Oltipraz regulates different categories of genes relevant to chemoprevention in human hepatocytes
Amélie Piton,
Eric Le Ferrec,
Sophie Langouët,
Claudine Rauch,
Elise Petit,
Frédérick Le Goff,
André Guillouzo and
Fabrice Morel*
INSERM U620, Faculté des Sciences Pharmaceutiques et Biologiques, Université de Rennes I, 35043 Rennes, France
* To whom correspondence should be addressed Email: fabrice.morel{at}rennes.inserm.fr
 |
Abstract
|
---|
Numerous chemical compounds are cytotoxic or carcinogenic to human beings and attention is now focusing on preventative strategies. One agent, oltipraz (OPZ), regarded as one of the most promising chemoprotectors, has been shown to be a potent inducer of phase II enzymes involved in the detoxification of carcinogens, including aflatoxins. However, little is known about its effects on global gene expression in human cells. Thus, we used microarrays and reverse transcriptionquantitative polymerase chain reaction to test the effects of OPZ on the overall pattern of mRNA expression of multiple metabolic pathways in human hepatocytes in primary culture. Our results show for the first time that OPZ significantly alters the expression of human genes within different functional categories (detoxification of xenobiotics, antioxidant defences, xenobiotic transport, cell cycle and stress responses), at both the mRNA and protein levels, some of which are highly relevant to chemoprevention. Amongst these genes, several have never been described as being regulated by OPZ before. We also demonstrate variations in response to OPZ, depending on the individual from whom the cells were derived, that might potentially contribute to differences in efficacy of chemopreventive treatments between individuals. Moreover, comparison of our results with those obtained in rodents demonstrates species differences in response to OPZ for some genes, underlying the importance of studies on human cells to predict the effects of chemopreventive agents.
Abbreviations: CYP, cytochrome P450; GST, glutathione S-transferase; HL, human liver; OPZ, oltipraz; ROS, reactive oxygen species; RT-qPCR, reverse transcriptionquantitative polymerase chain reaction; TBS, Tris-buffered saline
 |
Introduction
|
---|
About 35% of total cancers are estimated to be diet related, but there is also accumulating evidence from population as well as laboratory studies to support an inverse relationship between regular consumption of fruit and vegetables and the risk of specific cancers. Several organizations, such as the WHO, the American Cancer Society, the American Institute of Cancer Research (AICR) and the National Cancer Institute (NCI), have established dietary guidelines to help people to reduce the cancer risk (for further information see the 1997 World Cancer Research Fund and AICR report at: http://www.aicr.org/exreport.html). Among the dietary constituents that are particularly efficient in reducing the development of chemically induced cancers in rodents, dithiole thiones, present in cruciferous vegetables, have received most attention. However, one of the most powerful compounds appears to be oltipraz (OPZ), a substituted 1,2-dithiole-3-thione originally developed as an antischistosomal agent. This molecule is recognized as exerting chemoprotective activity against different classes of carcinogens targeting multiple organs. OPZ is an effective chemopreventive agent in at least 12 different tissues when given before and during carcinogen exposure (for a review see 1). The anticarcinogenic effects of OPZ in rodents were at first attributed to its induction of phase II detoxifying enzymes, e.g. microsomal epoxide hydrolase and glutathione S-transferase (GST), as well as inhibition of certain cytochromes P450 (CYPs), e.g. CYP1A2 and CYP3A4 (24). Nevertheless, induction of phase II enzymes and inhibition of phase I enzymes are probably not the only mechanisms involved in its chemopreventive efficacy. In particular, several studies performed with rodents have demonstrated that OPZ and 1,2-dithiole-3-thione affect several functions (510) in rat liver. To more directly test the hypothesis that OPZ modulates the metabolism of carcinogens in humans, a placebo controlled, double blind clinical trial of OPZ was conducted in residents of Qidong, China, who were continuously exposed to dietary aflatoxins and who are at high risk of the development of liver cancer (11,12). OPZ significantly enhanced excretion of a phase II product, aflatoxinmercapturic acid, in the urine of study participants administered 125 mg/day OPZ orally (13), suggesting an increase in detoxification processes. A clinical trial also demonstrated elevations in GSH and GST levels above baseline in lymphocytes of subjects receiving 100 and 125 mg OPZ (14). These observations are encouraging, however, many questions still have to be considered due to the fact that little is known about the effects of this chemopreventive agent on human gene expression. Moreover, huge variations in detoxification enzyme activities can be observed in response to chemical compounds both between animals and humans and in humans among individuals, indicating that meaningful extrapolation of existing animal data to the human situation is not necessarily possible. Since in vivo experimentation in humans is not possible for ethical reasons, the aim of our study was to investigate the effects of OPZ on the pattern of mRNA expression of multiple metabolic pathways in primary cultures of human hepatocytes, a target cell type which represents a unique model system. Our results show that OPZ significantly alters the expression of several human genes within different functional categories, some of them being highly relevant to chemoprevention.
