Establishment and validation of a sensitive LC-MS/MS method for quantification of KRASG12C protein PROTAC molecule LC-2 in rat plasma and its application to in vivo pharmacokinetic studies of LC-2 PEGylated liposomes.

Abstract

LC-2, is a molecule of Proteolysis Targeting Chimeras (PROTACs), with a large molecular weight, poor water solubility and low system bioavailability, which was designed to degrade KRASG12C protein. In this study, LC-2 PEGylated liposomes were developed and characterized. Moreover, a rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method in rat plasma was established and effectively utilized to an in vivo pharmacokinetic investigation. LC-2 PEGylated liposomes with better properties were prepared by an improved ethanol injection method. The chromatographic separation was achieved on an Agilent Eclipse XDB-CN column (100 × 2.1 mm, 3.5μm.) with acetonitrile-ammonium deionized water (5 mM) (80:20, v/v) at a flow rate of 0.5 mL/min. The mass spectra of LC-2 and the IS (gefitinib) were obtained at m/z 1132.5→626.4 and 447.1→128.2, respectively. The pharmacokinetic study was carried out by analyzing plasma concentrations of LC-2 solution or produced LC-2 PEGylated liposomes in rats using the developed and validated method. The pharmacokinetic results indicate that PEGylated liposome-encapsulation protected LC-2 from the influence of endogenous protein binding, improved insolubility, prolonged t1/2 and increased system bioavailability. This study provides a feasible solution for future preclinical and clinical studies of LC-2 and/or other PROTACs.