ClinVar Genomic variation as it relates to human health

This variant is classified as a variant of uncertain significance as there is insufficient evidence to determine its impact on protein function and/or cancer risk.

This sequence change replaces alanine, which is neutral and non-polar, with serine, which is neutral and polar, at codon 29 of the MLH1 protein (p.Ala29Ser). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individuals with Lynch syndrome, and in many individuals the variant has been reported on the same chromosome with the c.-27C>A promoter MLH1 variant. (PMID: 2408575, 16083711, 21840485, 22878509). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 90397). 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) has been performed at Invitae for this missense variant, however the output from this modeling did not meet the statistical confidence thresholds required to predict the impact of this variant on MLH1 protein function. Experimental studies have shown that this missense change does not substantially affect MLH1 function (PMID: 16083711, 17510385, 21840485). In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance.

MLH1 c.-27C>A has been reported in cis with MLH1 c.85G>T (p.Ala29Ser) in patients with Lynch-related cancers and tumor studies consistent with pathogenic variants in this gene (Raevaara 2005, Hitchins 2011, Ward 2013, Kwok 2014); Published functional studies, when MLH1 c.85G>T (p.Ala29Ser) is evaluated in isolation, demonstrate no damaging effect; however, the MLH1 c.[-27C>A;85G>T] haplotype results in constitutional MLH1 promoter methylation and reduced allelic expression (Raevaara 2005, Takahashi 2007, Hitchins 2011, Kwok 2014); While segregation data of MLH1 c.-27C>A alone is not available, the MLH1 c.[-27C>A;85G>T] haplotype segregates with disease in affected individuals from several unrelated families in published literature (Raevaara 2005, Hitchins 2011, Ward 2013, Kwok 2014); Not observed in large population cohorts (Lek 2016); In silico analysis supports that this missense variant does not alter protein structure/function; This variant is associated with the following publications: (PMID: 24084575, 18383312, 12658575, 16083711, 17594722, 21120944, 22949387, 17370310, 17192056, 16995940, 22878509, 17510385, 21840485, 25345868, 27435373, 28152038, 29341452, 30283143)

****************HAPTOTYPE RD************************ The p.A29S alteration (also known as c.85G>T) is located in coding exon 1 of the MLH1 gene. This alteration results from a G to T substitution at nucleotide position 85. The alanine at codon 29 is replaced by serine, an amino acid with similar properties. The c.-27C>A alteration is located in the 5' untranslated region (5’UTR) of the MLH1 gene. This variant results from a C to A substitution 27 nucleotides upstream from the first translated codon. The c.-27C>A and c.85G>T alterations are linked in cis, as both are part of a European ancestral haplotype. The c.[-27C>A;85G>T] haplotype has been reported to be linked to a constitutional MLH1 epimutation in several unrelated families that either met Amsterdam I/II criteria or were suspected to have HNPCC/Lynch syndrome (Kwok CT et al. Eur. J. Hum. Genet. 2014 May;22(5):617-24; Hitchins MP et al. Cancer Cell 2011;20:200-13; Ward RL et al. Genet. Med. 2013;15:25-35). Within these families, the c.[-27C>A;85G>T] haplotype was associated with differing levels of MLH1 promoter constitutional methylation and with loss of PMS2 and/or MLH1 staining on immunohistochemistry in tumor samples (Kwok CT et al. Eur. J. Hum. Genet. 2014 May;22(5):617-24; Hitchins MP et al. Cancer Cell 2011;20:200-13; Ward RL et al. Genet. Med. 2013;15:25-35). Luciferase reporter assays performed for the c.-27C>A alteration demonstrated reduced promoter activity (Hitchins MP et al. Cancer Cell 2011;20:200-13; Ward RL et al. Genet. Med. 2013;15:25-35). In contrast, luciferase reporter assays performed for the c.85G>T alteration showed little to no reduction in promoter activity compared to wild type suggesting the c.-27C>A alteration accounts for allelic repression of MLH1 transcription resulting in loss of MLH1 expression; however, a direct causal relationship has yet to be clarified (Hitchins MP et al. Cancer Cell 2011;20:200-13). Based on the supporting evidence, the c.[-27C>A;85G>T] haplotype is interpreted as a disease-causing mutation.