Development and Validation of a Simultaneous Quantification Method for 12 Targeted Therapies and 3 Active Metabolites in Human Plasma Using Liquid Chromatography–Tandem Mass Spectrometry: An Application to Therapeutic Drug Monitoring in Patients With Hematological Malignancies

BACKGROUND: Treatment strategies for hematological cancers and immunological diseases increasingly incorporate targeted oral therapies. These drugs provide improved quality of life but exhibit complex pharmacokinetics. Therapeutic drug monitoring (TDM) may help optimize treatment in various clinical situations, including managing drug-drug interactions, assessing adherence, evaluating exposure-response relationships, and investigating suspected drug toxicities. In this study, we developed and validated a sensitive liquid chromatography-tandem mass spectrometry method for simultaneous quantification of multiple targeted therapies and applied this method to clinical samples for TDM. METHODS: After simple protein precipitation of plasma samples, chromatographic separation was performed on a UPLC system coupled with MS/MS in positive ionization mode. The mobile phase consisted of a gradient elution using 10 mM of ammonium formate with 0.1% (v/v) formic acid (phase A) and acetonitrile with 0.1% (v/v) formic acid (phase B), at a flow rate of 300 µL/min. RESULTS: The analysis time was 7.0 minutes per run. Calibration curves were linear over the ranges of 0.5-500 ng/mL for ruxolitinib, tofacitinib, baricitinib, and hydroxyquizartinib AC488; 5-2500 ng/mL for asciminib, gilteritinib, and quizartinib; 50-10000 ng/mL for ivosidenib, venetoclax, midostaurin, CGP52421, CGP62221, and pacritinib; 100-100000 ng/mL for enasidenib; and 500-100000 ng/mL for eltrombopag. All analytes showed correlation coefficients above 0.99. Intra- and interday precision values were below 14.67%. CONCLUSIONS: We developed and validated a sensitive liquid chromatography-tandem mass spectrometry method requiring only 50 µL of plasma volume for the quantification of 12 targeted oral anticancer drugs and 3 active metabolites. This multianalyte assay offers strong potential for TDM in patients receiving contemporary anticancer treatments.