2-47803419-G-C

Position:

• chr2-47803419-G-C

Variant summary

Our verdict is Pathogenic. Variant got 22 ACMG points: 22P and 0B. PVS1 PM2 PP3_Strong PP5_Very_Strong

The NM_000179.3(MSH6):​c.3173-1G>C variant causes a splice acceptor change involving the alteration of a conserved nucleotide. The variant allele was found at a frequency of 0.00000137 in 1,461,880 control chromosomes in the GnomAD database, with no homozygous occurrence. In-silico tool predicts a pathogenic outcome for this variant. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Likely pathogenic (★★).

• Frequency:
  • Genomes: not found (cov: 32)
  • Exomes 𝑓: 0.0000014 (0 hom.)
• Consequence: MSH6 NM_000179.3 splice_acceptor
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:9
• Conservation: PhyloP100: 9.97

Genes affected

MSH6 (HGNC:7329): (mutS homolog 6) This gene encodes a member of the DNA mismatch repair MutS family. In E. coli, the MutS protein helps in the recognition of mismatched nucleotides prior to their repair. A highly conserved region of approximately 150 aa, called the Walker-A adenine nucleotide binding motif, exists in MutS homologs. The encoded protein heterodimerizes with MSH2 to form a mismatch recognition complex that functions as a bidirectional molecular switch that exchanges ADP and ATP as DNA mismatches are bound and dissociated. Mutations in this gene may be associated with hereditary nonpolyposis colon cancer, colorectal cancer, and endometrial cancer. Transcripts variants encoding different isoforms have been described. [provided by RefSeq, Jul 2013]

FBXO11 (HGNC:13590): (F-box protein 11) This gene encodes a member of the F-box protein family which is characterized by an approximately 40 amino acid motif, the F-box. The F-box proteins constitute one of the four subunits of ubiquitin protein ligase complex called SCFs (SKP1-cullin-F-box), which function in phosphorylation-dependent ubiquitination. The F-box proteins are divided into 3 classes: Fbws containing WD-40 domains, Fbls containing leucine-rich repeats, and Fbxs containing either different protein-protein interaction modules or no recognizable motifs. The protein encoded by this gene belongs to the Fbxs class. It can function as an arginine methyltransferase that symmetrically dimethylates arginine residues, and it acts as an adaptor protein to mediate the neddylation of p53, which leads to the suppression of p53 function. This gene is known to be down-regulated in melanocytes from patients with vitiligo, a skin disorder that results in depigmentation. Polymorphisms in this gene are associated with chronic otitis media with effusion and recurrent otitis media (COME/ROM), a hearing loss disorder, and the knockout of the homologous mouse gene results in the deaf mouse mutant Jeff (Jf), a single gene model of otitis media. Alternatively spliced transcript variants encoding distinct isoforms have been identified for this gene. [provided by RefSeq, Jun 2010]

ACMG classification

Verdict is Pathogenic. Variant got 22 ACMG points.

• PVS1: Splicing +-2 bp (donor or acceptor) variant, LoF is a know mechanism of disease, Cryptic splice site detected, with MaxEntScore 3.4, offset of 28, new splice context is: ctgtgcctggctaactatAGtcg. Cryptic site results in frameshift change. If cryptic site found is not functional and variant results in exon loss, it results in frameshift change.
• PM2: Very rare variant in population databases, with high coverage
• PP3: Splicing scoreres supports a deletorius effect: Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF, max_spliceai. No scorers claiming Uncertain. No scorers claiming Benign.
• PP5: Variant 2-47803419-G-C is Pathogenic according to our data. Variant chr2-47803419-G-C is described in ClinVar as [Likely_pathogenic]. Clinvar id is 42469.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars. Variant chr2-47803419-G-C is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
MSH6	NM_000179.3	c.3173-1G>C		splice_acceptor_variant		ENST00000234420.11

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
MSH6	ENST00000234420.11	c.3173-1G>C		splice_acceptor_variant		1	NM_000179.3	P4

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome AF: 0.00000137 AC: 2 AN: 1461880 Hom.: 0 Cov.: 33 AF XY: 0.00000138 AC XY: 1 AN XY: 727238

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.00

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.00

Gnomad4 SAS exome AF: 0.00

Gnomad4 FIN exome AF: 0.00

Gnomad4 NFE exome AF: 8.99e-7

Gnomad4 OTH exome AF: 0.0000166

GnomAD4 genome Cov.: 32

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:9

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

not provided Pathogenic:3

Pathogenic, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Dec 01, 2022	This variant disrupts a canonical splice-acceptor site and interferes with normal MSH6 mRNA splicing. The variant has not been reported in individuals with MSH6-related diseases in the published literature. It also has not been reported in large, multi-ethnic general populations (http://gnomad.broadinstitute.org). Based on the available information, this variant is classified as pathogenic. -
Likely pathogenic, no assertion criteria provided	research	Mayo Clinic Laboratories, Mayo Clinic	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	ARUP Laboratories, Molecular Genetics and Genomics, ARUP Laboratories	Mar 15, 2021	The MSH6 c.3173-1G>C variant (rs397515875), to our knowledge, is not reported in the medical literature but is reported as pathogenic/likely pathogenic in ClinVar (Variation ID: 42469). This variant is absent from general population databases (1000 Genomes Project, Exome Variant Server, and Genome Aggregation Database), indicating it is not a common polymorphism. This variant abolishes the canonical splice acceptor site of intron 4, which is likely to disrupt gene function. Based on available information, this variant is considered to be pathogenic. -

