Recommendations for application of the functional evidence PS3/BS3 criterion using the ACMG/AMP sequence variant...

Area of Science:

• Genetics
• Molecular Biology
• Clinical Diagnostics

Background:

• Current clinical variant interpretation guidelines lack detailed methods for evaluating functional assay evidence.
• Inconsistent application of functional evidence codes (PS3/BS3) contributes to inter-laboratory variant interpretation discrepancies.
• There is a need for a standardized framework to assess the clinical validity and strength of functional assay data.

Purpose of the Study:

• To develop a structured approach for evaluating functional assays in clinical variant interpretation.
• To refine the criteria for using strong evidence codes (PS3/BS3) based on functional assay validation.
• To provide guidance on determining the appropriate strength of evidence from functional assays.

Main Methods:

• The Clinical Genome Resource (ClinGen) Sequence Variant Interpretation (SVI) Working Group convened experts to refine PS3/BS3 criteria.
• Functional evidence was curated, and the odds of pathogenicity were estimated for assays with varying numbers of variant controls.
• Recommendations were developed through iterative feedback from expert panels and steering committees.

Main Results:

• A four-step framework was established for assessing functional data, including defining disease mechanisms, evaluating assay applicability and validity, and applying evidence to variants.
• A minimum of 11 total pathogenic and benign variant controls are necessary to achieve moderate-level evidence without rigorous statistical analysis.
• Recommendations provide clarity on evaluating functional assay data for clinical variant interpretation.

Conclusions:

• The developed recommendations offer a clearer process for functional evidence evaluation in clinical variant interpretation.
• This work aims to improve consistency and reduce discordance in variant classification.
• The study encourages collaboration between clinical geneticists and basic scientists developing functional assays.