rs63751657

chr3-37042331-G-Achr3-37042331-G-Cchr3-37042331-G-T

Variant summary

Our verdict is Pathogenic. Variant got 11 ACMG points: 11P and 0B. PM2 PP3 PP5_Very_Strong

The NM_000249.4(MLH1):c.1731G>A(p.Ser577=) variant causes a splice region, synonymous change involving the alteration of a non-conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. In-silico tool predicts a benign outcome for this variant. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Pathogenic (★★★). Synonymous variant affecting the same amino acid position (i.e. S577S) has been classified as Likely pathogenic.

• Frequency:
  • Genomes: not found (cov: 32)
  • Exomes 𝑓: 0.0 (0 hom.)
  • Failed GnomAD Quality Control
• Consequence: MLH1 NM_000249.4 splice_region, synonymous
• Scores: Splicing: ADA: 0.9900
• Clinical Significance: Pathogenic reviewed by expert panel P:16 U:1 O:1
• Conservation: PhyloP100: 0.502

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 11 ACMG points.

• PM2: Very rare variant in population databases, with high coverage
• PP3: Splicing scoreres supports a deletorius effect: Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF, max_spliceai. No scorers claiming Uncertain. No scorers claiming Benign.
• PP5: Variant 3-37042331-G-A is Pathogenic according to our data. Variant chr3-37042331-G-A is described in ClinVar as [Pathogenic]. Clinvar id is 89857.Status of the report is reviewed_by_expert_panel, 3 stars. Variant chr3-37042331-G-A is described in Lovd as [Pathogenic]. Variant chr3-37042331-G-A is described in Lovd as [Benign]. Variant chr3-37042331-G-A is described in Lovd as [Likely_pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
MLH1	NM_000249.4	c.1731G>A	p.Ser577=	splice_region_variant, synonymous_variant	15/19	ENST00000231790.8

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
MLH1	ENST00000231790.8	c.1731G>A	p.Ser577=	splice_region_variant, synonymous_variant	15/19	1	NM_000249.4	P1

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Data not reliable, filtered out with message: AC0 AF: 0.00 AC: 0 AN: 1443524 Hom.: 0 Cov.: 29 AF XY: 0.00 AC XY: 0 AN XY: 719466

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.00

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.00

Gnomad4 SAS exome AF: 0.00

Gnomad4 FIN exome AF: 0.00

Gnomad4 NFE exome AF: 0.00

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome Cov.: 32

EpiCase AF: 0.0000545

EpiControl AF: 0.00

ClinVar

Significance: Pathogenic

Submissions summary: Pathogenic:16 Uncertain:1 Other:1

Revision: reviewed by expert panel

Submissions by phenotype

Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:4

Pathogenic, criteria provided, single submitter	clinical testing	Human Genome Sequencing Center Clinical Lab, Baylor College of Medicine	Oct 12, 2018	The c.1731G>A (p.Ser577=) variant in the MLH1 gene has been reported in multiple unrelated patients with hereditary nonpolyposis colorectal cancer [PMID: 8808596, 16451135, 16341550, 21598002, 26300997]. This variant has not been observed in the general population according to the gnomAD database. This variant is next to the exon 15 splicing donor site and functional study showed that this variant causes exon 15 skipping [PMID:18561205]. Therefore, this c.1731G>A (p.Ser577=) variant in the MLH1 gene is classified as pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Counsyl	Oct 06, 2017	- -
Pathogenic, criteria provided, single submitter	clinical testing	3billion	Mar 22, 2022	The variant has been observed in multiple (>3) similarly affected unrelated individuals (PMID: 16341550, 15849733, 20223024, 18931482, 26300997, 14635101, 16216036). Functional studies provide strong evidence of the variant having a damaging effect on the gene or gene product (PMID: 16341550, 18561205, 16451135). The variant has been reported multiple times as an established pathogenic/likely pathogenic variant (ClinVar ID: VCV000089857). It is not observed in the gnomAD v2.1.1 dataset. Therefore, this variant is classified as pathogenic according to the recommendation of ACMG/AMP guideline. -
Pathogenic, criteria provided, single submitter	clinical testing	Myriad Genetics, Inc.	Mar 15, 2023	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. Functional studies indicate this variant impacts protein function [PMID: 16341550]. This variant has been reported in multiple individuals with clinical features of gene-specific disease [8808596, 16341550]. This variant is strongly associated with more severe personal and family histories of cancer, typical for individuals with pathogenic variants in this gene [PMID: 25085752]. -

