rs63750266

Variant names:

• chr3-37001046-G-A
• rs63750266
• NM_000249.4:c.299G>A

Your query was ambiguous. Multiple possible variants found:

• chr3-37001046-G-A
• chr3-37001046-G-C
• chr3-37001046-G-T

Variant summary

Our verdict is Uncertain significance. The variant received 5 ACMG points: 5P and 0B. PM1 PM5 PP3

The NM_000249.4(MLH1):​c.299G>A​(p.Arg100Gln) variant causes a missense change involving the alteration of a conserved nucleotide. The variant allele was found at a frequency of 0.0000175 in 1,595,510 control chromosomes in the GnomAD database, with no homozygous occurrence. In-silico tool predicts a pathogenic outcome for this variant. 11/18 in silico tools predict a damaging outcome for this variant. Variant has been reported in ClinVar as Uncertain significance (★★). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. R100P) has been classified as Pathogenic.

• Frequency:
  • Genomes: 𝑓 0.000039 (0 hom., cov: 32)
  • Exomes 𝑓: 0.000015 (0 hom.)
• Consequence: MLH1 NM_000249.4 missense
• Clinical Significance: Uncertain significance criteria provided, multiple submitters, no conflicts U:15 O:1
• Conservation: PhyloP100: 9.42
• Publications: 22 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 5 ACMG points.

• PM1: In a hotspot region, there are 16 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 2 benign, 35 uncertain in NM_000249.4
• PM5: Other missense variant is known to change same aminoacid residue: Variant chr3-37001046-G-C is described in ClinVar as Pathogenic. ClinVar VariationId is 90133.Status of the report is reviewed_by_expert_panel, 3 stars.
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.833

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
MLH1	NM_000249.4	c.299G>A	p.Arg100Gln	missense_variant	Exon 3 of 19	ENST00000231790.8	NP_000240.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8	c.299G>A	p.Arg100Gln	missense_variant	Exon 3 of 19	1	NM_000249.4	ENSP00000231790.3

Frequencies

GnomAD3 genomes AF: 0.0000394 AC: 6 AN: 152100 Hom.: 0 Cov.: 32

Gnomad AFR AF: 0.0000241

Gnomad AMI AF: 0.00

Gnomad AMR AF: 0.00

Gnomad ASJ AF: 0.00

Gnomad EAS AF: 0.00

Gnomad SAS AF: 0.00

Gnomad FIN AF: 0.00

Gnomad MID AF: 0.00

Gnomad NFE AF: 0.0000735

Gnomad OTH AF: 0.00

GnomAD2 exomes AF: 0.0000159 AC: 4 AN: 251360 AF XY: 0.00000736

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.00

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.00

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.0000264

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.0000152 AC: 22 AN: 1443292 Hom.: 0 Cov.: 27 AF XY: 0.0000153 AC XY: 11 AN XY: 719186

African (AFR) AF: 0.00 AC: 0 AN: 33048

American (AMR) AF: 0.0000224 AC: 1 AN: 44682

Ashkenazi Jewish (ASJ) AF: 0.00 AC: 0 AN: 26038

East Asian (EAS) AF: 0.00 AC: 0 AN: 39520

South Asian (SAS) AF: 0.0000349 AC: 3 AN: 85876

European-Finnish (FIN) AF: 0.00 AC: 0 AN: 53394

Middle Eastern (MID) AF: 0.00 AC: 0 AN: 5726

European-Non Finnish (NFE) AF: 0.0000155 AC: 17 AN: 1095196

Other (OTH) AF: 0.0000167 AC: 1 AN: 59812

GnomAD4 genome AF: 0.0000394 AC: 6 AN: 152218 Hom.: 0 Cov.: 32 AF XY: 0.0000403 AC XY: 3 AN XY: 74432

