Sulforaphane (SFN), an isothiocyanate found in cruciferous vegetables, exerts many beneficial effects on human health such as antioxidant, anti-inflammatory, and anticancer effects. The effect of SFN alone on drug-metabolizing enzymes (DMEs) has been investigated in numerous in vitro and in vivo models, but little is known about the effect of SFN in combination with cytochrome P450 (CYP) inducer. The aim of our study was to evaluate the effect of SFN on the activity and gene expression of selected DMEs in primary cultures of rat hepatocytes treated or non-treated with β-naphthoflavone (BNF), the model CYP1A inducer. In our study, SFN alone did not significantly alter the activity and expression of the studied DMEs, except for the glutathione S-transferase (GSTA1) mRNA level, which was significantly enhanced. Co-treatment of hepatocytes with SFN and BNF led to a substantial increase in sulfotransferase, aldoketoreductase 1C, carbonylreductase 1 and NAD(P)H:quinone oxidoreductase 1 activity and a marked decrease in cytochrome P450 (CYP) Cyp1a1, Cyp2b and Cyp3a4 expression in comparison to the treatment with BNF alone. Sulforaphane is able to modulate the activity and/or expression of DMEs, thus shifting the balance of carcinogen metabolism toward deactivation, which could represent an important mechanism of its chemopreventive activity.

Department of Biochemical Sciences Faculty of Pharmacy Charles University Heyrovského 1203 50005 Hradec Králové Czech Republic

Zobrazit více v PubMed

Royston K.J., Tollefsbol T.O. The Epigenetic Impact of Cruciferous Vegetables on Cancer Prevention. Curr. Pharmacol. Rep. 2015;1:46–51. doi: 10.1007/s40495-014-0003-9. PubMed DOI PMC

Tarozzi A., Angeloni C., Malaguti M., Morroni F., Hrelia S., Hrelia P. Sulforaphane as a Potential Protective Phytochemical against Neurodegenerative Diseases. Oxid. Med. Cell. Longev. 2013;2013:415078. doi: 10.1155/2013/415078. PubMed DOI PMC

Angelino D., Jeffery E. Glucosinolate hydrolysis and bioavailability of resulting isothiocyanates: Focus on glucoraphanin. J. Funct. Foods. 2014;7:67–76. doi: 10.1016/j.jff.2013.09.029. DOI

Atwell L.L., Hsu A., Wong C.P., Stevens J.F., Bella D., Yu T.W., Pereira C.B., Lohr C.V., Christensen J.M., Dashwood R.H., et al. Absorption and chemopreventive targets of sulforaphane in humans following consumption of broccoli sprouts or a myrosinase-treated broccoli sprout extract. Mol. Nutr. Food Res. 2015;59:424–433. doi: 10.1002/mnfr.201400674. PubMed DOI PMC

Gasper A.V., Al-Janobi A., Smith J.A., Bacon J.R., Fortun P., Atherton C., Taylor M.A., Hawkey C.J., Barrett D.A., Mithen R.F. Glutathione S-transferase M1 polymorphism and metabolism of sulforaphane from standard and high-glucosinolate broccoli. Am. J. Clin. Nutr. 2005;82:1283–1291. PubMed

La Marca M., Beffy P., Della Croce C., Gervasi P.G., Iori R., Puccinelli E., Longo V. Structural influence of isothiocyanates on expression of cytochrome P450, phase II enzymes, and activation of Nrf2 in primary rat hepatocytes. Food Chem. Toxicol. 2012;50:2822–2830. doi: 10.1016/j.fct.2012.05.044. PubMed DOI

Cheung K.L., Kong A.N. Molecular targets of dietary phenethyl isothiocyanate and sulforaphane for cancer chemoprevention. AAPS J. 2010;12:87–97. doi: 10.1208/s12248-009-9162-8. PubMed DOI PMC

Petri N., Tannergren C., Holst B., Mellon F.A., Bao Y., Plumb G.W., Bacon J., O’Leary K.A., Kroon P.A., Knutson L., et al. Absorption/metabolism of sulforaphane and quercetin, and regulation of phase II enzymes, in human jejunum in vivo. Drug. Metab. Dispos. 2003;31:805–813. doi: 10.1124/dmd.31.6.805. PubMed DOI

