Simultaneous and rapid determination of 12 tyrosine kinase inhibitors by LC-MS/MS in human plasma: Application to therapeutic drug monitoring in patients with non-small cell lung cancer

Abstract

In recent years, more than 50 tyrosine kinase inhibitors (TKIs) was indicated against numerous cancers, especially outstanding advantages in the treatment of non-small cell lung cancer (NSCLC), and several studies have shown that therapeutic drug monitoring (TDM) of TKIs can improve treatment efficacy and safety. The present study aimed to develop and validate a LC-MS/MS method for the TDM of 12 TKIs (gefitinib, erlotinib, afatinib, dacomitinib, icotinib, osimertinib, crizotinib, ceritinib, alectinib, dabrafenib, trametinib, anlotinib) in patients with NSCLC. The analytes of interest and internal standard were extracted from human plasma. Salting-out assisted liquid-liquid extraction (SALLE) with 5 M ammonium acetate solution was optimized for method validation and compared to simple protein precipitation (PPT). Chromatographic separation was conducted on Waters X bridge C18 column (100 × 4.6 mm, 3.5 μm) using a gradient elution of acetonitrile/5mM ammonium acetate in pure water with 0.1% (v/v) formic acid at 40 °C within 6 min. The total flow was maintained at 1 mL/min, 30% of the post column flow was split into the mass spectrometer and the rest to waste via a 3-way tee. The mass analysis was performed by positive ion electrospray ionization (ESI) in multiple-reaction monitoring (MRM) mode. The assay was validated based on the guidelines on bioanalytical methods by FDA. This quantification method was proved to be satisfactory in selectivity, accuracy, precision, linearity (r² > 0.995), recovery, matrix effect and stability and the accuracy was further assessed in plasma with a degree of hemolysis of 4%. The described method to simultaneously quantify the 12 selected anticancer drugs in human plasma was successfully validated and applied to routine TDM of gefitinib, erlotinib, icotinib, osimertinib, crizotinib and anlotinib in cancer patients. TKIs plasma monitoring helps to individualize dose adjustment and manage adverse effects in NSCLC patients.

Keywords: LC-MS/MS; Salting-out assisted liquid-liquid extraction; Therapeutic drug monitoring; Tyrosine kinase inhibitors.