chr3-36993616-A-T

Variant names:

• chr3-36993616-A-T
• rs63750555
• NM_000249.4:c.69A>T

Variant summary

Our verdict is Pathogenic. The variant received 13 ACMG points: 13P and 0B. PS1 PM1 PM2 PP3_Strong PP5

The NM_000249.4(MLH1):​c.69A>T​(p.Glu23Asp) variant causes a missense change involving the alteration of a non-conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. In-silico tool predicts a pathogenic outcome for this variant. Variant has been reported in ClinVar as Conflicting classifications of pathogenicity (no stars). Another nucleotide change resulting in the same amino acid substitution has been previously reported as Likely pathogenic in ClinVar. Another variant affecting the same amino acid position, but resulting in a different missense (i.e. E23G) has been classified as Uncertain significance.

• Frequency: Genomes: not found (cov: 33)
• Consequence: MLH1 NM_000249.4 missense
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:1 U:4
• Conservation: PhyloP100: 1.17
• Publications: 7 publications found

Genes affected

MLH1 (HGNC:7127): (mutL homolog 1) The protein encoded by this gene can heterodimerize with mismatch repair endonuclease PMS2 to form MutL alpha, part of the DNA mismatch repair system. When MutL alpha is bound by MutS beta and some accessory proteins, the PMS2 subunit of MutL alpha introduces a single-strand break near DNA mismatches, providing an entry point for exonuclease degradation. The encoded protein is also involved in DNA damage signaling and can heterodimerize with DNA mismatch repair protein MLH3 to form MutL gamma, which is involved in meiosis. This gene was identified as a locus frequently mutated in hereditary nonpolyposis colon cancer (HNPCC). [provided by RefSeq, Aug 2017]

MLH1 Gene-Disease associations (from GenCC):

• Lynch syndrome

  Inheritance: AD Classification: DEFINITIVE, SUPPORTIVE Submitted by: G2P, ClinGen, Orphanet
• Lynch syndrome 2

  Inheritance: AD Classification: DEFINITIVE, STRONG Submitted by: Ambry Genetics, Genomics England PanelApp
• Muir-Torre syndrome

  Inheritance: AD Classification: DEFINITIVE, STRONG, MODERATE, SUPPORTIVE Submitted by: Genomics England PanelApp, Ambry Genetics, G2P, Orphanet
• mismatch repair cancer syndrome 1

  Inheritance: AR Classification: DEFINITIVE, STRONG, SUPPORTIVE Submitted by: Ambry Genetics, G2P, Labcorp Genetics (formerly Invitae), Orphanet, ClinGen
• Lynch syndrome 1

  Inheritance: AD Classification: STRONG Submitted by: Labcorp Genetics (formerly Invitae)
• ovarian cancer

  Inheritance: AD Classification: STRONG Submitted by: Genomics England PanelApp
• malignant pancreatic neoplasm

  Inheritance: AD Classification: MODERATE Submitted by: Genomics England PanelApp
• rhabdomyosarcoma

  Inheritance: AR Classification: MODERATE Submitted by: Genomics England PanelApp
• prostate cancer

  Inheritance: AD Classification: LIMITED Submitted by: Ambry Genetics
• breast cancer

  Inheritance: AD Classification: NO_KNOWN Submitted by: Ambry Genetics
• hereditary breast carcinoma

  Inheritance: AD Classification: NO_KNOWN Submitted by: ClinGen

ACMG classification

Our verdict: Pathogenic. The variant received 13 ACMG points.

• PS1: Transcript NM_000249.4 (MLH1) is affected with MISSENSE_VARIANT having same AA change as one Pathogenic present in ClinVar.
• PM1: In a hotspot region, there are 14 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 4 benign, 56 uncertain in NM_000249.4
• PM2: Very rare variant in population databases, with high coverage
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.977
• PP5: Variant 3-36993616-A-T is Pathogenic according to our data. Variant chr3-36993616-A-T is described in ClinVar as Conflicting_classifications_of_pathogenicity. ClinVar VariationId is 90335.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
MLH1	NM_000249.4	c.69A>T	p.Glu23Asp	missense_variant	Exon 1 of 19	ENST00000231790.8	NP_000240.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8	c.69A>T	p.Glu23Asp	missense_variant	Exon 1 of 19	1	NM_000249.4	ENSP00000231790.3