not provided Pathogenic:3 Uncertain:1

Pathogenic, no assertion criteria provided	research	Mayo Clinic Laboratories, Mayo Clinic	-	- -
Uncertain significance, no assertion criteria provided	clinical testing	Department of Pathology and Laboratory Medicine, Sinai Health System	-	- -
Likely pathogenic, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Aug 11, 2023	The MLH1 c.1731G>A (p.Ser577=) synonymous variant has been reported in the published literature in individuals and families with Lynch syndrome (PMIDs: 26300997 (2015), 24278394 (2013), 20223024 (2006), 16395668 (2006)). This variant was shown to interfere with normal MLH1 mRNA splicing (PMIDs: 19669161 (2010), 18561205 (2008), 16341550 (2006)). This variant has not been reported in large, multi-ethnic general populations (Genome Aggregation Database, http://gnomad.broadinstitute.org). Based on the available information, this variant is classified as likely pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Dec 23, 2019	Exonic splice site variant demonstrated to result in protein truncation or nonsense mediated decay in a gene for which loss-of-function is a known mechanism of disease (Liu 1995, Kohonen-Corish 1996, Plaschke 1999, Pistorious 2000, Pagenstecher 2006); Not observed in large population cohorts (Lek et al., 2016); This variant is associated with the following publications: (PMID: 18561205, 21598002, 16451135, 24278394, 19731080, 9087566, 10596954, 11112663, 18772310, 7704024, 21286823, 17576681, 8808596, 16395668, 19669161, 11151427, 14514376, 15046089, 16341550, 11601928, 18726168, 19072991, 21387278, 18931482, 26300997, 25345868, 15849733, 16216036, 20007843, 27064304, 28050887, 27601186, 22949379, 27007491, 26681312, 29706640, 29506128, 24456667, 14635101, 29887214, 30521064, 31054147, 20223024, 31447099, 32658311) -

Lynch syndrome Pathogenic:2

Pathogenic, reviewed by expert panel	research	International Society for Gastrointestinal Hereditary Tumours (InSiGHT)	Sep 05, 2013	Variant causes splicing aberration leading to truncated protein: complete inactivation of variant allele -
Pathogenic, criteria provided, single submitter	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Oct 14, 2019	The p.Ser577Ser variant in MLH1 has been previously reported in at least 17 individuals with Lynch syndrome (Auclair 2006, Tournier 2008, Bozzao 2011, Kurzawski 2006, Kohonen-Corish 1996, Mangold 2005, Hinrichsen 2014, Betz 2010, Simbolo 2015, Pagenstecher 2006, De Lellis 2013) and was absent from large population studies. This variant was classified as Pathogenic by several clinical labs in ClinVar and on September 5, 2013 by the ClinGen-approved InSiGHT expert panel (Variation ID 89857). This variant is located in the last three bases of the exon, which is part of the 5’ splice region. Computational tools do not predict a splicing impact, though this information is not predictive enough to rule out pathogenicity. In vitro and in vivo functional studies support an impact on protein function, due to out-of-frame exon skipping (Auclair 2006, Tournier 2008, Kohonen-Corish 1996, Betz 2010, Pagenstecher 2006, De Lellis 2013). Heterozygous loss-of-function of the MLH1 gene is an established disease mechanism in individuals with Lynch syndrome. In summary, this variant meets criteria to be classified as pathogenic for autosomal dominant Lynch syndrome. ACMG/AMP criteria applied: PS3, PS4, PM2. -

