Validated LC-MS/MS Method for Simultaneous Quantitation of Three PI3K Inhibitors, Copanlisib, Duvelisib and Idelalisib in Mouse Plasma: Application to a Pharmacokinetic Study in Mice

Phosphatidylinositol 3-kinase (PI3K) inhibitors are emerging novel class of anticancer drugs. In this work, we report the development and validation of an LC-MS/MS method for the simultaneous quantitation of three PI3K inhibitors namely copanlisib, duvelisib and idelalisib in mouse plasma as per the FDA regulatory guidelines. Liquid-liquid extraction was used to extract copanlisib, duvelisib and idelalisib along with internal standard (I.S; filgotinib) from mouse plasma. Thereafter, chromatographic separation was achieved on an Ascentis Express RP-amide column using an isocratic mobile phase (5 mM ammonium acetate and acetonitrile in the ratio of 25:75, v/v) at a flow-rate of 0.5 mL/min. The analytes and I.S were detected by positive electrospray ionization tandem mass spectrometry (ESI-MS/MS) operated in multiple-reaction-monitoring (MRM) mode. The analytical time was 2.50 min in total. The calibration curve ranged from 4.41-1029, 0.70-1395 and 0.77-1544 ng/mL for copanlisib, duvelisib and idelalisib, respectively. The intra-assay accuracy and precision were & LE;108 % and & LE;6.23 %; & LE;95.8 % and & LE;13.1 %; and & LE;101 % and & LE;7.92 % for copanlisib, duvelisib and idelalisib, respectively and the inter-assay accuracy and precision were & LE;107 % and & LE;6.13 %; & LE;97.6 % and & LE;9.41 %; and & LE;100% and & LE;7.79 % for copanlisib, duvelisib and idelalisib, respectively. The validated method was successfully applied to a pharmacokinetic study in mice.