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 33

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Pathogenic:1 Uncertain:4

Revision: criteria provided, conflicting classifications

Submissions by phenotype

Hereditary cancer-predisposing syndrome Pathogenic:1 Uncertain:1

Jul 23, 2021

Color Diagnostics, LLC DBA Color Health

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This missense variant replaces glutamic acid with aspartic acid at codon 23 of the MLH1 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 have shown that the equivalent yeast variant had a mutator phenotype (PMID: 17210669), and that this variant had 25.2% mismatch repair activity in a yeast-based assay (PMID: 17510385). To our knowledge, this variant has not been reported in individuals affected with hereditary cancer in the literature. This variant has not been identified 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. -

Nov 29, 2021

Ambry Genetics

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The p.E23D variant (also known as c.69A>T), located in coding exon 1 of the MLH1 gene, results from an A to T substitution at nucleotide position 69. The glutamic acid at codon 23 is replaced by aspartic acid, an amino acid with highly similar properties. A functional study showed only 25% MMR activity in yeast with the p.E23D variant; however, immunohistochemistry analyses (IHC) indicated >75% expression when compared to wild type. These findings suggest that loss of MLH1 expression may not be detected by IHC in those with p.E23D (Takahashi et al. Cancer Res. 2007 May 15;67(10):4595-604). In another functional study, this variant demonstrated a mutator phenotype in a yeast reversion assay (Wanat JJ et al. Hum. Mol. Genet. 2007 Feb;16:445-52). This alteration has been observed in at least one individual with a personal and/or family history that is consistent with MLH1-related disease (Ambry internal data). This amino acid position is highly conserved in available vertebrate species. In addition, this alteration is predicted to be deleterious by in silico analysis. Based on the majority of available evidence to date, this variant is likely to be pathogenic. -

Colorectal cancer, hereditary nonpolyposis, type 2 Uncertain:1

Nov 08, 2016

Counsyl

Significance: Uncertain significance

Review Status: no assertion criteria provided

Collection Method: clinical testing

This submission and the accompanying classification are no longer maintained by the submitter. For more information on current observations and classification, please contact variantquestions@myriad.com. -

Lynch syndrome Uncertain:1

Sep 05, 2023

All of Us Research Program, National Institutes of Health

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This missense variant replaces glutamic acid with aspartic acid at codon 23 of the MLH1 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 have shown that the equivalent yeast variant had a mutator phenotype (PMID: 17210669), and that this variant had 25.2% mismatch repair activity in a yeast-based assay (PMID: 17510385). To our knowledge, this variant has not been reported in individuals affected with hereditary cancer in the literature. This variant has not been identified 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. -

Hereditary nonpolyposis colorectal neoplasms Uncertain:1

Dec 05, 2024

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This sequence change replaces glutamic acid, which is acidic and polar, with aspartic acid, which is acidic and polar, at codon 23 of the MLH1 protein (p.Glu23Asp). This variant is not present in population databases (gnomAD no frequency). This variant has not been reported in the literature in individuals affected with MLH1-related conditions. ClinVar contains an entry for this variant (Variation ID: 90335). Invitae Evidence Modeling incorporating data from in vitro experimental studies (internal data) indicates that this missense variant is expected to disrupt MLH1 function with a positive predictive value of 95%. Experimental studies have shown that this missense change affects MLH1 function (PMID: 17210669, 17510385). 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. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	1.0
BayesDel_addAF	Pathogenic	0.45	D
BayesDel_noAF	Pathogenic	0.41
CADD	Uncertain	24
DANN	Uncertain	1.0
DEOGEN2	Pathogenic	0.87	D
Eigen	Benign	-0.13
Eigen_PC	Benign	-0.15
FATHMM_MKL	Uncertain	0.91	D
LIST_S2	Pathogenic	1.0	D
M_CAP	Pathogenic	0.67	D
MetaRNN	Pathogenic	0.98	D
MetaSVM	Uncertain	0.059	D
MutationAssessor	Pathogenic	4.2	H
PhyloP100		1.2
PrimateAI	Pathogenic	0.80	T
PROVEAN	Benign	-2.3	N
REVEL	Pathogenic	0.85
Sift	Pathogenic	0.0	D
Sift4G	Uncertain	0.016	D
Polyphen		1.0	D
Vest4		0.81
MutPred		0.88		Loss of helix (P = 0.1706);
MVP		0.99
MPC		0.36
ClinPred		1.0	D
GERP RS		2.4
PromoterAI		0.069	Neutral
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.6
Varity_R		0.96
gMVP		0.80
Mutation Taster		=13/87	disease causing (ClinVar)

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.010		Details are displayed if max score is > 0.2

Find out detailed SpliceAI scores and Pangolin per-transcript scores at spliceailookup.broadinstitute.org

Other links and lift over

dbSNP: rs63750555; hg19: chr3-37035107;