title: Development and validation of a quantitative assay for the determination of tamoxifen and its five main phase I metabolites in human serum using liquid chromatography coupled with tandem mass spectrometry.
abstract: A sensitive bioanalytical assay for the quantitative determination of tamoxifen and five of its phase I metabolites (N-desmethyltamoxifen, N-desmethyl-4-hydroxytamoxifen, N-desmethyl-4'-hydroxytamoxifen, 4-hydroxytamoxifen and 4'-hydroxytamoxifen) in serum is described. The method has been fully validated at ranges covering steady-state serum concentrations in patients receiving therapeutic dosages of tamoxifen. The bioanalytical assay is based on reversed phase liquid chromatography coupled with tandem mass spectrometry in the positive ion mode using multiple reaction monitoring for drug (-metabolite) quantification. The sample pretreatment consists of protein precipitation with acetonitrile using only 50 μL of serum. In the past, numerous assays have been developed by other groups for the quantification of tamoxifen and its phase I metabolites. However, the number of metabolites included in these studies is very limited and only very few of these assays have been fully validated. A liquid chromatography tandem mass spectrometry assay for the quantification of tamoxifen and four phase I metabolites in human serum that was previously developed by our group is now explicitly improved and described herein. Time of analysis has been reduced by 50% and sensitivity was increased by a reduction of the lower limit of quantification from 1.0 to 0.2 ng/mL for 4-hydroxytamoxifen and 4'-hydroxytamoxifen. Additionally, two phase I metabolites that have never been quantified in human serum hitherto, namely 4'-hydroxytamoxifen and N-desmethyl-4'-hydroxytamoxifen, were included in this assay. Validation results demonstrate an accurate and precise quantification of tamoxifen, N-desmethyltamoxifen, N-desmethyl-4-hydroxytamoxifen, N-desmethyl-4'-hydroxytamoxifen, 4-hydroxytamoxifen and 4'-hydroxytamoxifen in human serum. The applicability of the assay was demonstrated and it is now successfully used to support clinical studies in which patient-specific dose optimization is performed based on serum concentrations of tamoxifen metabolites.
authors.affiliation: Department of Pharmacy & Pharmacology, Slotervaart Hospital/The Netherlands Cancer Institute, Amsterdam, The Netherlands. Bas.Teunissen@slz.nl
authors.forename: S F
authors.initials: SF
authors.lastname: Teunissen
authors.forename: N G L
authors.initials: NG
authors.lastname: Jager
authors.forename: H
authors.initials: H
authors.lastname: Rosing
authors.forename: A H
authors.initials: AH
authors.lastname: Schinkel
authors.forename: J H M
authors.initials: JH
authors.lastname: Schellens
authors.forename: J H
authors.initials: JH
authors.lastname: Beijnen
mesh_terms: D056148:Chromatography, Reverse-Phase / Q000379:methods*; D004355:Drug Stability; D006801:Humans; D016018:Least-Squares Analysis; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013629:Tamoxifen / Q000031:analogs & derivatives* / Q000097:blood* / Q000737:chemistry / Q000302:isolation & purification; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D013629:Tamoxifen