ClinVar Genomic variation as it relates to human health

The p.Ile41Thr variant in FIG4 has been reported in at least 23 individuals with Charcot Marie Tooth type 4J and segregated with disease in at least 14 affected individuals (first reported by Chow 2007 PMID: 17572665; >15 publications including de Leeuw 2008 PMID: 18261132, Ikonomov 2010 PMID: 20630877, Lenk 2011 PMID: 21655088, Cottenie 2013 PMID: 23489662, Lassuthova 2016 PMID: 27549087, Gentil 2017 PMID: 28859335, Morgan 2017 PMID: 28430856, Bacquet 2018 PMID: 30373780, Orengo 2018 PMID: 29468183). It has also been identified in 0.189% (244/128994) of European chromosomes by gnomAD (http://gnomad.broadinstitute.org). However, this frequency is low enough to be consistent with a recessive carrier frequency. This variant has also been reported in ClinVar as pathogenic by multiple submitters (Variation ID 1721). Computational prediction tools and conservation analyses suggest that this variant may impact the protein, though this information is not predictive enough to determine pathogenicity. In vitro and in vivo functional studies both provide some evidence that this variant impacts protein function (first by de Leeuw 2008 PMID: 18261132) and animal models in mice have shown that this variant causes peripheral neuropathy (Gentil 2017: 28859335). This variant was observed in trans with >10 different nonsense variants in FIG4, which are classified as pathogenic by other laboratories. In summary, this variant meets criteria to be classified as pathogenic for autosomal recessive CMT4J. ACMG/AMP Criteria applied: PM3_VeryStrong, PS3_Moderate, PP3, PP1_Strong.

The variant is observed at an extremely low frequency in the gnomAD v2.1.1 dataset (total allele frequency: 0.101%). While this variant results in missense change, protein truncation variants are a common disease-causing mechanism. In silico tool predictions suggest damaging effect of the variant on gene or gene product (REVEL: 0.84; 3Cnet: 0.60). Same nucleotide change resulting in same amino acid change has been previously reported as pathogenic/likely pathogenic with strong evidence (ClinVar ID: VCV000001721). Therefore, this variant is classified as Likely pathogenic according to the recommendation of ACMG/AMP guideline.

This sequence variant is a single nucleotide substitution (T>C) which results in an isoleucine to threonine amino acid change at residue 41 of the FIG4 protein. This is a previously reported variant (ClinVar) which has been observed in homozygous state (PMID: 33405357, 31743256) or compound heterozygous state with a null allele in individuals with Charcot-Marie-Tooth Type 4J (PMID: 17572665, 18556664 , 21705420, 23489662, 24878229); though the variant has also been observed on occasion in individuals with amyotrophic lateral sclerosis (PMID: 28430856, 25382069, 28051077), evidence suggests that the variant is not associated with that disease. This variant is rare in the gnomAD control population dataset (284/282242 alleles or 0.1%). Multiple functiol studies indicate that the variant protein has decreased function, stability, and ability to interact with scaffold proteins (PMID: 20630877, 21655088, 17572665). Given the available evidence, we consider this variant to be pathogenic. ACMG Criteria: PM1, PM3, PP3, PS3, PS4

The FIG4 c.122T>C (p.Ile41Thr) missense variant has been reported in four studies in which it is found in a total of 36 individuals with Charcot-Marie-Tooth, type 4, including in 18 in a compound heterozygous state, all in combination with a null variant, and in 18 in a heterozygous state in whom the second variant has not been found (Chow et al. 2007; Nicholson et al. 2011; Cottenie et al. 2013; Menezes et al. 2014). The p.Ile41Thr variant was found in 13 of 6064 controls, and is reported at a frequency of 0.001889 in the European (non-Finnish) population of the Genome Aggregation Database. Functional studies in proband fibroblasts, cultured cells, and yeast demonstrated that the p.Ile41Thr variant leads to low levels of protein due to impaired interaction with the scaffold protein, VAC14, which stabilizes FIG4 and impairs kinase activation (Chow et al. 2007; Zhang et al. 2008; Ikonomov et al. 2010; Lenk et al. 2011). In a mouse model, overexpression (5-fold) of variant p.Ile41Thr in Fig4-null mice rescued the severe neurodegenerative phenotype and lethality, whereas lesser overexpression (2-fold) resulted in partial rescue and a CMT-like phenotype (Lenk et al. 2011). Based on the collective evidence, the p.Ile41Thr variant is classified as pathogenic for Charcot-Marie-Tooth, type 4. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population.

