NM_000249.3:c.1738G>A (p.Ala580Thr) is a missense variant in MLH1 exon 16. It is extremely rare in population databases (gnomAD v4.1 grpmax FAF = 2.8e-07), meeting PM2_Supporting per the InSiGHT VCEP v2.0 specification.1 In silico analysis yields an HCI prior probability of 0.8146, which falls in the PP3_Supporting range (>0.68 and ≤0.81) per the InSiGHT VCEP v2.0 specification. SpliceAI predicts no splicing impact (max delta score = 0.00). REVEL score is 0.703.2 No variant-specific functional data, co-segregation data, tumor phenotype data, or de novo observations were identified in the reviewed literature or ClinVar submissions. ClinVar reports this variant as Uncertain Significance (6 submissions; review status: criteria provided, single submitter).3 Under the InSiGHT VCEP v2.0 framework, the only criteria met are PM2_Supporting and PP3_Supporting. Multiple criteria are designated as Not Applicable per the VCEP specification (PS4, PM1, PM6, PP2, PP5, BP1, BP2, BP6, PM4) or are not applicable to a missense substitution (PVS1, BP3, BP7). No benign criteria are met. With only two supporting pathogenic criteria and no moderate, strong, or very strong criteria met, the variant remains as Uncertain Significance per the VCEP combination rules.4
MLH1
Final classification
VUS
MLH1 c.1738G>A · p.Ala580Thr
MLH1
NM_000249.3:c.1738G>A (p.Ala580Thr) is a missense variant in MLH1 exon 16. It is extremely rare in population databases (gnomAD v4.1 grpmax FAF = 2.8e-07), meeting PM2_Supporting per the InSiGHT VCEP v2.0 specification.
ClinGen InSiGHT Hereditary Colorectal Cancer/Polyposis Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines for MLH1 Version 2.0 v2.0 criteria-combination framework was evaluated deterministically with applied criteria: PM2 supporting, PP3 supporting; no rule matched the adjudicated criteria.
Classification rationale
PM2PP3
VUS
MLH1 c.1738G>A
PM2 + PP3
→
VUS
Gene diagram
· NM_000249.3 · variants mapped to exon structure
MLH1
NM_000249.3
Fetching transcript structure from UCSC…
Applied criteria · 2 applied · 13 assessed
Applied · 2
Strength
Supporting
Moderate
Strong
Very strong
✓
PM2
supporting
Pathogenic
The variant is extremely rare in gnomAD v4.1 with a grpmax filtering allele frequency of 2.8e-07 (2/1,614,160 alleles, 0 homozygotes), which is well below the VCEP PM2_Supporting threshold of <0.00002 (<1 in 50,000 alleles). Absent from gnomAD v2.1.
gnomAD v4.1 grpmax FAF = 2.8e-07well below the VCEP threshold of 0.00002.Absent from gnomAD v2.1.
✓
PP3
supporting
Pathogenic
The HCI prior probability for pathogenicity for p.Ala580Thr is 0.8146, which falls in the PP3_Supporting range (>0.68 and ≤0.81) per the InSiGHT VCEP v2.0 specification. SpliceAI predicts no splicing impact (max delta score 0.00), so the splice-based PP3 rule does not apply. REVEL score is 0.703 and BayesDel score is 0.291.
HCI prior probability = 0.8146meeting VCEP PP3_Supporting threshold (>0.68≤0.81).
Assessed · not applied
Pathogenic
PS1
No evidence that a different underlying nucleotide change producing the same amino acid substitution (p.Ala580Thr) has been classified as Pathogenic or Likely Pathogenic by this VCEP.
PS2
No de novo occurrence data were identified for NM_000249.3:c.1738G>A in the reviewed literature or ClinVar submissions.
PS3
No variant-specific functional data were identified for p.Ala580Thr (NM_000249.3:c.1738G>A) in the reviewed literature, the VCEP calibrated functional assay documentation, or OncoKB.
PM5
No different missense change at amino acid residue 580 was identified that has been classified as Pathogenic or Likely Pathogenic by this VCEP.
PP1
No co-segregation data were identified for this variant in the reviewed literature or ClinVar submissions.
PP4
No tumor data (MSI status, MMR protein expression by IHC) were identified for patients carrying NM_000249.3:c.1738G>A in the reviewed literature or ClinVar submissions.
