the GGT motif, which is the motif and major mutation found in codon 12 of Ki-ras from NNK-induced lung tumors in strain A mice. Since NNK may be metabolized via different pathways to pyridyloxobutylate or methylate DNA, the data suggest that methylation damage causes the major mutagenic events in AS52-E8 cells when NNK is activated by human CYP2A6.
Carcinogenesis 1994 Dec
PMID:Human CYP2A6 activation of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK): mutational specificity in the gpt gene of AS52 cells. 800 Dec 47

Formation of 2-bromoacrolein (2BA) from tris(2,3-dibromopropyl)phosphate (Tris-BP) by cytochrome P450 (cyt. P450) activity is most likely responsible for the high mutagenicity of Tris-BP for bacteria in vitro when rat liver microsomes are used for metabolic activation. As yet, it has not been established whether cyt. P450 plays a role in the formation of genotoxic metabolites from Tris-BP in higher organisms in vivo. This was studied by comparing the effects of completely deuterated Tris-BP (D15-Tris-BP) with normal Tris-BP in various in vivo test systems for genotoxicity: deuterium substitution should decrease cyt. P450-dependent bioactivation of Tris-BP. Three test systems in Drosophila were used to measure different types of genetic damage: (I) the ring-X chromosome loss (CL) test to detect clastogenicity, (II) the sex-linked recessive lethal (RL) test to detect forward mutations and deletions and (III) the white/white+ (w/w+) eye mosaic assay to detect predominantly mitotic recombination. Tris-BP was positive in all test systems, while D15-Tris-BP was without effect. The relative clastogenic efficiency of a compound, defined as the ratio of CL over RL, can be used to distinguish monofunctional agents (CL/RL ratio < or = 1) from those having cross-linking potential (CL/RL ratio > 2). The CL/RL ratio of Tris-BP was 2.4, indicating that Tris-BP has cross-linking potential. No CL/RL ratio for D15-Tris-BP could be determined, because it was negative in both tests. The putative Tris-BP metabolite 2BA was also tested in the Drosophila test systems. 2BA was positive in the CL test at a high dose, but it showed no response in the RL test. The relative clastogenic efficiency (the ratio of CL/RL) could not be determined accurately, but the data nevertheless argue in favour of cross-linking properties of 2BA. Further, 2BA was clearly positive in the w/w+ eye mosaic test system, which indicates that it is a recombinagen. Genotoxicity of Tris-BP in rats was determined by the induction of micronuclei in hepatocytes. Tris-BP administered intraperitoneally 17 h after 2/3 partial hepatectomy (PH), induced a high frequency of micronuclei at days 2 and 3 after injection (26% and 22%, respectively). D15-Tris-BP, however did not increase the micronuclei frequency.(ABSTRACT TRUNCATED AT 400 WORDS)
Carcinogenesis 1994 Jun
PMID:Genotoxicity of the flame retardant tris(2,3-dibromopropyl)phosphate in the rat and Drosophila: effects of deuterium substitution. 802 Jan 56

