3-37025635-A-G

Position:

• chr3-37025635-A-G

Variant summary

Our verdict is Pathogenic. Variant got 22 ACMG points: 22P and 0B. PVS1 PM2 PP3_Strong PP5_Very_Strong

The NM_000249.4(MLH1):​c.1039-2A>G variant causes a splice acceptor 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: 26)
  • Exomes 𝑓: 0.0 (0 hom.)
  • Failed GnomAD Quality Control
• Consequence: MLH1 NM_000249.4 splice_acceptor
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Likely pathogenic reviewed by expert panel P:6
• Conservation: PhyloP100: 8.52

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 22 ACMG points.

• PVS1: Splicing variant, LoF is a know mechanism of disease, Cryptic splice site detected, with MaxEntScore 5.2, offset of 13, new splice context is: aatacggactttgctaccAGgac. 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
• 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 3-37025635-A-G is Pathogenic according to our data. Variant chr3-37025635-A-G is described in ClinVar as [Likely_pathogenic]. Clinvar id is 89621.Status of the report is reviewed_by_expert_panel, 3 stars. Variant chr3-37025635-A-G is described in Lovd as [Likely_pathogenic]. Variant chr3-37025635-A-G is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
MLH1	NM_000249.4	c.1039-2A>G		splice_acceptor_variant		ENST00000231790.8

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
MLH1	ENST00000231790.8	c.1039-2A>G		splice_acceptor_variant		1	NM_000249.4	P1

Frequencies

GnomAD3 genomes Cov.: 26

GnomAD4 exome Data not reliable, filtered out with message: AC0 AF: 0.00 AC: 0 AN: 1224776 Hom.: 0 Cov.: 25 AF XY: 0.00 AC XY: 0 AN XY: 603684

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.: 26

ClinVar

Significance: Likely pathogenic

Submissions summary: Pathogenic:6

Revision: reviewed by expert panel

Submissions by phenotype

Lynch syndrome Pathogenic:2

Likely pathogenic, reviewed by expert panel	curation	International Society for Gastrointestinal Hereditary Tumours (InSiGHT)	Jun 21, 2019	Interrupts canonical donor splice site -
Pathogenic, criteria provided, single submitter	clinical testing	University of Washington Department of Laboratory Medicine, University of Washington	May 01, 2018	MLH1 NM_000249.3:c.1039-2A>G has a 99.98% probability of pathogenicity based on combining prior probability from public data with a likelihood ratio of 26.5 to 1, generated from evidence of seeing this as a somatic mutation in a tumor with loss of heterozygosity at the MLH1 locus. See Shirts et al 2018, PMID 29887214. -

Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Myriad Genetics, Inc.

Dec 29, 2022

This variant is considered likely 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. -

MLH1-related disorder Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

PreventionGenetics, part of Exact Sciences

Jun 20, 2023

The MLH1 c.1039-2A>G variant is predicted to disrupt the AG splice acceptor site and interfere with normal splicing. This variant has been reported in at least two individuals with hereditary non-polyposis colorectal cancer (HNPCC) (Mangold et al. 2005. PubMed ID: 15849733; Betz et al. 2010. PubMed ID: 19669161). mRNA studies showed that this variant results in the skipping of exon 12 leading to a frameshift and premature protein termination (Betz et al. 2010. PubMed ID: 19669161). This variant has not been reported in a large population database (http://gnomad.broadinstitute.org) and is interpreted as pathogenic or likely pathogenic in ClinVar (https://www.ncbi.nlm.nih.gov/clinvar/variation/89621/). Variants that disrupt the consensus splice acceptor site in MLH1 are expected to be pathogenic. This variant is interpreted as likely pathogenic. -

not provided Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

GeneDx

Apr 28, 2023

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); Published functional studies suggest a damaging effect: exon skipping resulting in truncated protein (Betz et al., 2010); Observed in at least one family reported to have features consistent with Lynch syndrome (Mangold et al., 2005); This variant is associated with the following publications: (PMID: 25525159, 19669161, Castillejo2016[article], 15849733, 29887214) -

Hereditary nonpolyposis colorectal neoplasms Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Invitae

Apr 22, 2022

This sequence change affects an acceptor splice site in intron 11 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: 15849733, 19669161; Invitae). ClinVar contains an entry for this variant (Variation ID: 89621). 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	0.16
CADD	Pathogenic	32
DANN	Uncertain	0.99
Eigen	Pathogenic	1.1
Eigen_PC	Pathogenic	0.96
FATHMM_MKL	Pathogenic	0.98	D
MutationTaster	Benign	1.0	D;D;D;D;D;D
GERP RS		5.7

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.92
SpliceAI score (max)		0.92		Details are displayed if max score is > 0.2
DS_AG_spliceai		0.24		Position offset: 15
DS_AL_spliceai		0.92		Position offset: 2

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

Publications

LitVar

Other links and lift over

dbSNP: rs267607815; hg19: chr3-37067126;