Enantioselective Chiral Separation and Quantitation of PI3Kγ Inhibitor Eganelisib in Rat Plasma Using UHPLC-MS/MS:...

Eganelisib (EGN, IPI-549), an investigational new drug, is a potent first-in-class PI3Kγ inhibitor with high selectivity compared to other class I PI3K isoforms, with one asymmetric center. Enantiomers have similar physicochemical properties but differ in their biological characteristics, like absorption, distribution, metabolism, and excretion. Hence, a suitable analytical approach is required to investigate the enantioselective behavior in ADME-Tox. In the current study, a sensitive, reliable, and precise chiral LC-MS/MS method has been developed for quantifying EGN enantiomers in rat plasma, in accordance with USFDA guidelines. Enantiomeric separation was achieved within 15 min on a Chiralpak-IK-3 (250 × 4.6 mm, 3 μm) immobilized polysaccharide column, containing 10 mM ammonium bicarbonate and a premixed mixture of acetonitrile and methanol (80:20 v/v) as the mobile phase in the ratio of 10:90 (v/v), with isocratic elution at a flow rate of 0.6 mL/min. Multiple reaction monitoring (MRM) mode was used to detect EGN and SOF in positive electrospray ionization (ESI) mode, characterized by their transitions at m/z 529.3→326.3, 161.1, and m/z 465.1→447.1, 270.2, 252.1. The method demonstrated a linear response in the range of 1-600 ng/mL for both enantiomers, with a lower limit of quantification of 1 ng/mL. The validated bioanalytical method was applied to evaluate the enantioselective pharmacokinetics of EGN enantiomers in rat plasma for the first time, utilizing a model-independent approach. The (R)-enantiomer showed relative fold differences in C_max, t_1/2, AUC_0-t, and AUC_0-∞ when compared to the (S)-enantiomer, indicating the enantioselective behavior of EGN enantiomers.