MLH1 c.2084C>T, p.Ser695Leu
NM_000249.4:c.2084C>T
COSMIC ID: COSM5609387
Variant of Uncertain Significance (VUS)
This MLH1 S695L variant is absent from population databases (PM2) and computational analyses predict no impact (BP4), but there is no additional functional, segregation, de novo, or case-control evidence. Insufficient evidence for pathogenicity or benignity leads to a classification of Variant of Uncertain Significance.
ACMG/AMP Criteria Applied
PM2
BP4
Genetic Information
Gene & Transcript Details
Gene
MLH1
Transcript
NM_000249.4
MANE Select
Total Exons
19
Strand
Forward (+)
Reference Sequence
NC_000003.11
Alternative Transcripts
| ID | Status | Details |
|---|---|---|
| NM_000249.3 | RefSeq Select | 19 exons | Forward |
| NM_000249.2 | Alternative | 19 exons | Forward |
Variant Details
HGVS Notation
NM_000249.4:c.2084C>T
Protein Change
S695L
Location
Exon 18
(Exon 18 of 19)
5'Exon Structure (19 total)3'
Functional Consequence
Loss of Function
Related Variants
No evidence of other pathogenic variants at position 695 in gene MLH1
Alternate Identifiers
COSM5609387
Variant interpretation based on transcript NM_000249.4
Genome Browser
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HGVS InputNM_000249:c.2084C>T
Active Tracks
ConservationRefSeqClinVargnomAD
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Clinical Data
Global Frequency
0.0 in 100,000
Extremely Rare
Global: 0.0%
0%
0.05%
0.1%
1%
5%
10%+
Allele Information
Total: 251010Alt: 0Homozygotes: 0
ACMG Criteria Applied
PM2
This variant is present in gnomAD (MAF= 0%, 0/251010 alleles, homozygotes = 0) but does not appear at a higher frequency in any of the selected populations. The variant is rare (MAF < 0.1%), supporting PM2 criterion application.
Classification
2 publications
Uncertain Significance (VUS)
Based on 5 submitter reviews in ClinVar
Submitter Breakdown
5 VUS
Pathogenic
Likely Path.
VUS
Likely Benign
Benign
Publications (2)
The p.S695L variant (also known as c.2084C>T), located in coding exon 18 of the MLH1 gene, results from a C to T substitution at nucleotide position 2084. The serine at codon 695 is replaced by leucine, an amino acid with dissimilar properties. This amino acid position is well conserved in available vertebrate species. In addition, the in silico prediction for this alteration is inconclusive. Based on the available evidence, the clinical significance of this variant remains unclear.
This sequence change replaces serine, which is neutral and polar, with leucine, which is neutral and non-polar, at codon 695 of the MLH1 protein (p.Ser695Leu). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individual(s) with multiple cancers (PMID: 33850299). ClinVar contains an entry for this variant (Variation ID: 483566). 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) performed at Invitae indicates that this missense variant is not expected to disrupt MLH1 protein function with a negative predictive value of 95%. 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.
Clinical Statement
This variant has been reported in ClinVar as Uncertain significance (5 clinical laboratories) and as Uncertain Significance (1 clinical laboratories).
Functional Impact
Functional Domain
Hotspot Status
Not a hotspot
Domain Summary
This variant is not located in a mutational hotspot or critical domain (0 mutations).
Related Variants in This Domain
No evidence of other pathogenic variants at position 695 in gene MLH1
Computational Analysis
Pathogenicity Predictions
REVEL Score
0.517
0.517
Likely Benign0.0
Uncertain (Low)0.2
Uncertain (Med)0.5
Likely Pathogenic0.75
REVEL scores ≥ 0.75 are strong evidence (PP3)
Predictor Consensus
Mixed/VUS
PP3 Applied
No
Additional Predictors
Pathogenic:
metasvm: D
Benign:
CADD: 4.18polyphen_prediction: benignmetalr: Tprimateai: T
Neutral: Show all
VCEP Guidelines
Applied ACMG/AMP Criteria (VCEP Specific) VCEP Guidelines
PVS1
PVS1 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for PVS1 is: "Very Strong Nonsense/frameshift variant introducing Premature Termination Codon at or before codon 753 in MLH1, or canonical ±1/2 splice variant." The evidence for this variant shows: c.2084C>T results in a missense change (S695L), not a null or canonical splice variant. Therefore, this criterion is not applied because the variant is not predicted to introduce a premature stop or disrupt splicing at canonical sites.
