ClinVar Genomic variation as it relates to human health

The NM_000152.5:c.671G>A variant in GAA is a missense variant predicted to cause substitution of arginine by glutamine at amino acid 224 (p.Arg224Gln). This variant has been reported in one individual with documented GAA deficiency (PMID: 23000108), meeting criteria for PP4_moderate. This individual is compound heterozygous for the variant and a pathogenic variant based on the ClinGen LSD VCEP's specifications. It has also been reported in two individuals described as having late-onset Pompe disease diagnosed after positive newborn screening in Taiwan, though no phenotype is reported. One individual is compound heterozygous for the variant and a pathogenic variant based on the ClinGen LSD VCEP's specifications (PM3) and the other is compound heterozygous for the variant and a pathogenic or likely pathogenic variant that has not been assessed according to the ClinGen LSD VCEP's specifications. The highest population minor allele frequency in gnomAD is 0.00004 (3/74944 alleles) in the African/African-American population, which is lower than the ClinGen LSD VCEP threshold (<0.001) for PM2, meeting this criterion. When expressed in COS cells, this variant was classified as Class B ("potentially less severe") by Kroos et al, 2012 (PMID:22644586). This includes 0.7% GAA activity in cells and 0.1% in medium, and evidence of abnormal synthesis and processing on Western blot. This meets the ClinGen LSD VCEP specifications for PS3. The computational predictor REVEL gives a score of 0.757 which is above the threshold of 0.7, evidence that correlates with impact to GAA function (PP3). Another missense variant [c.670C>T, p.Arg224Trp] [ClinVar Variation ID:186994] in the same codon has been classified as pathogenic for Pompe disease by the ClinGen Lysosomal Diseases VCEP (PM5). There is a ClinVar entry for this variant (Variation ID: 646875, 2-star review status) with 5 submitters classifying the variant as likely pathogenic (3) or pathogenic (2). In summary, this variant meets the criteria to be classified as likely pathogenic for Pompe disease based on the ACMG/AMP criteria applied, as specified by the ClinGen Lysosomal Diseases Variant Curation Expert panel (specifications Version 2.0): PP4_moderate, PM3, PM2_supporting, PS3_moderate, PP3, PM5 (Classification approved by the ClinGen Lysosomal Diseases Variant Curation Expert Panel on December 20, 2023).

The p.Arg224Gln variant in GAA has been reported in 4 individuals with glycogen storage disease II (PMID: 23147228, 23000108, 21488293, 22644586), and has been identified in 0.008% (2/24814) of African chromosomes, 0.005% (1/19926) of East Asian chromosomes, and 0.003% (1/35358) of Latino chromosomes by the Genome Aggregation Database (gnomAD, http://gnomad.broadinstitute.org; dbSNP rs200210219). Although this variant has been seen in the general population, its frequency is low enough to be consistent with a recessive carrier frequency. In vitro functional studies with COS cells transfected with the variant provide some evidence that the p.Arg224Gln variant may impact protein function (PMID: 22644586). However, these types of assays may not accurately represent biological function. Computational prediction tools and conservation analyses suggest that this variant may impact the protein, though this information is not predictive enough to determine pathogenicity. The phenotype of an individual heterozygous for this variant is highly specific for glycogen storage disease II based on GAA enzyme activity being <1% of normal in fibroblasts and lymphocytes, consistent with disease (PMID: 23000108). One additional likely pathogenic variant, resulting in a different amino acid change at the same position, p.Arg224Pro, has been reported in association with disease in the literature, slightly supporting that a change at this position may not be tolerated (PMID: 22644586, 20080426). Additionally, the presence of this variant in combination with reported pathogenic variant c.525delT (VariationID: 4033) and in two individuals with glycogen storage disease II increases the likelihood that the p.Arg224Gln variant is pathogenic. In summary, this variant meets criteria to be classified as pathogenic for glycogen storage disease in an autosomal recessive manner based on the predicted impact on the protein based on in vitro functional studies and computational evidence, the low frequency of the variant in the general population, and the detection of the variant in affected individuals also harboring another pathogenic variant. ACMG/AMP Criteria applied: PS3, PM2, PM3, PP3, PM5_Supporting, PP4 (Richards 2015).

This sequence change replaces arginine, which is basic and polar, with glutamine, which is neutral and polar, at codon 224 of the GAA protein (p.Arg224Gln). This variant is present in population databases (rs200210219, gnomAD 0.008%). This missense change has been observed in individual(s) with Pompe disease (PMID: 23000108). ClinVar contains an entry for this variant (Variation ID: 664582). Advanced modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) performed at Invitae indicates that this missense variant is expected to disrupt GAA protein function. Experimental studies have shown that this missense change affects GAA function (PMID: 22644586, 23000108). This variant disrupts the p.Arg224 amino acid residue in GAA. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 8429042, 12923862, 14643388, 25026126). This suggests that this residue is clinically significant, and that variants that disrupt this residue are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic.

Variant summary: GAA c.671G>A (p.Arg224Gln) results in a conservative amino acid change located in the Galactose mutarotase, N-terminal barrel domain (IPR031727) of the encoded protein sequence. Four of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 2.5e-05 in 243266 control chromosomes (gnomAD). c.671G>A has been reported in the literature in individuals affected with Glycogen Storage Disease, Type 2 (Pompe Disease) (van der Beek_2011, Kroos_2012, Liao_2017). At least one functional study reports experimental evidence evaluating an impact on protein function and variant effect results in <10% of normal activity (Kroos_2012). A different variant affecting this residue has been classified pathogenic internally. ClinVar contains an entry for this variant (Variation ID: 664582). Based on the evidence outlined above, the variant was classified as pathogenic.