This FIG4 variant (rs121908287) is rare in a large population dataset (284/282242 total alleles; 0.1%; no homozygotes), and has an entry in ClinVar. Three bioinformatic tools queried predict that p.Ile41Thr would be damaging, and the isoleucine residue at this position is strongly conserved across the species assessed. Functional studies in proband fibroblasts, cultured cells, and yeast demonstrate that the p.Ile41Thr substitution leads to low levels of FIG4 protein. This variant is not predicted to affect normal exon 2 splicing, although this has not been confirmed experimentally to our knowledge. This variant in the FIG4 gene has been reported previously in association with CMT4J in individuals with a second FIG4 pathogenic variant on the opposite chromosome. A small number of patients with clinically diagnosed Charcot-Marie-Tooth disease (0.45%) are heterozygous carriers of the c.122T>C variant; some of these patients might carry an undetected non-coding variant that was missed by exon sequencing. We consider c.122T>C to be pathogenic.

This variant was determined to be pathogenic according to ACMG Guidelines, 2015 [PMID:25741868]. This variant has been previously reported as pathogenic [PMID: 28051077, 28859335, 28430856, 18261132, 25617005, 21655088, 24878229, 17572665, 20630877, 23489662, 27549087, ClinVar ID: 1721]

The c.122T>C;p.(Ile41Thr) missense variant has been observed in affected individual(s) and ClinVar contains an entry for this variant (ClinVar ID: 1721; OMIM: 609390.0001; PMID: 20301641; 17572665) - PS4_moderate. Well-established in vitro or in vivo functional studies support a damaging effect on the gene or gene product ( PMID: 17572665; 21655088) - PS3_moderate. Multiple lines of computational evidence support a deleterious effect on the gene or gene product - PP3. and allele frequency is greater than expected for disorder -BS1. In summary, the currently available evidence indicates that the variant is pathogenic.

Variant summary: FIG4 c.122T>C (p.Ile41Thr) results in a non-conservative amino acid change in 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 0.00098 in 250852 control chromosomes. This frequency does not allow any conclusion about variant significance. c.122T>C has been reported in the literature as a compound heterozygous genotpye in multiple individuals affected with Charcot-Marie Disease Type 4J (example, Chow_2007, Gentil_2017, Michaelidou_2020). These data indicate that the variant is very likely to be associated with disease. At least one publication reports experimental evidence evaluating an impact on protein function (Chow_2007). The most pronounced variant effect results in impaired activation of the Fab1/PIKfyve kinase resulting from decreased levels of phosphatidylinositol-3,5-bisphosphate (PtdIns(3,5)P2). Multiple clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation and a majority consensus as Pathogenic (n=15). Based on the evidence outlined above, the variant was classified as pathogenic.

Published functional studies demonstrate protein instability resulting in low levels of FIG4 protein (Lenk et al., 2011); Observed in the heterozygous state in multiple individuals with ALS or PLS (Cady et al., 2015; Osmanovic et al., 2017; Morgan et al., 2017); 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: 30373780, 24878229, 23489662, 18261132, 20630877, 25617005, 22131434, 19118816, 17572665, 21705420, 21655088, 27549087, 28051077, 25382069, 28859335, 29468183, 32022442, 31980526, 32268254, 28430856, 31589614, 33096303, 32376792)

This variant is one of the most common variants associated with autosomal recessive Charcot-Marie-Tooth disease, type 4J (PMID 21705420), therefore the frequency of this variant in the general population is consistent with pathogenicity (http://gnomad.broadinstitute.org). In multiple individuals with CMT, this variant has been seen with a single recessive pathogenic variant in the same gene, suggesting this variant may also be pathogenic. This variant segregates with CMT in at least one family. Assessment of experimental evidence suggests this variant results in abnormal protein function. Expression of this variant in mice resulted in reduced protein stability (PMID 21655088).

This sequence change replaces isoleucine, which is neutral and non-polar, with threonine, which is neutral and polar, at codon 41 of the FIG4 protein (p.Ile41Thr). This variant is present in population databases (rs121908287, gnomAD 0.2%), and has an allele count higher than expected for a pathogenic variant. This missense change has been observed in individual(s) with Charcot-Marie-Tooth disease type 4 (PMID: 17572665, 18556664, 21705420, 23489662, 24878229; Invitae). In at least one individual the data is consistent with being in trans (on the opposite chromosome) from a pathogenic variant. ClinVar contains an entry for this variant (Variation ID: 1721). 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 FIG4 function (PMID: 17572665, 20630877, 21655088). For these reasons, this variant has been classified as Pathogenic.

