ClinVar Genomic variation as it relates to human health

Conflicting evidence has been reported regarding the effect of this variant on protein function and structure (PMID: 25456137, 29603717); In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; This variant is associated with the following publications: (PMID: 33289110, 35960392, 37541188, 27539236, 29030706, 31692161, 24262145, 27290639, 28139822, 23001123, 29221912, 29603717, 30609409, 31664448, 31737037, 30729177, 31772029, 31980526, 34758253, 27943079, 34426522, 33307271, 34778129, 34631961, Liang[article]2022, 38292175, 35859177, 25456137, 37421629, 33528536)

The ADAR c.577C>G (p.Pro193Ala) missense variant has been reported in at least three studies in which it is found in a compound heterozygous state in at least 15 probands with ADAR-related disorders, including two sets of siblings and eight affected individuals from five families (Rice et al. 2012; Livingston et al. 2014; Schmelzer et al. 2018). The p.Pro193Ala variant has not been found in any affected individuals with dyschromatosis symmetrica hereditaria. Affected probands exhibited acute or subacute onset of striatal necrosis, early-onset encephalopathy, intracranial calcifications, developmental delay, severe and progressive dystonia, and often exhibited bilateral striatal necrosis (Rice et al. 2012; Livingston et al. 2014). Control data are unavailable for this variant, which is reported at a frequency of 0.003976 in the European population of the 1000 Genomes Project and observed in one homozygote in the Genome Aggregation Database. The p.Pro193Ala variant is reasonably common in the general population but still present at a frequency consistent with autosomal recessive disease. It is also noted that variants in the ADAR gene may have an atypical or milder presentation (Crow et al. 2016). Expression analysis of p.Pro193Ala in HEK293 cells showed a significant reduction in RNA-editing enzyme activity (Mannion et al. 2014). The loss of ADAR in hematopoietic stem cells in mice was associated with upregulation of type-I and type-II interferon-inducible transcripts and rapid apoptosis (Hartner et al. 2009). Based on the evidence, the p.Pro193Ala variant is classified as pathogenic for ADAR-Related disorders. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population.

The p.Pro193Ala (NM_001111.4 c.577C>G) variant in ADAR has been reported in at l east 8 affected individuals (two of whom were identical twins) from 5 families w ith Aicardi-Goutieres syndrome (Rice 2012) and 6 individuals from 5 families wit h bilateral striatal necrosis (Livingston 2014). All affected individuals were c ompound heterozygotes with a second ADAR variant. This variant has also been rep orted in ClinVar (Variation ID# 126395). The p.Pro193Ala variant has also been i dentified in 0.3% (199/66,740) of European chromosomes by the Exome Aggregation Consortium (ExAC, http://exac.broadinstitute.org; dbSNP rs145588689). Although t his variant has been seen in the general population, its frequency is low enough to be consistent with a recessive carrier frequency (of note, the prevalence of the disease is unknown). Computational prediction tools and conservation analys is suggest that the p.Pro193Ala variant may impact the protein, though this info rmation is not predictive enough to determine pathogenicity. In summary, althoug h additional studies are required to fully establish its clinical significance, the p.Pro193Ala variant is likely pathogenic based on its occurrence in individu als with this disease.

DNA sequence analysis of the ADAR gene demonstrated a sequence change, c.577C>G, in exon 2 that results in an amino acid change, p.Pro193Ala. The p.Pro193Ala change affects a moderately conserved amino acid residue located in a domain of the ADAR protein that is known to be functional. In-silico pathogenicity prediction tools (SIFT, PolyPhen2, Align GVGD, REVEL) provide contradictory results for the p.Pro193Ala substitution. The variant has been reported previously in the compound heterozygous state with another ADAR pathogenic variant in several individuals with Aicardi-Goutieres syndrome (Rice et al., 2012). Functional studies in HEK293 cells transfected with the p.Pro193Ala variant exhibit a significant decrease in RNA-editing enzyme activity (Mannion et al., 2014). This sequence change has been described in the gnomAD database with a population frequency of 0.21% (dbSNP rs145588689). We classify this sequence change as likely pathogenic.

