Genetic Information
Gene & Transcript Details
| ID | Status | Details |
|---|---|---|
| NM_000249.3 | RefSeq Select | 2662 nt | 199–2469 |
| NM_000249.2 | Alternative | 2524 nt | 61–2331 |
| NM_000249.4 | MANE Select | 2494 nt | 31–2301 |
Variant Details
Clinical & Population Data
Population Frequency
gnomADClinVar
OpenThe 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.
"This variant has been reported in ClinVar as Uncertain significance (5 clinical laboratories) and as Uncertain Significance (1 clinical laboratories)."
COSMIC Somatic Evidence
Open
Functional Impact & Domains
Functional Domain
Computational Analysis
Pathogenicity Predictions
SpliceAISpliceAI Scores
Window: ±500bp| Effect Type | Score | Position |
|---|---|---|
| Acceptor Loss (AL) | 0.03 | -116 bp |
| Donor Loss (DL) | 0.01 | 19 bp |
| Acceptor Gain (AG) | 0.0 | 12 bp |
| Donor Gain (DG) | 0.0 | 132 bp |
VCEP Guidelines
Applied ACMG/AMP Criteria (VCEP Specific)
PVS1 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Supporting)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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 (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 (Not Applied)
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 (Not Applied)
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 (Not Applied)
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.