A validated LC–MS/MS assay for the simultaneous determination of the anti-leukemic agent dasatinib and two pharmacologically active metabolites in human plasma: Application to a clinical pharmacokinetic study

Dasatinib (Sprycel®) is a potent antitumor agent prescribed for patients with chronic myeloid leukemia (CML). To enable reliable quantification of dasatinib and its pharmacologically active metabolites in human plasma during clinical testing, a sensitive and reliable liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated. Samples were prepared using solid phase extraction on Oasis HLB 96-well plates. Chromatographic separation was achieved isocratically on a Luna phenyl–hexyl analytical column. Analytes and the stable labeled internal standards were detected by positive ion electrospray tandem mass spectrometry. The assay was validated over a concentration range of 1.00–1000 ng/mL for dasatinib and its two active metabolites. Intra- and inter-assay precision values for replicate QC control samples were within 5.3% for all analytes during the assay validation. Mean QC control accuracy values were within ±9.0% of nominal values for all analytes. Assay recoveries were high (>79%) and internal standard normalized matrix effects were minimal. The three analytes were stable in human plasma for at least 22 h at room temperature, for at least 123 days at −20 °C, and following at least six freeze–thaw cycles. The validated method was successfully applied to the quantification of dasatinib and two active metabolites in a human pharmacokinetic study.

► Dasatinib and active metabolites LC–MS/MS assay validation.
► Application of the assay to a human pharmacokinetic study.
► Active metabolite exposure in humans is low compared to dasatinib.