3-37014530-T-C

Variant names:

• chr3-37014530-T-C
• rs56250509
• NM_000249.4:c.776T>C

Variant summary

Our verdict is Uncertain significance. The variant received 2 ACMG points: 2P and 0B. PM1

The NM_000249.4(MLH1):​c.776T>C​(p.Leu259Ser) variant causes a missense change involving the alteration of a conserved nucleotide. The variant allele was found at a frequency of 0.00000626 in 1,437,850 control chromosomes in the GnomAD database, with no homozygous occurrence. 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. L259V) has been classified as Uncertain significance.

• Frequency:
  • Genomes: not found (cov: 33)
  • Exomes 𝑓: 0.0000063 (0 hom.)
• Consequence: MLH1 NM_000249.4 missense
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications U:9 B:2
• Conservation: PhyloP100: 7.68
• Publications: 15 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: Uncertain_significance. The variant received 2 ACMG points.

• PM1: In a hotspot region, there are 8 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 1 benign, 47 uncertain in NM_000249.4

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
MLH1	NM_000249.4	c.776T>C	p.Leu259Ser	missense_variant	Exon 9 of 19	ENST00000231790.8	NP_000240.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8	c.776T>C	p.Leu259Ser	missense_variant	Exon 9 of 19	1	NM_000249.4	ENSP00000231790.3

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD2 exomes AF: 0.0000199 AC: 5 AN: 250852 AF XY: 0.0000147

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.00

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.000272

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.00

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.00000626 AC: 9 AN: 1437850 Hom.: 0 Cov.: 26 AF XY: 0.00000418 AC XY: 3 AN XY: 716868

African (AFR) AF: 0.00 AC: 0 AN: 32994

American (AMR) AF: 0.00 AC: 0 AN: 44684

Ashkenazi Jewish (ASJ) AF: 0.00 AC: 0 AN: 25988

East Asian (EAS) AF: 0.000227 AC: 9 AN: 39586

South Asian (SAS) AF: 0.00 AC: 0 AN: 85760

European-Finnish (FIN) AF: 0.00 AC: 0 AN: 53392

Middle Eastern (MID) AF: 0.00 AC: 0 AN: 5722

European-Non Finnish (NFE) AF: 0.00 AC: 0 AN: 1090114

Other (OTH) AF: 0.00 AC: 0 AN: 59610

GnomAD4 genome Cov.: 33

Alfa AF: 0.00 Hom.: 0

Bravo AF: 0.0000227

ExAC AF: 0.0000412 AC: 5

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Uncertain:9 Benign:2

Revision: criteria provided, conflicting classifications

Submissions by phenotype

Colorectal cancer, hereditary nonpolyposis, type 2 Uncertain:3

Mar 13, 2023

Myriad Genetics, Inc.

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant is classified as a variant of uncertain significance as there is insufficient evidence to determine its impact on protein function and/or cancer risk.

Dec 18, 2015

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.

Mar 17, 2024

Baylor Genetics

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

not provided Uncertain:3

Feb 04, 2020

GeneDx

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; Published functional studies demonstrate no damaging effect: retains PMS2 binding ability (Wang 2012); Observed in an individual with a pediatric solid tumor in published literature (Chan 2018); This variant is associated with the following publications: (PMID: 30866919, 9057658, 24362816, 22252508, 30455982)

Jun 07, 2024

Quest Diagnostics Nichols Institute San Juan Capistrano

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The MLH1 c.776T>C (p.Leu259Ser) variant has been reported in the published literature in individuals with colorectal cancer (PMID: 37743058 (2023)), gastrointestinal stromal tumor (PMID: 35171259 (2022)), and breast cancer (PMID: 32091409 (2020), 33471991 (2021), see also LOVD (http://databases.lovd.nl/shared)). This variant is also found in reportedly healthy individuals (PMID: 33471991 (2021), see also LOVD (http://databases.lovd.nl/shared)). A yeast-based study showed this variant has neutral effects on PMS2 binding, however other MLH1 protein functions were not examined (PMID: 22252508 (2012)). The frequency of this variant in the general population, 0.00027 (5/18390 chromosomes in East Asian subpopulation (Genome Aggregation Database, http://gnomad.broadinstitute.org)), is higher than would generally be expected for pathogenic variants in this gene. Analysis of this variant using bioinformatics tools for the prediction of the effect of amino acid changes on protein structure and function yielded predictions that this variant is damaging. Based on the available information, we are unable to determine the clinical significance of this variant.

