3-37040294-G-C

Position:

chr3-37040294-G-C

Variant summary

Our verdict is Likely pathogenic. Variant got 7 ACMG points: 7P and 0B. PM1PM2PM5PP5

The NM_000249.4(MLH1):c.1667G>C(p.Ser556Thr) variant causes a missense, splice region change involving the alteration of a conserved nucleotide. The variant allele was found at a frequency of 0.00000311 in 1,608,450 control chromosomes in the GnomAD database, with no homozygous occurrence. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Conflicting classifications of pathogenicity (no stars). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. S556I) has been classified as Pathogenic.

Frequency

Genomes: 𝑓 0.0000066 ( 0 hom., cov: 32)

Exomes 𝑓: 0.0000027 ( 0 hom. )

Consequence

MLH1
NM_000249.4 missense, splice_region

Scores

Splicing: ADA: 1.000

Clinical Significance

Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:7U:1

Conservation

PhyloP100: 9.74

Variant links:

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 links:

MLH1 (HGNC:):

MLH1 links:

Genome browser will be placed here

ACMG classification

Classification made for transcript

Verdict is Likely_pathogenic. Variant got 7 ACMG points.

PM1

In a helix (size 14) in uniprot entity MLH1_HUMAN there are 9 pathogenic changes around while only 1 benign (90%) in NM_000249.4

PM2

Very rare variant in population databases, with high coverage;

PM5

Other missense variant is known to change same aminoacid residue: Variant chr3-37040294-G-T is described in Lovd as [Pathogenic].

PP5

Variant 3-37040294-G-C is Pathogenic according to our data. Variant chr3-37040294-G-C is described in ClinVar as [Conflicting_classifications_of_pathogenicity]. Clinvar id is 185187.We mark this variant Likely_pathogenic, oryginal submissions are: {Uncertain_significance=1, Pathogenic=2, Likely_pathogenic=5}. Variant chr3-37040294-G-C is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
MLH1	NM_000249.4 linkuse as main transcript	c.1667G>C	p.Ser556Thr	missense_variant, splice_region_variant	14/19	ENST00000231790.8	NP_000240.1	P40692-1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8 linkuse as main transcript	c.1667G>C	p.Ser556Thr	missense_variant, splice_region_variant	14/19	1	NM_000249.4	ENSP00000231790.3		P40692-1

Frequencies

GnomAD3 genomes

AF:

0.00000657

AC:

AN:

152106

Hom.:

Cov.:

show subpopulations

Gnomad AFR

AF:

0.00

Gnomad AMI

AF:

0.00

Gnomad AMR

AF:

0.00

Gnomad ASJ

AF:

0.00

Gnomad EAS

AF:

0.00

Gnomad SAS

AF:

0.00

Gnomad FIN

AF:

0.00

Gnomad MID

AF:

0.00

Gnomad NFE

AF:

0.0000147

Gnomad OTH

AF:

0.00

GnomAD3 exomes

AF:

0.00000398

AC:

AN:

251200

Hom.:

AF XY:

0.00000737

AC XY:

AN XY:

135768

show subpopulations

Gnomad AFR exome

AF:

0.00

Gnomad AMR exome

AF:

0.00

Gnomad ASJ exome

AF:

0.00

Gnomad EAS exome

AF:

0.00

Gnomad SAS exome

AF:

0.00

Gnomad FIN exome

AF:

0.00

Gnomad NFE exome

AF:

0.00000880

Gnomad OTH exome

AF:

0.00

GnomAD4 exome

AF:

0.00000275

AC:

AN:

1456344

Hom.:

Cov.:

AF XY:

0.00000552

AC XY:

AN XY:

724946

show subpopulations

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:

0.0000232

Gnomad4 FIN exome

AF:

0.00

Gnomad4 NFE exome

AF:

0.00000181

Gnomad4 OTH exome

AF:

0.00

GnomAD4 genome

AF:

0.00000657

AC:

AN:

152106

Hom.:

Cov.:

AF XY:

0.0000135

AC XY:

AN XY:

74284

show subpopulations

Gnomad4 AFR

AF:

0.00

Gnomad4 AMR

AF:

0.00

Gnomad4 ASJ

AF:

0.00

Gnomad4 EAS

AF:

0.00

Gnomad4 SAS

AF:

0.00

Gnomad4 FIN

AF:

0.00

Gnomad4 NFE

AF:

0.0000147

Gnomad4 OTH

AF:

0.00

Alfa

AF:

0.0000712

Hom.:

Bravo

AF:

0.00000756

ExAC

AF:

0.00000824

AC:

