title: Phase 2 enzyme induction by the major metabolite of oltipraz.
abstract: Treatment for 48 h of murine Hepa 1c1c7 cells in culture with the cancer chemopreventive oltipraz (1) followed by addition of CD(3)I and immediate cell lysis yields, by LC/MS analysis, three isotopomers of the methylated pyrrolopyrazine (2), a known human metabolite of oltipraz. The major isotopomer (58%) is the one containing two CD(3)- groups attached to the pendant sulfur atoms of the pyrrolopyrazine ring, the others containing one CD(3)- and one CH(3)- group or two CH(3)- groups. It is concluded from this that the unmethylated pyrrolopyrazine (4) is the major metabolite of oltipraz. Prodrugs 5 and 6, which have been shown to rapidly generate 4 in the presence of GSH at physiological pH, induce the phase 2 enzyme NQO1 in Hepa 1c1c7 cells with potencies on par with oltipraz itself: CD(NQO1) = 14.4 +/- 1.3, 20.1 +/- 4.6, and 23.6 +/- 1.6 microM for oltipraz, 5, and 6, respectively. Pretreatment of oltipraz, 5, and 6 in cell culture media with 1 mM GSH, which is shown to immediately convert 5 and 6 to 4, followed by incubation with Hepa 1c1c7 cells shows similar potencies for oltipraz and the (decomposed) produrgs, with CD(NQO1) = 18.0 +/- 4.4 microM for 5, 17.8 +/- 0.2 microM for 6, and 13.5 +/- 1.4 microM for oltipraz. Treatment with compound 6 of murine hepatoma cells containing a luciferase gene under the control of the antioxidant response element (ARE) from the mouse heme oxygenase (ho-1) gene elicits induction of luciferase activity, CD = 35.8 +/- 2.8 microM, somewhat greater than the potency than oltipraz itself. Western blots of nuclear proteins isolated from Hepa 1c1c7 cells and probed with anti-Nrf2 indicate that as compared to vehicle DMSO, compound 6 stimulates nuclear translocation of Nrf2 from the cytosol. From this study, it is concluded that the major metabolite of the cancer chemopreventive oltipraz is a phase 2 enzyme inducer of comparable potency that activates the ARE and initiates nuclear translocation of transcription factor Nrf 2.
authors: Department of Chemistry and Biochemistry, University of Maryland, Baltimore County, 1000 Hilltop Circle, Baltimore, Maryland 21250, USA.
authors: Jacobus P
authors: JP
authors: Petzer
authors: Mettachit
authors: M
authors: Navamal
authors: Jesse K
authors: JK
authors: Johnson
authors: Mi-Kyoung
authors: MK
authors: Kwak
authors: Thomas W
authors: TW
authors: Kensler
authors: James C
authors: JC
authors: Fishbein
mesh_terms: D000818:Animals; D015153:Blotting, Western; D016923:Cell Death / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology; D004790:Enzyme Induction / Q000187:drug effects* / Q000502:physiology; D005978:Glutathione / Q000187:drug effects / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006419:Heme Oxygenase (Decyclizing) / Q000378:metabolism; D008156:Luciferases / Q000235:genetics; D008745:Methylation; D051379:Mice; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000494:pharmacology; D051267:NF-E2-Related Factor 2; D011355:Prodrugs / Q000378:metabolism; D011719:Pyrazines / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D011758:Pyrroles / Q000737:chemistry / Q000378:metabolism; D013871:Thiones / Q000737:chemistry / Q000378:metabolism; D013876:Thiophenes; D013997:Time Factors; D015534:Trans-Activators / Q000235:genetics / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: C495786:7-mercapto-6-methylpyrrolo(1,2-a)pyrazine-2H,5H-5-thione; D004268:DNA-Binding Proteins; D051267:NF-E2-Related Factor 2; C495636:Nfe2l2 protein, mouse; D011355:Prodrugs; D011719:Pyrazines; D011758:Pyrroles; D013871:Thiones; D013876:Thiophenes; D015534:Trans-Activators; D014157:Transcription Factors; C026209:oltipraz; D008156:Luciferases; D006419:Heme Oxygenase (Decyclizing); D016660:NAD(P)H Dehydrogenase (Quinone); D005982:Glutathione Transferase; D005978:Glutathione