Jan 08, 2018

PreventionGenetics, part of Exact Sciences

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Hereditary cancer-predisposing syndrome Uncertain:1 Benign:1

Dec 19, 2022

Ambry Genetics

Significance: Likely benign

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This alteration is classified as likely benign based on a combination of the following: seen in unaffected individuals, population frequency, intact protein function, lack of segregation with disease, co-occurrence, RNA analysis, in silico models, amino acid conservation, lack of disease association in case-control studies, and/or the mechanism of disease or impacted region is inconsistent with a known cause of pathogenicity.

Nov 16, 2020

Color Diagnostics, LLC DBA Color Health

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This missense variant replaces leucine with serine at codon 259 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). This variant protein retains the ability to bind PMS2 in a yeast two-hybrid assay (PMID: 22252508). This variant has been observed in at least two unrelated individuals with a likely pathogenic MLH1 covariant, however, the in cis or in trans phasing of the two variants is unknown (UMD record ID: 1120; Color internal data). This variant has been identified in 5/250852 chromosomes 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.

not specified Uncertain:1

Aug 13, 2018

Women's Health and Genetics/Laboratory Corporation of America, LabCorp

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Variant summary: MLH1 c.776T>C (p.Leu259Ser) results in a non-conservative amino acid change located in the DNA mismatch repair protein, S5 domain 2-like domain of the encoded protein sequence. Four of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 2e-05 in 245718 control chromosomes (gnomAD), predominantly at a frequency of 0.00029 within the East Asian subpopulation in the gnomAD database. The available data on variant occurrences in the general population are insufficient to allow any conclusion about variant significance. The variant, c.776T>C has been reported in the literature in a CML cell line (Hangaishi_1997). This report does not provide an unequivocal conclusion about association of the variant with Lynch Syndrome. Co-occurrences with other pathogenic variant(s) have been reported (MLH1 c.1989+1G>T (2X)), providing supporting evidence for a benign role. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. Five ClinVar submissions from other clinical diagnostic laboratories (evaluation after 2014) cite the variant as "uncertain significance." Based on the evidence outlined above, the variant was classified as VUS-possibly benign.

Lynch syndrome Uncertain:1

Aug 15, 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 leucine with serine at codon 259 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). This variant protein retains the ability to bind PMS2 in a yeast two-hybrid assay (PMID: 22252508). This variant has been observed in at least two unrelated individuals with a likely pathogenic MLH1 covariant, however, the in cis or in trans phasing of the two variants is unknown (UMD record ID: 1120; Color internal data). This variant has been identified in 5/250852 chromosomes 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 Benign:1

Dec 02, 2024

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Likely benign

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.83
BayesDel_addAF	Uncertain	0.097	D
BayesDel_noAF	Pathogenic	0.20
CADD	Pathogenic	26
DANN	Uncertain	1.0
DEOGEN2	Uncertain	0.76	D;.;.;.;.;.;.
Eigen	Uncertain	0.40
Eigen_PC	Uncertain	0.49
FATHMM_MKL	Pathogenic	0.99	D
LIST_S2	Uncertain	0.96	D;D;.;.;.;D;D
M_CAP	Benign	0.066	D
MetaRNN	Uncertain	0.72	D;D;D;D;D;D;D
MetaSVM	Benign	-0.61	T
MutationAssessor	Benign	1.8	L;.;.;.;.;.;.
PhyloP100		7.7
PrimateAI	Uncertain	0.67	T
PROVEAN	Pathogenic	-5.4	D;D;D;D;D;D;D
REVEL	Pathogenic	0.82
Sift	Pathogenic	0.0	D;D;D;D;D;D;D
Sift4G	Uncertain	0.045	D;D;D;D;D;T;D
Vest4		0.83
ClinPred		0.77	D
GERP RS		5.4
Varity_R		0.92
gMVP		0.86
Mutation Taster		=10/90	disease causing

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

Other links and lift over

dbSNP: rs56250509; hg19: chr3-37056021; COSMIC: COSV51615676;