Much progress has been made in elucidating the biochemical and molecular mechanisms that underlie aflatoxin carcinogenesis. In humans, biotransformation of AFB1 to the putative carcinogenic intermediate. AFB-8,9-exo-epoxide, occurs predominantly by cytochromes P450 1A2 and 3A4, with the relative importance of each dependent upon the relative magnitude of expression of the respective enzymes in liver. Genetic variability in the expression of these and other cytochromes P450 may result in substantial interindividual differences in susceptibility to the carcinogenic effects of aflatoxins. Detoxification of AFB-8,9-epoxide by a specific alpha class glutathione S-transferase is an important protective mechanism in mice, and it accounts for the resistance of this species to the carcinogenic effects of AFB. This particular form of GST is expressed constitutively only at low levels in rats, but it is inducible by antioxidants such as ethoxyquin, and it accounts for much of the chemoprotective effects of a variety of substances, including natural dietary components that putatively act via an "antioxidant response element" (ARE). In humans, the constitutively expressed GSTs have very little activity toward AFB1-8,9-exo-epoxide, suggesting that--on a biochemical basis--humans should be quite sensitive to the genotoxic effects of aflatoxins. If a gene encoding a high aflatoxin-active form of GST is present in the human genome, but is not constitutively expressed, and is inducible by dietary antioxidants (as occurs in rats), then chemo- and/or dietary intervention measures aimed at inducing this enzyme could be highly effective. However, as it is possible that human CYP 1A2 may also be inducible by these same chemicals (because of the possible presence of an ARE in this gene), the ultimate consequence of dietary treatment with chemicals that induce biotransformation enzymes via an ARE is uncertain. The balance of the rate of activation (exo-epoxide production) to inactivation (GST conjugation plus other P450-mediated non-epoxide oxidations) may be a strong indicator of individual and species susceptibility to aflatoxin carcinogenesis, if the experimental conditions are reflective of true dietary exposures. There is strong evidence that AFB-8,9-exo-epoxide binds to G:C rich regions of DNA, forming an adduct at the N7-position of guanine. Substantial evidence demonstrates that AFB1-8,9-epoxide can induce activating mutations in the ras oncogene in experimental animals, primarily at codon 12.(ABSTRACT TRUNCATED AT 400 WORDS)
 ...
PMID:Mechanisms of aflatoxin carcinogenesis. 804 48

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-1,2-dihydro-1,2-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-acetyltransferase-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 beta-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 alpha-naphthoflavone, a specific inhibitor of P4501A-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 cell 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.
Carcinogenesis 1994 Mar
PMID:Activation of trans-1,2-dihydro-1,2-dihydroxy-6-aminochrysene to genotoxic metabolites by rat and human cytochromes P450. 811 30

Using microsomal preparations from rat and human liver, we investigated the activation of the anti-estrogen compound tamoxifen (TMX) to form DNA adducts. Pretreatment of rats with phenobarbital increased DNA adduct formation by microsomal activation of TMX 3- to 6-fold, depending on the cofactors used. When reduced nicotinamide-adenine dinucleotide phosphate (NADPH) was used as a cofactor in human and rat microsomal activation systems, the relative DNA adduct levels were 2.9 and 5.2 x 10(-8) respectively and 1-3 TMX-DNA adducts were detected by 32P-postlabeling; DNA adduct 1 was the same in both microsomal systems. When cumene hydroperoxide (CuOOH) was used as a cofactor, activation of TMX produced four major DNA adducts and several minor DNA adducts in both rat and human liver microsomes; the relative adduct levels were 11.1 and 23.1 x 10(-8) respectively. TMX-DNA adducts 1, 4, 5 and 6 were similar in both human and rat microsomal systems with CuOOH as the cofactor. The TMX-DNA adducts formed with NADPH as the cofactor were clearly different from those formed with CuOOH as the cofactor, which implies that the metabolites leading to the individual DNA adducts were different. Addition of a P450 inhibitor, either n-octylamine or alpha-naphthylisothiocyanate, to the activation system reduced adduct formation by 70-93%. We propose that the TMX-DNA adducts formed with NADPH as the cofactor result from P450 acting as a mono-oxygenase, whereas the adducts formed with CuOOH as the cofactor result from P450 acting as a peroxidase. Our findings suggest that further studies may be required to establish the safety of TMX treatment of women for purposes other than chemotherapy.
Carcinogenesis 1994 Mar
PMID:DNA adduct formation by tamoxifen with rat and human liver microsomal activation systems. 811 38

