Methodology for clinical genotyping of CYP2D6 and CYP2C19

Beatriz Carvalho Henriques,Michael S Carr,Amanda Buchner ,Daniel Souery,Yabing Wang,Bahareh Behroozi Asl,Mojca Zvezdana Dernovšek ,Stacey Hume,Joshua Hague,Wolfgang Maier,Joanna Hauser,Sudhakar Sivapalan,Kristina Martens,Annamaria Cattaneo,Rachael Dong,Xiuying Hu,Vasyl Yavorskyy,Keanna Wallace,Neven Henigsberg,Ole Mors,Marcella Rietschel,Gerald Pfeffer,Katherine J Aitchison

doi:10.1038/s41398-021-01717-9

Abstract

Many antidepressants, atomoxetine, and several antipsychotics are metabolized by the cytochrome P450 enzymes CYP2D6 and CYP2C19, and guidelines for prescribers based on genetic variants exist. Although some laboratories offer such testing, there is no consensus regarding validated methodology for clinical genotyping of CYP2D6 and CYP2C19. The aim of this paper was to cross-validate multiple technologies for genotyping CYP2D6 and CYP2C19 against each other, and to contribute to feasibility for clinical implementation by providing an enhanced range of assay options, customizable automated translation of data into haplotypes, and a workflow algorithm. AmpliChip CYP450 and some TaqMan single nucleotide variant (SNV) and copy number variant (CNV) data in the Genome-based therapeutic drugs for depression (GENDEP) study were used to select 95 samples (out of 853) to represent as broad a range of CYP2D6 and CYP2C19 genotypes as possible. These 95 included a larger range of CYP2D6 hybrid configurations than have previously been reported using inter-technology data. Genotyping techniques employed were: further TaqMan CNV and SNV assays, xTAGv3 Luminex CYP2D6 and CYP2C19, PharmacoScan, the Ion AmpliSeq Pharmacogenomics Panel, and, for samples with CYP2D6 hybrid configurations, long-range polymerase chain reactions (L-PCRs) with Sanger sequencing and Luminex. Agena MassARRAY was also used for CYP2C19. This study has led to the development of a broader range of TaqMan SNV assays, haplotype phasing methodology with TaqMan adaptable for other technologies, a multiplex genotyping method for efficient identification of some hybrid haplotypes, a customizable automated translation of SNV and CNV data into haplotypes, and a clinical workflow algorithm.

Highlights

Many antidepressants, atomoxetine, and several antipsychotics are metabolized by CYP2D6 and CYP2C19 [1,2,3,4,5,6,7]
CYP2D6 Comparative genotypic and copy number variant (CNV) data across the technologies for samples with one and three copies of the CYP2D6 gene are shown in Supplementary Tables 2 and 3, respectively
Owing to the “no calls” in the AmpliSeq CNV data, we revised these to manual calls, where possible, after reviewing the vcf files

Summary

Introduction

Atomoxetine, and several antipsychotics are metabolized by CYP2D6 and CYP2C19 [1,2,3,4,5,6,7]. The high homology between CYP2D6 and these pseudogenes and the presence of flanking transposable genetic elements [11] makes the region vulnerable to the generation of variable copy numbers of the CYP2D6 gene and hybrid genes made up of sequence derived in part from CYP2D7 and in part from CYP2D6 [12,13,14,15,16,17,18,19,20,21]. While structural variants of CYP2C19 have recently been identified [26], the more commonly studied haplotypes result from single nucleotide variants (SNVs) [27] Haplotypes in both genes are referred to as “star alleles,” e.g., *2, *3, etc. Haplotypes in both genes are referred to as “star alleles,” e.g., *2, *3, etc. as defined by PharmVar [13], a consortium which maintains a curated catalog of allelic variation in genes impacting drug metabolism, disposition, and response

Objectives

Results

Conclusion