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

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 C18 column using an isocratic mobile phase of methanol:5 mM 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.

► Simultaneously determine flumatinib and its metabolites in CML patient plasma.
► Flumatinib and its metabolites have great differences in physicochemical properties.
► Each analyte was retained on the C18 column by using strong acidic mobile phase.
► A simple one-step protein precipitation increased throughput and efficiency.
► The method was applied to clinical studies of flumatinib mesylate in CML patients.