3-36993616-A-T

Position:

• chr3-36993616-A-T

Variant summary

Our verdict is Likely pathogenic. Variant got 6 ACMG points: 6P and 0B. PM2 PP3_Strong

The NM_001167617.3(MLH1):​c.-448A>T variant causes a 5 prime UTR premature start codon gain 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).

• Frequency: Genomes: not found (cov: 33)
• Consequence: MLH1 NM_001167617.3 5_prime_UTR_premature_start_codon_gain
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:1 U:4
• Conservation: PhyloP100: 1.17

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 (HGNC:):

ACMG classification

Verdict is Likely_pathogenic. Variant got 6 ACMG points.

• PM2: Very rare variant in population databases, with high coverage
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.977

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
MLH1	NM_000249.4	c.69A>T	p.Glu23Asp	missense_variant	1/19	ENST00000231790.8	NP_000240.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8	c.69A>T	p.Glu23Asp	missense_variant	1/19	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

Likely pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	Nov 29, 2021	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. -
Uncertain significance, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Jul 23, 2021	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. -

Colorectal cancer, hereditary nonpolyposis, type 2 Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Counsyl

Nov 08, 2016

- -

Lynch syndrome Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

All of Us Research Program, National Institutes of Health

Sep 05, 2023

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

Uncertain significance, criteria provided, single submitter

clinical testing

Labcorp Genetics (formerly Invitae), Labcorp

Jan 03, 2023

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. Experimental studies have shown that this missense change affects MLH1 function (PMID: 17210669, 17510385). 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 expected to disrupt MLH1 protein function. ClinVar contains an entry for this variant (Variation ID: 90335). This variant has not been reported in the literature in individuals affected with MLH1-related conditions. This variant is not present in population databases (gnomAD no frequency). 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). -

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
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
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.6
Varity_R		0.96
gMVP		0.80

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

Publications

LitVar

Other links and lift over

dbSNP: rs63750555; hg19: chr3-37035107;