Quantitation of SU1 1248, an oral multi-target tyrosine kinase inhibitor, and its metabolite in monkey tissues by liquid chromatograph with tandem mass spectrometry following semi-automated liquid-liquid extraction.
SU11248 is a potent inhibitor of PDGFR, VEGFR, KIT, and Flt3, and is currently under Phase I clinical evaluation as an anticancer drug. A sensitive and specific analytical method for the quantitation of SU11248 and its metabolite in several monkey tissues (liver, kidney, brain and white fat) using LC-MS-MS following semi-automated liquid-liquid extraction (LLE) was developed and validated. Amounts of 50 mg of tissue were homogenized using an ultrasonic processor. After addition of the stable labelled internal standard (IS) and ammonium hydroxide (0.3%), samples were extracted with 2.5 ml of tert-butyl methyl ether. Following centrifugation, aliquots of 1.8 ml of the organic phase were transferred into a 96-well plate. The Packard Multiprobe II robotic liquid handler was used to perform all steps mentioned above. The organic phase was dried and the residue was reconstituted with 800 microl of 15 mM ammonium formate buffer solution (pH 3.25) using a Tomtec Quadra 96 workstation. Aliquots of 10 microl of the resulting solution were injected into the LC-MS-MS system. A Symmetry Shield C8 column (50 mm x 2.1 mm, 3.5 microm) was used to perform the chromatographic analysis. The mobile phase was 15 mM ammonium formate buffer solution (pH 3.25)-acetonitrile (74:26 (v/v)) with a flow-rate of 0.35 ml/min. Retention times of the metabolite and SU11248 were about 2.5 and 3.5 min, respectively. Total cycle time was 5 min. MS detection used the Applied Biosystems-MDS Sciex API 3000 with TurbolonSpray interface and multiple reaction monitoring (MRM) operated in positive ion mode. The method was validated for both compounds over the calibration range of about 2 and 2000 ng/g. The suitability and robustness of the method for in vivo samples were confirmed by analysis of monkey tissues from animals dosed with SU11248.
Barattè
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Global Drug Metabolism, Pharmacia, Viale Pasteur 10, 20014 Nerviano, Italy. simona.baratte@pharmacia.com
Sarati
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Frigerio
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James
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Ye
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Zhang
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D000818:Animals; D000970:Antineoplastic Agents / Q000032:analysis* / Q000378:metabolism / Q000493:pharmacokinetics; D002853:Chromatography, Liquid / Q000379:methods*; D004791:Enzyme Inhibitors / Q000032:analysis* / Q000378:metabolism / Q000493:pharmacokinetics; D000882:Haplorhini; D007211:Indoles / Q000032:analysis* / Q000378:metabolism / Q000493:pharmacokinetics; D013058:Mass Spectrometry / Q000379:methods*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D011758:Pyrroles / Q000032:analysis* / Q000378:metabolism / Q000493:pharmacokinetics; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D000077210:Sunitinib; D014018:Tissue Distribution
D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D007211:Indoles; D011758:Pyrroles; D011505:Protein-Tyrosine Kinases; D000077210:Sunitinib