 |
Materials and methods
|
---|
Chemicals
Culture media, glutamine and penicilinstreptomycin were obtained from Gibco BRL Life Technologies (Rockland, MD), fetal calf serum from PAN Biotechnology (Stanford, USA), bovine insulin from Sigma (St Louis, MO), bovine serum albumin from Eurobio (Les Ulis, France) and hydrocortisone hemisuccinate from Roche (Basel, Switzerland). OPZ was kindly supplied by Dr F.Ballet (Aventis, Evry, France).
Cell isolation and culture
Human liver samples were obtained in France from 19 patients undergoing liver resection for primary or secondary hepatomas. Hepatocytes were isolated by a two-step collagenase perfusion procedure. The experimental procedures used were done in compliance with French laws and approved by the National Ethic Committee. Cell viability was 7085%, as estimated by the trypan blue exclusion test while cell yields varied with the size of the fragment. Liver parenchymal cells were seeded at a density of 107 viable cells/75 cm2 dish in a nutrient medium consisting of Williams E medium supplemented with 0.2% bovine serum albumin, 0.01% bovine insulin, 2 mM glutamine, 100 U/ml penicillin, 10 µg/ml streptomycin and 10% fetal calf serum. Medium supplemented with 0.1 µM hydrocortisone hemisuccinate but lacking serum was renewed daily. OPZ, dissolved in dimethylsulfoxide, was added 3648 h after cell seeding and at each medium renewal to give a final concentration of 50 µM (in 0.1% dimethylsulfoxide). Control cultures received the same concentration of solvent.
Isolation of RNA
Total RNA were prepared by one of two different procedures depending on whether it was to be used for microarray or reverse transcriptionquantitative polymerase chain reaction (RT-qPCR) analysis. For microarray experiments, 107 cultured hepatocytes were washed and total RNA was extracted from cells by the guanidium thiocyanate/cesium chloride method, followed by a DNase treatment step. For RT-qPCR analysis, total RNA was extracted from 106 hepatocytes with a SV total RNA isolation system (Promega, Madison, WI), which includes a DNase treatment step.
Atlas cDNA array
cDNA array analysis was performed using the Atlas Human Toxicology arrays 1.2 (Clontech, Palo Alto, CA) containing 1176 unique cDNAs spotted on a nylon membrane. Probing of cDNA arrays was performed according to the manufacturer's directions with [
-33P]dATP (Amersham, Little Chalfont, UK). The hybridized array membranes were washed and exposed to a phosphor screen for 4896 h. The signal intensity was obtained by scanning the screen using a Storm 840 (Molecular Dynamics, Watertown, USA). Gene-specific binding to each spot was quantified using Atlas Image 2.01 software, corrected for background and normalized using the sum of intensities method. In order to limit the numbers of false positives and as recommended by Atlas Image, a gene was considered to be differentially expressed at a threshold ratio of 2. Two independent hybridizations were carried out with hepatocytes obtained from different donors.
RT-qPCR analysis
RNAs were reversed transcribed into cDNA using Superscript II (Gibco BRL Life Technologies, Rockland, MD) following the manufacturer's protocols with the following exceptions: 1 µg total RNA and a 500 ng mixture of random hexamers (Promega) were used in each RT reaction.
Real-time quantitative PCR was performed by the fluorescent dye SYBR Green methodology using the qPCRTM Core Kit for SybrTM Green I (Eurogentec, Seraing, Belgium) and an ABI Prism 7000 (Applied Biosystem, Foster City, CA). Table I shows primer pairs for each transcript chosen with the Primer3 program (http://frodo.wi.mit.edu/cgi-bin/primer3/primer3_www.cgi). Gene specificity of each pair of primers was checked by comparing their sequences to the GenBank database using the program BLASTN (http://www.ncbi.nlm.nih.gov/BLAST/). The curves of amplification were read with ABI Prism 7000 SDS software using the comparative cycle threshold method. Relative quantification of the steady-state target mRNA levels was calculated after normalization of the total amount of cDNA tested to an active reference, 18S. The specificity of each gene amplification was also verified at the end of each qPCR reaction in an ABI 7000 by performing a dissociation analysis of the amplified DNA. In each case we obtained a curve with only one peak, which is characteristic of a single amplification product.
