3-37008812-A-G

Variant names:

• chr3-37008812-A-G
• rs267607753
• NM_000249.4:c.454-2A>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.454-2A>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: 32)
• Consequence: MLH1 NM_000249.4 splice_acceptor, intron
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Likely pathogenic reviewed by expert panel P:3
• Conservation: PhyloP100: 8.15
• Publications: 2 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 5.1, offset of -41, new splice context is: tcttgggttttattttcaAGtac. 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 3-37008812-A-G is Pathogenic according to our data. Variant chr3-37008812-A-G is described in ClinVar as Likely_pathogenic. ClinVar VariationId is 90232.Status of the report is reviewed_by_expert_panel, 3 stars.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
MLH1	NM_000249.4	c.454-2A>G		splice_acceptor_variant, intron_variant	Intron 5 of 18	ENST00000231790.8	NP_000240.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8	c.454-2A>G		splice_acceptor_variant, intron_variant	Intron 5 of 18	1	NM_000249.4	ENSP00000231790.3

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Cov.: 30

GnomAD4 genome Cov.: 32

Alfa AF: 0.00000534 Hom.: 0

ClinVar

Significance: Likely pathogenic

Submissions summary: Pathogenic:3

Revision: reviewed by expert panel

Submissions by phenotype

Lynch syndrome Pathogenic:1

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

Hereditary nonpolyposis colorectal neoplasms Pathogenic:1

Mar 30, 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 5 of the MLH1 gene. RNA analysis indicates that disruption of this splice site induces altered splicing and may result in an absent or disrupted protein product. This variant is not present in population databases (gnomAD no frequency). Disruption of this splice site has been observed in individual(s) with clinical features of Lynch syndrome (PMID: 7584997, 8940269, 10829038, 14574010, 23544471, 27601186; Invitae). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 90232). Studies have shown that disruption of this splice site results in skipping of exon 6 and introduces a premature termination codon (PMID: 7584997, 15235038, 16395668). The resulting mRNA is expected to undergo nonsense-mediated decay. For these reasons, this variant has been classified as Pathogenic.

Hereditary cancer-predisposing syndrome Pathogenic:1

Jul 14, 2022

Ambry Genetics

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The c.454-2A>G intronic pathogenic mutation results from an A to G substitution two nucleotides upstream from coding exon 6 in the MLH1 gene. RNA studies have demonstrated that this alteration results in coding exon 6 skipping (Auclair J et al. Hum. Mutat. 2006 Feb; 27(2):145-54; Ambry internal data). In addition to the report in the literature, 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.

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.63	D
BayesDel_noAF	Pathogenic	0.36
CADD	Pathogenic	30
DANN	Uncertain	0.99
DEOGEN2	Benign	0.0	.;.;.
Eigen	Pathogenic	1.2
Eigen_PC	Pathogenic	1.0
FATHMM_MKL	Pathogenic	1.0	D
LIST_S2	Benign	0.0	.;.;.
MetaRNN	Benign	0.0	.;.;.
MutationAssessor	Benign	0.0	.;.;.
PhyloP100		8.2
PROVEAN	Benign	0.0	.;.;.
REVEL	Benign	0.0
Sift	Pathogenic	0.0	.;.;.
Sift4G	Pathogenic	0.0	.;.;.
Vest4		0.0
GERP RS		5.9
Mutation Taster		=0/100	disease causing (ClinVar)

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.95
Splicevardb		3.0
SpliceAI score (max)		0.98		Details are displayed if max score is > 0.2
DS_AL_spliceai		0.98		Position offset: 2

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

Other links and lift over

dbSNP: rs267607753; hg19: chr3-37050303;