Assuntos
Acetilcisteína/farmacologia , Glutationa/farmacologia , Isoniazida/farmacologia , Mycobacterium fortuitum/efeitos dos fármacos , Antioxidantes/farmacologia , Interações Medicamentosas , Testes de Sensibilidade Microbiana , Compostos de Sulfidrila/farmacologia , Ácido Tióctico/análogos & derivados , Ácido Tióctico/farmacologiaRESUMO
Resveratrol is a cancer preventative agent that is found in red wine. Piceatannol is a closely related stilbene that has antileukaemic activity and is also a tyrosine kinase inhibitor. Piceatannol differs from resveratrol by having an additional aromatic hydroxy group. The enzyme CYP1B1 is overexpressed in a wide variety of human tumours and catalyses aromatic hydroxylation reactions. We report here that the cancer preventative agent resveratrol undergoes metabolism by the cytochrome P450 enzyme CYP1B1 to give a metabolite which has been identified as the known antileukaemic agent piceatannol. The metabolite was identified by high performance liquid chromatography analysis using fluorescence detection and the identity of the metabolite was further confirmed by derivatisation followed by gas chromatography-mass spectrometry studies using authentic piceatannol for comparison. This observation provides a novel explanation for the cancer preventative properties of resveratrol. It demonstrates that a natural dietary cancer preventative agent can be converted to a compound with known anticancer activity by an enzyme that is found in human tumours. Importantly this result gives insight into the functional role of CYP1B1 and provides evidence for the concept that CYP1B1 in tumours may be functioning as a growth suppressor enzyme.
Assuntos
Antineoplásicos Fitogênicos/metabolismo , Antineoplásicos/química , Hidrocarboneto de Aril Hidroxilases , Sistema Enzimático do Citocromo P-450/metabolismo , Estilbenos/química , Estilbenos/metabolismo , Antineoplásicos Fitogênicos/farmacologia , Quimioprevenção , Cromatografia Líquida de Alta Pressão , Citocromo P-450 CYP1B1 , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Neoplasias/enzimologia , Neoplasias/prevenção & controle , Resveratrol , Estilbenos/farmacologia , Células Tumorais Cultivadas , VinhoRESUMO
1. The bioreductive activation of the alkylaminoanthraquinone di-N-oxide prodrug AQ4N has been characterized in rat hepatic tissue using HPLC. 2. AQ4N was shown to be metabolized to two products, namely AQM, the two electron reduced mono-N-oxide, and AQ4, the four electron reduced active cytotoxic agent. 3. Metabolism was shown to occur in microsomes with an apparent Km = 30.29 microM and Vmax = 1.05 nmol/mg/min. 4. Bioreduction was dependent on anaerobic conditions and the presence of the reduced cofactor NADPH. Ketoconazole (100 microM) and carbon monoxide both inhibited AQ4N metabolism inferring a role for cytochrome P450 (CYP). 5. Microsomes from phenobarbitone and isoniazid-pretreated animals significantly (p < 0.05) enhanced the formation of AQ4 from AQ4N indicating a role for CYP2B and 2E respectively. The involvement of both CYP2B and 2E was confirmed by the use of CYP-specific inhibitors. 6. In conclusion, the involvement of rat hepatic CYP in the reductive bioactivation of the novel antitumour prodrug AQ4N has been established in detail for the first time. These findings highlight an important interspecies difference between the metabolism of AQ4N in rat and man which was shown earlier to be mediated by CYP3A enzymes. The pharmacological significance of this is discussed.
Assuntos
Antraquinonas/metabolismo , Antineoplásicos/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Pró-Fármacos/metabolismo , Animais , Biotransformação , Cromatografia Líquida de Alta Pressão , Cinética , Masculino , Oxirredução , Proteínas/metabolismo , Ratos , Ratos Sprague-Dawley , Frações Subcelulares/metabolismoRESUMO
PURPOSE: To establish the role of the human cytochromes P450 (CYPs) in the reductive metabolism of the novel anthraquinone di-N-oxide prodrug AQ4N. METHODS AND MATERIALS: Metabolism of AQ4N was conducted in a panel of 17 human phenotyped liver microsomes. AQ4N and metabolites were detected by reverse phase isocratic HPLC. CYP inhibitors and Spearman rank correlation were used to determine the significance of AQ4N metabolism versus specific CYP activity and/or expression. RESULTS: Anaerobic metabolism of AQ4N to the 2-electron reduction product, AQM, and the 4-electron reduced tertiary amine, AQ4, occurred in all 17 human liver microsome preparations. The range (+/- SE) for total AQ4N turnover was 14.26 +/- 1.43 nmol/incubate (highest) to 3.65 +/- 1.05 nmol/incubate (lowest). Metabolism was not detected in the absence of NADPH or microsomes. In aerobic incubates, AQM was less than 4% of anaerobic values whereas AQ4 was undetectable. CYP-mediated metabolism of AQ4N was inhibited completely by ketoconazole (KET) and carbon monoxide (CO), two global inhibitors of CYP-mediated metabolism. AQ4N metabolism correlated significantly with probes for CYP 3A, specifically benzoxylresorufin O-dealkylation [r(s) = 0.70,p <0.01] and tamoxifen N-demethylation (r(s) = 0.85, p < 0.01), but not with probes for CYPs 2C, 2D, and 1A. CYP 3A involvement was confirmed by the use of the CYP 3A specific inhibitor, triacetyloleandomycin (TAO), which repressed the formation of AQM to 13% of the uninhibited value and abolished completely the formation of AQ4. Alpha-naphthoflavone (ANF), an inhibitor of CYP 2C and 1A, had no significant effect on AQ4N metabolism. CONCLUSIONS: These data suggest that the human CYP 3A enzymes can contribute to the reductive metabolism of AQ4N. CYP 3A enzymes are highly expressed in a broad spectrum of human cancers. The results show that AQ4N requires anaerobic conditions for CYP 3A-mediated reduction and hence this subfamily of enzymes is likely to selectively activate AQ4N in hypoxic tumors.