ClinVar Genomic variation as it relates to human health

This variant is denoted MSH6 c.3961A>G at the cDNA level, p.Arg1321Gly (R1321G) at the protein level, and results in the change of an Arginine to a Glycine (AGA>GGA). This variant has been reported in individuals with early-onset colorectal cancer (CRC), pancreatic cancer, and in one individual with a personal history of Lynch syndrome-associated cancer and/or polyps (Pinto 2006, Barneston 2008, Yurgelun 2015, Jansen 2016, Shindo 2017). This variant was also reported to co-occur with an MSH2 pathogenic deletion in a patient with mismatch repair-deficient CRC (Le 2017). MSH6 Arg1321Gly was observed at an allele frequency of 0.03% (35/124,972) in individuals of European ancestry in large population cohorts (Lek 2016). MSH6 Arg1321Gly is located within an MSH2 binding site and the ATPase domain (Kariola 2002, Warren 2007, Kansikas 2011). In silico analysis, which includes protein predictors and evolutionary conservation, supports a deleterious effect. Based on currently available evidence, it is unclear whether MSH6 Arg1321Gly is a pathogenic or benign variant. We consider it to be a variant of uncertain significance.

This variant is considered benign. This variant is strongly associated with less severe personal and family histories of cancer, typical for individuals without pathogenic variants in this gene [PMID: 25085752]. Homozygosity for this variant has been confirmed in one or more individuals lacking clinical features consistent with gene-specific recessive disease, indicating that this variant is unlikely to be pathogenic.

This missense variant replaces arginine with glycine at codon 1321 of the MSH6 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). To our knowledge, functional studies have not been performed for this variant. This variant has been reported in individuals affected with Lynch syndrome-associated cancer and/or polyps, pancreatic, kidney, uterine, omentum cancer, breast cancer, or glioblastoma (PMID: 16940983, 18033691, 25980754, 26648449, 26689913, 28767289, 29596542, 29684080, 31422818, 31391288; DOI: 10.1101/2021.04.15.21255554, 33471991), including one case with a pathogenic MSH2 covariant (PMID: 25980754). In a large breast cancer case-control study, the variant was observed in 31/60466 cases and 37/53461 unaffected controls (PMID: 33471991). This variant has also been identified in 39/279718 chromosomes in the general population by the Genome Aggregation Database (gnomAD). The available evidence is insufficient to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance.

This missense variant replaces arginine with glycine at codon 1321 of the MSH6 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). To our knowledge, functional studies have not been performed for this variant. This variant has been reported in individuals affected with Lynch syndrome-associated cancer and/or polyps, pancreatic, kidney, uterine, omentum cancer, breast cancer, or glioblastoma (PMID: 16940983, 18033691, 25980754, 26648449, 26689913, 28767289, 29596542, 29684080, 31422818, 31391288; DOI: 10.1101/2021.04.15.21255554, 33471991), including one case with a pathogenic MSH2 covariant (PMID: 25980754). In a large breast cancer case-control study, the variant was observed in 31/60466 cases and 37/53461 unaffected controls (PMID: 33471991). This variant has also been identified in 39/279718 chromosomes in the general population by the Genome Aggregation Database (gnomAD). The available evidence is insufficient to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance.

proposed classification - variant undergoing re-assessment, contact laboratory

The MSH6 c.3961A>G (p.Arg1321Gly) missense change has a maximum subpopulation frequency of 0.027% in gnomAD v2.1.1 (https://gnomad.broadinstitute.org/variant/2-48033750-A-G). Five of seven in silico tools predict a damaging effect of this variant on protein function (PP3), but these predictions have not been confirmed by functional studies. This variant has been reported in individuals with colorectal cancer (PMID: 16940983, 18033691) and an individual with suspected Lynch syndrome (PMID: 25980754). It has also been reported in an individual with pancreatic ductal adenocarcinoma and family history of miscellaneous cancer types (PMID: 28767289). In summary, this variant meets criteria to be classified as of uncertain significance based on the ACMG/AMP criteria: PP3.

