rs63750271

chr3-37040279-A-Cchr3-37040279-A-Gchr3-37040279-A-T

Variant summary

Our verdict is Likely pathogenic. Variant got 8 ACMG points: 8P and 0B. PM1 PM2 PP3_Strong

The NM_000249.4(MLH1):c.1652A>C(p.Asn551Thr) 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 Conflicting classifications of pathogenicity (no stars). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. N551S) has been classified as Benign.

• Frequency: Genomes: not found (cov: 32)
• Consequence: MLH1 NM_000249.4 missense
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:1 U:2
• Conservation: PhyloP100: 9.08

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 Likely_pathogenic. Variant got 8 ACMG points.

• PM1: In a hotspot region, there are 5 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 0 benign, 15 uncertain in NM_000249.4
• PM2: Very rare variant in population databases, with high coverage
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.971

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
MLH1	NM_000249.4	c.1652A>C	p.Asn551Thr	missense_variant	14/19	ENST00000231790.8

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
MLH1	ENST00000231790.8	c.1652A>C	p.Asn551Thr	missense_variant	14/19	1	NM_000249.4	P1

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Cov.: 30

GnomAD4 genome Cov.: 32

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Pathogenic:1 Uncertain:2

Revision: criteria provided, conflicting classifications

Submissions by phenotype

Hereditary cancer-predisposing syndrome Pathogenic:1 Uncertain:1

Uncertain significance, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Jan 01, 2022	This missense variant replaces asparagine with threonine at codon 551 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 mutant protein shows partially reduced expression levels, mildly reduced mismatch repair activity, and normal interaction with PMS2 (PMID: 17510385, 20533529, 23403630). This variant has been reported in individuals affected with Lynch syndrome or Lynch syndrome-associated cancers (PMID: 9833759, 25133505, 25712738). This variant has been shown to segregate with colorectal cancer in several individuals from a consanguineous family (PMID: 9399661). Three additional affected family members from this family were not genotyped. This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). The available functional evidence is inconsistent with the clinical findings that suggest this variant may have a pathogenic role. Additional evidence is necessary to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance. -
Likely pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	Sep 25, 2018	The p.N551T variant (also known as c.1652A>C), located in coding exon 14 of the MLH1 gene, results from an A to C substitution at nucleotide position 1652. The asparagine at codon 551 is replaced by threonine, an amino acid with similar properties. This alteration was identified in a Swiss proband diagnosed with colorectal cancer as well as stomach cancer at age 46 whose family history met Amsterdam criteria for Lynch syndrome and tumor demonstrated high microsatellite instability (MSI-H) (Hutter P et al. Int. J. Cancer, 1998 Dec;78:680-4). This alteration was also identified in a Saudi Arabian proband with a MSI-H colorectal tumor who met Amsterdam criteria for Lynch syndrome (Siraj AK et al. Cancer, 2015 Jun;121:1762-71). In a large consanguineous Turkish family that met Amsterdam criteria for Lynch syndrome, this alteration segregated with disease in 5 individuals affected with colorectal cancer, two of whom were obligate carriers (Wang Q et al. Int. J. Cancer, 1997 Dec;73:831-6). In two in vitro mismatch repair (MMR) complementation assays, this variant demonstrated proficient MMR activity and interaction with PMS2 was intact in a co-immunoprecipitation study (Takahashi M et al. Cancer Res., 2007 May;67:4595-604; Kosinski J et al. Hum. Mutat., 2010 Aug;31:975-82; Hinrichsen I et al. Clin. Cancer Res., 2013 May;19:2432-41). However, in reversion assays performed in yeast, no dominant mutator effect was seen with this variant (Takahashi M et al. Cancer Res., 2007 May;67:4595-604). Also, although expression levels were variable, several studies report reduced relative expression in human cell lines by Western blotting for this variant (Takahashi M et al. Cancer Res., 2007 May;67:4595-604; Kosinski J et al. Hum. Mutat., 2010 Aug;31:975-82; Hinrichsen I et al. Clin. Cancer Res., 2013 May;19:2432-41). Furthermore, this variant did not demonstrate a deleterious effect on splicing by mini gene assay or by RT-PCR using patient RNA (Auclair J et al. Hum. Mutat., 2006 Feb;27:145-54; Tournier I et al. Hum. Mutat., 2008 Dec;29:1412-24). This variant was not reported in population-based cohorts in the Genome Aggregation Database (gnomAD). This amino acid position is highly conserved in available vertebrate species. In addition, this alteration is predicted to be deleterious by in silico analysis. In addition, this alteration is predicted to be deleterious by MAPP-MMR in silico analysis (Chao EC et al. Hum. Mutat. 2008 Jun;29:852-60). Based on the majority of available evidence to date, this variant is likely to be pathogenic. -

Hereditary nonpolyposis colorectal neoplasms Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Invitae

Nov 17, 2023

This sequence change replaces asparagine, which is neutral and polar, with threonine, which is neutral and polar, at codon 551 of the MLH1 protein (p.Asn551Thr). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individual(s) with clinical features of Lynch syndrome (PMID: 9399661, 9833759, 25133505, 33309985). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 89819). 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) has been performed at Invitae for this missense variant, however the output from this modeling did not meet the statistical confidence thresholds required to predict the impact of this variant on MLH1 protein function. Studies have shown that this missense change alters MLH1 gene expression (PMID: 20533529, 23403630, 31784484). RNA analysis performed to evaluate the impact of this missense change on mRNA splicing indicates it does not significantly alter splicing (PMID: 16395668). 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	Uncertain	0.54
BayesDel_addAF	Pathogenic	0.30	D
BayesDel_noAF	Pathogenic	0.20
Cadd	Uncertain	25
Dann	Uncertain	0.99
DEOGEN2	Pathogenic	0.93	D;.;.;.;.;.;D
Eigen	Uncertain	0.53
Eigen_PC	Uncertain	0.51
FATHMM_MKL	Pathogenic	0.99	D
LIST_S2	Uncertain	0.97	D;D;.;.;.;D;D
M_CAP	Uncertain	0.26	D
MetaRNN	Pathogenic	0.97	D;D;D;D;D;D;D
MetaSVM	Uncertain	0.26	D
MutationAssessor	Pathogenic	3.3	M;.;.;.;.;.;.
MutationTaster	Benign	1.0	D;D;D;D;D;D
PrimateAI	Uncertain	0.77	T
PROVEAN	Uncertain	-4.1	D;D;D;D;D;D;D
REVEL	Pathogenic	0.80
Sift	Uncertain	0.011	D;D;D;D;D;D;D
Sift4G	Uncertain	0.022	D;D;D;D;D;D;D
Polyphen		0.99	D;.;.;.;.;.;.
Vest4		0.95
MutPred		0.91		Loss of sheet (P = 0.007);.;.;.;.;.;.;
MVP		0.95
MPC		0.38
ClinPred		0.98	D
GERP RS		5.7
Varity_R		0.91
gMVP		0.69

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.020		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: rs63750271; hg19: chr3-37081770;