Background: EAI045 is the fourth-generation epidermal growth factor receptor tyrosine
kinase inhibitor (EGFR TKI), which can overcome acquired resistance to the third-generation EGFR
TKIs and is the first allosteric inhibitor that targets T790M and C797S EGFR mutants.
Methods: A rapid and sensitive LC-MS/MS method was established and validated for the quantification
of EAI045 in rat plasma. Chromatographic separation was carried out at 25°C on a Hypersil GOLD C18
column (50 mm × 2.1 mm, 3 μm) and eluted on a gradient mobile phase of water (containing 0.1% formic
acid) and acetonitrile at a flow rate of 0.5 mL/min. The mass spectrometer was operated in the positive
ESI mode and selected reaction monitoring mode.
Results: The assay was validated over a concentration range of 1.0 - 1000 ng/mL for EAI045 with a
lower limit of quantification (LLOQ) of 1.0 ng/mL. The intra- and inter-batch accuracy for the EAI045
ranged from 92.25% to 97.18% and 95.94% to 102.69%, and the intra- and inter-batch precision for the
EAI045 ranged from 1.41% to 4.57% and 5.18% to 6.37%, respectively. The extraction recovery, matrix
effect and stability met all requirements of the guidelines for bioanalytical method validation.
Conclusion: The rapid and sensitive LC-MS/MS method was successfully applied in a pharmacokinetic
study of EAI045 following oral administration (5 mg/kg) to rats.