High-Throughput Reclassification of SCN5A Variants.

Area of Science:

• Cardiovascular Genetics
• Molecular Cardiology
• Genetic Diagnostics

Background:

• Loss-of-function variants in SCN5A are a primary genetic cause of Brugada syndrome (BrS).
• A significant majority (80%) of identified SCN5A missense variants are classified as variants of uncertain significance (VUSs), hindering clinical application.
• Resolving the pathogenicity of SCN5A VUSs is crucial for accurate genetic diagnosis and patient management in BrS.

Purpose of the Study:

• To functionally reclassify a set of 83 SCN5A variants using high-throughput automated patch clamping.
• To determine the pathogenicity of suspected benign and Brugada syndrome-associated SCN5A variants.
• To investigate the structural mechanisms underlying SCN5A variant loss-of-function.

Main Methods:

• Selected 83 SCN5A variants, including controls, suspected benign, and suspected BrS variants.
• Employed high-throughput automated patch clamping to assess ion channel function for each variant.
• Utilized structural modeling to elucidate mechanisms of functional impairment.

Main Results:

• Functional studies confirmed the pathogenicity of known control and suspected benign variants.
• Identified 22 variants with loss-of-function and 22 with partial loss-of-function.
• Reclassified 61 initial VUSs, with 45 now classified as pathogenic/likely pathogenic and 16 as benign/likely benign, leaving only 12 VUSs.

Conclusions:

• High-throughput patch clamping is effective for functional reclassification of SCN5A variants.
• This method significantly reduces the number of VUSs, improving diagnostic yield for Brugada syndrome.
• Structural modeling provides insights into the molecular basis of SCN5A-related channel dysfunction.