Variant summary: MSH6 c.3961A>G (p.Arg1321Gly) results in a non-conservative amino acid change located in the DNA mismatch repair protein MutS, C-terminal (IPR000432) domain, comprised of the ATPase domain and the HTH (helix-turn-helix) domain, the latter being involved in dimer contacts (InterPro) of the encoded protein sequence. Four in-silico tools predict a damaging effect of the variant on protein function, while the mismatch repair in-silico tool PON-MMR2 predicts a benign effect (Niroula_2015). The variant allele was found at a frequency of 0.00014 in 252294 control chromosomes, predominantly at a frequency of 0.00028 within the Non-Finnish European subpopulation in the gnomAD database. The observed variant frequency within Non-Finnish European control individuals in the gnomAD database is approximately 2 fold of the estimated maximal expected allele frequency for a pathogenic variant in MSH6 causing Hereditary Nonpolyposis Colorectal Cancer phenotype (0.00014), strongly suggesting that the variant is a benign polymorphism found primarily in populations of Non-Finnish European origin. c.3961A>G has been reported in the literature in individuals affected with Lynch Syndrome, colorectal cancer (Pinto_2006, Barneston_2008, Yurgelun_2015, Jansen_2016, Gordon_2019), Cowden/Cowden-like (CS/CS-like) or Bannayan-Riley-Ruvalcaba syndromes (Yehia 2018), Tubo-ovarian cancer (Delahunty_2022), and breast cancer (Faldoni_2020, Guindalini_2022). These reports do not provide unequivocal conclusions about association of the variant with Hereditary Nonpolyposis Colorectal Cancer.- The UMD database reported the variant in 3 patients, one of which was indicated to carry another pathogenic MSH6 variant (c.3514dup, p.Arg1172LysfsX5), providing supporting evidence for a benign role. 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: 26689913, 31422818, 31391288, 29684080, 29596542, 28767289, 26648449, 26333163, 35263119, 33007869, 26332594, 25980754, 23621914, 24055113, 18033691, 16940983, 25637381, 36845387, 35264596). 16 submitters have cited clinical-significance assessments for this variant to ClinVar after 2014 and classified this variant as uncertain significance (n=12), likely benign (n=3) and benign (n=1). Based on the evidence outlined above, the variant was classified as likely benign.

This alteration is classified as likely benign based on a combination of the following: seen in unaffected individuals, population frequency, intact protein function, lack of segregation with disease, co-occurrence, RNA analysis, in silico models, amino acid conservation, lack of disease association in case-control studies, and/or the mechanism of disease or impacted region is inconsistent with a known cause of pathogenicity.

The MSH6 p.Arg1321Gly variant was identified in 3 of 2112 proband chromosomes (frequency: 0.0014) from individuals or families with colorectal cancer and was not identified in 2676 control chromosomes from healthy individuals (Bameston 2008, Le 2017, Pinto 2006). The variant was also identified in the following databases: dbSNP (ID: rs41295278) as With Uncertain significance allele, ClinVar (classified as uncertain significance by InSight, Genedx, Ambry Genetics, Counsyl, Mayo Clinic; as likely benign by Invitae), Clinvitae (classified as uncertain significance by ClinVar), MutDB (classified as polymorphism), UMD-LSDB (3X neutral), Insight Colon Cancer Gene Variant Database (8X class3), Mismatch Repair Genes Variant Database, and the Insight Hereditary Tumors Database (9X). In UMD the variant was identified with a co-occurring pathogenic MSH6 variant (c.3514dup (p.Arg1172LysfsX5)), increasing the likelihood that the p.Arg1321Gly variant does not have clinical significance. The variant was not identified in the COGR, Cosmic, or Zhejiang Colon Cancer Databases. The variant was identified in control databases in 40 of 274126 chromosomes at a frequency of 0.0002 in the following populations: African in 1 of 23864 chromosomes (freq. 0.00004), other in 2 of 6418 chromosomes (freq. 0.0003), European in 35 of 124972 chromosomes (freq. 0.0003), increasing the likelihood that this may be a low frequency increasing the likelihood that this may be a low frequency variant in certain populations of origin (Genome Aggregation Consortium Feb 27, 2017). The p.Arg1321 residue is conserved in mammals but not in more distantly related organisms, and computational analyses (PolyPhen-2, SIFT, AlignGVGD, BLOSUM, MutationTaster) suggest that the variant may impact the protein; however, this information is not predictive enough to assume pathogenicity. The variant occurs outside of the splicing consensus sequence and 2 of 5 in silico or computational prediction software programs (SpliceSiteFinder, MaxEntScan, NNSPLICE, GeneSplicer, HumanSpliceFinder) predict a greater than 10% difference in splicing; this is not very predictive of pathogenicity. The variant is located with the DNA mismatch repair protein MutS, C-terminal P-loop containing nucleoside triphosphate hydrolase and DNA mismatch repair Msh6 functional domains. In summary, based on the above information, the clinical significance of this variant cannot be determined with certainty at this time. This variant is classified as a variant of uncertain significance.

The MSH6 c.3961A>G variant is predicted to result in the amino acid substitution p.Arg1321Gly. This variant has been reported in individuals with Lynch syndrome and sporadic, early-onset colorectal cancer (Barnetson et al. 2008. PubMed ID: 18033691; Pinto et al. 2006. PubMed ID: 16940983, Yurgelun et al. 2015. PubMed ID: 25980754). This variant was also found with a MSH2 deletion variant in a patient with colorectal cancer (Le et al. 2017. PubMed ID: 28596308). Another missense variant involving the same amino acid residue (p.Arg1321Ser) has been reported in at least one individual with endometrial cancer (Devlin et al. 2008. PubMed ID: 18269114). This variant is reported in 0.027% of alleles in individuals of European (Non-Finnish) descent in gnomAD and has conflicting interpretations regarding its pathogenicity in ClinVar, ranging from likely benign to uncertain (https://www.ncbi.nlm.nih.gov/clinvar/variation/89490/). At this time, the clinical significance of this variant is uncertain due to the absence of conclusive functional and genetic evidence.