ClinVar Genomic variation as it relates to human health

The c.1268G>A (p.R423H) alteration is located in exon 2 (coding exon 2) of the KCNC3 gene. This alteration results from a G to A substitution at nucleotide position 1268, causing the arginine (R) at amino acid position 423 to be replaced by a histidine (H). This variant was not reported in population-based cohorts in the Genome Aggregation Database (gnomAD). This alteration has been reported in multiple unrelated patients with early-onset slowly-progressive ataxia (Figueroa, 2010; Duarri, 2015; Khare, 2017). This amino acid position is highly conserved in available vertebrate species. The p.R423H alteration has been shown in multiple independent studies to negatively affect Kv3.3 channel functioning by suppressing current amplitude (Figueroa, 2010), significantly altering gating (Minassian, 2012), or resulting in altered glycosylation and aberrant retention of the mutant channels in the anterograde and/or endosomal vesicles instead of at the plasma membrane (Khare, 2017). This alteration is predicted to be deleterious by in silico analysis. Based on the available evidence, this alteration is classified as pathogenic.

The variant is not observed in the gnomAD v2.1.1 dataset. Missense changes are a common disease-causing mechanism. Functional studies provide strong evidence of the variant having a damaging effect on the gene or gene product (PMID:19953606, 22289912, 25756792, 28467418). In silico tool predictions suggest damaging effect of the variant on gene or gene product (REVEL: 0.92; 3Cnet: 0.72). Same nucleotide change resulting in same amino acid change has been previously reported as pathogenic/likely pathogenic with strong evidence (ClinVar ID: VCV000208671). The variant has been previously reported as de novo (PMID: 28467418). and observed in multiple (>3) similarly affected unrelated individuals (PMID:19953606, 28467418). Therefore, this variant is classified as pathogenic according to the recommendation of ACMG/AMP guideline.

The missense c.1268G>A(p.Arg423His) variant in KCNC3 gene has been reported in heterozygous state in multiple individuals affected with KCNC3 related disorders (Khare S, et. al., 2017; Coutelier M, et. al., 2017; Duarri A, et. al., 2015). Functional studies show p.Arg423His disrupts the channel activity of the KCNC3 protein and demonstrates a dominant-negative effect (Figueroa KP, et. al., 2010). The p.Arg423His variant is novel (not in any individuals) in gnomAD Exomes and 1000 Genomes. This variant has been reported to the ClinVar database as Pathogenic (multiple submissions). The amino acid change p.Arg423His in KCNC3 is predicted as conserved by GERP++ and PhyloP across 100 vertebrates. The amino acid Arg at position 423 is changed to a His changing protein sequence and it might alter its composition and physico-chemical properties. For these reasons, this variant has been classified as Pathogenic.

This sequence change is predicted to replace arginine with histidine at codon 423 of the KCNC3 protein (p.(Arg423His)). The arginine residue is invariant across species (100 vertebrates, UCSC), and is located in the S4 transmembrane segment in the ion transport domain. There is a small physicochemical difference between arginine and histidine. The variant is absent in a large population cohort (rs797044872, gnomAD v2.1 and v3.0). This variant has been identified in multiple individuals with early-onset non-progressive spinocerebellar ataxia, with at least two individuals assumed de novo (PMID: 19953606 , 25756792, 28216058, 28467418). The variant segregates with affected status in multiple families (PMID: 19953606, 28216058, 28467418). A zebrafish model of the variant recapitulates the cerebellar degeneration, and demonstrated differential affects on Purkinje cell excitability, maturation, and viability to an adult-onset associated variant (PMID: 32644043). Multiple lines of computational evidence predict a deleterious effect for the missense substitution (5/5 algorithms). Based on the classification scheme RMH Modified ACMG Guidelines v1.3.0, this variant is classified as PATHOGENIC. Following criteria are met: PS3, PM6_Strong, PP1_Strong, PS4_Moderate, PM2_Supporting, PP3.

Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as Pathogenic. Following criteria are met: 0103 - Dominant negative and gain of function are known mechanisms of disease in this gene and are associated with Spinocerebellar ataxia 13 (MIM#605259). (PMIDs: 22289912, 26442672, GeneReviews). (I) 0107 - This gene is associated with autosomal dominant disease. (I) 0115 - Variants in this gene are known to have variable expressivity. Age of onset (congenital or adult-onset), phenotype and disease progression depend on the location of the variant and its action on the channel, Kv3.3. (PMID: 25756792; 26442672, GeneReviews). (I) 0200 - Variant is predicted to result in a missense amino acid change from arginine to histidine. (I) 0251 - This variant is heterozygous. (I) 0301 - Variant is absent from gnomAD (both v2 and v3). (SP) 0501 - Missense variant consistently predicted to be damaging by multiple in silico tools. Very high conservation with a minor amino acid change. (SP) 0600 - Variant is located in the annotated transmembrane S4 domain (UniProt, PMID: 26442672). (I) 0705 - No comparable missense variants have previous evidence for pathogenicity. (I) 0801 - This variant has strong previous evidence of pathogenicity in unrelated individuals. This variant has been reported as pathogenic in multiple individuals with early onset non-progressive spinocerebellar ataxia. It has been shown to segregate with disease in several families, and there have been at least two reports of de novo cases (ClinVar, LOVD3, PMID: 19953606, 21479265, 25756792, 28467418). (SP) 0902 - This variant has moderate evidence for segregation with disease (PMID: 28467418). (SP) 1002 - This variant has moderate functional evidence supporting abnormal protein function. Targeted expression in zebrafish and drosophila models have demonstrated disrupted axonal pathfinding and neurodegeneration consistent with the neurodevelopmental presentations of disease in patients (PMIDs: 21543613, 30862666, 28467418). In addition, patch clamp studies of this variant expressed in Xenopus oocytes have shown significant loss of Kv3.3 channel activity, reduced current amplitude and cell surface expression. (PMIDs: 19953606, 22289912, 25756792). NB. However, patch clamp assays have been shown to be unreliable; therefore results from these studies are used with caution during variant classification. (SP) 1208 - Inheritance information for this variant is not currently available in this individual. (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign

DNA sequence analysis of the KCNC3 gene demonstrated a sequence change, c.1268G>A, in exon 2 that results in an amino acid change, p.Arg423His. This sequence change is absent in the gnomAD population database. This sequence change has been previously described in individuals and families with ataxia or spinocerebellar ataxia 13 (PMIDs: 19953606, 25756792, 28467418, 28216058). The p.Arg423His change affects a highly conserved amino acid residue located in the S4 transmembrane segment of the KCNC3 protein. It appears to be deleterious using several in-silico pathogenicity prediction tools (SIFT, PolyPhen2, Align GVGD, REVEL). Functional studies show p.Arg423His disrupts the channel activity of the KCNC3 protein and demonstrates a dominant-negative effect (PMIDs: 19953606, 25756792, 28467418). These collective evidences suggest this p.Arg423His change is pathogenic.

This sequence change replaces arginine, which is basic and polar, with histidine, which is basic and polar, at codon 423 of the KCNC3 protein (p.Arg423His). For these reasons, this variant has been classified as Pathogenic. Experimental studies have shown that this missense change affects KCNC3 function (PMID: 19953606, 22289912, 25756792, 28467418). 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 KCNC3 protein function. ClinVar contains an entry for this variant (Variation ID: 208671). This missense change has been observed in individual(s) with spinocerebellar ataxia (PMID: 19953606, 25756792, 28467418). In at least one individual the variant was observed to be de novo. It has also been observed to segregate with disease in related individuals. This variant is not present in population databases (gnomAD no frequency).

Published functional studies demonstrate a damaging effect, resulting in loss of channel activity consistent with a strong dominant negative effect (Figueroa et al., 2010; Duarri et al., 2015); Not observed at significant frequency in large population cohorts (gnomAD); This variant is associated with the following publications: (PMID: 25356970, 23734863, 22289912, 19953606, 26795593, 21479265, 28216058, 28467418, 25756792, 33624863)