African (AFR) AF: 0.0000241 AC: 1 AN: 41552

American (AMR) AF: 0.00 AC: 0 AN: 15286

Ashkenazi Jewish (ASJ) AF: 0.00 AC: 0 AN: 3468

East Asian (EAS) AF: 0.00 AC: 0 AN: 5182

South Asian (SAS) AF: 0.00 AC: 0 AN: 4830

European-Finnish (FIN) AF: 0.00 AC: 0 AN: 10586

Middle Eastern (MID) AF: 0.00 AC: 0 AN: 292

European-Non Finnish (NFE) AF: 0.0000735 AC: 5 AN: 67998

Other (OTH) AF: 0.00 AC: 0 AN: 2112

Alfa AF: 0.0000442 Hom.: 0

Bravo AF: 0.0000340

ESP6500AA AF: 0.00 AC: 0

ESP6500EA AF: 0.000116 AC: 1

ExAC AF: 0.0000165 AC: 2

ClinVar

Significance: Uncertain significance

Submissions summary: Uncertain:15 Other:1

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Colorectal cancer, hereditary nonpolyposis, type 2 Uncertain:4

Apr 01, 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. -

Feb 16, 2024

Baylor Genetics

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

- -

Mar 15, 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. -

Aug 14, 2024

St. Jude Molecular Pathology, St. Jude Children's Research Hospital

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The MLH1 c.299G>A (p.Arg100Gln) missense change has a maximum subpopulation frequency of 0.0033% in gnomAD v2.1.1 (https://gnomad.broadinstitute.org). The in silico tool REVEL predicts a deleterious effect on protein function for this variant, but functional studies have provided inconclusive evidence regarding its pathogenicity. One study using human embryonic stem cells reported normal protein stability, a moderate response to DNA repair, and a limited response to DNA damage (PMID: 36054288). Another study, conducted in a hybrid human-yeast MLH1 protein, indicated a partial loss of MMR function (PMID: 15475387). This variant has been reported in individuals with Lynch syndrome (PMID: 18809606, 22086678, 26845104), and it has also been reported in a large case-control study of breast cancer in 1 of 60,466 cases and 3 of 53,461 controls (PMID: 33471991). Another missense variant at the same amino acid residue, p.Arg100Pro, has been determined to be pathogenic by the International Society for Gastrointestinal Hereditary Tumours (InSiGHT). In summary, the evidence currently available is insufficient to determine the clinical significance of this variant. It has therefore been classified as of uncertain significance. -

not provided Uncertain:3

Apr 01, 2025

GeneDx

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Not observed at significant frequency in large population cohorts (gnomAD); In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; Published functional studies are inconclusive: protein stability, microsatellite instability, and resistance to MNNG similar to wildtype and intermediate loss of mismatch repair function (PMID: 36054288, 15475387); Observed in individuals with colorectal cancer with presence of MLH1 protein on tumor immunohistochemistry analysis (PMID: 18809606, 22086678, 26845104, 28765196); This variant is associated with the following publications: (PMID: 26333163, 12824425, 26845104, 12202775, 28466842, 25871441, 22086678, 15475387, 18809606, 28765196, 36054288, 29212164, 31697235, 32849802, 17510385, 17210669, 22753075, 16083711, 21120944, 31391288) -

Dec 20, 2023

Quest Diagnostics Nichols Institute San Juan Capistrano

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The MLH1 c.299G>A (p.Arg100Gln) variant has been reported in the published literature in individuals with colorectal cancer (PMIDs: 31391288 (2020), 28765196 (2017), 26845104 (2016), 22086678 (2012), and 18809606 (2008)). Additionally, this variant was reported as a somatic variant in a brain metastasis of a breast cancer patient (PMID: 29755676 (2018)). In a study measuring DNA mismatch repair capacity, this variant showed intermediate function (PMID: 15475387 (2004)). In a study measuring DNA mismatch repair capacity, this variant showed intermediate function (PMID: 15475387 (2004)). The frequency of this variant in the general population, 0.000026 (3/113664 chromosomes (Genome Aggregation Database, http://gnomad.broadinstitute.org)), is uninformative in the assessment of its pathogenicity. 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. -

