A rapid, accurate and robust UHPLC-MS/MS method for quantitative determination of BMS-927711, a CGRP receptor antagonist, in plasma in support of non-clinical toxicokinetic studies.

Abstract BMS-927711 is a calcitonin gene-related peptide (CGRP) receptor antagonist that is being developed for the treatment of migraine. A rapid, accurate and robust assay was developed and validated for the quantitation of BMS-927711 in rat, monkey, rabbit and mouse plasma using ultra high performance liquid chromatography with tandem mass spectrometry (UHPLC–MS/MS). A simplified method screening strategy was utilized that included a liquid–liquid extraction (LLE) methodology and eleven LC columns (ten sub-2 μm UHPLC columns and one 2.6 μm HPLC column) for screening with emphasis on the removal of phospholipids, avoidance of metabolite interference and ruggedness of LC conditions. A stable isotope labeled [ 13 C 2 , D 4 ]-BMS-927711 was used as the internal standard, and 50 μL of plasma samples were used for extraction by automated LLE with methyl tert -butyl ether (MTBE) in 96-well format. Chromatographic separation was achieved with an isocratic elution and a gradient column wash on a Waters Acuity UPLC ® BEH C18 column (2.1 mm × 50 mm, 1.7 μm) with run time of 3.7 min. Positive electrospray ionization was performed using selected reaction monitoring (SRM) with transitions of m/z 535 > 256 for BMS-927711 and m/z 541 > 256 for [ 13 C 2 , D 4 ]-BMS-927711. The standard curve, which ranged from 3.00 to 3000 ng/mL for BMS-927711, was fitted to a 1/ x 2 weighted linear regression model. The intra-assay precision was within 5.2% CV, inter-assay precision was within 5.9% CV, and the assay accuracy was within ±5.2% deviation (%Dev) of the nominal values in all the species. The stability of an N-carbamoyl glucuronide metabolite was carefully investigated, and the conversion of this metabolite to BMS-927711 was minimal and manageable without a stabilization procedure. The method was successfully applied to multiple non-clinical toxicokinetic studies in different species in support of the investigative new drug (IND) filing.