ClinVar Genomic variation as it relates to human health

The p.Arg356Gln variant in GLA has been reported in multiple individuals with Fabry disease (PMID: 19621417, 27238910, 23826564, 28615118), and has been identified in 0.0076% (1/13161) African chromosomes, including one hemizygote, and 0.0012% (1/81755) European (non-Finnish) chromosomes by the Genome Aggregation Database (gnomAD, http://gnomad.broadinstitute.org; dbSNP rs869312163). Although this variant has been seen in the general population, its frequency is low enough to be consistent with Fabry disease. Please note that for diseases with clinical variability, or reduced penetrance, pathogenic variants may be present at a low frequency in the general population. This variant has also been reported in ClinVar as likely pathogenic by Invitae and as a VUS by EGL Genetic Diagnostics (Variation ID:222135). In vitro functional studies provide some evidence that the p.Arg356Gln variant may slightly impact protein function through decreased GLA enzyme activity (PMID: 28615118, 27238910, 19621417). However, these types of assays may not accurately represent biological function. Computational prediction tools and conservation analyses do not provide strong support for or against an impact to the protein. One additional likely pathogenic variant, causing a different amino acid change at the same position, (p.Arg356Trp), has been reported in association with disease in the literature and ClinVar, slightly supporting that a change at this position may not be tolerated (PMID: 23935525, 22773828, 17555407, 17532296, 20031620, 23537685, 2539398, 21598360, 22551898, 26415523;Variation ID:217411,10713). In summary, while there is some suspicion for a pathogenic role, the clinical significance of this variant is uncertain. ACMG/AMP Criteria applied: PM2_supporting, PM5_supporting, PS3_supporting (Richards 2015).

This missense variant replaces arginine with glutamine at codon 356 of the GLA protein. Computational prediction is inconclusive regarding the impact of this variant on protein structure and function (internally defined REVEL score threshold 0.5 < inconclusive < 0.7, PMID: 27666373). This variant has also been reported in over 20 individuals in several Fabry disease screening studies (PMID: 19621417, 20864368, 27238910, 28615118, 29330335, Coutinho et al., 2017, Burlina et al., 2019). Some studies have shown that this variant causes a partial reduction of alpha-galactosidase activity in carriers (PMID: 19621417, 27238910, 28615118, Burlina et al., 2019), while other studies have shown no impact on the enzyme activity (PMID: 23935525, 28615118, 29330335). This variant has been reported in a male individual affected with Fabry disease who also carried another variant of unknown significance in the GLA gene (PMID: 31634893). The proband's sister and daughter both carried these two variants. The sister was affected with left ventricular hypertrophy and microalbuminuria, while the daughter was asymptomatic. This variant has been reported in eight individuals including four males in a family, none of whom had clinical manifestations related to Fabry disease (PMID: 28615118). This variant has been reported in multiple healthy adults in the UK Biobank (PMID: 35977816). This variant has been identified in 2/183292 chromosomes in the general population by the Genome Aggregation Database (gnomAD). A different missense variant at the same codon, p.Arg356Trp, has been associated with Fabry disease and hypertrophic cardiomyopathy (Clinvar variation ID: 10713), suggesting that arginine at this position is important for GLA function. n summary, this variant has been reported in individuals affected with Fabry disease and/or showing reduced GLA enzyme activity and it has also been observed in multiple unaffected individuals. Additional studies are necessary to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance.

This sequence change replaces arginine, which is basic and polar, with glutamine, which is neutral and polar, at codon 356 of the GLA protein (p.Arg356Gln). This variant is present in population databases (no rsID available, gnomAD 0.008%). This missense change has been observed in individual(s) with GLA-related conditions (PMID: 19621417, 27238910, 28615118, 30477121, 31956509, 31996269). ClinVar contains an entry for this variant (Variation ID: 222135). 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 not expected to disrupt GLA protein function with a negative predictive value of 80%. Experimental studies are conflicting or provide insufficient evidence to determine the effect of this variant on GLA function (PMID: 19621417, 23935525, 28615118). This variant disrupts the p.Arg356 amino acid residue in GLA. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 2539398, 17532296, 17555407, 21598360, 24582695, 25611685). 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.

Reported in numerous newborn screening cohorts, in which reduced enzyme activity levels were detected but clinical features varied, including multiple hemizygous adult individuals without clinical manifestations (Hwu et al., 2009, Elliott et al., 2016, Liao et al., 2018; Gilchrist M et al., 2023); Previously reported in males with suspected Fabry disease; detailed clinical and segregation information was not provided (Duro G et al., 2018; Varela P et al., 2020); Published functional studies found this variant retains significant residual enzyme activity (Lukas et al., 2013, Liao et al., 2018; Benjamin ER et al., 2017); In silico analysis supports that this missense variant does not alter protein structure/function; This variant is associated with the following publications: (PMID: 25382311, 27238910, 28615118, 19621417, 33597575, 31956509, 30477121, 31634893, 36695159, 27657681, 23935525, 31996269, 35977816, 32531501)

