Targeted next generation sequencing as a tool for precision medicine

Area of Science:

• Genomics
• Pharmacogenomics
• Molecular Biology

Background:

• Targeted next-generation sequencing (NGS) is crucial for identifying genetic variations influencing drug response and adverse effects, essential for personalized pharmacotherapy.
• Challenges exist in applying NGS to pharmacogenes with complex structures, high homology, and pseudogenes, particularly for detecting copy number variation (CNV) in CYP2D6 and specific polymorphisms like UGT1A1*28.
• The PGxSeq panel was developed to address these challenges in pharmacogene analysis.

Purpose of the Study:

• To develop and validate PGxSeq, a targeted exome sequencing panel for pharmacogenes.
• To assess the panel's ability to detect single nucleotide variants (SNVs), copy number variations (CNVs), and specific polymorphisms in pharmacogenes.
• To evaluate the clinical utility of PGxSeq for identifying individuals requiring altered pharmacotherapy.

Main Methods:

• A capture probe panel was designed to target 422 kb of coding regions in 100 pharmacogenes.
• Next-generation sequencing (NGS) was performed on 235 subjects.
• Bioinformatics tools were used for variant calling, including CYP2D6 CNV detection, and in silico analysis of SNV frequency and functional impact.

Main Results:

• The PGxSeq panel achieved a depth-of-coverage (DOC) ≥ 20× for over 94% of the target sequence.
• Accurate detection of 39 clinically relevant gene variants was achieved with 99.9% concordance compared to standard genotyping, including CYP2D6 CNV and UGT1A1*28.
• A significant proportion of subjects (78%) carried variants necessitating altered pharmacotherapy, with allele frequencies of novel variants aligning with large genomic datasets.

Conclusions:

• PGxSeq provides a comprehensive, rapid, and reliable method for detecting common and novel SNVs in pharmacogenes.
• The panel supports the advancement of precision medicine by facilitating individualized pharmacotherapy.
• PGxSeq demonstrates high accuracy in identifying complex genetic variations relevant to drug response.