A Variant Detection Pipeline for Inherited Cardiomyopathy-Associated Genes Using Next-Generation Sequencing

Area of Science:

• Cardiovascular Genetics
• Molecular Diagnostics
• Genomic Medicine

Background:

• Genetic testing is crucial for diagnosing inherited cardiomyopathies.
• Next-generation sequencing (NGS) offers a reliable approach for identifying disease-causing mutations.
• A validated NGS pipeline is essential for routine clinical application.

Purpose of the Study:

• To validate a novel NGS pipeline for detecting cardiomyopathy-related genetic mutations.
• To assess the analytical performance of the pipeline using a targeted gene panel.
• To determine the feasibility of this assay for diagnostic use in inherited cardiac conditions.

Main Methods:

• Development and validation of an NGS pipeline targeting 74 cardiomyopathy genes.
• Processing of 52 hypertrophic cardiomyopathy patient samples and one reference sample (NA12878).
• Assessment of sequencing coverage, variant call sensitivity, positive predictive values, and reproducibility.

Main Results:

• Satisfactory sequencing coverage achieved (mean 250×, 95.2% ≥10×).
• Maximum variant call sensitivity reached 96.7% for single-nucleotide variants and 28.5% for indels.
• High positive predictive values (>0.959) and good inter/intra-assay reproducibility (89.5%/87.3%) were observed.

Conclusions:

• The validated NGS pipeline demonstrates strong analytical performance.
• The assay is feasible for routine diagnostic use in inherited cardiomyopathies.
• The pipeline offers minimal time requirements and a simple bioinformatics structure for clinical implementation.