Submissions for variant NM_000249.4(MLH1):c.677G>T (p.Arg226Leu)

dbSNP: rs63751711

Total submissions: 13

Submitter	RCV	SCV	Clinical significance	Condition	Last evaluated	Review status	Method	Comment
International Society for Gastrointestinal Hereditary Tumours (InSiGHT)	RCV000075810	SCV000106824	likely pathogenic	Lynch syndrome	2019-06-21	reviewed by expert panel	curation	G>non-G at last base of exon with first 6 bases of the intron not GTRRGT (splicing aberration reported, but not quantified)
GeneDx	RCV000160555	SCV000211133	pathogenic	not provided	2018-11-19	criteria provided, single submitter	clinical testing	This variant is denoted MLH1 c.677G>T at the cDNA level. Located in the last nucleotide of exon 8, it destroys a natural splice site and causes abnormal splicing. Multiple studies report, but do not quantify, aberrant splicing caused by this variant that results in skipping of exon 8 (Maliaka 1996, Kurzawski 2006, Hardt 2011). This is concordant with multiple protein and RNA analyses of an alternate variant at this position, MLH1 c.677G>A, which have consistently demonstrated that it causes complete skipping of exon 8 (Leung 1998, Sharp 2004, Pagenstecher 2006, Tournier 2008). In addition, in vitro functional assays of MLH1 c.677G>T showed reduced mismatch repair activity and possibly reduced protein expression (Takahashi 2007). MLH1 c.677G>T has been observed in multiple individuals with early-onset colorectal, endometrial, or gastric cancer whose family histories fulfilled Bethesda or Amsterdam Criteria for Hereditary Nonpolyposis Colorectal Cancer, but also in a sporadic gastric tumor (Maliaka 1996, Evans 2001, Kurzawski 2002, Bartosova 2003, Wagner 2003, Kurzawski 2006, Zavodna 2006, Alemayehu 2008, Hardt 2011, Schofield 2012, Raskin 2017, Martin-Morales 2018). Tumor testing in many of these individuals showed microsatellite instability and loss of MLH1 protein expression, and Bujalkova et al. (2008) found tumoral loss of heterozygosity. In addition, MLH1 c. 677G>T co-occurred with two other pathogenic variants, one in BRCA2 and one in MSH6, in an individual with early-onset endometrial cancer and bilinear family history (Gong 2012). Although the nucleotide substitution results in the change of an Arginine to a Leucine at codon 226, and is called Arg226Leu in the literature, we are using only the nucleotide nomenclature to refer to the variant since the defect is determined to be one of splicing rather than a resulting missense variant. MLH1 c.677G>T was not observed in large population cohorts (Lek 2016). The nucleotide which is altered, a guanine (G) at base 677, is conserved across species. Based on the current evidence, we consider this variant to be pathogenic.
Invitae	RCV000524313	SCV000543530	pathogenic	Hereditary nonpolyposis colorectal neoplasms	2023-10-22	criteria provided, single submitter	clinical testing	This sequence change replaces arginine, which is basic and polar, with leucine, which is neutral and non-polar, at codon 226 of the MLH1 protein (p.Arg226Leu). RNA analysis indicates that this missense change 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 missense change has been observed in individual(s) with clinical features of Lynch syndrome (PMID: 8566964, 12655568, 16830052, 17510385, 18383312, 20223024; Invitae). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 90319). An algorithm developed specifically for the MLH1 gene suggests that this missense change is likely to be deleterious (PMID: 18383312). Experimental studies have shown that this missense change affects MLH1 function (PMID: 17510385). 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 results in skipping of exon 8 and introduces a premature termination codon (Invitae). The resulting mRNA is expected to undergo nonsense-mediated decay. This variant disrupts the c.677G nucleotide in the MLH1 gene. Other variant(s) that disrupt this nucleotide have been determined to be pathogenic (PMID: 12547705, 15300854, 16341550, 18561205, 21034533). This suggests that this nucleotide is clinically significant, and that variants that disrupt this position are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic.
GeneKor MSA	RCV000708610	SCV000821736	likely pathogenic	Hereditary cancer-predisposing syndrome	2020-01-01	criteria provided, single submitter	clinical testing	This variation is a single nucleotide substitution, resulting in the replacement of Arginine with Leucine at codon 226 of the MLH1 protein. The arginine residue is conserved among species and is located in a functional domain of the protein. There is a large physiochemical difference between leucine and leucine (Grantham Score 102).This variant has been reported in the literature in in multiple individuals affected with Lynch syndrome (PMID: 18383312, PMID: 20223024, PMID: 16830052, PMID: 12655568). Loss of MLH1 protein and microsatellite instability has been detected in many of these cases in the tumor tissue (PMID: 18772310). Algorithms developed to predict the effect of missense changes on protein structure and function suggest that this variant may be damaging to the protein. In vitro functional assays showed reduced mismatch repair activity and possibly reduced protein expression (PMID: 17510385 ).The mutation databases (ClinVar, InSiGHT) contain entries for this variant.
