Abstract
Comparative metabolite profiling of geldanamycin and 17-allylamino-17-demethoxygeldanamycin (17AAG) using human liver microsomes in normoxia and hypoxia was conducted to understand their differential metabolic fates. Geldanamycin bearing a 17-methoxy group primarily underwent reductive metabolism, generating the corresponding hydroquinone under both conditions. The formed hydroquinone resists further metabolism and serves as a reservoir. On exposure to oxygen, this hydroquinone slowly reverts to geldanamycin. In the presence of glutathione, geldanamycin was rapidly converted to 19-glutathionyl geldanamycin hydroquinone, suggesting its reactive nature. In contrast, the counterpart (17AAG) preferentially remained as its quinone form, which underwent extensive oxidative metabolism on both the 17-allylamino sidechain and the ansa ring. Only a small amount (<1%) of 19-glutathione conjugate of 17AAG was detected in the incubation of 17AAG with glutathione at 37°C for 60 min. To confirm the differential nature of quinone-hydroquinone conversion between the two compounds, hypoxic incubations with human cytochrome P450 reductase at 37°C and direct injection analysis were performed. Approximately 89% of hydroquinone, 5% of quinone, and 6% of 17-O-demethylgeldanamycin were observed after 1-min incubation of geldanamycin, whereas about 1% of hydroquinone and 99% of quinone were found in the 60-min incubation of 17AAG. The results provide direct evidence for understanding the 17-substituent effects of these benzoquinone ansamycins on their phase I metabolism, reactivity with glutathione, and acute hepatotoxicity.
Footnotes
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Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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doi:10.1124/dmd.106.009639.
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ABBREVIATIONS: 17AAG, 17-allylamino-17-demethoxygeldanamycin; HSP90, heat shock protein 90; HLM, human liver microsome; hP450R, human NADPH/cytochrome P450 reductase; LC/QTOF-MS, liquid chromatography and quadrupole time-of-flight mass spectrometry; LC/QTOF-MS/MS, liquid chromatography and quadrupole time-of-flight tandem mass spectrometry; HPLC, high-performance liquid chromatography; DAD, diode array detector; GQH2, geldanamycin hydroquinone; M1, 17AAG hydroquinone; M2, 6-O-demethyl-17-(2′,3′-dihydroxypropylamino)-geldanamycin; M3, 12-O-demethyl-17-(2′,3′-dihydroxypropylamino)-geldanamycin; M4, 22-hydroxyl-17-(2′,3′-dihydroxypropylamino)-geldanamycin; M5, 17-(2′,3′-dihydroxypropylamino)-geldanamycin; M6, 17-(didehydroallylamino)-geldanamycin; M7, 17-(3′-hydroxyallylamino)-geldanamycin; M8, 17-aminogeldanamycin; M9, 12-O-demethyl-17AAG; M10, 22-hydroxyl-17AAG; 17DMG, 17-O-demethylgeldanamycin; GSGQH2, 19-glutathionyl geldanamycin hydroquinone.
- Received February 2, 2006.
- Accepted September 22, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
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