The FIG4 c.122T>C; Ile41Thr variant (rs121908287) is reported in several unrelated individuals with a clinical diagnosis Charcot-Marie-Tooth disease (Chow 2007, Cottenie 2013, Menezes 2014, Nicholson 2011, Zhang 2008), often observed in trans to a truncating FIG4 variant (Chow 2007, Nicholson 2011). The variant is reported in the ClinVar database (Variation ID: 1721) and is reported in the general population with an overall allele frequency of 0.1% (284/282,242 alleles) in the Genome Aggregation Database. The isoleucine at codon 41 is highly conserved and computational analyses predict that this variant is deleterious (REVEL: 0.84). In support of this prediction, the p.Ile41Thr variant has been shown to impart instability to FIG4 protein (Ikonomov 2010, Lenk 2011). A FIG4 mouse model created using a null FIG4 allele and transgenic expression of the human p.Ile41Thr protein corroborates the conclusion that protein instability is likely the basis of the pathogenicity of the p.Ile41Thr variant (Lenk 2011, Winters 2011). Based on available information, this variant is classified as pathogenic. References: Chow CY et al. Mutation of FIG4 causes neurodegeneration in the pale tremor mouse and patients with CMT4J. Nature. 2007; 448(7149): 68-72. Cottenie E et al. Rapidly progressive asymmetrical weakness in Charcot-Marie-Tooth disease type 4J resembles chronic inflammatory demyelinating polyneuropathy. Neuromuscul Disord. 2013; 23(5): 399-403. Ikonomov OC et al. ArPIKfyve regulates Sac3 protein abundance and turnover: disruption of the mechanism by Sac3I41T mutation causing Charcot-Marie-Tooth 4J disorder. J Biol Chem. 2010; 285(35): 26760-26774. Lenk GM et al. Pathogenic mechanism of the FIG4 mutation responsible for Charcot-Marie-Tooth disease CMT4J. PLoS Genet. 2011; 7(6): e1002104. Menezes MP et al. Whole exome sequencing identifies three recessive FIG4 mutations in an apparently dominant pedigree with Charcot-Marie-Tooth disease. Neuromuscul Disord. 2014; 24(8): 666-670. Nicholson G et al. Distinctive genetic and clinical features of CMT4J: a severe neuropathy caused by mutations in the PI(3,5)P2 phosphatase FIG4. Brain. 2011; 134(Pt 7): 1959-1971. Winters JJ et al. Congenital CNS hypomyelination in the Fig4 null mouse is rescued by neuronal expression of the PI(3,5)P(2) phosphatase Fig4. J Neurosci. 2011; 31(48): 17736-17751. Zhang X et al. Mutation of FIG4 causes a rapidly progressive, asymmetric neuronal degeneration. Brain. 2008; 131(Pt 8): 1990-2001.

The p.I41T pathogenic mutation (also known as c.122T>C), located in coding exon 2 of the FIG4 gene, results from a T to C substitution at nucleotide position 122. The isoleucine at codon 41 is replaced by threonine, an amino acid with similar properties. This mutation has been reported in multiple individuals with Charcot-Marie-Tooth disease (Nicholson G et al. Brain, 2011 Jul;134:1959-71; Lenk GM et al. PLoS Genet., 2011 Jun;7:e1002104). Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation.

PS3, PM3_Very Strong

FIG4: PM3:Very Strong, PS3:Moderate, PM2:Supporting, PP3

The FIG4 c.122T>C variant is predicted to result in the amino acid substitution p.Ile41Thr. This variant has been reported in individuals with autosomal recessive Charcot-Marie-Tooth (CMT) disease type 4J (Chow et al. 2007. PubMed ID: 17572665; Cottenie et al. 2013. PubMed ID: 23489662; Menezes et al. 2014. PubMed ID: 24878229). This variant has also been reported in patients with ALS, though one study found that the frequency of the variant was actually higher in controls (ALS: 3 in 1126, 0.27%; controls: 5 in 613, 0.82%) (Morgan et al. 2017. PubMed ID: 28430856). Therefore, this variant's role in causing an ALS phenotype remains unclear. This variant has been found in 0.19% of Non-Finnish Europeans. This variant has been classified as pathogenic by multiple independent submitters to the ClinVar database (https://www.ncbi.nlm.nih.gov/clinvar/variation/1721/). Given the evidence, we interpret this variant as pathogenic for CMT related disease.

Converted during submission to Benign.

This variant was classified as: Uncertain significance. The available evidence favors the pathogenic nature of this variant, however the currently available data is insufficient to conclusively support its pathogenic nature. Thus this variant is classified as Uncertain significance - favor pathogenic. The following ACMG criteria were applied in classifying this variant: PS1,PS3,PP3,BP1,BS1.

Variant interpreted as Pathogenic and reported on 07-09-2018 by Invitae. GenomeConnect-Invitae Patient Insights Network assertions are reported exactly as they appear on the patient-provided report from the testing laboratory. Registry team members make no attempt to reinterpret the clinical significance of the variant. Phenotypic details are available under supporting information.