ClinGen Myeloid Malignancy Variant Curation Expert Panel recommendations for germline RUNX1 variants

Abstract

Standardized variant curation is essential for clinical care recommendations for patients with inherited disorders. Clinical Genome Resource (ClinGen) variant curation expert panels are developing disease-associated gene specifications using the 2015 American College of Medical Genetics and Genomics (ACMG) and Association for Molecular Pathology (AMP) guidelines to reduce curation discrepancies. The ClinGen Myeloid Malignancy Variant Curation Expert Panel (MM-VCEP) was created collaboratively between the American Society of Hematology and ClinGen to perform gene- and disease-specific modifications for inherited myeloid malignancies. The MM-VCEP began optimizing ACMG/AMP rules for RUNX1 because many germline variants have been described in patients with familial platelet disorder with a predisposition to acute myeloid leukemia, characterized by thrombocytopenia, platelet functional/ultrastructural defects, and a predisposition to hematologic malignancies. The 28 ACMG/AMP codes were tailored for RUNX1 variants by modifying gene/disease specifications, incorporating strength adjustments of existing rules, or both. Key specifications included calculation of minor allele frequency thresholds, formulating a semi-quantitative approach to counting multiple independent variant occurrences, identifying functional domains and mutational hotspots, establishing functional assay thresholds, and characterizing phenotype-specific guidelines. Preliminary rules were tested by using a pilot set of 52 variants; among these, 50 were previously classified as benign/likely benign, pathogenic/likely pathogenic, variant of unknown significance (VUS), or conflicting interpretations (CONF) in ClinVar. The application of RUNX1-specific criteria resulted in a reduction in CONF and VUS variants by 33%, emphasizing the benefit of gene-specific criteria and sharing internal laboratory data.

Graphical abstract

Figure 1.

Schematic of RUNX1 exonic distribution, protein isoforms, and functional domain structure with all 52 pilot variants and their final MM-VCEP classification. (A) Isoform C with RHD, transactivation domain (TAD), and the VWRPY motif and location of all 49 single-nucleotide pilot variants with their final MM-VCEP classification. PATH and LPATH variants are shown at the top, and VUS, LBEN, and BEN variants are shown at the bottom. The exonic distribution of isoform C is displayed below. (B) Schematic of RUNX1 isoforms A, B, and C and their functional domains. Regions in gray are unique to 1 isoform. The 3 pilot CNVs are shown at the bottom, with the deletion of exons 2 and 3 exclusively affecting the N-terminal 33 AA of isoform C.

Figure 2.

PVS1 decision tree for SNVs/indels. Application of different levels of strength for PVS1 depending on the prediction of nonsense-mediated decay (NMD), the location within a known critical protein domain, and the expression of alternative isoforms. The splicing effects table is given in supplemental Data.

Figure 3.

Comparison of ClinVar and MM-VCEP classifications. Fifty previously asserted and ClinVar-deposited RUNX1 variants are shown on the x-axis. Final MM-VCEP classifications are color-coded (see legend on the right). ClinVar variants with previous LPATH, CONF, and VUS assertions were most often reclassified by using MM-VCEP–specified rules for RUNX1.