Variant summary: ADAR c.577C>G (p.Pro193Ala) results in a non-conservative amino acid change located in the Z-binding domain (IPR042371) of the encoded protein sequence. Five of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 0.0021 in 282848 control chromosomes, predominantly at a frequency of 0.0033 within the Non-Finnish European subpopulation in the gnomAD database, including one homozygote (of note, this homozygous carrier was also part of the non-neuro dataset of gnomAD v2.1). The observed relatively high frequency in non-Finnish Europeans, together with the homozygous occurrence, might indicate a benign nature for the variant. However, the variant, c.577C>G has also been reported in the literature in numerous compound heterozygous individuals, who were affected with phenotypes that belong to the Aicardi-Goutieres Syndrome 6 (AGS6) spectrum, including atypical (late-infantile and juvenile-onset) AGS, non-syndromic bilateral striatal necrosis (BSN), and calcifying cardiac valve disease (e.g. Rice_2012, Livingston_2014, Piekutowska-Abramczuk_2016, Kono_2018, Sathishkumar_2021, Crow_2020, Piccoli_2021). Of note, in several of these reported patients the signs of dyschromatosis symmetrica hereditaria (DSH) were also described (e.g. in Livingston_2014, Piekutowska-Abramczuk_2016, Kono_2018, Sathishkumar_2021, Crow_2020). In many patients, upregulation of IFN-stimulated genes (ISGs) were demonstrated, which is consistent with the proposed disease mechanism (e.g. Rice_2012, Livingston_2014). These data indicate that the variant is very likely to be associated with disease. A recent study noted that over 60% of AGS patients with ADAR mutations carry the P193A allele in compound heterozygous state with either a loss of function (LoF) mutation or a missense in the deaminase domain of ADAR1, however no homozygous patients have been reported (Maurano_2021). Authors of this study generated a knock-in mouse model for the variant, and found that P195A/P195A mice (P195A is homologous to human P193A) were indistinguishable from wild type controls, but compound heterozygous mice carrying the variant together with a more severe (i.e. null) ADAR allele, recapitulated the human AGS phenotype, with marked upregulation of ISGs (Maurano_2021); notably, homozygous null alleles of Adar in mice are known to result in embryonic lethality (OMIM). A publication also reported experimental evidence evaluating an impact on protein function, and demonstrated a partial loss of function for the variant protein (Mannion_2014). These data suggest that the pathogenicity of this variant is genotype-dependent, i.e. highly dependent on the variant observed in trans. The following publications have been ascertained in the context of this evaluation (PMID: 25456137, 28139822, 31772029, 29603717, 24262145, 34343497, 33307271, 23001123, 33289110, 33723056). 18 submitters have cited clinical-significance assessments for this variant to ClinVar after 2014, and reported the variant with conflicting assessments. Based on the evidence outlined above, the variant likely represents a hypomorphic allele that is subject to interallelic interactions, which in compound heterozygous state, together with other (more severe) variants in trans, can cause AGS6 and related phenotypes, therefore, this variant was classified as pathogenic.

The ADAR c.577C>G variant is predicted to result in the amino acid substitution p.Pro193Ala. This variant has been reported in the compound heterozygous state in multiple families with Aicardi-Goutières syndrome and is predicted to disrupt DNA binding (Rice et al. 2012. PubMed ID: 23001123; Livingston et al. 2015. PubMed ID: 24262145; Piekutowska-Abramczuk et al. 2016. PubMed ID: 28139822). It was also reported in the compound heterozygous state in an individual who possibly had a mitochondrial disorder (Pronicka et al. 2016. PubMed ID: 27290639). Based on these observations, we interpret this variant as likely pathogenic.

This sequence change replaces proline, which is neutral and non-polar, with alanine, which is neutral and non-polar, at codon 193 of the ADAR protein (p.Pro193Ala). This variant is present in population databases (rs145588689, gnomAD 0.3%), including at least one homozygous and/or hemizygous individual. This missense change has been observed in individual(s) with autosomal recessive ADAR-related diseases (PMID: 23001123, 26629815). ClinVar contains an entry for this variant (Variation ID: 126395). An algorithm developed to predict the effect of missense changes on protein structure and function (PolyPhen-2) suggests that this variant is likely to be disruptive. Experimental studies have shown that this missense change affects ADAR function (PMID: 9889202, 25456137, 34343497). In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance.