Trp-P-1 (3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole) is a carcinogen metabolized by hepatic cytochrome P4501A (P4501A). This study showed that there was a highly selective solvent-resistant binding of radioactive substance in endothelial cells of heart and kidney 1 day following injection of [3H]-Trp-P-1 (0.1 or 1.5 mg/kg) in NMRI mice treated with the P450-inducing agent beta-naphthoflavone (BNF). In the heart, the binding was highest in capillaries and coronary vessels. In the kidney, the binding was highest in afferent and efferent arterioles and glomerular and peritubular capillaries. A corresponding localization of radioactivity did not occur in corn oil-treated mice injected with [3H]-Trp-P-1. On incubation of heart and kidney slices with [3H]-Trp-P-1, there was a binding of radioactivity in endothelial cells of BNF-treated mice, but not in corn oil-treated mice. The P4501A inhibitor ellipticine abolished the BNF-induced endothelial binding of [3H]-Trp-P-1 in vivo and in vitro, whereas the effects of alpha-naphthoflavone were inconsistent. The results indicate an in situ metabolism of [3H]-Trp-P-1 to a reactive species, catalysed by a BNF-inducible enzyme, possibly P4501A1, in endothelial cells in the heart and kidney. Since related heterocyclic amines induce tumors and other lesions in the rodent vascular system, the results raise the possibility that Trp-P-1 and other food mutagens may play a role in cardiovascular disease.
Carcinogenesis 1994 Apr
PMID:Metabolic activation of the food mutagen Trp-P-1 in endothelial cells of heart and kidney in cytochrome P450-induced mice. 814 78

Cultured human mammary carcinoma (MCF-7) cells exhibited constitutive cytochrome P450-dependent metabolism of 7,12-dimethylbenz[a]anthracene (DMBA) (45-75 pmol/mg microsomal protein). Exposure of the cells to 2,3,7,8-tetrachloro-dibenzo-p-dioxin (TCDD), which is known to induce CYP1A1, not only resulted in a 30-fold increase in the total microsomal metabolism of DMBA but produced substantial differences in the distribution of DMBA metabolites formed. This suggested that different cytochrome P450 (P450) forms predominated in untreated and induced cells. Comparative studies with TCDD-induced human hepatoblastoma (HepG2) and skin cell carcinoma (SCC-13) cells and also recombinantly expressed human CYP1A1, confirmed that the DMBA-metabolite profile in TCDD-induced MCF-7 cells was that of human CYP1A1. This distribution, however, differed substantially from the regioselectivity of rat CYP1A1 and mouse Cyp1a-1. Rabbit antibodies to rat CYP1A1 completely inhibited the DMBA-metabolizing activity of TCDD-induced MCF-7 cells but had no inhibitory effect on constitutive DMBA metabolism which was, however, completely inhibited by chicken antibodies to the novel P450 in mouse embryo fibroblasts (P450-EF). Anti-P450-EF inhibited only 10% of the DMBA-metabolizing activity in the TCDD-induced MCF-7 cell microsomes. Microsomes from untreated MCF-7 cells expressed a 52 kDa protein that was immunodetectable by rabbit anti-P450-EF and failed to express immunodetectable levels of human CYP1A1. DMBA metabolism, therefore, switches from P450-EF in uninduced microsomes to CYP1A1 in TCDD-induced microsomes. The mobility of the P450-EF-like protein in MCF-7 cells was higher than that of P450-EF from C3H/10T1/2CL8 (10T1/2) cells (55 kDa). The 52 kDa protein from MCF-7 cells was induced approximately 8-fold by TCDD while CYP1A1 immunodetectable protein was increased to much higher levels. The SCC-13 cell line exhibited a similar pattern of expression of a 52 kDa P450-EF-like protein and CYP1A1. HepG2 cells expressed the highest levels of CYP1A1 in response to TCDD without expression of the 52 kDa protein.
Carcinogenesis 1994 Apr
PMID:Co-expression of human CYP1A1 and a human analog of cytochrome P450-EF in response to 2,3,7,8-tetrachloro-dibenzo-p-dioxin in the human mammary carcinoma-derived MCF-7 cells. 814 87


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