Submissions for variant NM_000249.4(MLH1):c.545+3A>G

dbSNP: rs267607760

Total submissions: 14

Submitter	RCV	SCV	Clinical significance	Condition	Last evaluated	Review status	Method	Comment
International Society for Gastrointestinal Hereditary Tumours (InSiGHT)	RCV000075749	SCV000106757	pathogenic	Lynch syndrome	2013-09-05	reviewed by expert panel	research	Variant causes splicing aberration, >2 MSI-H, co-segregation with disease & absent in 1000 genomes
GeneDx	RCV000215515	SCV000279598	pathogenic	not provided	2017-11-10	criteria provided, single submitter	clinical testing	This variant is denoted MLH1 c.545+3A>G or IVS6+3A>G and consists of an A>G nucleotide substitution at the +3 position of intron 6 of the MLH1 gene. In-silico analyses, including splice predictors and evolutionary conservation, support a deleterious effect. RNA studies and protein truncation testing confirm this prediction, demonstrating abnormal splicing leading to a frameshift that results in a truncated protein (Pensotti 1997, Thiffault 2004). This variant was observed in multiple individuals with colon cancer, including two who had absent MLH1 on immunohistochemistry, and is considered an Italian pathogenic founder variant (Pensotti 1997, Hendriks 2003, Liu 2004, Thiffault 2004, Mangold 2005, Valentin 2011). This variant has been shown to segregate with disease in two affected siblings in one family and with three affected relatives in another family (Thiffault 2004). MLH1 c.545+3A>G was not observed in large population cohorts (Lek 2016). The adenine (A) nucleotide that is altered is conserved across species. Based on the current evidence, we consider this variant to be pathogenic.
Ambry Genetics	RCV000564669	SCV000676016	pathogenic	Hereditary cancer-predisposing syndrome	2023-11-03	criteria provided, single submitter	clinical testing	The c.545+3A>G intronic pathogenic mutation results from an A to G substitution 3 nucleotides after coding exon 6 in the MLH1 gene. This mutation has been identified in probands whose Lynch syndrome-associated tumor demonstrated high microsatellite instability and/or loss of MLH1 or MLH1/PMS2 expression by immunohistochemistry (Hendriks Y et al. Am. J. Pathol., 2003 Feb;162:469-77; Thiffault I et al. Clin. Genet., 2004 Aug;66:137-43; Carneiro da Silva F et al. PLoS ONE, 2015 Oct;10:e0139753; Ambry internal data). This alteration has been shown to cause a deleterious impact on splicing by RT-PCR and cDNA analyses (Pensotti V et al. Genes Chromosomes Cancer, 1997 Jul;19:135-42; Morak M et al. Eur J Hum Genet, 2019 Dec;27:1808-1820). In addition, RNA studies have demonstrated that this alteration results in abnormal splicing in the set of samples tested (Ambry internal data). This nucleotide position is highly conserved in available vertebrate species. In silico splice site analysis predicts that this alteration will weaken the native splice donor site. 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.
Women's Health and Genetics/Laboratory Corporation of America, LabCorp	RCV000075749	SCV000696170	pathogenic	Lynch syndrome	2016-09-19	criteria provided, single submitter	clinical testing	Variant summary: The MLH1 c.545+3A>G variant involves the alteration of a conserved nucleotide located at the intron/exon boundary. One in silico tool predicts a damaging outcome for this variant. 4/4 splice prediction tools predict loss/weakening effect on the canonical splicing donor site. This variant was absent in 121426 control chromosomes but has been reported in multiple affected individuals/families, and some families showed evidence of co-segregation of the variant with disease. In addition, multiple clinical diagnostic laboratories/reputable databases classified this variant as pathogenic. RT-PCR showed the variant lead to an aberrant splicing product (Pensotti_GCC_1997). Taken together, this variant was classified as pathogenic.
A.C.Camargo Cancer Center / LGBM, A.C.Camargo Cancer Center	RCV000075749	SCV000914314	pathogenic	Lynch syndrome	2019-01-30	criteria provided, single submitter	research
Color Diagnostics, LLC DBA Color Health	RCV000564669	SCV001340977	pathogenic	Hereditary cancer-predisposing syndrome	2020-05-26	criteria provided, single submitter	clinical testing	This variant causes an A to G nucleotide substitution at the +3 position of intron 6 of the MLH1 gene. A functional RNA study has shown that this variant causes aberrant splicing and a significant truncation of exon 6 due to the use of a cryptic donor site leaving all but the first 3 nucleotides of exon 6 (PMID: 9218993). This variant has been reported in multiple individuals affected with hereditary nonpolyposis colorectal cancer (PMID: 9218993, 12547705, 15253764, 15309712) and shown to segregate with disease in family studies (PMID: 9218993, 15253764). 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.
Invitae	RCV001202202	SCV001373307	pathogenic	Hereditary nonpolyposis colorectal neoplasms	2023-10-28	criteria provided, single submitter	clinical testing	This sequence change falls in intron 6 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 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: 9218993, 12547705, 15253764, 15309712, 15849733, 26437257, 28874130). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 90260). 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 activation of a cryptic splice site and introduces a premature termination codon (PMID: 9218993). The resulting mRNA is expected to undergo nonsense-mediated decay. For these reasons, this variant has been classified as Pathogenic.
Quest Diagnostics Nichols Institute San Juan Capistrano	RCV000215515	SCV002046091	pathogenic	not provided	2021-03-16	criteria provided, single submitter	clinical testing	The variant has been reported in individuals and families with Lynch syndrome in the published literature (PMIDs: 28874130 (2017), 26437257 (2015), 21681552 (2011), 15849733 (2005)). In addition, published studies indicate that this variant causes a damaging effect on normal MLH1 splicing (PMIDs: 15253764 (2004), 9218993 (1997)). This variant has not been reported in large, multi-ethnic general populations (http://gnomad.broadinstitute.org). Based on the available information, this variant is classified as pathogenic.
Sema4, Sema4	RCV000564669	SCV002528750	pathogenic	Hereditary cancer-predisposing syndrome	2021-04-11	criteria provided, single submitter	curation
Diagnostic Laboratory, Department of Genetics, University Medical Center Groningen	RCV000609647	SCV000734263	pathogenic	Colorectal cancer, hereditary nonpolyposis, type 2		no assertion criteria provided	clinical testing
Constitutional Genetics Lab, Leon Berard Cancer Center	RCV001249949	SCV001423891	pathogenic	Lynch-like syndrome	2019-07-01	no assertion criteria provided	clinical testing
Clinical Genetics DNA and cytogenetics Diagnostics Lab, Erasmus MC, Erasmus Medical Center	RCV000215515	SCV001975824	pathogenic	not provided		no assertion criteria provided	clinical testing
Joint Genome Diagnostic Labs from Nijmegen and Maastricht, Radboudumc and MUMC+	RCV000215515	SCV001979522	pathogenic	not provided		no assertion criteria provided	clinical testing
GeneReviews	RCV001804822	SCV002054095	not provided	Lynch syndrome 1		no assertion provided	literature only