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Pathogenic:7Uncertain:1

Revision: criteria provided, conflicting classifications

LINK: link

Submissions by phenotype

not provided

Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	CeGaT Center for Human Genetics Tuebingen	Feb 01, 2024	MLH1: PM1, PM2, PM5, PS4:Moderate, PP3 -
Likely pathogenic, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Feb 13, 2023	The frequency of this variant in the general population, 0.0000071 (2/282602 chromosomes, http://gnomad.broadinstitute.org), is uninformative in assessment of its pathogenicity. In the published literature, the variant has been reported in individuals with Lynch syndrome (PMID: 21404117 (2011), or suspected of having hereditary breast cancer/Lynch syndrome (PMID: 28514183 (2017)). This variant is located at the last nucleotide of exon 14. In particular, the c.1667G>T (p.Ser556Ile) variant has been shown in experimental study to cause disruption of the adjacent splice site and incorporation of intronic sequences into the transcript, including a premature stop codon (PMID: 15300854 (2004)). Consistent with this finding, analysis of the c.1667G>C (p.Ser556Thr) variant using software algorithms for the prediction of the effect of nucleotide changes on splicing yielded predictions that this variant may affect proper MLH1 mRNA splicing . Based on the available information, this variant is classified as likely pathogenic. -

Hereditary cancer-predisposing syndrome

Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Aug 09, 2021	This missense variant replaces serine with threonine at codon 556 of the MLH1 protein. Computational prediction is inconclusive regarding the impact of this variant on protein structure and function (internally defined REVEL score threshold 0.5 < inconclusive < 0.7, PMID: 27666373). This variant (c.1667G>C) alters the conserved G nucleotide at the last nucleotide position of exon 14 of the MLH1 gene, and is predicted to disrupt RNA splicing. To our knowledge, functional studies have not been reported for this variant. This variant has been reported in individuals affected with or suspected of having Lynch syndrome (PMID: 21404117, 28514183). This variant has been identified in 1/245968 chromosomes in the general population by the Genome Aggregation Database (gnomAD). Different variants affecting the same nucleotide position c.1667G>T and c.1667G>A have been shown to impact RNA splicing and are associated with disease (Clinvar variation ID: 89825, 449776). Based on the available evidence, this variant is classified as Likely Pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	May 08, 2024	The c.1667G>C (p.S556T) alteration is located in exon 14 (coding exon 14) of the MLH1 gene. This alteration results from a G to C substitution at nucleotide position 1667, causing the serine (S) at amino acid position 556 to be replaced by a threonine (T). However, this change occurs in the last base pair of coding exon 14, which makes it likely to have some effect on normal mRNA splicing. Based on data from gnomAD, the C allele has an overall frequency of 0.001% (2/282602) total alleles studied. The highest observed frequency was 0.002% (2/129026) of European (non-Finnish) alleles. This alteration has been detected in multiple patients with personal and/or family histories of Lynch syndrome (LS)-associated cancers including a patient diagnosed with a MSI-H colorectal cancer whose tumor demonstrated loss of MLH1 and PMS2 on immunohistochemistry (Ambry internal data). This variant has also been reported in an individual affected with MSI-H colorectal cancer and had a family history that met Amsterdam II criteria (Yurgelun, 2017). Another alteration at the same last nucleotide position, p.S556I (c.1667G>T), has been reported in a family with colon and breast cancer meeting Amsterdam Criteria for HNPCC/LS. This nucleotide position is highly conserved in available vertebrate species. The p.S556I (c.1667G>T) alteration resulted in partial read through into intron 14 and the inclusion of an additional 88 bp of intronic sequence into the mRNA by RT-PCR analysis (Sharp, 2004). In silico splice site analysis predicts that this alteration will weaken the native splice donor site; however, direct evidence is insufficient at this time (Ambry internal data). In addition, as a missense substitution, this variant is predicted to be deleterious by in silico analysis. Based on the available evidence, this alteration is classified as pathogenic. -

Lynch syndrome

Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

All of Us Research Program, National Institutes of Health

Mar 25, 2024

This missense variant replaces serine with threonine at codon 556 of the MLH1 protein. Computational prediction is inconclusive regarding the impact of this variant on protein structure and function (internally defined REVEL score threshold 0.5 < inconclusive < 0.7, PMID: 27666373). This variant (c.1667G>C) alters the conserved G nucleotide at the last nucleotide position of exon 14 of the MLH1 gene, and is predicted to disrupt RNA splicing. To our knowledge, functional studies have not been reported for this variant. This variant has been reported in individuals affected with or suspected of having Lynch syndrome (PMID: 21404117, 28514183). This variant has been identified in 1/245968 chromosomes in the general population by the Genome Aggregation Database (gnomAD). Different variants affecting the same nucleotide position c.1667G>T and c.1667G>A have been shown to impact RNA splicing and are associated with disease (Clinvar variation ID: 89825, 449776). Based on the available evidence, this variant is classified as Likely Pathogenic. -

Hereditary nonpolyposis colorectal neoplasms

Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Labcorp Genetics (formerly Invitae), Labcorp

