Apatinib, also known as YN968D1, is a novel antiangiogenic agent that selectively inhibits vascular endothelial growth factor receptor-2. Currently, apatinib is
undergoing phase II/III clinical trials in China for the treatment of solid tumors. Apatinib is extensively metabolized in humans, and its major metabolites in circulation include cis-3-hydroxy-apatinib (M1-1), trans-3-hydroxy-apatinib (M1-2), apatinib-25-N-oxide (M1-6), and cis-3-hydroxy-apatinib-O-glucuronide (M9-2). To investigate the pharmacokinetics of apatinib and its four major metabolites in patients with advanced colorectal cancer, a sensitive and selective liquid chromatography-tandem mass spectrometry method was developed and validated for the simultaneous determination of apatinib, M1-1, M1-2, M1-6, and M9-2 in human plasma. After a simple protein precipitation using acetonitrile as the precipitation solvent, all the analytes and the internal standard vatalanib were separated on a Zorbax Eclipse XDB C(18) column (50 mm × 4.6 mm, 1.8 μm, Agilent) using acetonitrile: 5 mmol/L ammonium acetate with 0.1% formic acid as the mobile phase with gradient elution. A chromatographic total run time of 9 min was achieved. Mass spectrometry detection was conducted through electrospray ionization in positive ion multiple reaction monitoring modes. The method was linear over the concentration range of 3.00-2000 ng/mL for each analyte. The lower limit of quantification for each analyte was 3.00 ng/mL. The intra-assay precision for all the analytes was less than 11.3%, the inter-assay precision was less than 13.8%, and the accuracy was between -5.8% and 3.3%. The validated method was successfully applied to a clinical pharmacokinetic study following oral administration of 500 mg apatinib mesylate in patients with advanced colorectal cancer.