PS1
PS1 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for PS1 is: "Strong: A predicted missense substitution that encodes the same amino acid change with a different underlying nucleotide change previously established by this VCEP as Pathogenic." The evidence for this variant shows: no other nucleotide change at codon 695 has been classified as Pathogenic. Therefore, this criterion is not applied because there is no established pathogenic variant encoding S695L by another nucleotide change.
PS2
PS2 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for PS2 is: "Very Strong ≥ 4 de novo points; Strong 2–3 points; Moderate 1 point; Supporting 0.5 points for de novo occurrences with maternity and paternity confirmed." The evidence for this variant shows: no information on de novo occurrence or parental testing. Therefore, this criterion is not applied because de novo status cannot be evaluated without Trio data.
PS3
PS3 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for PS3 is: "Strong: Calibrated functional assays with functional odds for Pathogenicity >18.7; Moderate: odds >4.3–18.7; Supporting: odds >2.08–4.3." The evidence for this variant shows: no functional studies have been performed for S695L. Therefore, this criterion is not applied because functional assay data are not available.
PS4
PS4 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for PS4 is: "The prevalence of the variant in affected individuals is significantly increased compared to controls." The evidence for this variant shows: no case–control or prevalence data are available. Therefore, this criterion is not applied because population-based enrichment in cases versus controls cannot be evaluated.
PM1
PM1 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for PM1 is: "Located in a mutational hot spot and/or critical and well-established functional domain without benign variation." The evidence for this variant shows: no data indicating that codon 695 resides in a defined hotspot or uniquely critical domain lacking benign variation. Therefore, this criterion is not applied because hotspot/domain evidence is not present.
PM2
PM2 (Supporting) Strength Modified
According to VCEP guidelines, the rule for PM2 is: "Supporting: Absent/extremely rare (<1 in 50,000 alleles) in gnomAD v4 dataset." The evidence for this variant shows: absent from gnomAD (0/251,010 alleles). Therefore, this criterion is applied at Supporting strength because the variant is absent from population databases meeting the VCEP threshold.
PM3
PM3 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for PM3 is: "Points for detected in trans with a pathogenic variant in recessive disorders." The evidence for this variant shows: no data on allelic configuration or co-occurrence with another MLH1 variant. Therefore, this criterion is not applied because allelic phase and recessive context cannot be assessed.
PM4
PM4 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for PM4 is: "Protein length changes due to in-frame deletions/insertions or stop-loss variants." The evidence for this variant shows: S695L is a missense change without alteration of protein length. Therefore, this criterion is not applied because there is no change in protein length.
PM5
PM5 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for PM5 is: "Moderate: Missense change at a residue where a different missense change was classified as Pathogenic by this VCEP." The evidence for this variant shows: no other missense at codon 695 has been classified as Pathogenic. Therefore, this criterion is not applied because no pathogenic missense variant at the same residue is known.
PM6
PM6 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for PM6 is: "Assumed de novo, but without confirmation of paternity or maternity." The evidence for this variant shows: no de novo data are available. Therefore, this criterion is not applied because unconfirmed de novo occurrence cannot be assessed.
PP1
PP1 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for PP1 is: "Co-segregation with disease in multiple affected family members." The evidence for this variant shows: no segregation data are provided. Therefore, this criterion is not applied because familial segregation has not been evaluated.
PP2
PP2 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for PP2 is: "Missense variant in a gene with low rate of benign missense variation and where missense are a common mechanism of disease." The evidence for this variant shows: MLH1 has both loss-of-function and missense pathogenic variants reported, but it is not a gene characterized by exclusively benign missense tolerance. Therefore, this criterion is not applied.