Western blot analysis
Human hepatocytes were homogenized in 50 mM TrisHCl buffer (pH 7.4) containing 0.25 M sucrose, 1 mM EDTA, 25 µM phenylmethylsulfonyl fluoride and 1 mM dithiothreitol to obtain total proteins. In the case of microsomal or cytosolic preparations, nuclei and mitochondria were eliminated by centrifugation at 3000 g for 10 min and at 8000 g for 20 min. The supernatant containing microsomes and cytosol was subjected to centrifugation at 30 000 g for 1 h. Cytosols were recovered and stored at 80°C while microsomal pellets were dissolved in 0.1 M phosphate buffer (pH 7.4) containing 10% glycerol and stored at 80°C until use.
Microsomal, cytosolic and total protein fractions were subjected to electrophoresis on a polyacrylamide slab gel and electroblotted overnight onto Hybond enhanced chemiluminescence (ECL) nitrocellulose membranes (Amersham). Filters were blocked in 5% low fat milk in Tris-buffered saline (TBS) (25 mM Tris, pH 7.5, 0.9% NaCl, 0.04%) and then incubated with the first antibody diluted at the appropriate concentration in 5% low fat milk in TBS containing 1% Tween 20. The filters were then washed with TBS containing 1% Tween 20 and incubated with peroxidase-conjugated secondary antibody in 5% low fat milk in TBS containing 1% Tween 20. All incubations were done at room temperature for 2 h. Peroxidase activity was detected using an ECL western blotting detection procedure (Amersham). Equal transfer of proteins was confirmed by staining the nitrocellulose with Ponceau Red (0.2% w/v H2O diluted 99:1 in 10% trifluoroacetic acid).
The primary antibodies used were: rabbit anti-CYP2B6 antibody (a gift from F.Guenguerich); rabbit anti-glutathione peroxidase 2 antibody (a gift from R.Brigelius-Flohé); rabbit anti-CYP2E1 antibody and rabbit anti-microsomal epoxide hydrolase antibody (gifts from I.De Waziers); rabbit anti-asparagine synthase antibody (a gift from M.Kilberg); goat anti-thioredoxin reductase 1 antibody, goat anti-thioredoxin antibody, rabbit anti-nucleoside diphosphate kinase antibody, mouse anti-c-myc antibody (Santa-Cruz Biotechnology, Santa Cruz, CA); rabbit anti-CYP4A11 (Research Diagnostics, Flanders, NJ); rabbit anti-glutamate dehydrogenase 1 antibody (Rockland, Gilbertsville, PA). CYPs and epoxide hydrolase were analyzed in microsomal fractions; glutathione peroxidase 2, asparagine synthase, thioredoxin and thioredoxin reductase 1 were detected in cytosolic fractions; analysis of c-myc, glutamate dehydrogenase 1 and nucleoside diphosphate kinase was performed on total proteins.
 |
Results
|
---|
To monitor changes in gene expression due to OPZ treatment in human hepatocytes we carried out expression profiling using commercial cDNA-based arrays (Clontech Atlas human toxicology array 1.2). Cultured hepatocytes obtained from eight different donors were treated with either vehicle control (0.1% dimethylsulfoxide) or 50 µM OPZ for 72 h. Table II presents results obtained with total RNA isolated from one hepatocyte population and from a pool of RNA prepared from seven different populations of hepatocytes in order to be free of inter-individual variations. Analysis of our acquired data set showed that 342 of the 1176 genes on the arrays yielded signal intensities above background levels. Applying the arbitrary threshold of 2, we found 35 genes that are candidates for OPZ-regulated genes (Table II). The list contains 25 up-regulated and 10 down-regulated genes. Because the Clontech array is based on hybridization to cDNA, we wished to confirm differential expression of the mRNAs detected by this method by RT-qPCR analysis, which confers increased specificity (specific primers used for each gene) and precise quantification. For 10 genes, no confirmation of the data obtained with the cDNA arrays was possible. Although not fully understood, the false positive signals on arrays most likely resulted from cross-hybridization with related sequences.
