PacBio long-read amplicon sequencing enables scalable high-resolution population allele typing of the complex CYP2D6 locus

Sarah Charnaud,Jessica Brewster,Shazia Ruybal-Pesántez,Lucie Semenec,Robert James,Dulcie Lautu-Gumal,Harin Karunajeewa,Ivo Mueller,Ramin Mazhari,Caitlin Bourke,Melanie Bahlo,Jacob E Munro

doi:10.1038/s42003-022-03102-8

Abstract

The CYP2D6 enzyme is estimated to metabolize 25% of commonly used pharmaceuticals and is of intense pharmacogenetic interest due to the polymorphic nature of the CYP2D6 gene. Accurate allele typing of CYP2D6 has proved challenging due to frequent copy number variants (CNVs) and paralogous pseudogenes. SNP-arrays, qPCR and short-read sequencing have been employed to interrogate CYP2D6, however these technologies are unable to capture longer range information. Long-read sequencing using the PacBio Single Molecule Real Time (SMRT) sequencing platform has yielded promising results for CYP2D6 allele typing. However, previous studies have been limited in scale and have employed nascent data processing pipelines. We present a robust data processing pipeline “PLASTER” for accurate allele typing of SMRT sequenced amplicons. We demonstrate the pipeline by typing CYP2D6 alleles in a large cohort of 377 Solomon Islanders. This pharmacogenetic method will improve drug safety and efficacy through screening prior to drug administration.

Highlights

The cytochrome P450-2D6 (CYP2D6) enzyme is estimated to metabolize 25% of commonly used pharmaceuticals and is of intense pharmacogenetic interest due to the polymorphic nature of the CYP2D6 gene
The *10 allele is reportedly frequently observed in a tandem arrangement with the CYP2D6-D7 fusion allele *3629,30, which suggests that NA17058 may be harboring a *36-*10 tandem allele on each haplotype
We have developed a robust pipeline to sequence and phase alleles of the pharmacogenetically important gene CYP2D6 using PacBio long-read amplicon sequencing

Summary

Introduction

The CYP2D6 enzyme is estimated to metabolize 25% of commonly used pharmaceuticals and is of intense pharmacogenetic interest due to the polymorphic nature of the CYP2D6 gene. We demonstrate the pipeline by typing CYP2D6 alleles in a large cohort of 377 Solomon Islanders. This pharmacogenetic method will improve drug safety and efficacy through screening prior to drug administration. 1234567890():,; Pharmacogenetics is the study of the association between genetic variation and the differences in an individual’s metabolic response to certain drugs. It has been established that the frequencies of pharmacogenetic variants may vary widely between populations with different ancestries[1]. CYP2D6 is highly polymorphic, with different alleles displaying a wide spectrum of enzymatic activity. Estimates of metabolizer phenotype frequencies across several distinct populations, inferred from genotyping data, range from 0.4 to 5.4% for PMs, 0.4 to 11% for IMs, 67 to 90% for EMs, and 1 to 21% for UMs6.

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Conclusion