Jul 17, 2023

In summary, the currently available evidence indicates that the variant is pathogenic, but additional data are needed to prove that conclusively. Therefore, this variant has been classified as Likely Pathogenic. This variant disrupts the c.1667G nucleotide in the MLH1 gene. Other variant(s) that disrupt this nucleotide have been determined to be pathogenic (PMID: 27064304, 28135145; Invitae; external communication). This suggests that this nucleotide is clinically significant, and that variants that disrupt this position are likely to be disease-causing. Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Studies have shown that this missense change is associated with altered splicing resulting in unknown protein product impact (Invitae). An algorithm developed to predict the effect of missense changes on protein structure and function (PolyPhen-2) suggests that this variant¬†is likely to be tolerated. ClinVar contains an entry for this variant (Variation ID: 185187). This missense change has been observed in individual(s) with clinical features of Lynch syndrome (PMID: 21404117, 28514183). The frequency data for this variant in the population databases is considered unreliable, as metrics indicate poor data quality at this position in the gnomAD database. This sequence change replaces serine, which is neutral and polar, with threonine, which is neutral and polar, at codon 556 of the MLH1 protein (p.Ser556Thr). This variant also falls at the last nucleotide of exon 14, which is part of the consensus splice site for this exon. -

Familial cancer of breast

Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Human Genetics Bochum, Ruhr University Bochum

Feb 02, 2022

ACMG criteria used to clasify this variant: PVS1, PM2, PM5, PM1, PP3 -

not specified

Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

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

Jul 20, 2020

Variant summary: MLH1 c.1667G>C (p.Ser556Thr) results in a conservative amino acid change located in the DNA mismatch repair protein MLH1, C-terminal domain of the encoded protein sequence. Four of five in-silico tools predict a benign effect of the variant on protein function. Additionally, c.1667G>C alters a conserved nucleotide located within the confines of a putative splice region at the end of exon 14. Several computational tools predict a significant impact on normal splicing: Three predict the variant weakens the canonical 5' donor site. One predict the variant abolishes the canonical 5 splicing donor site. However, these predictions have yet to be confirmed by functional studies. Nevertheless, a variant located at the same nucleotide position but causing a different change (MLH1 c.1667G>T) is predicted by computational tools to cause a very similar effect on splicing as the variant of interest. c.1667G>T (not the variant of interest) was reported to cause the inclusion of an additional 88bp of intronic sequence into the mRNA resulting in a frameshift and the inclusion of a stop codon in the additional intronic sequence (Sharp_2004). Therefore, the possibility of this variant (c.1667G>C) having a similar splicing outcome cannot be ruled out. The variant allele was found at a frequency of 4e-06 in 251200 control chromosomes. The available data on variant occurrences in the general population are insufficient to allow any conclusion about variant significance. c.1667G>C has been reported in the literature in individuals affected with features of Lynch syndrome whose tumors were MSI-high and showed positive staining for MLH1 by IHC (Alter_2018, Hardt_2011). Specifically, a non-peer reviewed case report of co-occurrence with a putative pathogenic variant in MSH2 gene was ascertained at the time of this classification (MSH2 deletion of exon 9) (Alter_2018). This index patient had a MSI-high mucinous carcinoma in the colon ascendens at the age of 38. Immunohistochemical staining showed a loss of MSH2 and MSH6 expression. These reports do not provide unequivocal conclusions about association of the variant with Lynch Syndrome. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. Two clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation and both laboratories classified the variant as likely pathogenic. Based on the evidence outlined above, the variant was classified as VUS-possibly pathogenic. -

Computational scores

Source: dbNSFP v4.3

Name

Calibrated prediction

Score

Prediction

AlphaMissense

Benign

0.12

BayesDel_addAF

Pathogenic

0.27

BayesDel_noAF

Pathogenic

0.16

CADD

Pathogenic

DANN

Benign

0.93

DEOGEN2

Benign

0.37

Eigen

Uncertain

0.44

Eigen_PC

Uncertain

0.36

FATHMM_MKL

Pathogenic

0.99

LIST_S2

Benign

0.29

M_CAP

Pathogenic

0.30

MetaRNN

Uncertain

0.51

MetaSVM

Benign

-0.36

PROVEAN

Benign

1.5

REVEL

Uncertain

0.44

Sift

Benign

0.11

Sift4G

Pathogenic

0.0

MutPred

0.42

Loss of helix (P = 0.0123);

MVP

0.94

ClinPred

0.63

GERP RS

4.5

RBP_binding_hub_radar

0.0

RBP_regulation_power_radar

1.7

Splicing

Name

Calibrated prediction

Score

Prediction

dbscSNV1_ADA

Pathogenic

1.0

dbscSNV1_RF

Pathogenic

0.99

SpliceAI score (max)

0.92

Details are displayed if max score is > 0.2

DS_DL_spliceai

0.92

Position offset: 0

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

Publications

LitVar

Below is the list of publications found by LitVar. It may be empty.

Variant summary

Frequency

Consequence

Scores

Clinical Significance

Conservation

ACMG classification

Transcripts

RefSeq

Ensembl

Frequencies

ClinVar

Submissions by phenotype

Computational scores

Splicing

Publications

LitVar

Other links and lift over