NM_000249.4:c.332C>T

Variant names:

• chr3-37004426-C-T
• rs63750539
• NM_000249.4:c.332C>T

Variant summary

Our verdict is Pathogenic. Variant got 16 ACMG points: 16P and 0B. PM2 PM5 PP3_Strong PP5_Very_Strong

The NM_000249.4(MLH1):​c.332C>T​(p.Ala111Val) variant causes a missense change involving the alteration of a 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 Likely pathogenic (★★★). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. A111P) has been classified as Likely pathogenic.

• Frequency: Genomes: not found (cov: 33)
• Consequence: MLH1 NM_000249.4 missense
• Clinical Significance: Likely pathogenic reviewed by expert panel P:6
• Conservation: PhyloP100: 7.59

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]

ACMG classification

Verdict is Pathogenic. Variant got 16 ACMG points.

• PM2: Very rare variant in population databases, with high coverage
• PM5: Other missense variant is known to change same aminoacid residue: Variant chrnull-null-null-null is described in UniProt as null.
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.956
• PP5: Variant 3-37004426-C-T is Pathogenic according to our data. Variant chr3-37004426-C-T is described in ClinVar as [Likely_pathogenic]. Clinvar id is 90171.Status of the report is reviewed_by_expert_panel, 3 stars. Variant chr3-37004426-C-T is described in Lovd as [Likely_pathogenic]. Variant chr3-37004426-C-T is described in Lovd as [Pathogenic]. Variant chr3-37004426-C-T is described in Lovd as [Benign].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
MLH1	NM_000249.4	c.332C>T	p.Ala111Val	missense_variant	Exon 4 of 19	ENST00000231790.8	NP_000240.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8	c.332C>T	p.Ala111Val	missense_variant	Exon 4 of 19	1	NM_000249.4	ENSP00000231790.3

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome Cov.: 30

GnomAD4 genome Cov.: 33

ClinVar

Significance: Likely pathogenic

Submissions summary: Pathogenic:6

Revision: reviewed by expert panel

Submissions by phenotype

Hereditary cancer-predisposing syndrome Pathogenic:2

Sep 20, 2018

Ambry Genetics

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The p.A111V pathogenic mutation (also known as c.332C>T), located in coding exon 4 of the MLH1 gene, results from a C to T substitution at nucleotide position 332. The alanine at codon 111 is replaced by valine, an amino acid with similar properties. This mutation has been identified in multiple families fulfilling Amsterdam Criteria II (AC II) for a clinical diagnosis of hereditary non-polyposis colorectal cancer (HNPCC) syndrome (Nomura S et al. Biochem. Biophys. Res. Commun. 2000 Apr; 271(1):120-9. Caldes T et al. Int. J. Cancer 2002 Apr; 98(5):774-9). In particular, this mutation was identified in one individual from a family meeting AC II whose tumor studies demonstrated microsatellite instability and loss of MLH1 and PMS2 proteins on immunohistochemistry (Sanchez de Abajo A et al. Oncogene. 2006 Mar 30;25(14):2124-30). In addition to the clinical data, functional analysis of this mutation demonstrated decreased MMR activity in vitro when compared to wildtype (Takahashi M et al. Cancer Res. 2007 May; 67(10):4595-604). Furthermore, this alteration has been classified as likely pathogenic by multifactorial analysis, which integrates the following lines of evidence to produce a quantitative likelihood of pathogenicity: in silico prediction models, segregation with disease, tumor characteristics, mutation co-occurrence, and functional assay results (Thompson B et al. Nat Genet. 2014 Feb;46(2):107-15; available at [www.insight-group.org/variants/classifications/]). Based on the available evidence, this alteration is classified as a pathogenic mutation. -

Nov 24, 2024

Molecular Diagnostics Laboratory, Catalan Institute of Oncology

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

PS3, PM2_Supporting, PP1_Moderate, PP3_Moderate, PM3_Supporting c.332C>T located in exon 4 of the MLH1 gene, is predicted to result in the substitution of alanine by valine at codon 111, p.(Ala111Val). It is not present in the population database gnomAD v2.1.1, non-cancer dataset (PM2_supporting). Computational tools for this variant suggests a deleterious effect of the variant on protein function (MAPP+PolyPhen-2 prior probability for pathogenicity: 0.958)(PP3_Moderate). This variant was analyzed in a clinically calibrated functional assay, the cell-free in vitro MMR activity (CIMRA) assay from Drost 2019 (PMID: 30504929), showing a functional odds for pathogenicity = 21.908 (PS3). Its defective MMR function was also evidenced in other studies (PMID: 17510385). The variant co-segregates in several individuals with clinical features of Lynch syndrome in a family with a LR=3.75 (PMID: 16083711) and others from our internal cohort (PP1_moderate). The variant c.332C>T has also been identified in homozygous state in a patient diagnosed with T-cell lymphoblastic lymphoma at 7months of age (internal data)(PM3_Supporting). Based on currently available information, the variant c.332C>T is classified as a pathogenic variant according to ACMG guidelines. -

Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:1

Jul 13, 2023

Myriad Genetics, Inc.

