NM_000249.4:c.1896G>T

Variant names:

• chr3-37047683-G-T
• rs63751632
• NM_000249.4:c.1896G>T

Variant summary

Our verdict is Pathogenic. The variant received 16 ACMG points: 16P and 0B. PM1 PM2 PP3_Strong PP5_Very_Strong

The NM_000249.4(MLH1):​c.1896G>T​(p.Glu632Asp) variant causes a missense, splice region change. The variant was absent in control chromosomes in GnomAD project. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Likely pathogenic (★★). Another nucleotide change resulting in the same amino acid substitution has been previously reported as Likely pathogenic in ClinVar. Another variant affecting the same amino acid position, but resulting in a different missense (i.e. E632G) has been classified as Uncertain significance.

• Frequency: Genomes: not found (cov: 31)
• Consequence: MLH1 NM_000249.4 missense, splice_region
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Likely pathogenic criteria provided, multiple submitters, no conflicts P:2
• Conservation: PhyloP100: 6.64
• 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 16 ACMG points.

• PM1: In a hotspot region, there are 10 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 2 benign, 42 uncertain in NM_000249.4
• PM2: Very rare variant in population databases, with high coverage
• PP3: Splicing predictors support a deleterious effect. Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF, max_spliceai. No scorers claiming Uncertain. No scorers claiming Benign.
• PP5: Variant 3-37047683-G-T is Pathogenic according to our data. Variant chr3-37047683-G-T is described in ClinVar as Likely_pathogenic. ClinVar VariationId is 1782160.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 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.1896G>T	p.Glu632Asp	missense splice_region	Exon 16 of 19	NP_000240.1
	MLH1	NM_001354628.2		c.1896G>T	p.Glu632Asp	missense splice_region	Exon 16 of 18	NP_001341557.1
	MLH1	NM_001354629.2		c.1797G>T	p.Glu599Asp	missense splice_region	Exon 15 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.1896G>T	p.Glu632Asp	missense splice_region	Exon 16 of 19	ENSP00000231790.3
	MLH1	ENST00000456676.7	TSL:1	c.1896G>T	p.Glu632Asp	missense splice_region	Exon 16 of 17	ENSP00000416687.3
	MLH1	ENST00000458205.6	TSL:1	c.1173G>T	p.Glu391Asp	missense splice_region	Exon 17 of 20	ENSP00000402667.2

Frequencies

GnomAD3 genomes Cov.: 31

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 31

ClinVar

Significance: Likely pathogenic

Submissions summary: Pathogenic:2

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:1

Nov 06, 2023

Myriad Genetics, Inc.

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant is considered likely pathogenic. mRNA analysis has demonstrated abnormal mRNA splicing occurs [PMID: 8571956].

Hereditary cancer-predisposing syndrome Pathogenic:1

Mar 15, 2021

Ambry Genetics

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The p.E632D variant (also known as c.1896G>T), located in coding exon 16 of the MLH1 gene, results from a G to T substitution at nucleotide position 1896. The amino acid change results in glutamic acid to aspartic acid at codon 632, an amino acid with highly similar properties. However, this change occurs in the last base pair of coding exon 16, which makes it likely to have some effect on normal mRNA splicing. A different variant at the same nucleotide position, c.1896G>A, has been classified as pathogenic based on identification in Lynch syndrome families and reportedly demonstrated aberrant splicing by RT-PCR analysis (Wijnen J et al. Am J Hum Genet. 1996 Feb;58(2):300-7; Wijnen J et al. Am J Hum Genet. 1997 Aug;61(2):329-35; Wagner A et al. J Med Genet. 2002 Nov;39(11):833-7; Ramsoekh D et al. Gut. 2008 Nov;57(11):1539-44; van Lier MG et al. J Pathol. 2012 Apr;226(5):764-74; De Lellis L et al. PLoS One. 2013 Nov 20;8(11):e81194). A different alteration (c.1896G>C) resulting in the same amino acid substitution (p.E632D) was identified in a Balkan patient with suspected Lynch syndrome (Goldberg Y et al. Clin. Genet., 2015 Jun;87:549-53) and in an individual diagnosed with MLH1-absent uterine cancer at age 46 (South SA et al. Obstet Gynecol, 2007 Aug;110:543-5). This nucleotide position is well conserved in available vertebrate species. In silico splice site analysis predicts that this alteration will weaken the native splice donor site. This amino acid position is not well conserved in available vertebrate species. This variant was not reported in population-based cohorts in the Genome Aggregation Database (gnomAD). In addition, the in silico prediction for this variant as a missense substitution is inconclusive. Based on the majority of available evidence to date, this variant is likely to be pathogenic.

Computational scores

Source: dbNSFP v4.9

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Benign	0.095
BayesDel_addAF	Pathogenic	0.24	D
BayesDel_noAF	Uncertain	0.11
CADD	Uncertain	24
DANN	Uncertain	0.99
DEOGEN2	Uncertain	0.60	D
Eigen	Uncertain	0.60
Eigen_PC	Uncertain	0.59
FATHMM_MKL	Pathogenic	0.98	D
LIST_S2	Uncertain	0.91	D
M_CAP	Benign	0.032	D
MetaRNN	Uncertain	0.50	T
MetaSVM	Benign	-0.80	T
MutationAssessor	Benign	1.5	L
PhyloP100		6.6
PrimateAI	Benign	0.38	T
PROVEAN	Benign	-0.62	N
REVEL	Uncertain	0.46
Sift	Benign	0.23	T
Sift4G	Benign	0.49	T
Polyphen		0.0	B
Vest4		0.53
MutPred		0.32		Loss of disorder (P = 0.2235)
MVP		0.90
MPC		0.061
ClinPred		0.70	D
GERP RS		4.6
Varity_R		0.30
gMVP		0.53
Mutation Taster		=0/100	disease causing

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	1.0
SpliceAI score (max)		0.73		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.73		Position offset: 0

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

Other links and lift over

dbSNP: rs63751632; hg19: chr3-37089174;