3-37048614-G-T

Variant names:

• chr3-37048614-G-T
• rs267607878
• NM_000249.4:c.1989+5G>T

Variant summary

Our verdict is Pathogenic. The variant received 11 ACMG points: 11P and 0B. PM2 PP3 PP5_Very_Strong

The NM_000249.4(MLH1):​c.1989+5G>T variant causes a splice region, intron 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 Likely pathogenic (★★).

• Frequency: Genomes: not found (cov: 31)
• Consequence: MLH1 NM_000249.4 splice_region, intron
• Scores: Splicing: ADA: 0.9592
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:6
• Conservation: PhyloP100: 2.41

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

Our verdict: Pathogenic. The variant received 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 3-37048614-G-T is Pathogenic according to our data. Variant chr3-37048614-G-T is described in ClinVar as [Likely_pathogenic]. Clinvar id is 234540.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
MLH1	NM_000249.4	c.1989+5G>T		splice_region_variant, intron_variant	Intron 17 of 18	ENST00000231790.8	NP_000240.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
MLH1	ENST00000231790.8	c.1989+5G>T		splice_region_variant, intron_variant	Intron 17 of 18	1	NM_000249.4	ENSP00000231790.3

Frequencies

GnomAD3 genomes Cov.: 31

GnomAD4 exome Cov.: 29

GnomAD4 genome Cov.: 31

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:6

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Colorectal cancer, hereditary nonpolyposis, type 2 Pathogenic:2

Jul 25, 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: 8993976]. -

Mar 20, 2024

Baylor Genetics

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

- -

not provided Pathogenic:2

Dec 13, 2024

GeneDx

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant is denoted MLH1 c.1989+5G>T or IVS17+5G>T and consists of a G>T nucleotide substitution at the +5 position of intron 17 of the MLH1 gene. Multiple in silico models predict this variant to destroy the nearby natural donor site, and to possibly cause abnormal gene splicing. However, in the absence of RNA or functional studies, the actual effect of this variant is unknown. This variant was not observed in approximately 6,500 individuals of European and African American ancestry in the NHLBI Exome Sequencing Project, suggesting it is not a common benign variant in these populations. MLH1 c.1989+5G>T has not, to our knowledge, been published in the literature as pathogenic or benign. The guanine (G) nucleotide that is altered is not conserved across species. Based on currently available information, it is unclear whether MLH1 c.1989+5G>T is pathogenic or benign. -

Sep 11, 2024

Quest Diagnostics Nichols Institute San Juan Capistrano

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The MLH1 c.1989+5G>T variant (also known as IVS17+5G>T) has been reported in the published literature in individuals with Lynch syndrome or Lynch-associated disorders where loss of MLH1 and PMS2 has been reported (PMIDs: 28640387 (2017) and 28449805 (2017) and personal communication with Ambry Genetics and Invitae related to ClinVar ID: 234540). In addition, RNA studies have shown that this variant results in aberrant splicing and exon 17 skipping (personal communication with Ambry Genetics and Invitae related to ClinVar ID: 234540). In the published literature, another deleterious variant at the same position, c.1989+5G>C, has also resulted in loss of exon 17 as shown by RNA splicing analysis (PMIDs: 8993976 (1997) and 32363481 (2020)). This variant has not been reported in large, multi-ethnic general populations (Genome Aggregation Database, http://gnomad.broadinstitute.org). Analysis of this 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 nonpolyposis colorectal neoplasms Pathogenic:1

May 19, 2024

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This sequence change falls in intron 17 of the MLH1 gene. It does not directly change the encoded amino acid sequence of the MLH1 protein. RNA analysis indicates that this variant induces altered splicing and likely results in a shortened protein product. This variant is not present in population databases (gnomAD no frequency). This variant has been observed in individuals with Lynch syndrome (PMID: 28449805; Invitae). ClinVar contains an entry for this variant (Variation ID: 234540). Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Studies have shown that this variant results in skipping of exon 17, but is expected to preserve the integrity of the reading-frame (Invitae). This variant disrupts a region of the MLH1 protein in which other variant(s) (p.Thr662Pro) have been determined to be pathogenic (PMID: 11726306, 11754112, 16341550, 17510385, 19621678, 20533529, 21404117, 23403630). This suggests that this is a clinically significant region of the protein, and that variants that disrupt it are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic. -

Hereditary cancer-predisposing syndrome Pathogenic:1

Dec 02, 2024

Ambry Genetics

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The c.1989+5G>T intronic pathogenic mutation results from a G to T substitution 5 nucleotides after coding exon 17 in the MLH1 gene. This variant has been identified in a proband(s) who met Amsterdam I/II criteria for Lynch syndrome and tumor demonstrated loss of MLH1/PMS2 expression by immunohistochemistry (IHC) (Li S et al. J. Med. Genet. 2020 Jan;57:62-69; Ambry internal data). In addition, this alteration has been identified in a proband that also had somatic MLH1 copy neutral loss of heterozygosity (CN-LOH) in a MSI-H colon tumor with loss of MLH1 and PMS2 expression by IHC where MLH1 promotor hypermethylation was negative (Ambry internal data). In silico splice site analysis predicts that this alteration may weaken the native splice donor site and will result in the creation or strengthening of a novel splice donor site. RNA studies have demonstrated that this alteration results in abnormal splicing in the set of samples tested (Ambry internal data). The resulting transcript is in-frame and is not expected to trigger nonsense-mediated mRNAdecay. The exact functional effect of the altered amino acid sequence is unknown; however, the impacted region is critical for protein function (Ambry internal data). This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_noAF	Benign	-0.78
CADD	Pathogenic	32
DANN	Benign	0.74
Mutation Taster		=0/100	disease causing

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	0.96
dbscSNV1_RF	Pathogenic	0.85
SpliceAI score (max)		0.87		Details are displayed if max score is > 0.2
DS_DG_spliceai		0.87		Position offset: 26
DS_DL_spliceai		0.35		Position offset: -5

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

Publications

LitVar

Other links and lift over

dbSNP: rs267607878; hg19: chr3-37090105;