Benign
BA1
The gnomAD v4.1 grpmax filtering allele frequency is 2.8e-07, which is far below the VCEP BA1 threshold of ≥0.001 (0.1%).
BS1
The gnomAD v4.1 grpmax filtering allele frequency is 2.8e-07, which is below the VCEP BS1 threshold of ≥0.0001 (0.01%).
BS2
No observation of co-occurrence in trans with a known pathogenic MLH1 variant was identified in the reviewed literature or ClinVar submissions.
BS3
No functional data demonstrating a benign effect for p.Ala580Thr were identified in the reviewed literature, the VCEP calibrated functional assay documentation, or OncoKB.
BS4
No lack of co-segregation data were identified for this variant.
BP4
The HCI prior probability for pathogenicity for p.Ala580Thr is 0.8146, which is well above the VCEP BP4_Supporting threshold of <0.11.
BP5
No tumor data demonstrating MSS status or retained MMR protein expression were identified for patients carrying this variant.
N/A · 13
PVS1 · PS4 · PM1 · PM3 · PM4 · PM6 · PP2 · PP5 · BP1 · BP2 · BP3 · BP6 · BP7
Research & evidence
Population frequency
gnomAD v4.1
gnomAD v2.1
v4.1
This variant is present in gnomAD v4.1 (AF= 1.23903e-06; MAF= 0.00012%, 2/1614160 alleles, homozygotes = 0) and has highest observed frequency in the European (non-Finnish) population (AF= 1.69493e-06; MAF= 0.00017%, 2/1179992 alleles, homozygotes = 0); grpmax FAF= 2.8e-07.
v2.1
Absent from gnomAD v2.1.
🇨🇦 CA
Absent from gnomAD-Canada v1.0.
Allele frequency by ancestry
three datasets · side by side
gnomAD v4.1
0.00012%
· 2 / 1,614,160
0 hom · FAF 2.8e-05%
0 hom · FAF 2.8e-05%
European (non-Finnish) 2 / 1,179,992 |
0.00017% |
+ 9 not observed (Remaining individuals, Admixed American, European (Finnish), Amish, East Asian, Middle Eastern, South Asian, Ashkenazi Jewish, African/African American)
gnomAD v2.1
Absent
· 0 / ?
0 hom
0 hom
Not observed in any ancestry group.
gnomAD Canada 🇨🇦
Absent
· 0 / ?
0 hom
0 hom
Not observed in any ancestry group.
ClinVar
This variant has been reported in ClinVar as Uncertain significance (4 clinical laboratories) and as Uncertain Significance (1 clinical laboratory) and as Likely benign (1 clinical laboratory). (ClinVarID = 405428)
In silico
SpliceAI predicts no significant splice impact for this variant (max delta score = 0.00). REVEL score = 0.703. BayesDel score = 0.291065. HCI prior probability for pathogenicity = 0.8146. MAPP score = 33.24. Custom PP2 score = 0.64.
Functional
Unknown Oncogenic Effect
OncoKB did not identify variant-specific reviewed functional evidence for this variant; gene-level curated context is available for reviewer follow-up. MLH1, a DNA mismatch repair protein, is recurrently altered by deletion and mutation in various cancer types.
COSMIC
Cancer hotspots
Somatic evidence
Not in COSMIC / hotspots
COSMIC
This variant does not lie in a statistically significant hotspot. This variant has previously been reported in somatic cancers (COSMIC; COSV51614640, n = 2 times).
Hotspots
This variant does not lie in a statistically significant hotspot.
Sources & reference links
9Sources
Triaged references · 7 PMIDs not cited in assessment
25741868 ↗
Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology.
CLINVAR
15604628 ↗
Genetic cancer risk assessment and counseling: recommendations of the national society of genetic counselors.
CLINVAR
23788249 ↗
ACMG recommendations for reporting of incidental findings in clinical exome and genome sequencing.
CLINVAR
24310308 ↗
ACMG technical standards and guidelines for genetic testing for inherited colorectal cancer (Lynch syndrome, familial adenomatous polyposis, and MYH-associated polyposis).
CLINVAR
25394175 ↗
A practice guideline from the American College of Medical Genetics and Genomics and the National Society of Genetic Counselors: referral indications for cancer predisposition assessment.
CLINVAR
28492532 ↗
Sherloc: a comprehensive refinement of the ACMG-AMP variant classification criteria.
CLINVAR