View this table:
[in this window]
[in a new window]
|
Table II. Up- and down-regulated genes after 72 h treatment of human hepatocyte cultures with oltipraz as determined by microarrays
|
|
We extended the RT-qPCR studies to six additional human hepatocyte cultures, treated with OPZ for either 24 or 72 h. Furthermore, expression of a set of genes not detected on the arrays but known to be regulated by dithiole thiones in human (GSTA1/2, NAD(P)H-quinone reductase, c-jun, c-fos and ref-1) (4,15) or rodents [GSTA2, NAD(P)H-quinone reductase 1, multidrug resistance protein 3, microsomal epoxide hydrolase, manganese superoxide dismutase, ferritin, thioredoxin reductase, leukotriene B4 12-hydroxy dehydrogenase, heme oxygenase, aflatoxin B1 aldehyde reductase, catalytic subunit of glutamate-cysteine ligase and heme oxygenase] (6,9,10,1517) were also measured by RT-qPCR. Finally, in a recent unpublished work we found further OPZ-regulated genes in differentiated Caco-2 cells, a colonic carcinoma cell line (nucleoside diphosphate kinase 1 and 2, CYP4A11, tumor-associated antigen L6, asparagine synthase, nucleophosmin 1, glutamate dehydrogenase, c-jun, CD55 antigen and Na+/K+ transporting ATPase ß3 subunit). Therefore, expression of these genes in untreated and OPZ-treated hepatocytes was also analyzed by RT-qPCR (Table III).
The highest induction levels were found for CYP1A1, CYP1A2 and CYP2B6. RT-qPCR analysis revealed that the mRNAs encoding these CYP were increased in all hepatocyte populations tested after 24 and 72 h. Two other CYPs were also affected by OPZ at the mRNA level, although in a negative manner (CYP2E1 and CYP4A11). In contrast to its rat counterpart, which is induced by OPZ (5), human CYP2E1 mRNA was down-regulated in all hepatocyte populations tested by RT-qPCR as early as 24 h of treatment.
Regarding phase II enzymes, GSTA1/2, UDP glucuronosyl transferases 1A1, 1A3, 1A4 and 1A6, NADP(H)-quinone reductase 1 and, to a lesser extent, microsomal epoxide hydrolase, were increased as early as 24 h of treatment with OPZ. In contrast, aflatoxin B1 reductase, which is induced by OPZ in rodents, was unaffected by this compound in human hepatocytes.
We also demonstrate that OPZ modulates the mRNA levels of several genes involved in protection against oxidative stress (thioredoxin, thioredoxin reductase 1 and glutathione peroxidases 2 and 3) and in the metabolism of glutathione, either directly (
-glutamylcysteine synthetase) or indirectly (glutamate dehydrogenase, betaine homocysteine transferase and asparagine synthetase). Finally, we also showed an increase in the proto-oncogene c-myc and other genes involved in DNA replication and cell cycle control, i.e. nucleoside diphosphate kinases 1 and 2, prothymosin
and nucleophosmin 1.
To determine whether changes in steady-state mRNA levels result in an increase in the corresponding protein levels, several gene products were tested by western blotting, including CYP2B6, CYP4A11, CYP2E1, epoxide hydrolase, asparagine synthase, glutathione peroxidase, glutamate dehydrogenase, thioredoxin, thioredoxin reductase, c-Myc and nucleoside diphosphate kinases 1 and 2. With the exception of CYP4A11 and nucleoside diphosphate kinases 1 and 2, variations observed at the mRNA level correlated with the corresponding protein levels (Figure 1).

View larger version (56K):
[in this window]
[in a new window]
|
Fig. 1. Western blot of selected up- or down-regulated gene products in human hepatocytes treated with OPZ. Microsomal, cytosolic or total proteins (10100 µg) were prepared from control and OPZ-treated (24 or 72 h) human hepatocytes, electrophoresed in a polyacrylamide slab gel and electroblotted overnight onto Hybond ECL nitrocellulose membranes before incubation with primary and secondary antibodies. CYPs and epoxide hydrolase were analyzed in microsomal fractions; glutathione peroxidase 2, asparagine synthase, thioredoxin and thioredoxin reductase 1 were detected in cytosolic fractions; analysis of c-myc, glutamate dehydrogenase 1 and nucleoside diphosphate kinase was performed on total proteins. For each analyzed protein results obtained from three hepatocyte populations are shown. Gender and age of each donor are indicated in parentheses.
|
|
Both qPCR and western blot analyses demonstrate some differences between hepatocyte populations in response to OPZ treatment. However, no correlation was found between levels of induction by OPZ and age or sex of donors.
 |
Discussion
|
---|
Although several studies have been performed on the effect of chemopreventive agents against carcinogens in rodents, little is known about the modification of gene expression in human cells treated with such compounds. We herein describe, for the first time, the effects of OPZ, a powerful chemoprotector in rodent tissues, on the expression of several genes in human hepatocytes in primary culture. Our results demonstrate that OPZ increases the steady-state mRNA levels of several genes, including carcinogen metabolizing phase I and phase II enzymes as well as enzymes involved in protection against oxidative stress.
