LC-MS/MS method for quantification of avacopan in human plasma from patients treated for antineutrophil cytoplasmic antibody-associated vasculitis

Avacopan is a new treatment for antineutrophil cytoplasmic antibody-associated vasculitis (ANCA-AAV). To date, there has been a lack of published bioanalytical methods to assay it in plasma, resulting in sparse pharmacokinetic data in clinical settings. The objective of this study was to develop and validate a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the quantification of avacopan in human plasma. [ 2 H 4 ]-Avacopan was as used as internal standard (IS). Samples were prepared by protein precipitation and separated was on an Accurore® C18 column (2.1×50 mm, 2.6 µm), with an elution gradient at a flow rate of 0.5 mL/min. The mobile phase consisted of acetonitrile (0.1% formic acid) and water (0.1% formic acid). The analysis run time was 6 minutes. Avacopan was detected by electrospray ionization on a TSQ Quantis® triple quadrupole mass spectrometer (ThermoFisher Scientific). The linearity of method ranged from 10 to 800 ng/mL. The within-run and between-run relative standard deviations were <10.2%. The within-run and between-run relative errors ranged from 2.4 to 14.4%. The IS-normalized matrix effect ranged from 2.2% to 5.1%, and the IS-normalized extraction recovery ranged from 104.3% to 109.7%. The method was fully validated, including checks on linearity, dilution integrity and carry-over. Plasma samples from 16 patients undergoing treatment for ANCA-AAV were then successfully treated with the method. The reanalysis of the samples incurred was below 14%. This method is suitable for plasma drug monitoring of avacopan because it is both accurate and precise, and meets all validation criteria. • Rapid quantification of avacopan in human plasma • First fully validated LC-MS/MS method to quantify avacopan in human plasma • Suitable for plasma drug monitoring and PK/PD studies.