Simultaneous Determination of Osimertinib and its Metabolites in Human Plasma for Clinical Applications Using High-Performance Liquid Chromatography–Tandem Mass Spectrometry

Background: Osimertinib, the first-line treatment for nonsmall cell lung cancer, has 2 main active metabolites, AZ5104 and AZ7550, which demonstrate potency against wild-type epidermal growth factor receptor mutations. In this study, a simple and sensitive analytical method using high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/mass spectrometry) was developed and validated to simultaneously quantify osimertinib and its active metabolites in human plasma. Methods: Plasma samples were subjected to protein precipitation with zinc sulfate solution and acetonitrile using osimertinib- 13 C, d 3 as an internal standard. The separation was performed on a ThermoScientific Hypersil GOLD C18 column (4.6 × 50 mm, 5 μm) with a mobile phase composed of 0.1% formic acid water–acetonitrile through gradient elution. The compounds were monitored using electrospray ionization in the positive mode with multiple reaction monitoring. Results: Excellent linearity of calibration curves was observed over the ranges of 10–1000 ng·mL −1 for osimertinib, and 1.5–120 ng·mL −1 for AZ7550 and AZ5104 in human plasma. Both within-run and between-run accuracies were within ±15%, with the coefficient of variation <15%. Conclusions: The method met all the validation criteria and was successfully applied to clinical samples from patients with nonsmall cell lung cancer. Furthermore, the correlation between steady-state trough concentrations of osimertinib, AZ5104, and AZ7550 and patients' baseline characteristics were explored. Women exhibited a significantly higher C min, ss for AZ7550 ( P = 0.023). In addition, body mass index was associated with the C min, ss of both AZ5104 ( P = 0.047) and AZ7550 ( P = 0.001).