An LC–MS/MS method for determination ofthe bromodomain inhibitor ZEN-3694 and itsmetabolite ZEN-3791 in human plasma: supplementary data

We have developed and validated a novel LC–MS/MS method for the simultaneous quantification of ZEN-
3694 and its active metabolite ZEN-3791 in human plasma after protein precipitation. Stable isotope labeled
versions were used as internal standards. Chromatographic separation was achieved on a Kinetex
C18 column using 0.1% formic acid in H2O and 0.1% formic acid in MeOH as mobile phases. Detection
was performed via positive electrospray ionization mode with multiple reaction monitoring. The assay
exhibited linearity in the concentration range of 5–5000 ng/ml for both analytes. Intra- and inter-assay
precision and accuracy were within ±11%. ZEN-3694 and ZEN-3791 recoveries were between 93 and 105%.
This LC–MS/MS assay is an essential tool to study ZEN-3694 in an ongoing clinical trial (NCT04840589).