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant is considered likely pathogenic. Functional studies indicate this variant impacts protein function [PMID: 17510385, 30504929]. This variant is expected to disrupt protein structure [Myriad internal data]. This variant has been reported in multiple individuals with clinical features of gene-specific disease [PMID: 20007843, 11920650, 16451135, 28874130, 16288214, 30013564]. -

Carcinoma of colon Pathogenic:1

-

Department of Pathology and Laboratory Medicine, Sinai Health System

Significance: Likely pathogenic

Review Status: no assertion criteria provided

Collection Method: clinical testing

The p.Ala111Val variant has been reported in the literature in 4/594 proband chromosomes from individuals with HNPCC or suspected HNPCC, all of whom met either the Amsterdam criteria II or modified HNPCC criteria. It was not reported in any of the 800 control chromosomes evaluated (Caldes 2002, Chao 2008, Kurzawski 2006, Nomura 2000, Takahashi 2007). In one of these studies, all colorectal tumors belonging to a family with the variant had microsatellite instability at one of the loci evaluated (Caldes 2002). In a functional study that examined the MMR repair ability of the variant, the mutant was found to be have severely compromised repair capacity compared to the wild-type (only 25%). MLH1 expression was found to be between 25-75% (Takahashi 2007). This variant is listed in the dbSNP database (ID#:rs28897759) as coming from a clinical source, but no frequency information was provided, and so the prevalence of this variant in the population is not known. It is not listed in the LOVD or UMD databases. The p.Ala111 residue is conserved across mammals and computational analyses (PolyPhen, SIFT, AlignGVGD) suggest that the p.Ala111Val variant may impact the protein. However, this information is not predictive enough to assume pathogenicity. In addition, this individual was shown to have MLH1 deficient tumour by IHC, increasing the likelihood this variant may have clinical significance. In summary, based on the above information, the clinical significance of this variant cannot be determined with certainty at this time although we would lean towards a more pathogenic role for this variant. This variant is classified as predicted pathogenic. -

Lynch syndrome Pathogenic:1

Jun 21, 2019

International Society for Gastrointestinal Hereditary Tumours (InSiGHT)

Significance: Likely pathogenic

Review Status: reviewed by expert panel

Collection Method: curation

Multifactorial likelihood analysis posterior probability 0.95-0.99 -

Hereditary nonpolyposis colorectal neoplasms Pathogenic:1

Nov 15, 2024

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This sequence change replaces alanine, which is neutral and non-polar, with valine, which is neutral and non-polar, at codon 111 of the MLH1 protein (p.Ala111Val). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individuals with Lynch syndrome (PMID: 10777691, 11920650, 16451135, 17510385, 28449805; internal data). ClinVar contains an entry for this variant (Variation ID: 90171). 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: 17510385). This variant disrupts the p.Ala111 amino acid residue in MLH1. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 15475387). This suggests that this residue is clinically significant, and that variants that disrupt this residue are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.99
BayesDel_addAF	Pathogenic	0.48	D
BayesDel_noAF	Pathogenic	0.46
CADD	Pathogenic	30
DANN	Pathogenic	1.0
DEOGEN2	Pathogenic	0.95	D;.;D
Eigen	Pathogenic	0.82
Eigen_PC	Pathogenic	0.81
FATHMM_MKL	Pathogenic	0.99	D
LIST_S2	Pathogenic	0.99	D;D;D
M_CAP	Uncertain	0.20	D
MetaRNN	Pathogenic	0.96	D;D;D
MetaSVM	Uncertain	0.77	D
MutationAssessor	Pathogenic	3.9	H;.;.
PrimateAI	Uncertain	0.77	T
PROVEAN	Uncertain	-3.6	D;D;D
REVEL	Pathogenic	0.85
Sift	Uncertain	0.0020	D;D;D
Sift4G	Uncertain	0.0030	D;D;D
Polyphen		1.0	D;.;.
Vest4		0.96
MutPred		0.87		Loss of catalytic residue at A111 (P = 0.1586);.;.;
MVP		0.98
MPC		0.40
ClinPred		1.0	D
GERP RS		6.0
Varity_R		0.94
gMVP		0.92

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.0		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: rs63750539; hg19: chr3-37045917;