3-37001033-A-G

Variant summary

Our verdict is Likely pathogenic. Variant got 6 ACMG points: 6P and 0B. PM1 PM5 PP3_Moderate

The NM_000249.4(MLH1):c.286A>G(p.Thr96Ala) variant causes a missense change involving the alteration of a conserved nucleotide. The variant allele was found at a frequency of 0.00000549 in 1,456,706 control chromosomes in the GnomAD database, with no homozygous occurrence. In-silico tool predicts a pathogenic outcome for this variant. 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. T96A?) has been classified as Pathogenic.

• Frequency:
  • Genomes: not found (cov: 32)
  • Exomes 𝑓: 0.0000055 (0 hom.)
• Consequence: MLH1 NM_000249.4 missense
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications U:5 B:1
• Conservation: PhyloP100: 8.79

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 Likely_pathogenic. Variant got 6 ACMG points.

• PM1: In a hotspot region, there are 5 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 2 benign, 13 uncertain in NM_000249.4
• PM5: Other missense variant is known to change same aminoacid residue: Variant chr3-37001033-A-GC is described in ClinVar as [Pathogenic]. Clinvar id is 1797038.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars.
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.922

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
MLH1	NM_000249.4	c.286A>G	p.Thr96Ala	missense_variant	3/19	ENST00000231790.8

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
MLH1	ENST00000231790.8	c.286A>G	p.Thr96Ala	missense_variant	3/19	1	NM_000249.4	P1

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD3 exomes AF: 0.0000318 AC: 8 AN: 251398 Hom.: 0 AF XY: 0.0000294 AC XY: 4 AN XY: 135884

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.000231

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: 0.00

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.00000549 AC: 8 AN: 1456706 Hom.: 0 Cov.: 28 AF XY: 0.00000552 AC XY: 4 AN XY: 725148

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.000179

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.: 32

Alfa AF: 0.0000936 Hom.: 0

Bravo AF: 0.0000113

ExAC AF: 0.0000330 AC: 4

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Uncertain:5 Benign:1

Revision: criteria provided, conflicting classifications

Submissions by phenotype

not provided Uncertain:2

Uncertain significance, criteria provided, single submitter	clinical testing	GeneDx	Dec 08, 2016	This variant is denoted MLH1 c.286A>G at the cDNA level, p.Thr96Ala (T96A) at the protein level, and results in the change of a Threonine to an Alanine (ACC>GCC). This variant has not, to our knowledge, been published in the literature as pathogenic or benign. MLH1 Thr96Ala was observed at an allele frequency of 0.035% (4/11,548) in individuals of Latino ancestry in large population cohorts (NHLBI Exome Sequencing Project, The 1000 Genomes Consortium 2015, Lek 2016). Since Threonine and Alanine differ in polarity, charge, size or other properties, this is considered a non-conservative amino acid substitution. MLH1 Thr96Ala occurs at a position that is conserved across species and is located in the MLH and ATPase domains (Pang 1997, Hardt 2011). In silico analyses predict that this variant is probably damaging to protein structure and function. Based on currently available evidence, it is unclear whether MLH1 Thr96Ala is pathogenic or benign. We consider it to be a variant of uncertain significance. -
Uncertain significance, criteria provided, single submitter	clinical testing	Mayo Clinic Laboratories, Mayo Clinic	Sep 26, 2019	- -

Hereditary cancer-predisposing syndrome Uncertain:1 Benign:1

Likely benign, criteria provided, single submitter	clinical testing	Ambry Genetics	Jun 01, 2022	This alteration is classified as likely benign based on a combination of the following: seen in unaffected individuals, population frequency, intact protein function, lack of segregation with disease, co-occurrence, RNA analysis, in silico models, amino acid conservation, lack of disease association in case-control studies, and/or the mechanism of disease or impacted region is inconsistent with a known cause of pathogenicity. -
Uncertain significance, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Jan 03, 2019	- -

Colorectal cancer, hereditary nonpolyposis, type 2 Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Baylor Genetics

Oct 30, 2023

- -

Hereditary nonpolyposis colorectal neoplasms Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Invitae

Sep 29, 2023

In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance. Advanced modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) performed at Invitae indicates that this missense variant is expected to disrupt MLH1 protein function. ClinVar contains an entry for this variant (Variation ID: 188169). This variant has not been reported in the literature in individuals affected with MLH1-related conditions. This variant is present in population databases (rs770276731, gnomAD 0.02%). This sequence change replaces threonine, which is neutral and polar, with alanine, which is neutral and non-polar, at codon 96 of the MLH1 protein (p.Thr96Ala). -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.92
BayesDel_addAF	Pathogenic	0.35	D
BayesDel_noAF	Pathogenic	0.56
Cadd	Pathogenic	27
Dann	Uncertain	1.0
DEOGEN2	Pathogenic	0.96	D
Eigen	Pathogenic	1.1
Eigen_PC	Pathogenic	0.99
FATHMM_MKL	Pathogenic	0.98	D
LIST_S2	Pathogenic	0.99	D
M_CAP	Pathogenic	0.31	D
MetaRNN	Pathogenic	0.92	D
MetaSVM	Pathogenic	1.1	D
MutationAssessor	Pathogenic	4.2	H
MutationTaster	Benign	1.0	D;D;D;D;D;D
PrimateAI	Uncertain	0.63	T
PROVEAN	Uncertain	-4.1	D
REVEL	Pathogenic	0.97
Sift	Pathogenic	0.0	D
Sift4G	Uncertain	0.0040	D
Polyphen		1.0	D
Vest4		0.90
MutPred		0.70		Loss of glycosylation at T96 (P = 0.0501);
MVP		0.99
MPC		0.40
ClinPred		1.0	D
GERP RS		5.9
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.7
Varity_R		0.87
gMVP		0.89

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.0		Details are displayed if max score is > 0.2

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

Publications

LitVar

Other links and lift over

dbSNP: rs770276731; hg19: chr3-37042524;