title: Liquid chromatography-tandem mass spectrometric assay for ponatinib and N-desmethyl ponatinib in mouse plasma.
abstract: Ponatinib is a multi-targeted third generation BCR-ABL1 tyrosine-kinase inhibitor approved for specific types of leukemia. A bioanalytical assay for this drug and its N-desmethyl metabolite in mouse plasma was developed and validated using liquid chromatography-tandem mass spectrometric (LC-MS/MS) with liquid-liquid extraction as sample pre-treatment procedure. After extraction with tert-butyl methyl ether of both analytes with their isotopically labeled internal standards and evaporation and reconstitution of the extract, compounds were separated by reversed-phase liquid chromatography under alkaline conditions. After electrospray ionization, both compounds were quantified in the selected reaction monitoring mode of a triple quadrupole mass spectrometer. The linear assay was validated in the ranges 5-5000ng/ml for ponatinib and 1-1000ng/ml for N-desmethyl ponatinib. Within-run (n=18) and between-run (3 runs; n=18) precisions were 10% and 12% at the lower limit of quantification for the metabolite, all other precisions were ≤8% for the metabolite and ≤6% for ponatinib. Accuracies were between 92 and 108% for both compounds in the whole calibration range. The drug was sufficiently stable under most relevant analytical conditions, only ponatinib showed more than 15% hydrolytic degradation after storage for 6h and longer at ambient temperature in mouse plasma. Finally, the assay was successfully applied to determine plasma drug levels and study pharmacokinetics after oral administration of ponatinib to female FVB mice.
authors.affiliation: Utrecht University, Faculty of Science, Department of Pharmaceutical Sciences, Division of Pharmacoepidemiology & Clinical Pharmacology, Universiteitsweg 99, 3584CG Utrecht, The Netherlands; Utrecht University, Faculty of Science, Department of Pharmaceutical Sciences, Division of Chemical Biology & Drug Discovery, Universiteitsweg 99, 3584CG Utrecht, The Netherlands. Electronic address: R.W.Sparidans@uu.nl.
authors.forename: Rolf W
authors.initials: RW
authors.lastname: Sparidans
authors.affiliation: The Netherlands Cancer Institute, Division of Molecular Oncology, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.
authors.forename: Anita
authors.initials: A
authors.lastname: Kort
authors.affiliation: The Netherlands Cancer Institute, Division of Molecular Oncology, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.
authors.forename: Alfred H
authors.initials: AH
authors.lastname: Schinkel
authors.affiliation: Utrecht University, Faculty of Science, Department of Pharmaceutical Sciences, Division of Pharmacoepidemiology & Clinical Pharmacology, Universiteitsweg 99, 3584CG Utrecht, The Netherlands; The Netherlands Cancer Institute, Department of Clinical Pharmacology, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.
authors.forename: Jan H M
authors.initials: JH
authors.lastname: Schellens
authors.affiliation: Utrecht University, Faculty of Science, Department of Pharmaceutical Sciences, Division of Pharmacoepidemiology & Clinical Pharmacology, Universiteitsweg 99, 3584CG Utrecht, The Netherlands; The Netherlands Cancer Institute, Department of Clinical Pharmacology, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands; MC Slotervaart, Department of Pharmacy & Pharmacology, Louwesweg 6, 1066 EC Amsterdam, The Netherlands.
authors.forename: Jos H
authors.initials: JH
authors.lastname: Beijnen
mesh_terms: D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods*; D005260:Female; D007093:Imidazoles / Q000097:blood* / Q000737:chemistry; D057230:Limit of Detection; D016014:Linear Models; D051379:Mice; D011724:Pyridazines / Q000097:blood* / Q000737:chemistry; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D007093:Imidazoles; D011724:Pyridazines; C545373:ponatinib
keywords: LC−MS/MS; Liquid-liquid extraction; Mouse plasma; N-desmethyl ponatinib; Ponatinib