Human Genome Sequencing Center Clinical Lab, Baylor College of Medicine	RCV000709741	SCV000840010	likely pathogenic	Colorectal cancer, hereditary nonpolyposis, type 2	2018-04-27	criteria provided, single submitter	clinical testing	A heterozygous c. 677G>T (p.Arg226Leu) pathogenic variant in the MLH1 gene was detected in this individual. This variant has been previously described as disease-causing (PMID: 18383312, 16830052, 8566964, 12655568). Therefore, we consider this variant to be likely pathogenic. [alaimo, 2018-01-24]
Ambry Genetics	RCV000708610	SCV001187870	pathogenic	Hereditary cancer-predisposing syndrome	2022-03-29	criteria provided, single submitter	clinical testing	The c.677G>T pathogenic mutation (also known as p.R226L), located in coding exon 8 of the MLH1 gene, results from a G to T substitution at nucleotide position 677. This change occurs in the last base pair of coding exon 8, which makes it likely to have some effect on normal mRNA splicing. This mutation has been reported in multiple families meeting either Amsterdam diagnostic criteria or Bethesda guidelines for Lynch syndrome (Maliaka YK et al. Hum. Genet., 1996 Feb;97:251-5; Kurzawski G et al. Clin. Genet., 2006 Jan;69:40-7; Alemayehu A et al. Genes Chromosomes Cancer. 2008 Oct;47(10):906-14; Wagner A et al. Am J Hum Genet. 2003 May;72(5):1088-100; Bartosova Z et al. Hum. Mutat. 2003 Apr;21(4):449; Zavodna K et al. Neoplasma 2006 ;53(4):269-76; Evans DG et al. J. Natl. Cancer Inst. 2001 May;93(9):716-7). Functional RNA studies have demonstrated abnormal splicing for this variant with out-of-frame exon 8 skipping (Ambry internal data; Kurzawski G et al. Clin. Genet., 2006 Jan;69:40-7). In addition, in an in vitro complementation assay, the p.R226L variant showed decreased MMR activity (Takahashi et al. Cancer Res. 2007. 67(10):4595-4604). Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation.
CeGaT Center for Human Genetics Tuebingen	RCV000160555	SCV001248017	pathogenic	not provided	2017-05-01	criteria provided, single submitter	clinical testing
Quest Diagnostics Nichols Institute San Juan Capistrano	RCV000160555	SCV002047319	pathogenic	not provided	2021-02-16	criteria provided, single submitter	clinical testing	This variant has been reported in individuals affected with Lynch syndrome or related cancers in the published literature (PMID: 18772310 (2008), 20223024 (2006), 16830052 (2006), 12655568 (2003), 8566964 (1996)). Additionally, functional studies have determined that this variant interferes with normal MLH1 mRNA splicing and is damaging to the mismatch repair (MMR) activity of the MLH1 protein (PMID: 29505604 (2018), 18561205 (2008), 17510385 (2007)). Based on the available information, the variant is classified as pathogenic.
Sema4, Sema4	RCV000708610	SCV002528769	pathogenic	Hereditary cancer-predisposing syndrome	2020-10-20	criteria provided, single submitter	curation
Baylor Genetics	RCV000709741	SCV004193067	pathogenic	Colorectal cancer, hereditary nonpolyposis, type 2	2021-08-28	criteria provided, single submitter	clinical testing
Color Diagnostics, LLC DBA Color Health	RCV000708610	SCV004359196	pathogenic	Hereditary cancer-predisposing syndrome	2021-11-22	criteria provided, single submitter	clinical testing	This missense variant replaces arginine with leucine at codon 226 of the MLH1 protein. Computational prediction suggests that this variant may have deleterious impact on protein structure and function (internally defined REVEL score threshold >= 0.7, PMID: 27666373). Functional RNA studies or assays of patient RNA or cDNA have shown that this variant causes exon 8 deletion predicted to result in a premature truncation (PMID: 15300854,16341550, 18561205, 29505604). The variant protein in HCT116 cells was reported to have 32% of wild-type DNA mismatch repair (PMID: 17510385). This variant has been reported in individuals affected with Lynch syndrome (PMID: 8566964, 15300854, 16341550 16830052, 20223024, 21034533, 30256826). 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.
Department of Pathology and Laboratory Medicine, Sinai Health System	RCV000160555	SCV000592367	uncertain significance	not provided		no assertion criteria provided	clinical testing
Constitutional Genetics Lab, Leon Berard Cancer Center	RCV001249952	SCV001423894	likely pathogenic	Lynch-like syndrome	2019-07-01	no assertion criteria provided	clinical testing