Although we recently demonstrated CYP1A1 transcriptional induction by OPZ in Caco2 cells (18), this is the first time that an effect of OPZ on CYP1A2 and CYP2B6 mRNA has been observed in human cells. OPZ has also been found to be a potent inhibitor of the activities of human CYP1A1, CYP1A2, CYP3A4 and CYP1B1 in vitro (3), CYP1A and CYP2B in rat liver both in vivo and in vitro (8) and CYP1A2 and CYP3A4 in primary human hepatocyte cultures (19). Sofowora et al. (20) have demonstrated that OPZ is similarly a potent in vivo inhibitor of CYP1A2 in humans. Thus, OPZ has a dual effect on some CYP by activating their gene transcription and inhibiting activity of the corresponding protein. This was observed for CYP1A1 and CYP1A2 and further studies will be necessary to determine whether OPZ has similar effects on CYP2B6. We also showed down-regulation of CYP2E1 at both the mRNA and protein levels. Since CYP2E1 is involved in both activation of toxic compounds and production of reactive oxygen species (ROS), its inhibition by OPZ might lead to decreased levels of ROS in hepatocytes and participate in protecting against chemically induced hepatotoxicity. Interestingly, it has been demonstrated that another chemopreventive agent, 2-(allylthio)pyrazine, also reduces CYP2E1 protein level and is effective in protecting against toxicant-induced liver toxicity (21).
Phase II detoxifying enzymes like NAD(P)H-quinone oxidoreductase 1, glutathione transferases and UDP glucuronyltransferases, have been considered to play an important role in preventing carcinogen-induced cancers. These enzymes detoxify reactive metabolites which form DNA or protein adducts, thereby preventing their deleterious effects. Several groups demonstrated that increases in their expression is a key mechanism in chemoprevention (2224). Our results demonstrate induction of hGSTA1, hGSTA2 and several UDP glucuronyl transferase isoenzymes mRNA levels, as well as those of microsomal epoxide hydrolase and NADP(H)-quinone reductase 1. We suggest that up-regulation of these enzymes in human hepatocytes will be highly relevant to chemoprevention strategies.
Interestingly, we have demonstrated for the first time that OPZ regulates several enzymes involved in protection against oxidative stress in human hepatocytes. We observed up-regulation of enzymes involved in cellular reduction/oxidation status, which is controlled by the thioredoxin (thioredoxin and thioredoxin reductase) and glutathione (
-glutamylcysteine synthetase regulatory subunit and glutathione reductase) systems, which scavenge harmful intracellular ROS. This observation is in agreement with previous in vivo studies demonstrating up-regulation of
-glutamylcysteine synthetase in colon mucosa and peripheral blood lymphocytes in patients treated with OPZ (25). Moreover, we demonstrated down-regulation of two enzymes (glutamate dehydrogenase and betaine-homocysteine transferase) which are involved in the metabolic pathways of two amino acids necessary for the synthesis of glutathione, i.e. glutamate and homocysteine, a precursor of cysteine. An increase in asparagine synthetase, which catalyses the formation of asparagine in association with glutamate synthesis, was also observed. Together, modulation of these enzymes by OPZ might contribute to an increase in the availability of amino acids necessary for the synthesis of glutathione. Finally, glutathione peroxidase subunits 2 and 3 play an important role in the elimination of hydroperoxides and their induction by OPZ might contribute to protection against the toxicity of endogenous toxic compounds which are also associated with oxidative stress. This hypothesis is in agreement with a previous study demonstrating that dietary supplementation with OPZ protected cultured human retinal pigment epithelial cells against oxidant-induced injury (26).
Unexpectedly, an increase was observed in c-myc and other genes involved in DNA replication and cell cycle control, i.e. nucleoside diphosphate kinases 1 and 2, prothymosin
and nucleophosmin 1. These genes are closely related since they are known to be c-Myc target genes (27). Interestingly, western blot analysis demonstrates that c-Myc is also increased at the protein level. Further studies will have to be performed to determine whether OPZ has a direct effect on cell proliferation and DNA replication.