Variant summary: GLA c.1067G>A (p.Arg356Gln) results in a conservative amino acid change located in the Alpha galactosidase A, C-terminal beta-sandwich domain (IPR035373) of the encoded protein sequence. Four of five in-silico tools predict a benign effect of the variant on protein function. The variant allele was found at a frequency of 1.1e-05 in 183292 control chromosomes in gnomAD (including 5 hemizygotes in V4). The available data on variant occurrences in the general population are insufficient to allow any conclusion about variant significance. c.1067G>A has been reported in the literature in individuals from newborn screening cohorts with reduced enzyme activity (examples: Hwu_2009, Duro_2018, Sawada_2020). However, multiple studies have reported experimental evidence that this variant effect results in >50%-90% of normal activity in vitro (examples: Lukas_2012 and Liao_2018). One study reported that this variant was present in eight individuals from one family but none had clinical abnormalities (Liao_2018). Additionally, a second likely pathogenic variant (GLA p.G360R) was reported in a male patient affected with left ventricular hypertrophy and stroke (Carvalho_ 2019). These report(s) do not provide unequivocal conclusions about association of the variant with Fabry Disease. The following publications have been ascertained in the context of this evaluation (PMID: 19621417, 23935525, 30477121, 28615118, 31634893, 31956509). Eight submitters have cited clinical-significance assessments for this variant to ClinVar after 2014 and classified this variant as uncertain significance (n=7) and likely pathogenic (n=1). Based on the evidence outlined above, the variant was classified as uncertain significance.

The p.R356Q variant (also known as c.1067G>A), located in coding exon 7 of the GLA gene, results from a G to A substitution at nucleotide position 1067. The arginine at codon 356 is replaced by glutamine, an amino acid with highly similar properties. This variant has been reported in numerous lysosomal storage disease newborn screening cohorts, in which reduced &alpha;-Gal A enzyme activity levels were detected (Hwu WL et al. Hum Mutat, 2009 Oct;30:1397-405; Elliott S et al. Mol Genet Metab, 2016 08;118:304-9; Duro G et al. Int J Mol Sci, 2018 Nov;19; Liao HC et al. Mol Genet Metab, 2018 02;123:140-147; Wasserstein MP et al. Genet Med, 2019 03;21:631-640). However, functional studies demonstrate significant residual enzyme activity levels that tentatively support this variant might not cause disease or might cause only mild effects (Lukas J et al. PLoS Genet, 2013 Aug;9:e1003632; Liao HC et al. Mol Genet Metab, 2018 02;123:140-147). This amino acid position is poorly conserved in available vertebrate species, and glutamate is the reference amino acid in several vertebrate species. In addition, the in silico prediction for this alteration is inconclusive. Since supporting evidence is limited at this time, the clinical significance of this alteration remains unclear.

The GLA c.1067G>A variant is predicted to result in the amino acid substitution p.Arg356Gln. This variant was reported in individuals with positive newborn screening results for Fabry disease (Hwu et al. 2009. PubMed ID: 19621417; Sawada et al. 2020. PubMed ID: 31956509; Liao et al. 2018. PubMed ID: 28615118; Gragnaniello et al. 2021. PubMed ID: 34199132; Wasserstein MP et al 2018. PubMed ID: 30093709); male newborns hemizygous for the p.Arg356Gln variant retained 12.5 and 19.1% residual alpha-galactosidase activity in leukocytes however had no elevated lyso-Gb3 in urine ((Hwu et al. 2009. PubMed ID: 19621417). In another study newborns and their family members (7 females and 8 males from at least 3 generations) did not show clinical symptoms or elevated lyso-Gb3 (Figure 4b, Table 4, Liao et al. 2018. PubMed ID: 28615118). This variant was also described in a patient with classic Fabry phenotype (Duro et al. 2018. PubMed ID: 30477121). However, this variant was also detected in an individual with Fabry disease who harbored a second possible causative variant in GLA (Carvalho Silva et al. 2019. PubMed ID: 31634893). In vitro enzyme analysis indicates that the p.Arg356Gln change results in a protein with residual enzyme activity with conflicting results ranging from 15-89% in reported studies (Hwu et al. 2009. PubMed ID: 19621417; Liao et al. 2018. PubMed ID: 28615118; Table S1, Lukas et al. 2013. PubMed ID: 23935525; Table S1, Benjamin et al. 2017. PubMed ID: 27657681). This variant is reported in 0.0076% of alleles in individuals of African descent in gnomAD, including 1 hemizygote. Different amino acid changes at this position (p.Arg356Gly, p.Arg356Trp, and p.Arg356Pro) have been reported in individuals with suspected Fabry disease (Pan et al. 2016. PubMed ID: 27560961; Berstein et al. 1989. PubMed ID: 2539398; Lukas et al. 2016. PubMed ID: 26415523). In the ClinVar database, the p.Arg365Gln variant is reported with conflicting interpretations ranging from 'uncertain' to 'likely pathogenic' by outside laboratories (https://www.ncbi.nlm.nih.gov/clinvar/variation/222135/). At this time, the clinical significance of the c.1067G (p.Arg356Gln) variant is uncertain due to the absence of conclusive functional and genetic evidence.