Aug 01, 2020

CeGaT Center for Human Genetics Tuebingen

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

- -

not specified Uncertain:2

Jun 02, 2014

Department of Pathology and Laboratory Medicine, Sinai Health System

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

- -

Jun 07, 2024

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.299G>A (p.Arg100Gln) results in a conservative amino acid change located in the DNA mismatch repair protein family, N-terminal 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 1.6e-05 in 251360 control chromosomes. The available data on variant occurrences in the general population are insufficient to allow any conclusion about variant significance. c.299G>A has been reported in the literature in individuals affected with CRC or HBOC (Bartley_2011, Hampel_2008, Shirts_2016). These reports do not provide unequivocal conclusions about association of the variant with Hereditary Nonpolyposis Colorectal Cancer. One functional study shows 34-66% loss of MMR function but was performed in hybrid human-yeast genes (Ellison_2004). The following publications have been ascertained in the context of this evaluation (PMID: 15475387, 18809606, 22086678, 26845104). ClinVar contains an entry for this variant (Variation ID: 90132). Based on the evidence outlined above, the variant was classified as uncertain significance. -

Lynch syndrome Uncertain:2

Nov 20, 2015

University of Washington Department of Laboratory Medicine, University of Washington

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

- -

Jun 11, 2024

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 arginine with glutamine at codon 100 of the MLH1 protein. Computational prediction tools and conservation analyses suggest that this variant may have deleterious impact on protein structure and function. A functional study that examined this variant in the context of a human-yeast hybrid protein reported partial loss-of-function (PMID: 15475387). In another functional study done in human embryonic stem cells, the variant demonstrated a similar lack of response to DNA damage to a functionally abnormal variant, but resulted in limited microsatellite instability (PMID: 36054288). This variant has been reported in multiple individuals affected with colorectal cancer (PMID: 18809606, 22086678, 26845104). All tumors tested demonstrated no loss of mismatch repair protein via immunohistochemistry, but two tumor samples exhibited microsatellite instability (PMID: 18809606, 22086678, 26845104). This variant has also been reported in a homozygous or hemizygous individual who did not demonstrate expected clinical features (ClinVar: SCV000284057.10). This variant also has been reported in breast cancer case-control meta-analysis in 1 affected and 3 unaffected individuals (PMID: 33471991; LOVD DB-ID: MLH1_000129). This variant failed to exhibit a significant association with pancreatic cancer (PMID: 32980694). This variant has been identified in 4/251360 chromosomes in the general population by the Genome Aggregation Database (gnomAD). A different variant affecting the same position, p.Arg100Pro, is considered to be disease-causing (ClinVar variation ID: 90133), suggesting that arginine or similar amino acid at this position is important for the protein function. Although there is a suspicion that this variant may be associated with disease, additional studies are necessary to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance. -

Hereditary cancer-predisposing syndrome Uncertain:2

May 23, 2024

Ambry Genetics

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The p.R100Q variant (also known as c.299G>A), located in coding exon 3 of the MLH1 gene, results from a G to A substitution at nucleotide position 299. The arginine at codon 100 is replaced by glutamine, an amino acid with highly similar properties. This variant was previously reported in the germline of two individuals with MSI-H colorectal cancers who had no family history of colon cancer (Hampel H et al. J Clin Oncol. 2008; 26:5783-8; Bartley AN et al. Cancer Prev Res (Phila) 2012; 5:320-7). In a hybrid yeast-human experiment, this variant was previously found to be associated with a decrease in mismatch repair activity and was considered intermediate deficiency (Ellison AR et al. Nucleic Acids Res. 2004; 32:5321-38). This alteration has been classified as an uncertain variant 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/]). 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 available evidence, the clinical significance of this variant remains unclear. -

