ClinVar Genomic variation as it relates to human health

The p.Thr33Pro variant in MSH2 has been reported in 4 individuals with Lynch Syndrome-related cancers (Hedge 2005, Hampel 2006, Ollila 2006, Chubb 2015). Functional studies (microsatellite instability, immunohistochemistry, mismatch repair, expression and interaction assays) provide conflicting evidence on whether the p.Thr33Pro variant impacts protein function (Ollila 2006, Ollila 2008, Kansikas 2011, Martinez 2010). This variant has also been identified in 4/32028 Latino chromosomes by the Genome Aggregation Database (GnomAD, http://gnomad.broadinstitute.org). Computational prediction tools and conservation analysis suggest that the p.Thr33Pro variant may impact the protein, though this information is not predictive enough to determine pathogenicity. In addition, this variant was classified as a variant of uncertain significance on Sept. 5, 2013 by the ClinGen-approved InSiGHT expert panel (ClinVar SCV000107811.2). In summary, the clinical significance of the p.Thr33Pro variant is uncertain.

This variant is considered likely 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].

This missense variant replaces threonine with proline at codon 33 of the MSH2 protein. Computational prediction suggests that this variant may have deleterious impact on protein structure and function (internally defined REVEL score threshold >= 0.7, PMID: 27666373). Functional studies reported this variant as partially functional in mismatch repair assay in vitro (PMID: 17101317) and in yeast (PMID: 20176959). This variant is functional in a methylation tolerance assay (PMID: 30998989) and does not impact MSH2 function in a 6-thioguanine sensitivity assay in haploid human cells (internally defined LOF score threshold <= -1.32, PMID: 33357406). This variant has been reported in an individual affected with rectal cancer (PMID: 25559809) and at least two individuals affected with endometrial cancer (PMID: 16885385, 17101317, 32634176). This variant has been identified in 16/254182 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 alteration is classified as 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.

Published functional studies demonstrate no damaging effect: interaction with MSH6, expression in colon carcinoma cells, protein stability, mismatch binding, mismatch repair activity, and cell survival similar to wild type (PMID: 17101317, 18951462, 20176959, 30998989, 33357406, 26951660); Observed in patients with Lynch-related cancers; however, available tumor studies were inconsistent (PMID: 16885385, 25559809, 17101317, 32634176); 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: 17594722, 17101317, 18951462, 25559809, 22949387, 21120944, 16237223, 16885385, 17370310, 18383312, 20176959, 26951660, 30998989, 33357406, 35688932, 32634176, 26580448, 31391288, 32885271, 18822302)

Variant summary: MSH2 c.97A>C (p.Thr33Pro) results in a non-conservative amino acid change located in the DNA mismatch repair protein MutS-like, N-terminal 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 6.7e-05 in 222782 control chromosomes, predominantly at a frequency of 0.00013 within the Latino subpopulation in the gnomAD database. The available data on variant occurrences in the general population are insufficient to allow any conclusion about variant significance. c.97A>C has been reported in the literature in individuals affected with Lynch Syndrome, endometrial cancer and colorectal cancer (Hegde_2005, Hampel_2006, Ollila_2006, Chubb_2015, Li_2020, Singh_2020), without strong evidence for causality. Several publications have reported functional studies of the variant showing: normal interaction with MSH6 but decreased in vitro MMR activity (Ollila_2006), weak MMR defect (Martinez_2010), while other pubications reported the variant to be neutral via assays that measured proportions of cells that undergo death following exposure to a methylating agent (Bouvet_2019) and in human cell line models in which MSH2 deletion was complemented by libraries of variants comprising nearly every possible MSH2 missense allele (Jia_2021). The following publications have been ascertained in the context of this evaluation (PMID: 30998989, 25559809, 16885385, 16237223, 33357406, 31391288, 20176959, 17101317, 32634176, 26580448). ClinVar contains an entry for this variant (Variation ID: 91267). Based on the evidence outlined above, the variant was classified as uncertain significance.

This missense variant replaces threonine with proline at codon 33 of the MSH2 protein. Computational prediction suggests that this variant may have deleterious impact on protein structure and function (internally defined REVEL score threshold >= 0.7, PMID: 27666373). Functional studies reported this variant as partially functional in mismatch repair assay in vitro (PMID: 17101317) and in yeast (PMID: 20176959). This variant is functional in a methylation tolerance assay (PMID: 30998989) and does not impact MSH2 function in a 6-thioguanine sensitivity assay in haploid human cells (internally defined LOF score threshold <= -1.32, PMID: 33357406). This variant has been reported in individuals affected with Lynch syndrome-associated cancers (PMID: 16237223, 16885385, 17101317, 25559809, 32634176). This variant has been identified in 16/254182 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.

The MSH2 p.Thr33Pro variant was identified in 3 of 2424 proband chromosomes (frequency: 0.001) from individuals with colon and endometrial cancer (Hegde 2005, Hampel 2006, Chubb 2015). The variant was identified in dbSNP (rs63751107) as “with uncertain significance allele”, ClinVar (classified as "uncertain significance" by Invitae, GeneDx and five other submitters) and UMD-LSDB (observed 2x). The variant was identified in control databases in 14 of 248,976 chromosomes at a frequency of 0.00006 (Genome Aggregation Database Feb 27, 2017). The variant was observed in the following populations: Latino in 4 of 31,200 chromosomes (freq: 0.0001), European in 10 of 113,324 chromosomes (freq: 0.00009), but not in the African, Other, Ashkenazi Jewish, East Asian, Finnish, and South Asian populations. A cell line derived from a patient with the observed variant had normal levels of MSH2 and MSH6 protein as observed on a western blot (Ollila 2006). In an in vitro mismatch repair assay, the variant was generated by site-directed mutagenesis and decreased MMR efficiency by 23% compared to wild type (Ollila 2006). The variant is present in the DNA binding domain of MSH2 and in another study was demonstrated to bind mismatches as efficiently as wild type, and had a slightly reduced DNA release. These observations led to the conclusion that the variant causes a slight defect in MMR efficiency, and based on these experiments, it is unclear if the variant is pathogenic (Ollila 2008). The p.Thr33 residue is conserved across mammals and other organisms, and four out of five 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 in silico or computational prediction software programs (SpliceSiteFinder, MaxEntScan, NNSPLICE, GeneSplicer) do not predict a difference in splicing. 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.