The c.577C>G (p.P193A) alteration is located in coding exon 2 of the ADAR gene. This alteration results from a C to G substitution at nucleotide position 577, causing the proline (P) at amino acid position 193 to be replaced by an alanine (A). Based on the available evidence, the ADAR c.577C>G (p.P193A) alteration is classified as pathogenic for autosomal recessive Aicardi-Goutières syndrome; however, it is unlikely to be causative of autosomal dominant Aicardi-Goutières syndrome or dyschromatosis symmetrica hereditaria. Based on data from gnomAD, the G allele has an overall frequency of 0.21% (606/282848) total alleles studied. The highest observed frequency was 0.33% (426/129158) of European (non-Finnish) alleles. The ADAR c.577C>G (p.P193A) alteration is a recurrent Aicardi-Goutières syndrome (AGS)-associated alteration, which typically is found with a second null mutation (Crow, 2015; Rice, 2017). It has been reported in trans with other pathogenic ADAR variants in multiple unrelated individuals with AGS and ADAR-related disorders (Rice, 2012; Livingston, 2014; Rice, 2017; Schmelzer, 2018). This amino acid position is highly conserved in available vertebrate species. The p.P193 amino acid is located within the Z-DNA/Z-RNA binding domain. Structural modeling of the ADAR protein suggests that, in the wildtype protein, the p.P193 residue makes direct contact with the nucleic acid, and the p.P193A substitution removes important interactions between the ADAR protein and DNA/RNA (Rice, 2012). In one study, functional analysis in HEK293 cells demonstrated that the p.P193A alteration results in significantly decreased RNA-editing activity compared to wildtype (Mannion, 2014) while another study showed no significant effect on editing (Rice, 2012). Additionally, the P193A mutation, which is expressed only in the N-terminally extended ADAR1p150 isoform, was shown to increase the effect of other AGS mutations in the same protein (Mannion, 2014). The in silico prediction for this alteration is inconclusive. Based on the available evidence, this alteration is classified as pathogenic.

A heterozygous missense variant was identified, NM_001111.5(ADAR):c.577C>G in exon 2 of 15 of the ADAR gene (NB: this variant is non-coding in alternative transcripts). This substitution is predicted to create a minor amino acid change from a proline to an alanine at position 193 of the protein; NP_001102.3(ADAR):p.(Pro193Ala). The proline at this position has very high conservation (100 vertebrates, UCSC), and is located within the Z-DNA binding domain (Rice, G. I. et al. (2012)). In silico software predicts this variant to be damaging (Polyphen, SIFT, CADD, Mutation Taster). The variant is present in the gnomAD population database at a global population frequency at 0.21% (604 heterozygotes, 1 homozygote) and within the European sub-population frequency at 0.33%. This variant has been previously reported likely benign, likely pathogenic, pathogenic and VUS (ClinVar), but also as pathogenic in many patients with bilateral striatal necrosis or Aicardi-Goutieres syndrome (ClinVar, Piekutowska-Abramczuk, D. et al. (2016), Kono, M. et al. (2018)). It has also been shown to segregate with disease in several families (Rice, G. I. et al. (2012), Schmelzer, L. et al. (2018), Livingston, J. H. et al. (2014)). In addition, functional studies show that this variant causes reductions in RNA editing efficiency, but only in compound heterozygote with another pathogenic variant (Mannion, N. M. et al. (2014)). A different variant in the same codon resulting in a change to a serine has also been reported as a VUS (LOVD). Based on information available at the time of curation, this variant has been classified as LIKELY PATHOGENIC.

ADAR: PM3:Strong, PM2, PM5, PS3:Moderate

The variant is observed at an extremely low frequency in the gnomAD v2.1.1 dataset (total allele frequency: 0.214%). In silico tool predictions suggest damaging effect of the variant on gene or gene product (REVEL: 0.74; 3Cnet: 0.15). Same nucleotide change resulting in same amino acid change has been previously reported as pathogenic/likely pathogenic with strong evidence (ClinVar ID: VCV000126395). The variant has been reported to be in trans with pathogenic variants as either compound heterozygous or homozygous in at least 4 similarly affected unrelated individuals (PMID: 28561207). Therefore, this variant is classified as Pathogenic according to the recommendation of ACMG/AMP guideline.

This variant was identified as compound heterozygous.

This variant was determined to be of uncertain significance according to ACMG Guidelines, 2015 [PMID:25741868].

Variant reported in multiple GenomeConnect participants by multiple clinical testing laboratories. Variant classified as Likely pathogenic and reported most recently on 09-09-2022 by PerkinElmer Genomics and as Uncertain significance on 02-03-2017 by Baylor Medical Genetics Laboratories. Assertions are reported exactly as they appear on the patient provided laboratory report. GenomeConnect does not attempt to reinterpret the variant. The IDDRC-CTSA National Brain Gene Registry (BGR) is a study funded by the U.S. National Center for Advancing Translational Sciences (NCATS) and includes 13 Intellectual and Developmental Disability Research Center (IDDRC) institutions. The study is led by Principal Investigator Dr. Philip Payne from Washington University. The BGR is a data commons of gene variants paired with subject clinical information. This database helps scientists learn more about genetic changes and their impact on the brain and behavior. Participation in the Brain Gene Registry requires participation in GenomeConnect. More information about the Brain Gene Registry can be found on the study website - https://braingeneregistry.wustl.edu/.