3-37020308-A-G

Variant names:

• chr3-37020308-A-G
• rs267607805
• NM_000249.4:c.885-2A>G

Variant summary

Our verdict is Pathogenic. The variant received 18 ACMG points: 18P and 0B. PVS1 PM2 PP5_Very_Strong

The NM_000249.4(MLH1):​c.885-2A>G variant causes a splice acceptor, intron 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. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Likely pathogenic (★★★).

• Frequency: Genomes: not found (cov: 33)
• Consequence: MLH1 NM_000249.4 splice_acceptor, intron
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Likely pathogenic reviewed by expert panel P:10
• Conservation: PhyloP100: 8.88
• Publications: 6 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: Pathogenic. The variant received 18 ACMG points.

• PVS1: Splicing +-2 bp (donor or acceptor) variant, LoF is a know mechanism of disease, Cryptic splice site detected, with MaxEntScore 5.9, offset of 5, new splice context is: ttattttcctgacggtttAGaaa. Cryptic site results in frameshift change. If cryptic site found is not functional and variant results in exon loss, it results in frameshift change.
• PM2: Very rare variant in population databases, with high coverage
• PP5: Variant 3-37020308-A-G is Pathogenic according to our data. Variant chr3-37020308-A-G is described in ClinVar as Likely_pathogenic. ClinVar VariationId is 90421.Status of the report is reviewed_by_expert_panel, 3 stars.

Variant Effect in Transcripts

ACMG analysis was done for transcript: NM_000249.4. You can select a different transcript below to see updated ACMG assignments.

RefSeq Transcripts

Selected	Gene	Transcript	Tags	HGVSc	HGVSp	Effect	Exon Rank	Protein	UniProt
	MLH1	NM_000249.4	MANE Select	c.885-2A>G		splice_acceptor intron	N/A	NP_000240.1
	MLH1	NM_001354628.2		c.885-2A>G		splice_acceptor intron	N/A	NP_001341557.1
	MLH1	NM_001354629.2		c.786-2A>G		splice_acceptor intron	N/A	NP_001341558.1

Ensembl Transcripts

Selected	Gene	Transcript	Tags	HGVSc	HGVSp	Effect	Exon Rank	Protein	UniProt
	MLH1	ENST00000231790.8	TSL:1 MANE Select	c.885-2A>G		splice_acceptor intron	N/A	ENSP00000231790.3
	MLH1	ENST00000456676.7	TSL:1	c.885-2A>G		splice_acceptor intron	N/A	ENSP00000416687.3
	MLH1	ENST00000413740.2	TSL:1	c.885-2A>G		splice_acceptor intron	N/A	ENSP00000416476.2

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 33

Alfa AF: 0.00 Hom.: 0

ClinVar

Significance: Likely pathogenic

Submissions summary: Pathogenic:10

Revision: reviewed by expert panel

Submissions by phenotype

Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:2

Apr 13, 2022

Baylor Genetics

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Jul 18, 2023

Myriad Genetics, Inc.

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant is considered likely pathogenic. This variant occurs within a consensus splice junction and is predicted to result in abnormal mRNA splicing of either an out-of-frame exon or an in-frame exon necessary for protein stability and/or normal function.

Lynch syndrome Pathogenic:2

Jun 21, 2019

International Society for Gastrointestinal Hereditary Tumours (InSiGHT)

Significance: Likely pathogenic

Review Status: reviewed by expert panel

Collection Method: curation

Interrupts canonical donor splice site

Apr 02, 2019

Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The c.885-2A>G variant in MLH1 has been reported in 2 individuals with Lynch syndrome-related cancers (Clarke 2019, Terdiman 2001). It was absent from large population studies. This variant has also been reported in ClinVar (Variation ID: 90421). This variant occurs within the canonical splice site (+/- 1,2) and is predicted to cause altered splicing leading to an abnormal or absent protein. Loss of function of the MLH1 gene is an established disease mechanism in autosomal dominant Lynch syndrome. In summary, this variant meets criteria to be classified as pathogenic for autosomal dominant Lynch syndrome. ACMG/AMP Criteria applied: PVS1, PM2, PS4_Supporting.

not provided Pathogenic:2

Jul 16, 2020

Quest Diagnostics Nichols Institute San Juan Capistrano

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The variant disrupts a canonical splice site, and is therefore predicted to result in the loss of a functional protein. Found in at least one patient with expected phenotype for this gene, and not found in general population data.