Jones S.B., Brooks J.D. Modest induction of phase 2 enzyme activity in the F-344 rat prostate. BMC Cancer. 2006;6:62. doi: 10.1186/1471-2407-6-62. PubMed DOI PMC

Hayes J.D., Kelleher M.O., Eggleston I.M. The cancer chemopreventive actions of phytochemicals derived from glucosinolates. Eur. J. Nutr. 2008;47:73–88. doi: 10.1007/s00394-008-2009-8. PubMed DOI

Higgins L.G., Hayes J.D. Mechanisms of induction of cytosolic and microsomal glutathione transferase (GST) genes by xenobiotics and pro-inflammatory agents. Drug Metab. Rev. 2011;43:92–137. doi: 10.3109/03602532.2011.567391. PubMed DOI

McMahon M., Itoh K., Yamamoto M., Hayes J.D. Keap1-dependent proteasomal degradation of transcription factor Nrf2 contributes to the negative regulation of antioxidant response element-driven gene expression. J. Biol. Chem. 2003;278:21592–21600. doi: 10.1074/jbc.M300931200. PubMed DOI

Sahi J., Grepper S., Smith C. Hepatocytes as a tool in drug metabolism, transport and safety evaluations in drug discovery. Curr. Drug Discov. Technol. 2010;7:188–198. PubMed

Maheo K., Morel F., Langouet S., Kramer H., Le Ferrec E., Ketterer B., Guillouzo A. Inhibition of cytochromes P-450 and induction of glutathione S-transferases by sulforaphane in primary human and rat hepatocytes. Cancer Res. 1997;57:3649–3652. PubMed

Srovnalova A., Vanduchova A., Svecarova M., Anzenbacherova E., Tomankova V., Anzenbacher P., Dvorak Z. Effects of sulforaphane and its S- and R-enantiomers on the expression and activities of human drug-metabolizing cytochromes P450. J. Funct. Foods. 2015;14:487–501. doi: 10.1016/j.jff.2015.02.006. DOI

Yoxall V., Kentish P., Coldham N., Kuhnert N., Sauer M.J., Ioannides C. Modulation of hepatic cytochromes P450 and phase II enzymes by dietary doses of sulforaphane in rats: Implications for its chemopreventive activity. Int. J. Cancer. 2005;117:356–362. doi: 10.1002/ijc.21191. PubMed DOI

Matusheski N.V., Jeffery E.H. Comparison of the bioactivity of two glucoraphanin hydrolysis products found in broccoli, sulforaphane and sulforaphane nitrile. J. Agric. Food Chem. 2001;49:5743–5749. doi: 10.1021/jf010809a. PubMed DOI

Gross-Steinmeyer K., Stapleton P.L., Tracy J.H., Bammler T.K., Strom S.C., Eaton D.L. Sulforaphane- and Phenethyl Isothiocyanate-Induced Inhibition of Aflatoxin B1-Mediated Genotoxicity in Human Hepatocytes: Role of GSTM1 Genotype and CYP3A4 Gene Expression. Toxicol. Sci. 2010;116:422–432. doi: 10.1093/toxsci/kfq135. PubMed DOI PMC

Abdull Razis A.F., Hanlon N., Soltys E., Krizova V., Iori R., Plant K.E., Plant N., Ioannides C. The naturally occurring aliphatic isothiocyanates sulforaphane and erucin are weak agonists but potent non-competitive antagonists of the aryl hydrocarbon receptor. Arch. Toxicol. 2012;86:1505–1514. doi: 10.1007/s00204-012-0875-6. PubMed DOI

Zhou C., Poulton E.J., Grun F., Bammler T.K., Blumberg B., Thummel K.E., Eaton D.L. The dietary isothiocyanate sulforaphane is an antagonist of the human steroid and xenobiotic nuclear receptor. Mol. Pharmacol. 2007;71:220–229. doi: 10.1124/mol.106.029264. PubMed DOI