Apr 26, 2024

Color Diagnostics, LLC DBA Color Health

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This missense variant replaces arginine with glutamine at codon 100 of the MLH1 protein. Computational prediction tools and conservation analyses suggest that this variant may have deleterious impact on protein structure and function. A functional study that examined this variant in the context of a human-yeast hybrid protein reported partial loss-of-function (PMID: 15475387). In another functional study done in human embryonic stem cells, the variant demonstrated a similar lack of response to DNA damage to a functionally abnormal variant, but resulted in limited microsatellite instability (PMID: 36054288). This variant has been reported in multiple individuals affected with colorectal cancer (PMID: 18809606, 22086678, 26845104). All tumors tested demonstrated no loss of mismatch repair protein via immunohistochemistry, but two tumor samples exhibited microsatellite instability (PMID: 18809606, 22086678, 26845104). This variant has also been reported in a homozygous or hemizygous individual who did not demonstrate expected clinical features (ClinVar: SCV000284057.10). This variant also has been reported in breast cancer case-control meta-analysis in 1 affected and 3 unaffected individuals (PMID: 33471991; LOVD DB-ID: MLH1_000129). This variant failed to exhibit a significant association with pancreatic cancer (PMID: 32980694). This variant has been identified in 4/251360 chromosomes in the general population by the Genome Aggregation Database (gnomAD). A different variant affecting the same position, p.Arg100Pro, is considered to be disease-causing (ClinVar variation ID: 90133), suggesting that arginine or similar amino acid at this position is important for the protein function. Although there is a suspicion that this variant may be associated with disease, additional studies are necessary to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance. -

Muir-Torré syndrome;C1333991:Colorectal cancer, hereditary nonpolyposis, type 2;C5399763:Mismatch repair cancer syndrome 1 Uncertain:1

Dec 27, 2023

Fulgent Genetics, Fulgent Genetics

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

- -

Hereditary nonpolyposis colorectal neoplasms Uncertain:1

Jan 23, 2025

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Uncertain significance

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This sequence change replaces arginine, which is basic and polar, with glutamine, which is neutral and polar, at codon 100 of the MLH1 protein (p.Arg100Gln). This variant is present in population databases (rs63750266, gnomAD 0.003%). This missense change has been observed in individual(s) with Lynch syndrome (PMID: 18809606, 22086678, 26845104). This missense change has been observed to be homozygous or hemizygous in an individual who did not have the expected clinical features for that genetic result (internal data). ClinVar contains an entry for this variant (Variation ID: 90132). Invitae Evidence Modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) indicates that this missense variant is expected to disrupt MLH1 protein function with a positive predictive value of 95%. Experimental studies are conflicting or provide insufficient evidence to determine the effect of this variant on MLH1 function (PMID: 15475387). Studies have shown that this missense change is associated with inconclusive levels of altered splicing (internal data). 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. -

Colorectal cancer, hereditary nonpolyposis, type 2;C5399763:Mismatch repair cancer syndrome 1 Other:1

-

GenomeConnect - Invitae Patient Insights Network

Significance: not provided

Review Status: no classification provided

Collection Method: phenotyping only

Variant interpreted as Uncertain significance and reported on 11-01-2019 by Invitae. GenomeConnect-Invitae Patient Insights Network assertions are reported exactly as they appear on the patient-provided report from the testing laboratory. Registry team members make no attempt to reinterpret the clinical significance of the variant. Phenotypic details are available under supporting information. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.99
BayesDel_addAF	Pathogenic	0.40	D
BayesDel_noAF	Pathogenic	0.48
CADD	Pathogenic	34
DANN	Pathogenic	1.0
DEOGEN2	Uncertain	0.63	.;D
Eigen	Pathogenic	1.2
Eigen_PC	Pathogenic	1.1
FATHMM_MKL	Pathogenic	0.99	D
LIST_S2	Benign	0.41	T;T
M_CAP	Pathogenic	0.30	D
MetaRNN	Pathogenic	0.83	D;D
MetaSVM	Pathogenic	1.1	D
PhyloP100		9.4
PROVEAN	Uncertain	-3.5	D;D
REVEL	Pathogenic	0.73
Sift	Uncertain	0.0030	D;D
Sift4G	Uncertain	0.0050	D;D
Vest4		0.55
MVP		0.92
ClinPred		1.0	D
GERP RS		5.9
Varity_R		0.93
Mutation Taster		=27/73	disease causing

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.10		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: rs63750266; hg19: chr3-37042537;