PP3
PP3 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for PP3 is: "Supporting: Missense variant with HCI prior probability >0.68 & ≤0.81 or predicted splice defect with SpliceAI delta ≥0.2." The evidence for this variant shows: SpliceAI delta score 0.03 (no splicing impact) and mixed in silico predictions. Therefore, this criterion is not applied because computational evidence does not support a deleterious effect.
PP4
PP4 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for PP4 is: "Supporting/Moderate/Strong based on number of independent MSI-H or loss-of-MMR-expression tumors excluding MLH1 promoter methylation." The evidence for this variant shows: no tumor phenotype or MSI/MMR data are provided. Therefore, this criterion is not applied because phenotype-specific tumor data are unavailable.
PP5
PP5 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for PP5 is: "Reputable source reports variant as pathogenic without available evidence." The evidence for this variant shows: ClinVar reports Uncertain Significance. Therefore, this criterion is not applied because no reputable source calls this variant pathogenic.
BA1
BA1 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for BA1 is: "Allele frequency >5% in controls." The evidence for this variant shows: absent in population databases. Therefore, this criterion is not applied because the allele frequency is not above the established benign threshold.
BS1
BS1 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for BS1 is: "Strong: GnomAD v4 Grpmax filtering allele frequency ≥0.0001 and <0.001." The evidence for this variant shows: frequency = 0. Therefore, this criterion is not applied because the allele frequency does not reach the benign range.
BS2
BS2 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for BS2 is: "Strong: Co-occurrence in trans with a known pathogenic variant in a healthy adult without CMMRD." The evidence for this variant shows: no healthy adult co-occurrence data. Therefore, this criterion is not applied because co-occurrence data are not available.
BS3
BS3 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for BS3 is: "Strong: Calibrated functional assays with odds for Pathogenicity ≤0.05 or synonymous/intronic with no mRNA aberration." The evidence for this variant shows: no functional or mRNA assay data. Therefore, this criterion is not applied because functional assay data are lacking.
BS4
BS4 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for BS4 is: "Lack of segregation in affected members (combined LOD score)." The evidence for this variant shows: no segregation analysis. Therefore, this criterion is not applied because segregation data are not available.
BP1
BP1 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for BP1 is: "Missense variant in a gene for which primarily truncating variants cause disease." The evidence for this variant shows: MLH1 missense variants can be pathogenic. Therefore, this criterion is not applied because missense is an established mechanism of MLH1-related disease.
BP2
BP2 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for BP2 is: "Observed in trans with a pathogenic variant for a dominant disorder." The evidence for this variant shows: no trans co-occurrence data. Therefore, this criterion is not applied because allelic configuration with pathogenic variants is unknown.
BP3
BP3 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for BP3 is: "In-frame indels in repetitive regions without functional domains disrupted." The evidence for this variant shows: S695L is a missense change, not an in-frame indel. Therefore, this criterion is not applied because it is not applicable.
BP4
BP4 (Supporting)
According to VCEP guidelines, the rule for BP4 is: "Supporting: Missense variant with HCI-prior probability <0.11 or for intronic/synonymous variants SpliceAI predicts no splicing impact with delta score ≤0.1." The evidence for this variant shows: SpliceAI delta score 0.03 (no predicted splicing impact) and the majority of in silico tools predict benign effect. Therefore, this criterion is applied at Supporting strength because computational evidence predicts no impact on gene or gene product.
BP5
BP5 (Not Applied) Strength Modified
According to VCEP guidelines, the rule for BP5 is: "Strong: ≥4 tumors inconsistent with gene loss-of-function or BRAF/MLH1 methylation patterns; Supporting: 2–3 tumors." The evidence for this variant shows: no tumor co-occurrence or methylation data. Therefore, this criterion is not applied because tumor phenotype co-occurrence data are absent.
BP6
BP6 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for BP6 is: "Reputable source reports variant as benign without accessible evidence." The evidence for this variant shows: no source reports benign. Therefore, this criterion is not applied.
BP7
BP7 (Not Applied) Strength Modified
According to standard ACMG guidelines, the rule for BP7 is: "Synonymous or intronic variant at or beyond –21/+7 with no splice impact." The evidence for this variant shows: S695L is a missense variant. Therefore, this criterion is not applied because the variant is not synonymous.