title: CYP2D6 genotyping by liquid chromatography-electrospray ionization mass spectrometry.
abstract: Genetic polymorphisms can significantly affect the enzyme activity of the drug metabolizing enzyme Cytochrome P450 2D6 (CYP2D6; OMIM 124030). Accordingly, CYP2D6 genotyping is considered as a valid approach to predict the individual CYP2D6 metabolizing status. We introduce ion-pair reversed-phase high-performance liquid chromatography-electrospray ionization mass spectrometry (ICEMS) as method for the characterization of single base variants, small deletions, and insertions in the CYP2D6 gene. A two-step polymerase chain reaction (PCR) was developed for the simultaneous amplification of nine polymorphic regions within the CYP2D6 gene. Cleanup, separation, and denaturation of PCR amplicons were achieved by high-performance liquid chromatography. High-performance molecular mass measurements provided nucleotide composition profiles that principally enable the resolution of 37 reported CYP2D6 alleles. The developed assay was applied to the genotyping of 93 unrelated Austrian individuals. For validation, a selected number of samples and polymorphic sites were retyped by alternative genotyping technologies. The PCR-ICEMS assay turned out to be an accurate, robust, and cost-effective CYP2D6 genotyping strategy.
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Innsbruck, Austria.
authors.forename: Beate
authors.initials: B
authors.lastname: Beer
authors.forename: Robert
authors.initials: R
authors.lastname: Erb
authors.forename: Florian
authors.initials: F
authors.lastname: Pitterl
authors.forename: Harald
authors.initials: H
authors.lastname: Niederstätter
authors.forename: Olalla
authors.initials: O
authors.lastname: Maroñas
authors.forename: Alejandro
authors.initials: A
authors.lastname: Gesteira
authors.forename: Angel
authors.initials: A
authors.lastname: Carracedo
authors.forename: Irina
authors.initials: I
authors.lastname: Piatkov
authors.forename: Herbert
authors.initials: H
authors.lastname: Oberacher
mesh_terms: D002851:Chromatography, High Pressure Liquid; D019389:Cytochrome P-450 CYP2D6 / Q000235:genetics* / Q000378:metabolism; D005838:Genotype; D006801:Humans; D016133:Polymerase Chain Reaction; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D019389:Cytochrome P-450 CYP2D6