Therapeutic drug monitoring (TDM) help to improve treatment efficacy and safety. Therefore,
a simple and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method
was developed and validated for the simultaneous monitoring of 11 tyrosine kinase
inhibitors (TKIs) in human plasma. TKIs included in the assay are used in the treatment
of chronic myeloid leukemia (CML: imatinib, dasatinib, nilotinib, bosutinib, ponatinib),
polycythemia vera (ruxolitinib), chronic lymphocytic leukemia (ibrutinib) and rheumatoid
arthritis (filgotinib, tofacitinib, baricitinib, peficitinib). Caffeine was also included
in the method. Caffeine increases the acidity of the stomach and decreases its pH
as well as is a competitive inhibitor of cytochrome P450 isoenzymes. Thus, it may
influence absorption and metabolism of some TKIs, by modifying their plasma levels.
The analytes of interest and their stable isotope-labeled internal standards were
extracted from 200 μL of human plasma. Microelution-solid phase extraction (μ-SPE)
was optimized for method validation and compared to simple protein precipitation (PPT).
A gradient elution on a Poroshell 120 EC-C18 column at 60 °C and a flow rate of 0.5 mL/min
was applied for analyte separation. The analytical run lasted 8 min and it was followed
by a re-equilibration time of 4 min. Dynamic multiple reaction monitoring scan in
the positive ionization mode was applied to improve method sensitivity. Endogenous
plasma phospholipids can strongly affect MS analysis. Hence, the monitoring of endogenous
phospholipids was included in the assay. Full validation of the method was achieved,
including tests of precision, accuracy, trueness, linearity, extraction recovery,
matrix effect, process efficiency, stability, sensitivity (with excellent LLOQs),
selectivity, identity confirmation and carry-over effect. Regarding sample cleanup,
more than 91% of early eluting and more than 96% of late eluting endogenous phospholipids
were eliminated by μ-SPE when compared to PPT. This method enables the simultaneous
plasma monitoring of 11 TKIs and caffeine and ensures high effectiveness in phospholipids
elimination. The present approach is currently used in our clinical practice, being
applied to TDM of dasatinib, imatinib, nilotinib and ponatinib. TKIs plasma monitoring
helps to individualize dose adjustment and manage adverse effects in CML patients.