ClinGen's RASopathy Expert Panel consensus methods for variant interpretation

Abstract

Purpose: Standardized and accurate variant assessment is essential for effective medical care. To that end, Clinical Genome (ClinGen) Resource clinical domain working groups (CDWGs) are systematically reviewing disease-associated genes for sufficient evidence to support disease causality and creating disease-specific specifications of American College of Medical Genetics and Genomics-Association for Molecular Pathology (ACMG-AMP) guidelines for consistent and accurate variant classification.

Methods: The ClinGen RASopathy CDWG established an expert panel to curate gene information and generate gene- and disease-specific specifications to ACMG-AMP variant classification framework. These specifications were tested by classifying 37 exemplar pathogenic variants plus an additional 66 variants in ClinVar distributed across nine RASopathy genes.

Results: RASopathy-related specifications were applied to 16 ACMG-AMP criteria, with 5 also having adjustable strength with availability of additional evidence. Another 5 criteria were deemed not applicable. Key adjustments to minor allele frequency thresholds, multiple de novo occurrence events and/or segregation, and strength adjustments impacted 60% of variant classifications. Unpublished case-level data from participating laboratories impacted 45% of classifications supporting the need for data sharing.

Conclusion: RAS-specific ACMG-AMP specifications optimized the utility of available clinical evidence and Ras/MAPK pathway-specific characteristics to consistently classify RASopathy-associated variants. These specifications highlight how grouping genes by shared features promotes rapid multigenic variant assessment without sacrificing specificity and accuracy.

Keywords: ClinGen; Noonan; RASopathy; Ras/MAPK; variant interpretation.

Figure 1. Effect of Adjusted Allele Frequency Criteria on Variants from ClinVar

Variants with either concordant or discrepant classifications were assessed for their frequency in the general population. An additional 32% of variants met the RAS EP adjusted frequency threshold for BA1 versus the standard ACMG-AMP BA1 Threshold. One variant met the RAS EP adjusted frequency for BS1. Data points are colored by the ClinVar classification or discrepancy category of the variant. (Ben: benign, LBen: likely benign, VUS: uncertain significance, LPath: likely pathogenic, Path: pathogenic)

Figure 2. Assessment of usage of unmodified versus strength-modified pathogenic ACMG-AMP criteria in RAS EP classifications of variants

Typically, most variants had additional evidence to achieve higher strength specifications beyond the standard ACMG-AMP definitions. Note that all modified criteria increase in strength with additional evidence except PS4 (*) given it begins at the strong category. No variant met the PS4 threshold of at least five occurrences due to the requirement of extensive phenotypic data.

Figure 3. Comparison of Approved RAS EP Specific ACMG-AMP Variant Classifications to ClinVar Variant Classifications

Prior to determining concordance, ClinVar classes were grouped into three categories: 1.) pathogenic (Path) and likely pathogenic (LPath), 2.) benign (Ben) and likely benign (LBen), and 3.) VUS. Variant classifications were considered discordant if clinical laboratory submissions did not group into the same category. These grouped ClinVar classifications were compared to the classifications determined by using the RAS EP-Specific ACMG-AMP specifications.