NM_000249.4:c.2252_2253dupAA

Variant names:

• chr3-37050632-C-CAA
• rs267607901
• NM_000249.4:c.2252_2253dupAA

Variant summary

Our verdict is Likely pathogenic. The variant received 6 ACMG points: 6P and 0B. PVS1_Strong PM2

The NM_000249.4(MLH1):​c.2252_2253dupAA​(p.Val752LysfsTer32) variant causes a frameshift change involving the alteration of a conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. Variant has been reported in ClinVar as Uncertain significance (★★★). Synonymous variant affecting the same amino acid position (i.e. V752V) has been classified as Likely benign.

• Frequency: Genomes: not found (cov: 32)
• Consequence: MLH1 NM_000249.4 frameshift
• Clinical Significance: Uncertain significance reviewed by expert panel P:7 U:1
• Conservation: PhyloP100: 9.00
• Publications: 11 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: Likely_pathogenic. The variant received 6 ACMG points.

• PVS1: Loss of function variant, product does not undergo nonsense mediated mRNA decay. Variant is located in the 3'-most exon, not predicted to undergo nonsense mediated mRNA decay. There are 15 pathogenic variants in the truncated region.
• PM2: Very rare variant in population databases, with high coverage

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.2252_2253dupAA	p.Val752LysfsTer32	frameshift	Exon 19 of 19	NP_000240.1
	MLH1	NM_001354628.2		c.2159_2160dupAA	p.Val721LysfsTer32	frameshift	Exon 18 of 18	NP_001341557.1
	MLH1	NM_001354629.2		c.2153_2154dupAA	p.Val719LysfsTer32	frameshift	Exon 18 of 18	NP_001341558.1

Ensembl Transcripts

Selected	Gene	Transcript	Tags	HGVSc	HGVSp	Effect	Exon Rank	Protein	UniProt
	MLH1	ENST00000231790.8	TSL:1 MANE Select	c.2252_2253dupAA	p.Val752LysfsTer32	frameshift	Exon 19 of 19	ENSP00000231790.3
	MLH1	ENST00000456676.7	TSL:1	c.2045_2046dupAA	p.Val683LysfsTer32	frameshift	Exon 17 of 17	ENSP00000416687.3
	MLH1	ENST00000413740.2	TSL:1	c.1816_1817dupAA	p.Arg609fs	frameshift	Exon 15 of 15	ENSP00000416476.2

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 32

ClinVar

Significance: Uncertain significance

Submissions summary: Pathogenic:7 Uncertain:1

Revision: reviewed by expert panel

Submissions by phenotype

Hereditary cancer-predisposing syndrome Pathogenic:2

Jun 28, 2019

Color Diagnostics, LLC DBA Color Health

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant inserts 2 nucleotides in exon 19 of the MLH1 gene, creating a frameshift and premature translation stop signal. This variant is expected to result in an absent or non-functional protein product. Splice site prediction tools suggest that this variant may not impact RNA splicing. To our knowledge, functional studies have not been performed for this variant. This variant has been reported in multiple hereditary nonpolyposis colorectal cancer (HNPCC) families (PMID: 15849733, 15926618, 18931482). This variant has been reported to segregate with disease (InSiGHT database; http://insight-database.org/classifications/index.html?gene=MLH1&variant=c.2252_2253dup). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Based on the available evidence, this variant is classified as Likely Pathogenic.