Previously it has been demonstrated that induction by dithiole thiones of several genes is primarily due to transcriptional activation and is regulated by an enhancer, called an antioxidant-responsive element, which binds the transcription factor NF-E2-related factor 2 (10). The second mechanism involves the aryl hydrocarbon receptor xenobiotic-responsive element (18). Interestingly, use of Matinspector (http://www.genomatix.de/cgi-bin/matinspector/matinspector.pl) for in silico analysis of 5'-flanking regions of OPZ up-regulated genes demonstrates the presence of putative xenobiotic-responsive element and/or antioxidant-responsive element sequences in the majority of them (data not shown). Further studies will be necessary to determine whether these putative sites are involved or not in OPZ regulation of those genes. The NF
B and the CCAAT/enhancer binding protein pathways have also been reported to modulate OPZ gene regulation (28,29). Our results suggest that c-myc might represent a new pathway involved in OPZ gene induction. It is noteworthy that the RelA NF
B subunit and the aryl hydrocarbon receptor have been reported to cooperate to transactivate the c-myc promoter in mammary cells (30).
In conclusion, our study represents the first in vitro approach to investigate gene expression in human hepatocytes treated with OPZ. We provide evidence for up- and down-regulation of several categories of genes involved in detoxification, antioxidant defences, the stress response, transport of xenobiotics and cell cycle control. However, our results show important variations in the response to OPZ between hepatocyte populations. These variations in response to inducers is not unique, having already been observed with other enzymes in response to some inducers (4,3134). Obviously, factors related to the donor (i.e. genetic polymorphism, liver disease, premedication and/or nutritional status) and/or organ disruption can to some extent cause alterations in liver-specific functions in freshly isolated and short-term cultured hepatocytes. There was no correlation between levels of induction by OPZ and age or sex of donors. Although our data were obtained in vitro and their extrapolation to the in vivo human situation can be questioned, inter-individual variations in response to OPZ might potentially contribute to differences in efficacy of chemopreventive treatments between individuals. Although orthologous genes are similarly regulated in both rodents and human [GSTA1/2, CYP1A1/2, UDP glucuronyltransferase and NADP(H)-quinone reductase 1], we also demonstrated no or very low increases in other genes known to be induced by dithiole thiones in rodents and that could have potent protective effects (e.g. aflatoxin B1 aldehyde reductase, ferritin and Mn superoxide dismutase). Thus, we suggest that caution should be used when extrapolating data obtained with rodents to human beings in vivo. Although we found increases in CYP1A1, CYP1A2 and CYP2B6 gene expression induced by OPZ, the inhibitory effect of this agent on the activity of the corresponding proteins as well as induction of phase II enzymes, antioxidant enzymes and some transporters (multidrug resistance protein 2) all favor a protective role of OPZ against carcinogenesis and oxidative damage. Obviously, our study has extended our knowledge of the beneficial effects of OPZ on several detoxification systems in humans. Finally, it can be postulated that other chemopreventive agents will have different spectra of action on detoxifying functions leading to the conclusion that an association of several of them, by the consumption of fruits and vegetables, will increase their protective effects.
 |
Acknowledgments
|
---|
We thank the Biological Resource Centre (BRC) of Rennes for the supply of isolated human hepatocytes. This work was supported in part by the Institut National de la Santé et de la Recherche Médicale and the Association pour la Recherche sur le Cancer (ARC). Amélie Piton is the recipient of a fellowship from the Conseil Regional de Bretagne.
 |
References
|
---|
- Langouet,S., Morel,F. and Guillouzo,A. (2004) Effect of oltipraz on phase I and phase II xenobiotic metabolizing enzymes. In Bao,Y. and Fenwick,R. (eds) Phytochemicals in Health and Disease. Marcel Dekker, New York, NY, pp. 311329.
- Kwak,M.K., Egner,P.A., Dolan,P.M., Ramos-Gomez,M., Groopman,J.D., Itoh,K., Yamamoto,M. and Kensler,T.W. (2001) Role of phase 2 enzyme induction in chemoprotection by dithiolethiones. Mutat. Res., 480/481, 305315.
