Validated HPLC method for simultaneous quantification of mutant IDH1/2 inhibitors (enasidenib, ivosidenib and vorasidenib) in mouse plasma: Application to a pharmacokinetic study.

ABSTRACT

Isocitrate dehydrogenase (IDH) inhibitors comprise a novel class of anticancer drugs, which are approved to treat acute myeloid leukemia patients having mutations on IDH1/2. We report the development and validation of a high-performance liquid chromatography (HPLC) method for the simultaneous quantitation of IDH inhibitors, namely enasidenib (EDB), ivosidenib (IDB) and vorasidenib (VDB), in mouse plasma as per the US Food and Drug Administration regulatory guidelines. The method involves extraction of EDB, IDB and VDB along with internal standard (IS; phenacetin) from mouse plasma (100 µl) using a simple protein precipitation process. The chromatographic analysis was performed on an HPLC system using a gradient mobile phase (comprising 10 mm ammonium acetate and acetonitrile in a flow-gradient) and an X-Terra Phenyl column. The UV detection wave length was set at λmax 265 nm. EDB, IDB, VDB and the IS eluted at 7.36, 8.60, 9.50 and 5.12 min, respectively, with a total run time of 10 min. The calibration curve was linear over a concentration range of 0.20-12.5 µg/ml for EDB and 0.50-12.5 µg/ml for IDB and VDB (r2  = ≥0.998 for all of the analytes). Validation results met the acceptance criteria. The validated HPLC method was successfully applied to a pharmacokinetic study in mice.