3-37026008-G-T

Variant names:

• chr3-37026008-G-T
• rs267607825
• NM_000249.4:c.1409+1G>T

Variant summary

Our verdict is Pathogenic. Variant got 18 ACMG points: 18P and 0B. PVS1 PM2 PP5_Very_Strong

The NM_000249.4(MLH1):​c.1409+1G>T variant causes a splice donor, intron change involving the alteration of a non-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: 31)
• Consequence: MLH1 NM_000249.4 splice_donor, intron
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:8
• Conservation: PhyloP100: 1.97

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 Pathogenic. Variant got 18 ACMG points.

• PVS1: Splicing +-2 bp (donor or acceptor) variant, LoF is a know mechanism of disease, No cryptic splice site detected. Exon removal results in frameshift change.
• PM2: Very rare variant in population databases, with high coverage
• PP5: Variant 3-37026008-G-T is Pathogenic according to our data. Variant chr3-37026008-G-T is described in ClinVar as [Likely_pathogenic]. Clinvar id is 420226.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars. Variant chr3-37026008-G-T is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
MLH1	NM_000249.4	c.1409+1G>T		splice_donor_variant, intron_variant	Intron 12 of 18	ENST00000231790.8	NP_000240.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8	c.1409+1G>T		splice_donor_variant, intron_variant	Intron 12 of 18	1	NM_000249.4	ENSP00000231790.3

Frequencies

GnomAD3 genomes Cov.: 31

GnomAD4 exome Cov.: 34

GnomAD4 genome Cov.: 31

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:8

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Hereditary cancer-predisposing syndrome Pathogenic:3

May 13, 2024

Color Diagnostics, LLC DBA Color Health

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant causes a G>T nucleotide substitution at the +1 position of intron 12 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. To our knowledge, functional studies have not been reported for this variant. This variant has been reported in an individual whose tumor demonstrated loss of MLH1 and PMS2 protein via immunohistochemistry (ClinVar SCV002697829.2). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Different variants affecting the same splice donor site, c.1409+1G>A and c.1409+1G>C, are described to be disease-causing (ClinVar ID: 89718, 89719). Loss of MLH1 function is a known mechanism of disease. Based on the available evidence, this variant is classified as Pathogenic. -

Feb 05, 2022

Sema4, Sema4

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: curation

- -

Oct 28, 2021

Ambry Genetics

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The c.1409+1G>T intronic pathogenic mutation results from a G to T substitution one nucleotide after coding exon 12 of the MLH1 gene. This variant has been identified in a proband whose tumor demonstrated loss of MLH1 and PMS2 expression by immunohistochemistry with absent MLH1 promoter hypermethylation (Ambry internal data). This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). In silico splice site analysis predicts that this alteration will weaken the native splice donor site. Alterations that disrupt the canonical splice site are expected to cause aberrant splicing, resulting in an abnormal protein or a transcript that is subject to nonsense-mediated mRNA decay. As such, this alteration is classified as a disease-causing mutation. -

not provided Pathogenic:2

Apr 13, 2023

Quest Diagnostics Nichols Institute San Juan Capistrano

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant disrupts a canonical splice-donor site and is predicted to interfere with normal MLH1 mRNA splicing. It has not been reported in large, multi-ethnic general populations (http://gnomad.broadinstitute.org). This variant has not been reported in individuals in the published literature, however other variants at this same canonical splice-donor site (c.1409+1G>C and c.1409+1G>A) have been reported in individuals with Hereditary Non-polyposis Colorectal Cancer (HNPCC) (PMID: 17348456 (2007), PMID: 9322509 (1997), PMID: 16451135 (2006)). Based on the available information, this variant is classified as likely pathogenic. -

Nov 27, 2023

GeneDx

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Canonical splice site variant predicted to result in a null allele in a gene for which loss of function is a known mechanism of disease; Not observed at significant frequency in large population cohorts (gnomAD); Has not been previously published as pathogenic or benign to our knowledge; This variant is associated with the following publications: (PMID: 10200055, 16451135) -

Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:1

Jul 20, 2023

Myriad Genetics, Inc.

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant is considered 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. This variant is strongly associated with more severe personal and family histories of cancer, typical for individuals with pathogenic variants in this gene [PMID: 27363726]. -

Hereditary nonpolyposis colon cancer Pathogenic:1

Feb 28, 2022

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

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Variant summary: MLH1 c.1409+1G>T alters a conserved nucleotide located in a canonical splice-site and is predicted to affect mRNA splicing resulting in a significantly altered protein due to either exon skipping, shortening, or inclusion of intronic material. Several computational tools predict a significant impact on normal splicing: Five predict the variant abolishes the canonical 5' splicing donor site. However, these predictions have yet to be confirmed by functional studies. The variant was absent in 250196 control chromosomes. To our knowledge, no occurrence of c.1409+1G>T in individuals affected with Hereditary Nonpolyposis Colorectal Cancer/Lynch syndrome and no experimental evidence demonstrating its impact on protein function have been reported. However, two other nucleotide changes at the same location, namely c.1409+1G>A and c.1409+1G>C have been reported with an associated phenotype of Colorectal cancer, non-polyposis in the HGMD database, supporting the functional relevance of this variation. Three clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. All laboratories classified the variant as pathogenic/likely pathogenic. Based on the evidence outlined above, the variant was classified as likely pathogenic. -

Hereditary nonpolyposis colorectal neoplasms Pathogenic:1

Nov 06, 2024

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This sequence change affects a donor splice site in intron 12 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 individuals with Lynch syndrome (PMID: 9322509, 17348456; internal data). ClinVar contains an entry for this variant (Variation ID: 420226). 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 v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.63	D
BayesDel_noAF	Pathogenic	0.27
CADD	Pathogenic	32
DANN	Uncertain	0.99
Eigen	Pathogenic	0.98
Eigen_PC	Pathogenic	0.80
FATHMM_MKL	Uncertain	0.94	D
GERP RS		4.4

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.94
SpliceAI score (max)		0.93		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.93		Position offset: -1

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

Publications

LitVar

Other links and lift over

dbSNP: rs267607825; hg19: chr3-37067499; COSMIC: COSV99039975; COSMIC: COSV99039975;