May 31, 2022

GeneDx

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Canonical splice site variant in a gene for which loss-of-function is a known mechanism of disease; Not observed in large population cohorts (gnomAD); This variant is associated with the following publications: (PMID: 30264478, 11208710, 26648449, 28152038, 30729418, 30787465)

Hereditary cancer-predisposing syndrome Pathogenic:2

May 21, 2025

Ambry Genetics

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The c.885-2A>G intronic variant results from an A to G substitution two nucleotides upstream from coding exon 11 in the MLH1 gene. This variant has been previously reported in an individual from a high-risk colorectal family with a MSI-high colorectal tumor (Terdiman JP et al. Gastroenterology. 2001; 120:21-3). This variant was also identified in an individual diagnosed with early-onset MSI-H colorectal cancer that demonstrated absent MLH1/PMS2 expression on immunohistochemistry (IHC) (Ambry internal data). This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). This nucleotide position is highly conserved in available vertebrate species. In silico splice site analysis predicts that this alteration will weaken the native splice acceptor site and will result in the creation or strengthening of a novel splice acceptor site; however, direct evidence is insufficient at this time (Ambry internal data). Based on the majority of available evidence to date, this variant is likely to be pathogenic.

Jun 03, 2020

Color Diagnostics, LLC DBA Color Health

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant causes an A to G nucleotide substitution at the -2 position of intron 10 of the MLH1 gene. Splice site prediction tools suggest that this variant may have a significant impact on RNA splicing. Although this prediction has not been confirmed in published RNA studies, this variant is expected to result in an absent or disrupted protein product. This variant has been reported in individuals affected with Lynch syndrome-associated cancers (PMID: 11208710, 30729418). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Loss of MLH1 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Likely Pathogenic.

Muir-Torré syndrome;C1333991:Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:1

Dec 18, 2019

Department of Pathology and Laboratory Medicine, Sinai Health System

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Hereditary nonpolyposis colorectal neoplasms Pathogenic:1

Apr 10, 2024

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This sequence change affects an acceptor splice site in intron 10 of the MLH1 gene. It is expected to disrupt RNA splicing. Variants that disrupt the donor or acceptor splice site typically lead to a loss of protein function (PMID: 16199547), and loss-of-function variants in MLH1 are known to be pathogenic (PMID: 15713769, 24362816). This variant is not present in population databases (gnomAD no frequency). Disruption of this splice site has been observed in individual(s) with Lynch syndrome and a Lynch syndrome-associated cancer (PMID: 11208710; Invitae). In at least one individual the variant was observed to be de novo. Invitae’s Lynch syndrome clinical variant model, which takes into account the clinical and family history, age, sex, and reported ancestry of multiple individuals with this MLH1 variant, predicts that it is pathogenic with a positive predictive value of at least 99%. This is a validated machine learning model developed at Invitae that incorporates the clinical features of 1,370,736 individuals referred for testing at Invitae. This variant is also known as IVS10-2A>G. ClinVar contains an entry for this variant (Variation ID: 90421). Algorithms developed to predict the effect of sequence changes on RNA splicing suggest that this variant may disrupt the consensus splice site. For these reasons, this variant has been classified as Pathogenic.

Computational scores

Source: dbNSFP v4.9

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.63	D
BayesDel_noAF	Uncertain	0.13
CADD	Pathogenic	34
DANN	Uncertain	0.99
Eigen	Pathogenic	1.1
Eigen_PC	Pathogenic	0.94
FATHMM_MKL	Pathogenic	0.99	D
PhyloP100		8.9
GERP RS		4.0
PromoterAI		0.0036	Neutral
Mutation Taster		=0/100	disease causing (ClinVar)

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.93
SpliceAI score (max)		0.99		Details are displayed if max score is > 0.2
DS_AG_spliceai		0.68		Position offset: 7
DS_AL_spliceai		0.99		Position offset: 2

Find out detailed SpliceAI scores and Pangolin per-transcript scores at spliceailookup.broadinstitute.org

Other links and lift over

dbSNP: rs267607805; hg19: chr3-37061799; COSMIC: COSV51617154;