LC-MS/MS method for the quantitation of a dual PI3K/BRD4 inhibitor SF2523 in mouse plasma: application to plasma protein binding and metabolism studies.

A sensitive and selective liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantitation of dual PI3K/BRD4 inhibitor SF2523 in mouse plasma. The analysis was performed on a UPLC system connected to a Shimadzu 8060 mass spectrometer by electrospray in positive multiple reaction monitoring mode. Chromatographic separation was carried out on an ACE Excel C18 column with a gradient elution containing 0.1% formic acid and methanol as the mobile phase. The linearity was conducted in the concentration range of 0.1-500 ng/mL for SF2523 in 100μL plasma. The inter- and intra-batch precision (% RSD) were both lower than 13.5 %, with the accuracy (%Bias) ranged from varied from -10.03% to 11.56%. The validated method was successfully applied to plasma protein binding and in vitro metabolism studies. SF2523 was highly bound to mouse plasma proteins (>95% bound). Utilizing mouse S9 fractions, a total of seven phase I and II metabolites were identified with hydroxylation found to be the major metabolic pathway. Metabolites identification included analysis of retention behaviors, molecular weight changes and MS/MS fragment patterns of SF2523 and the metabolites. This newly developed and validated method allows the rapid and easy determination of the SF2523 concentration with high sensitivity in low sample volume and can be applied to future pre-clinical studies.