- Langouet,S., Furge,L.L., Kerriguy,N., Nakamura,K., Guillouzo,A. and Guengerich,F.P. (2000) Inhibition of human cytochrome P450 enzymes by 1,2-dithiole-3-thione, oltipraz and its derivatives and sulforaphane. Chem. Res. Toxicol., 13, 245252.[CrossRef][ISI][Medline]
- Morel,F., Fardel,O., Meyer,D.J. et al. (1993) Preferential increase of glutathione S-transferase class alpha transcripts in cultured human hepatocytes by phenobarbital, 3-methylcholanthrene and dithiolethiones. Cancer Res., 53, 231234.[Abstract]
- Maheo,K., Morel,F., Antras-Ferry,J., Langouet,S., Desmots,F., Corcos,L. and Guillouzo,A. (1998) Endotoxin suppresses the oltipraz-mediated induction of major hepatic glutathione transferases and cytochromes P450 in the rat. Hepatology, 28, 16551662.[ISI][Medline]
- Primiano,T., Li,Y., Kensler,T.W., Trush,M.A. and Sutter,T.R. (1998) Identification of dithiolethione-inducible gene-1 as a leukotriene B4 12-hydroxydehydrogenase: implications for chemoprevention. Carcinogenesis, 19, 9991005.[Abstract]
- Payen,L., Sparfel,L., Courtois,A., Vernhet,L., Guillouzo,A. and Fardel,O. (2002) The drug efflux pump MRP2: regulation of expression in physiopathological situations and by endogenous and exogenous compounds. Cell Biol. Toxicol., 18, 221233.[CrossRef][ISI][Medline]
- Langouet,S., Maheo,K., Berthou,F., Morel,F., Lagadic-Gossmann,D., Glaise,D., Coles,B., Ketterer,B. and Guillouzo,A. (1997) Effects of administration of the chemoprotective agent oltipraz on CYP1A and CYP2B in rat liver and rat hepatocytes in culture. Carcinogenesis, 18, 13431349.[Abstract]
- Antras-Ferry,J., Maheo,K., Chevanne,M., Dubos,M.P., Morel,F., Guillouzo,A., Cillard,P. and Cillard,J. (1997) Oltipraz stimulates the transcription of the manganese superoxide dismutase gene in rat hepatocytes. Carcinogenesis, 18, 21132117.[Abstract]
- Kwak,M.K., Wakabayashi,N., Itoh,K., Motohashi,H., Yamamoto,M. and Kensler,T.W. (2003) Modulation of gene expression by cancer chemopreventive dithiolethiones through the Keap1-Nrf2 pathway. Identification of novel gene clusters for cell survival. J. Biol. Chem., 278, 81358145.[Abstract/Free Full Text]
- Kensler,T.W., Egner,P.A., Wang,J.B. et al. (2002) Strategies for chemoprevention of liver cancer. Eur. J. Cancer. Prev., 11 (suppl. 2), S58S64.[ISI][Medline]
- Zhang,B.C., Zhu,Y.R., Wang,J.B. et al. (1997) Oltipraz chemoprevention trial in Qidong, Jiangsu Province, People's Republic of China. J. Cell. Biochem., 2829 (suppl.), 166173.
- Wang,J.S., Shen,X., He,X. et al. (1999) Protective alterations in phase 1 and 2 metabolism of aflatoxin B1 by oltipraz in residents of Qidong, People's Republic of China. J. Natl Cancer Inst., 91, 347354.[Abstract/Free Full Text]
- Gupta,E., Olopade,O.I., Ratain,M.J., Mick,R., Baker,T.M., Berezin,F.K., Benson,A.B.,III and Dolan,M.E. (1995) Pharmacokinetics and pharmacodynamics of oltipraz as a chemopreventive agent. Clin. Cancer Res., 1, 11331138.[Abstract]
- Yao,K.S. and O'Dwyer,P.J. (2003) Role of the AP-1 element and redox factor-1 (Ref-1) in mediating transcriptional induction of DT-diaphorase gene expression by oltipraz: a target for chemoprevention. Biochem. Pharmacol., 66, 1523.[CrossRef][ISI][Medline]
- Primiano,T., Kensler,T.W., Kuppusamy,P., Zweier,J.L. and Sutter,T.R. (1996) Induction of hepatic heme oxygenase-1 and ferritin in rats by cancer chemopreventive dithiolethiones. Carcinogenesis, 17, 22912296.[Abstract]
- Cherrington,N.J., Slitt,A.L., Maher,J.M., Zhang,X.X., Zhang,J., Huang,W., Wan,Y.J., Moore,D.D. and Klaassen,C.D. (2003) Induction of multidrug resistance protein 3 (mrp3) in vivo is independent of constitutive androstane receptor. Drug Metab. Dispos., 31, 13151319.[Abstract/Free Full Text]
- Le Ferrec,E., Lagadic-Gossmann,D., Rauch,C., Bardiau,C., Maheo,K., Massiere,F., Le Vee,M., Guillouzo,A. and Morel,F. (2002) Transcriptional induction of CYP1A1 by oltipraz in human Caco-2 cells is aryl hydrocarbon receptor- and calcium-dependent. J. Biol. Chem., 277, 2478024787.[Abstract/Free Full Text]