Wang D., Li L., Fuhrman J., Ferguson S., Wang H. The role of constitutive androstane receptor in oxazaphosphorine-mediated induction of drug-metabolizing enzymes in human hepatocytes. Pharm. Res. 2011;28:2034–2044. doi: 10.1007/s11095-011-0429-2. PubMed DOI PMC

Angeloni C., Leoncini E., Malaguti M., Angelini S., Hrelia P., Hrelia S. Modulation of phase II enzymes by sulforaphane: Implications for its cardioprotective potential. J. Agric. Food Chem. 2009;57:5615–5622. doi: 10.1021/jf900549c. PubMed DOI

Jiang Z.Q., Chen C., Yang B., Hebbar V., Kong A.N. Differential responses from seven mammalian cell lines to the treatments of detoxifying enzyme inducers. Life Sci. 2003;72:2243–2253. doi: 10.1016/S0024-3205(03)00101-2. PubMed DOI

Singh P., Sharma R., McElhanon K., Allen C.D., Megyesi J.K., Benes H., Singh S.P. Sulforaphane protects the heart from doxorubicin-induced toxicity. Free Radic. Biol. Med. 2015;86:90–101. doi: 10.1016/j.freeradbiomed.2015.05.028. PubMed DOI PMC

Bonnesen C., Eggleston I.M., Hayes J.D. Dietary indoles and isothiocyanates that are generated from cruciferous vegetables can both stimulate apoptosis and confer protection against DNA damage in human colon cell lines. Cancer Res. 2001;61:6120–6130. PubMed

Lakhman S.S., Chen X., Gonzalez-Covarrubias V., Schuetz E.G., Blanco J.G. Functional characterization of the promoter of human carbonyl reductase 1 (CBR1). Role of XRE elements in mediating the induction of CBR1 by ligands of the aryl hydrocarbon receptor. Mol. Pharmacol. 2007;72:734–743. doi: 10.1124/mol.107.035550. PubMed DOI

Nannelli A., Rossignolo F., Tolando R., Rossato P., Longo V., Gervasi P.G. Effect of beta-naphthoflavone on AhR-regulated genes (CYP1A1, 1A2, 1B1, 2S1, Nrf2, and GST) and antioxidant enzymes in various brain regions of pig. Toxicology. 2009;265:69–79. doi: 10.1016/j.tox.2009.09.010. PubMed DOI

Payen L., Courtois A., Loewert M., Guillouzo A., Fardel O. Reactive oxygen species-related induction of multidrug resistance-associated protein 2 expression in primary hepatocytes exposed to sulforaphane. Biochem. Biophys. Res. Commun. 2001;282:257–263. doi: 10.1006/bbrc.2001.4531. PubMed DOI

Buckley D.B., Klaassen C.D. Induction of Mouse UDP-Glucuronosyltransferase mRNA Expression in Liver and Intestine by Activators of Aryl-Hydrocarbon Receptor, Constitutive Androstane Receptor, Pregnane X Receptor, Peroxisome Proliferator-Activated Receptor alpha, and Nuclear Factor Erythroid 2-Related Factor 2. Drug Metab. Dispos. 2009;37:847–856. PubMed PMC

Falany C.N., Strom P., Swedmark S. Sulphation of o-desmethylnaproxen and related compounds by human cytosolic sulfotransferases. Br. J. Clin. Pharmacol. 2005;60:632–640. doi: 10.1111/j.1365-2125.2005.02506.x. PubMed DOI PMC

Maier T., Guell M., Serrano L. Correlation of mRNA and protein in complex biological samples. FEBS Lett. 2009;583:3966–3973. doi: 10.1016/j.febslet.2009.10.036. PubMed DOI

Bartels C.L., Tsongalis G.J. MicroRNAs: Novel biomarkers for human cancer. Clin. Chem. 2009;55:623–631. doi: 10.1373/clinchem.2008.112805. PubMed DOI

Richert L., Tuschl G., Abadie C., Blanchard N., Pekthong D., Mantion G., Weber J.C., Mueller S.O. Use of mRNA expression to detect the induction of drug metabolising enzymes in rat and human hepatocytes. Toxicol. Appl. Pharmacol. 2009;235:86–96. doi: 10.1016/j.taap.2008.11.019. PubMed DOI