Hereditary cancer-predisposing syndrome Pathogenic:2

Pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	Sep 03, 2021	The c.3173-1G>C intronic pathogenic mutation results from a G to C substitution one nucleotide upstream from coding exon 5 of the MSH6 gene. This alteration has been identified in multiple HNPCC families with probands' tumors exhibiting absent MSH2 and/or MSH6 on IHC (Ambry internal data). This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). In silico splice site analysis predicts that this alteration will weaken the native splice acceptor site. Alterations that disrupt the canonical splice site are expected to cause aberrant splicing, resulting in an abnormal protein or a transcript that is subject to nonsense-mediated mRNA decay. As such, this alteration is classified as a disease-causing mutation. -
Pathogenic, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Nov 10, 2022	This variant causes a G to C nucleotide substitution at the -1 position of intron 4 of the MSH6 gene. Splice site prediction tools suggest that this variant may have a significant impact on RNA splicing. Although this prediction has not been confirmed in published RNA studies, this variant is expected to result in an absent or disrupted protein product. This variant has been reported in multiple individuals and families affected with Lynch syndrome associated cancers (communication with external lab; SCV000216621.6). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). A different variant affecting the same splice acceptor site, c.3173-1G>A, is described to be disease-causing (ClinVar variation ID: 428387). Loss of MSH6 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Pathogenic. -

Lynch syndrome 5 Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Myriad Genetics, Inc.

Aug 22, 2023

This variant is considered likely pathogenic. This variant occurs within a consensus splice junction and is predicted to result in abnormal mRNA splicing of either an out-of-frame exon or an in-frame exon necessary for protein stability and/or normal function. -

Hereditary nonpolyposis colon cancer Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Women's Health and Genetics/Laboratory Corporation of America, LabCorp

Jul 31, 2023

Variant summary: MSH6 c.3173-1G>C is located in a canonical splice-site and is predicted to affect mRNA splicing resulting in a significantly altered protein due to either exon skipping, shortening, or inclusion of intronic material. Several computational tools predict a significant impact on normal splicing: Four predict the variant abolishes the canonical 3' acceptor site. However, these predictions have yet to be confirmed by functional studies. The variant was absent in 251374 control chromosomes (gnomAD). c.3173-1G>C has been reported in the literature in a setting of multigene panel testing in at least one individual affected with breast cancer (Lu_2019). These data does not allow for unequivocal conclusions about variant significance. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. The following publications have been ascertained in the context of this evaluation (PMID: 28514183, 30128536). Three submitters have cited clinical-significance assessments for this variant to ClinVar after 2014. All submitters, two of which cited internal observations of the variant in individuals with Hereditary Nonpolyposis Colorectal Cancer/Lynch syndrome-associated cancers, classified the variant as pathogenic. Based on the evidence outlined above, the variant was classified as likely pathogenic. -

Lynch syndrome Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine

Sep 21, 2011

The 3173-1G>C variant has not been previously reported and has not been previous ly identified by our laboratory. The 3173-1G>C variant is predicted to cause ab normal splicing because the nucleotide substitution occurs in the highly conserv ed splice consensus sequence. Computational splicing models predict this alterat ion to abolish the 3? acceptor splice site of intron 4 and result in both intron 4 retention and exon 5 skipping; however, the accuracy of these models is not k nown. Although this variant is consistent with a loss of function allele, additi onal family-based and functional studies are required to fully assess the impact of this variant. Together, the evidence suggests this variant is likely to be p athogenic. -

Hereditary nonpolyposis colorectal neoplasms Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Labcorp Genetics (formerly Invitae), Labcorp

Jan 10, 2024

This sequence change affects an acceptor splice site in intron 4 of the MSH6 gene. RNA analysis indicates that disruption of this splice site induces altered splicing and likely results in the loss of 57 amino acid residue(s), but is expected to preserve the integrity of the reading-frame. This variant is not present in population databases (gnomAD no frequency). Disruption of this splice site has been observed in individual(s) with Lynch syndrome-associated cancers (Invitae). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 42469). Studies have shown that disruption of this splice site results in the activation of a cryptic splice site in exon 5 (Invitae). This variant disrupts a region of the MSH6 protein in which other variant(s) (p.Leu1063Arg) have been determined to be pathogenic (PMID: 23733757, 26099011, 28531214; Invitae). This suggests that this is a clinically significant region of the protein, and that variants that disrupt it are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.37	D
BayesDel_noAF	Pathogenic	0.30
CADD	Pathogenic	33
DANN	Uncertain	0.99
Eigen	Pathogenic	1.2
Eigen_PC	Pathogenic	1.1
FATHMM_MKL	Pathogenic	0.98	D
MutationTaster	Benign	1.0	D;D;D;D
GERP RS		5.4
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.9

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.93
SpliceAI score (max)		0.98		Details are displayed if max score is > 0.2
DS_AL_spliceai		0.98		Position offset: 1

Find out detailed SpliceAI scores and Pangolin per-transcript scores at spliceailookup.broadinstitute.org

Publications

LitVar

Other links and lift over

dbSNP: rs397515875; hg19: chr2-48030558;