Abstract
Methylation of lysine by histone methyltransferases can be reversed by lysine demethylases (KDMs). Different KDMs have distinct oncogenic functions based on their cellular localization, stimulating cancer cell proliferation, reducing the expression of tumor suppressors, and/or promoting the development of drug resistance. JIB-04 is a small molecule that pan-selectively inhibits KDMs, showing maximal inhibitory activity against KDM5A, and as secondary targets, KDM4D/4B/4A/6B/4C. Recently, it was found that JIB-04 also potently and selectively blocks HIV-1 Tat expression, transactivation, and virus replication in T cell lines via the inhibition of a new target, serine hydroxymethyltransferase 2. Pharmacokinetic characterization and an analytical method for the quantification of JIB-04 are necessary for the further development of this small molecule. Herein, a sensitive, specific, fast and reliable UHPLC-MS/MS method for the quantification of JIB-04 in rat plasma samples was developed and fully validated using a SCIEX 6500+ triple QUAD LC-MS system equipped with an ExionLC UHPLC unit. The chromatographic separation was achieved on a reverse phase ACE Excel 2 Super C18 column with a flow rate of 0.5 mL/min under gradient elution. The calibration curves were linear (r2 > 0.999) over concentrations from 0.5 to 1000 ng/mL. The accuracy (RE%) was between -7.4% and 3.7%, and the precision (CV%) was 10.2% or less. The stability data showed that no significant degradation occurred under the experimental conditions. This method was successfully applied to the pharmacokinetic study of JIB-04 in rat plasma after intravenous and oral administration and the oral bioavailability of JIB-04 was found to be 44.4%.