Lnenickova K., Skalova L., Stuchlikova Raisova L., Szotakova B., Matouskova P. The induction of xenobiotic-metabolizing enzymes in hepatocytes by beta-naphthoflavone: Time-dependent changes in activities, protein and mRNA levels. Acta Pharm. 2017 in press. PubMed

Berry M., Edwards A., Barritt G. Isolated Hepatocytes: Preparation, Properties and Applications. Volume 21. Elsevier Science Publishers; Amsterdam, The Netherlands: 1991. p. 460.

Livak K.J., Schmittgen T.D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25:402–408. doi: 10.1006/meth.2001.1262. PubMed DOI

Burke M.D., Mayer R.T. Ethoxyresorufin: Direct fluorimetric assay of a microsomal O-dealkylation which is preferentially inducible by 3-methylcholanthrene. Drug Metab. Dispos. 1974;2:583–588. PubMed

Burke M.D., Thompson S., Elcombe C.R., Halpert J., Haaparanta T., Mayer R.T. Ethoxy-, pentoxy- and benzyloxyphenoxazones and homologues: A series of substrates to distinguish between different induced cytochromes P-450. Biochem. Pharmacol. 1985;34:3337–3345. doi: 10.1016/0006-2952(85)90355-7. PubMed DOI

Kobayashi K., Urashima K., Shimada N., Chiba K. Substrate specificity for rat cytochrome P450 (CYP) isoforms: Screening with cDNA-expressed systems of the rat. Biochem. Pharmacol. 2002;63:889–896. doi: 10.1016/S0006-2952(01)00843-7. PubMed DOI

Teel R.W., Huynh H. Modulation by phytochemicals of cytochrome P450-linked enzyme activity. Cancer Lett. 1998;133:135–141. doi: 10.1016/S0304-3835(98)00218-3. PubMed DOI

Elbarbry F., Attia A., Shoker A. Validation of a new HPLC method for determination of midazolam and its metabolites: Application to determine its pharmacokinetics in human and measure hepatic CYP3A activity in rabbits. J. Pharm. Biomed. Anal. 2009;50:987–993. doi: 10.1016/j.jpba.2009.07.004. PubMed DOI

Maté L., Virkel G., Lifschitz A., Ballent M., Lanusse C. Hepatic and extra-hepatic metabolic pathways involved in flubendazole biotransformation in sheep. Biochem. Pharmacol. 2008;76:773–783. doi: 10.1016/j.bcp.2008.07.002. PubMed DOI

Cullen J.J., Hinkhouse M.M., Grady M., Gaut A.W., Liu J., Zhang Y.P., Weydert C.J., Domann F.E., Oberley L.W. Dicumarol inhibition of NADPH:quinone oxidoreductase induces growth inhibition of pancreatic cancer via a superoxide-mediated mechanism. Cancer Res. 2003;63:5513–5520. PubMed

Mizuma T., Machida M., Hayashi M., Awazu S. Correlation of drug conjugative metabolism rates between in vivo and in vitro: Glucuronidation and sulfation of p-nitrophenol as a model compound in rat. J. Pharmacobiodyn. 1982;5:811–817. doi: 10.1248/bpb1978.5.811. PubMed DOI

Frame L.T., Ozawa S., Nowell S.A., Chou H.C., DeLongchamp R.R., Doerge D.R., Lang N.P., Kadlubar F.F. A simple colorimetric assay for phenotyping the major human thermostable phenol sulfotransferase (SULT1A1) using platelet cytosols. Drug Metab. Dispos. 2000;28:1063–1068. PubMed

Habig W.H., Pabst M.J., Jakoby W.B. Glutathione S-Transferases: The first enzymatic step in mercapturic acid formation. J. Biol. Chem. 1974;249:7130–7139. PubMed

The Modulation of Phase II Drug-Metabolizing Enzymes in Proliferating and Differentiated CaCo-2 Cells by Hop-Derived Prenylflavonoids