- Langouet,S., Coles,B., Morel,F., Becquemont,L., Beaune,P., Guengerich,F.P., Ketterer,B. and Guillouzo,A. (1995) Inhibition of CYP1A2 and CYP3A4 by oltipraz results in reduction of aflatoxin B1 metabolism in human hepatocytes in primary culture. Cancer Res., 55, 55745579.[Abstract]
- Sofowora,G.G., Choo,E.F., Mayo,G., Shyr,Y. and Wilkinson,G.R. (2001) In vivo inhibition of human CYP1A2 activity by oltipraz. Cancer Chemother. Pharmacol., 47, 505510.[CrossRef][ISI][Medline]
- Kim,N.D., Kwak,M.K. and Kim,S.G. (1997) Inhibition of cytochrome P450 2E1 expression by 2-(allylthio)pyrazine, a potential chemoprotective agent: hepatoprotective effects. Biochem. Pharmacol., 53, 261269.[CrossRef][ISI][Medline]
- Elegbede,J.A., Maltzman,T.H., Elson,C.E. and Gould,M.N. (1993) Effects of anticarcinogenic monoterpenes on phase II hepatic metabolizing enzymes. Carcinogenesis, 14, 12211223.[Abstract]
- Begleiter,A., Sivananthan,K., Curphey,T.J. and Bird,R.P. (2003) Induction of NAD(P)H quinone: oxidoreductase1 inhibits carcinogen-induced aberrant crypt foci in colons of Sprague-Dawley rats. Cancer Epidemiol. Biomarkers Prev., 12, 566572.[Abstract/Free Full Text]
- Kensler,T.W. (1997) Chemoprevention by inducers of carcinogen detoxication enzymes. Environ. Health Perspect., 105 (suppl. 4), 965970.[ISI][Medline]
- O'Dwyer,P.J., Szarka,C.E., Yao,K.S. et al. (1996) Modulation of gene expression in subjects at risk for colorectal cancer by the chemopreventive dithiolethione oltipraz. J. Clin. Invest., 98, 12101217.[Abstract/Free Full Text]
- Nelson,K.C., Armstrong,J.S., Moriarty,S., Cai,J., Wu,M.W., Sternberg,P.,Jr and Jones,D.P. (2002) Protection of retinal pigment epithelial cells from oxidative damage by oltipraz, a cancer chemopreventive agent. Invest. Ophthalmol. Vis. Sci., 43, 35503554.[Abstract/Free Full Text]
- Haggerty,T.J., Zeller,K.I., Osthus,R.C., Wonsey,D.R. and Dang,C.V. (2003) A strategy for identifying transcription factor binding sites reveals two classes of genomic c-Myc target sites. Proc. Natl Acad. Sci. USA, 100, 53135318.[Abstract/Free Full Text]
- Yao,K.S. and O'Dwyer,P.J. (1995) Involvement of NF-kappa B in the induction of NAD(P)H:quinone oxidoreductase (DT-diaphorase) by hypoxia, oltipraz and mitomycin C. Biochem. Pharmacol., 49, 275282.[CrossRef][ISI][Medline]
- Kang,K.W., Cho,I.J., Lee,C.H. and Kim,S.G. (2003) Essential role of phosphatidylinositol 3-kinase-dependent CCAAT/enhancer binding protein beta activation in the induction of glutathione S-transferase by oltipraz. J. Natl Cancer Inst., 95, 5366.[Abstract/Free Full Text]
- Kim,D.W., Gazourian,L., Quadri,S.A., Romieu-Mourez,R., Sherr,D.H. and Sonenshein,G.E. (2000) The RelA NF-kappaB subunit and the aryl hydrocarbon receptor (AhR) cooperate to transactivate the c-myc promoter in mammary cells. Oncogene, 19, 54985506.[CrossRef][ISI][Medline]
- Martelli,A., Mattioli,F., Angiola,M., Reimann,R. and Brambilla,G. (2003) Species, sex and inter-individual differences in DNA repair induced by nine sex steroids in primary cultures of rat and human hepatocytes. Mutat. Res., 536, 6978.[ISI][Medline]
- Morel,F., Beaune,P.H., Ratanasavanh,D., Flinois,J.P., Yang,C.S., Guengerich,F.P. and Guillouzo,A. (1990) Expression of cytochrome P-450 enzymes in cultured human hepatocytes. Eur. J. Biochem., 191, 437444.[Abstract]
- Raucy,J.L. (2003) Regulation of CYP3A4 expression in human hepatocytes by pharmaceuticals and natural products. Drug Metab. Dispos., 31, 533539.[Abstract/Free Full Text]
- Pichard,L., Fabre,I., Domergue,J., Joyeux,H. and Maurel,P. (1991) Effect of FK 506 on human hepatic cytochromes P-450: interaction with CyA. Transplant. Proc., 23, 27912793.[ISI][Medline]
Received May 18, 2004;
revised September 23, 2004;
accepted October 12, 2004.