Aug 26, 2025

Ambry Genetics

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The c.2252_2253dupAA variant, located in coding exon 19 of the MLH1 gene, results from a duplication of AA at nucleotide positions 2252 and 2253, causing a translational frameshift with a predicted alternate stop codon (p.Val752Lysfs*32). This alteration occurs at the 3' terminus of theMLH1 gene, is not expected to trigger nonsense-mediated mRNAdecay and results in the elongation of the protein by 26 amino acids. This frameshift impacts the last 5amino acids of the native protein. However, frameshifts are typically deleterious in nature and the impacted region is critical for protein function (Ambry internal data). This variant has been identified in HNPCC patients who fulfilled Bethesda and/or Amsterdam II guidelines with tumor analysis revealing MSI-H and loss of MLH1 and/or PMS2 by IHC (Pistorius SR et al. Int J Colorectal Dis. 2000 Nov;15(5-6):255-63; Sheng JQ et al. Cytogenet. Genome Res. 2008;122(1):22-7; Wolf B et al. Wien. Klin. Wochenschr. 2005 Apr;117(7-8):269-77; Mangold E et al. Int. J. Cancer. 2005 Sep;116(5):692-702; Fu L et al. Cell Oncol (Dordr). 2013 Jun;36:225-31; Rossi BM et al. BMC Cancer. 2017 Sep;17:623; Ambry internal data). Structural analysis shows that this alteration perturbs a known functional domain responsible for binding to PMS2 and removes a cysteine residue shown to be involved in metal binding as part of a functional domain in PMS2 (Mohd AB et al. DNA Repair (Amst.) 2006 Mar;5(3):347-61; Smith CE et al. PLoS Genet. 2013 Oct;9(10):e1003869). This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). Based on the majority of available evidence to date, this variant is likely to be pathogenic.

Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:1

Feb 25, 2021

Baylor Genetics

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Hereditary nonpolyposis colon cancer Pathogenic:1

Jan 23, 2023

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

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Variant summary: MLH1 c.2252_2253dupAA (p.Val752LysfsX32) causes a frameshift which results in an extension of the protein in the carboxy terminal homology domain (Sui_2019). The variant was absent in 251302 control chromosomes. c.2252_2253dupAA has been reported in the literature in multiple individuals affected with Hereditary Nonpolyposis Colorectal Cancer (Dominguez-Valentin_2013, Sheng_2008, Mangold_2005, Wolf_2005, Sui_2019). These data indicate that the variant is very likely to be associated with disease. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. Four (other) submitters including an expert panel have provided clinical-significance assessments for this variant to ClinVar after 2014, and classified it as pathogenic/likely pathogenic (n=3) or VUS (n=1). Based on the evidence outlined above, the variant was classified as pathogenic.

not provided Pathogenic:1

Aug 11, 2023

GeneDx

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Frameshift variant predicted to result in protein extension, as the last 5 amino acids are replaced with 31 different amino acids, and other loss-of-function variants have been reported downstream; Not observed at significant frequency in large population cohorts (gnomAD); Also known as c.2253_2254insAA; Observed in individuals with personal and/or family history of Lynch syndrome-related cancers (Pistorius et al., 2000; Mangold et al., 2005; Wolf et al., 2005; Sheng et al., 2008; Dominguez-Valentin et al., 2013; Sui et al., 2019); This variant is associated with the following publications: (PMID: 11151427, 16338176, 24204293, 12799449, 20533529, 22753075, 28874130, 18931482, 15926618, 30614234, 31830689, 23640085, 24344984, 15849733)

Hereditary nonpolyposis colorectal neoplasms Pathogenic:1

Jan 14, 2025

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This sequence change results in a frameshift in the MLH1 gene (p.Val752Lysfs*32). While this is not anticipated to result in nonsense mediated decay, it is expected to disrupt the last 5 amino acid(s) of the MLH1 protein. This variant is not present in population databases (gnomAD no frequency). This variant has been observed in individual(s) with clinical features of Lynch syndrome (PMID: 11151427, 15849733, 15926618, 18931482, 23640085, 24344984). This variant is also known as Ins AA at 2254 or c.2253_2254insAA. ClinVar contains an entry for this variant (Variation ID: 90102). This variant disrupts a region of the MLH1 protein in which other variant(s) (p.*757Leuext*33) have been determined to be pathogenic (PMID: 8128251, 9697702, 12810663, 14985405). This suggests that this is a clinically significant region of the protein, and that variants that disrupt it are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic.

Hereditary cancer Pathogenic:1

Aug 12, 2024

Mendelics

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Lynch syndrome Uncertain:1

Jun 21, 2019

International Society for Gastrointestinal Hereditary Tumours (InSiGHT)

Significance: Uncertain significance

Review Status: reviewed by expert panel

Collection Method: curation

Insufficient evidence: extends protein by 26 amino acids

Computational scores

Source: dbNSFP v4.9

Name	Calibrated prediction	Score	Prediction
PhyloP100		9.0
Mutation Taster		=0/200	disease causing

Splicing

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

Other links and lift over

dbSNP: rs267607901; hg19: chr3-37092123;