Simultaneous determination of flumatinib and its two major metabolites in plasma of chronic myelogenous leukemia patients by liquid chromatography-tandem mass spectrometry

Abstract

Flumatinib is an antineoplastic tyrosine kinase inhibitor used for the treatment of chronic myelogenous leukemia (CML). Its major metabolites in the circulation are N-desmethyl flumatinib (M1) and amide hydrolysis product (M3). To investigate the pharmacokinetics of flumatinib in CML patients, a simple, specific and rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous determination of flumatinib and its two major metabolites in patient plasma. After a simple, one-step protein precipitation with methanol, flumatinib, its two metabolites, and internal standard (HHGV-E) were separated on a C(18) column using an isocratic mobile phase of methanol:5mM ammonium acetate:formic acid (60:40:0.4, v/v/v). A total chromatographic run time of 4.2 min was achieved. The detection was performed in multiple reaction monitoring mode, using the transitions of m/z 563→m/z 463 for flumatinib, m/z 549→m/z 463 for M1, m/z 303→m/z 175 for M3, and m/z 529→m/z 429 for HHGV-E. The method was linear over the concentration ranges of 0.400-400 ng/mL for flumatinib, 0.100-100 ng/mL for M1, and 0.200-200 ng/mL for M3, using only 50 μL of plasma. The intra- and inter-day precisions were less than 8.5% for flumatinib, 9.8% for M1, and 10.6% for M3 in terms of the relative standard deviation. The accuracy was within ± 2.2% for flumatinib, ± 6.0% for M1, and ± 9.9% for M3 in terms of relative error. The validated method was successfully applied to clinical pharmacokinetic studies of flumatinib mesylate in CML patients following oral administration at all dosage regimens.