Hereditary cancer-predisposing syndrome Pathogenic:2

Pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	May 05, 2022	The c.1731G>A pathogenic mutation (also known as p.S577S), located in coding exon 15 of the MLH1 gene, results from a G to A substitution at nucleotide position 1731. This nucleotide substitution does not change the serine at codon 577. However, this change occurs in the last base pair of coding exon 15, which makes it likely to have some effect on normal mRNA splicing. This mutation has been reported in numerous individuals with HNPCC/Lynch syndrome or that have high suspicion for HNPCC/Lynch syndrome based on personal and/or family history (Pistorius SR et al. Int. J. Colorectal Dis. 2000 Nov;15(5-6):255-63; Mangold E et al. Int. J. Cancer. 2005 Sep;116(5):692-702; Pagenstecher C et al. Hum. Genet. 2006 Mar;119:9-22; Kurzawski G et al. Hered. Cancer Clin. Pract. 2006 Dec;4(4):197-205; Jasperson KW et al. Fam. Cancer, 2010 Jun;9:99-107; Bozzao C et al. Cancer. 2011 Sep;117(18):4325-35; De Lellis L et al. PLoS ONE. 2013 Nov;8(11):e81194; Simbolo M et al. Hered. Cancer Clin. Pract. 2015 Aug;13(1):18). This mutation has been shown in multiple RNA studies to result in skipping of coding exon 15, which is predicted to lead to a translational frameshift (Kohonen-Corish M et al. Am. J. Hum. Genet. 1996 Oct;59(4):818-24; Auclair J et al. Hum. Mutat. 2006 Feb;27(2):145-54; Pagenstecher C et.al. Hum. Genet. 2006 Mar;119:9-22; Tournier I et al. Hum. Mutat. 2008 Dec;29(12):1412-24; Ambry internal data). This nucleotide position is highly conserved through mammals. In silico splice site analysis predicts that this alteration will weaken the native splice donor site. Based on the available evidence, this alteration is classified as a pathogenic mutation. -
Pathogenic, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Jun 07, 2021	This variant alters the last nucleotide of exon 15 of the MLH1 gene. Functional studies using RNA derived from carrier individuals have shown that this variant causes a complete out-of-frame skipping of exon 15 and results in a premature protein truncation (PMID: 16341550, 16395668, 19669161). This variant has been reported in multiple individuals affected with Lynch syndrome (PMID: 8808596, 11151427, 14514376, 15849733, 16341550, 16451135, 20223024, 21598002, 24456667, 26300997, Insight-database.org). 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 Pathogenic. -

Breast and/or ovarian cancer Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

CHEO Genetics Diagnostic Laboratory, Children's Hospital of Eastern Ontario

Aug 31, 2022

- -

Hereditary nonpolyposis colon cancer Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

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

Jul 13, 2021

Variant summary: MLH1 c.1731G>A (p.Ser577Ser) alters a non-conserved nucleotide that is located at the last nucleotide of exon 15, therefore could affect mRNA splicing, leading to a significantly altered protein sequence. Several computational tools predicted a weak impact on normal splicing: 3/3 predict the variant slightly weakens a 5' donor site. However, multiple publications reported experimental evidence, demonstrating that this variant affects mRNA splicing, resulting in a complete skipping of exon 15, which is predicted to cause a frameshift at the protein level (Pagenstecher_2006, Auclair_2006, Tournier_2008, Betz_2010). The variant was absent in 251202 control chromosomes (gnomAD). The variant, c.1731G>A, has been reported in the literature in several individuals and families affected with Hereditary Nonpolyposis Colorectal Cancer (e.g. Kohonen-Corish_1996, Scott_2001, Raedle_2001, Pagenstecher_2006, Auclair_2006), and in numerous cases microsatellite instability and loss of the MLH1 protein was noted in the associated tumor. These data indicate that the variant is very likely to be associated with disease. Seven clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014, and all of them classified the variant as pathogenic. Based on the evidence outlined above, the variant was classified as pathogenic. -

Muir-Torré syndrome;C1333991:Colorectal cancer, hereditary nonpolyposis, type 2;C5399763:Mismatch repair cancer syndrome 1 Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Fulgent Genetics, Fulgent Genetics

Dec 08, 2021

- -

Hereditary nonpolyposis colorectal neoplasms Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Invitae

Dec 30, 2023

This sequence change affects codon 577 of the MLH1 mRNA. It is a 'silent' change, meaning that it does not change the encoded amino acid sequence of the MLH1 protein. RNA analysis indicates that this variant induces altered splicing and may result in an absent or disrupted protein product. This variant is not present in population databases (gnomAD no frequency). This variant has been observed in individuals with Lynch syndrome (PMID: 14635101, 15849733, 16216036, 16341550, 18931482, 20223024, 21598002, 26300997). ClinVar contains an entry for this variant (Variation ID: 89857). Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Studies have shown that this variant alters mRNA splicing and is expected to lead to the loss of protein expression (PMID: 16341550, 16451135, 18561205). For these reasons, this variant has been classified as Pathogenic. -

Colon cancer Pathogenic:1

Pathogenic, no assertion criteria provided

clinical testing

Medical Genetics Laboratory, Umraniye Training and Research Hospital, University of Health Sciences

Aug 23, 2021

Colon Cancer -

Lynch syndrome 1 Other:1

not provided, no classification provided

literature only

GeneReviews

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Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_noAF	Benign	-0.35
Cadd	Benign	22
Dann	Benign	0.87
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		2.8

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	0.99
dbscSNV1_RF	Pathogenic	0.92
SpliceAI score (max)		0.63		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.63		Position offset: 0

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

Publications

LitVar

Other links and lift over

dbSNP: rs63751657; hg19: chr3-37083822; COSMIC: COSV51613969; COSMIC: COSV51613969;