chrysin and 2-tert-butylhydroquinone

We have previously reported that breast cancer resistance protein (BCRP) is involved in the transport of phase II metabolites of the food carcinogen benzo[a]pyrene (BP) in the human intestinal cell line Caco-2. Furthermore, the expression of BCRP seemed most likely to be aryl hydrocarbon receptor (AhR) dependent. Since numerous plant-derived anticarcinogens with AhR-agonistic activity have been identified to date, in the present study we investigated the effects of naturally occurring dietary compounds and tert-butyl hydroquinone (TBHQ) for their effects on BCRP expression. In Caco-2 cells, the most pronounced induction of BCRP expression could be observed after treatment with TBHQ (100 microM), dibenzoylmethane (DBM, 50 microM), and quercetin (25 microM), while green tea component (-)-epicatechin (50 microM) decreased BCRP expression. On mRNA level, quercetin, chrysin, flavone, and indole-3-carbinol showed a strong inducing effect, while genistein had no effect on BCRP mRNA expression. Curcumin and resveratrol showed a strong effect on BCRP induction in MCF-7 wild-type cells but no response in AhR-deficient MCF-7AHR(200) cells, supporting our hypothesis that BCRP is regulated via AhR-dependent signaling pathways. Inhibition of proteasome-mediated degradation of ligand-activated AhR caused a "superinduction" of BCRP mRNA. Antioxidant responsive element activators sulforaphane and diethylmaleate (DEM) had no inducing effect on BCRP mRNA expression. Caco-2 cells pretreated with quercetin or DBM showed an enhancement of apically transported benzo[a]pyrene-3-sulfate, indicating that induced BCRP was functionally active. In conclusion, apart from the modulation of detoxifying enzymes in the intestine, induction of BCRP by dietary constituents may contribute to the detoxification of food-derived procarcinogens such as BP.

Topics: 7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; ATP Binding Cassette Transporter, Subfamily G, Member 2; ATP-Binding Cassette Transporters; Benzo(a)pyrene; Biological Transport; Caco-2 Cells; Catechin; Cell Line, Tumor; Chalcones; Flavonoids; Gene Expression; Humans; Hydroquinones; Indoles; Isothiocyanates; Maleates; Molecular Structure; Neoplasm Proteins; Plant Extracts; Quercetin; Receptors, Aryl Hydrocarbon; Resveratrol; RNA, Messenger; Silymarin; Stilbenes; Sulfoxides; Thiocyanates; Transfection

We identified the UDP-glucuronosyltransferase (UGT) 1A1 5'-upstream region that confers UGT1A1 induction by various agents, including flavonoids, on a luciferase reporter gene and has the properties of a transcriptional enhancer. Chrysin- and rifampicin-response activities were traced to the same element as a 290-bp distal enhancer module (-3483/-3194), in which the reporter activities were enhanced by activators of nuclear receptors [constitutive androstane receptor (CAR) and pregnane X receptor (PXR)] and transcription factor [aryl hydrocarbon receptor (AhR)]. Utilizing transactivation experiments with the UGT1A1 290-bp reporter gene, we assessed UGT1A1 induction by various flavonoids. 5,7-Dihydroxyflavones with varying substituents in the B-ring and gallocatechin dimers increased the reporter activity in a time- and dose-dependent manner. The treatment of HepG2 cells with the flavonoids for 24 hr elevated the expression of mRNAs and proteins of UGT1A1 and CYP1A1, while the mRNA levels of CYP2B6, CYP3A4, CAR, PXR and AhR was not altered. Chrysin and rifampicin induced the activation of the wild-type reporter gene and T-3263G-mutated gene to a similar extent in HepG2 cells cotransfected with expression vectors of CAR and PXR. Mutation of the AhR core binding region most prominently suppressed the activation of the 290-bp reporter gene by chrysin and baicalein, while mutations of four putative nuclear receptor motifs (DR4 element, PXRE, CARE and DR3 element) partly decreased its activation. Taken together, the results indicate that UGT1A1 was induced in response to flavonoids and xenobiotics through the transactivation of the 290-bp reporter gene, that was a multi-component enhancer containing CAR, PXR and AhR motifs.

Topics: Analysis of Variance; Aryl Hydrocarbon Hydroxylases; Basic Helix-Loop-Helix Transcription Factors; Benzo(a)pyrene; Biflavonoids; Calcium-Binding Proteins; Catechin; Cytochrome P-450 CYP1A1; Cytochrome P-450 CYP2B6; Cytochrome P-450 CYP3A; Cytochrome P-450 Enzyme System; Enhancer Elements, Genetic; Enzyme Induction; Eye Proteins; Flavonoids; Gene Expression Regulation, Enzymologic; Genes, Reporter; Glucuronosyltransferase; Hippocalcin; Humans; Hydroquinones; Lipoproteins; Molecular Conformation; Mutagenesis; Nerve Tissue Proteins; Oxidoreductases, N-Demethylating; Pregnane X Receptor; Proanthocyanidins; Quercetin; Receptors, Aryl Hydrocarbon; Receptors, Cytoplasmic and Nuclear; Receptors, Steroid; Recoverin; RNA, Messenger; Transfection; Tumor Cells, Cultured; Xenobiotics