Classification rationale

PM2 BP4 Uncertain Significance - Conflicting Evidence

MSH6 c.533G>T

c.533G>T (p.Arg178Leu) is a missense variant in exon 3 of MSH6. The variant is extremely rare in population databases (gnomAD v4.1 allele frequency = 1.24e-6; 2/1,614,132 alleles, 0 homozygotes), meeting PM2_Supporting per the InSiGHT MSH6 VCEP threshold of <0.00002.¹ Multiple lines of computational evidence support a benign effect: HCI prior probability for pathogenicity = 0.0003 (meeting BP4_Supporting threshold <0.11), REVEL score = 0.084, BayesDel score = -0.372, and SpliceAI max delta = 0.00.² In ClinVar (Variation ID 483842), the variant is classified as Uncertain Significance by the majority of clinical laboratories (3 VUS, 1 Likely Benign, 1 Benign), with review status 'criteria provided, single submitter' and no expert panel review.³ No variant-specific functional studies (PS3/BS3), segregation data (PP1/BS4), tumor phenotype data (PP4/BP5), de novo observations (PS2), or pathogenic comparator variants at the same residue (PM5/PS1) were identified. Applying the InSiGHT MSH6 VCEP v2.0.0 combining rules: one pathogenic supporting criterion (PM2_Supporting) and one benign supporting criterion (BP4_Supporting) yields conflicting evidence, classifying this variant as Uncertain Significance (Rule 31: >=1 Benign Supporting AND >=1 Pathogenic Supporting).⁴

PM2 + BP4 → Uncertain Significance - Conflicting Evidence

1 gnomad_v4 ↗

2 hci_priorrevelbayesdelspliceai ↗

3 clinvar ↗

4 cspec ↗

Gene diagram · NM_000179.3 · variants mapped to exon structure

MSH6 NM_000179.3

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 an allele frequency of 1.24e-6 (2/1,614,132 alleles), which is below the VCEP PM2_Supporting threshold of <0.00002 (<1 in 50,000 alleles). Absent in gnomAD-Canada.

gnomAD v4.1: AF = 1.23906e-6 (2/1614132 alleles

gnomad_v4 ↗gnomad_canada ↗

✓

BP4 supporting Benign

The HCI prior probability for pathogenicity is 0.0003 for p.Arg178Leu in MSH6, which is well below the VCEP BP4_Supporting threshold of <0.11. This is further supported by a REVEL score of 0.084, BayesDel score of -0.372, and SpliceAI max delta score of 0.00, all consistent with a benign computational profile.

HCI prior probability = 0.0003 (<0.11 = BP4_Supporting). REVEL = 0.084 (benign range). BayesDel = -0.372 (benign range). SpliceAI max delta = 0.00 (no predicted splicing impact).

spliceai ↗

Assessed · not applied

Pathogenic

PS1 No alternative nucleotide change encoding p.Arg178Leu has been previously classified as Pathogenic or Likely Pathogenic by the InSiGHT VCEP.

PS2 No de novo observations are documented for this variant.

PS3 No variant-specific calibrated functional assay data are available for p.Arg178Leu.

PM5 No different missense variant at amino acid residue Arg178 has been classified as Pathogenic or Likely Pathogenic by the InSiGHT VCEP.

PP1 No co-segregation data are available for c.533G>T (p.Arg178Leu).

PP3 The HCI prior probability for pathogenicity is 0.0003 for p.Arg178Leu in MSH6, which is well below the VCEP PP3_Supporting threshold (>0.68).

PP4 No tumor phenotype data are available for this variant.

Benign

BA1 The gnomAD v4.1 allele frequency is 1.24e-6 (0.000124%), which is far below the VCEP BA1 stand-alone threshold of >=0.0022 (0.22%).

BS1 The gnomAD v4.1 allele frequency is 1.24e-6 (0.000124%), which is below the VCEP BS1_Strong threshold range of >=0.00022 (0.022%) to <0.0022 (0.22%).

BS2 No evidence of co-occurrence in trans with a known pathogenic MSH6 variant has been reported.

BS3 No variant-specific functional studies demonstrating benign effect are available for p.Arg178Leu.

BS4 No segregation data are available to assess lack of co-segregation with disease.

BP5 No tumor data are available to apply BP5.

N/A · 11 PVS1 · PS4 · PM1 · 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= 1.23906e-06; MAF= 0.00012%, 2/1614132 alleles, homozygotes = 0) and has highest observed frequency in the Middle Eastern population (AF= 0.000164962; MAF= 0.01650%, 1/6062 alleles, homozygotes = 0).

v2.1

This variant is present in gnomAD v2.1 (AF= 7.95393e-06; MAF= 0.00080%, 2/251448 alleles, homozygotes = 0) and has highest observed frequency in the African/African American population (AF= 6.15233e-05; MAF= 0.00615%, 1/16254 alleles, homozygotes = 0).

🇨🇦 CA

Absent from gnomAD-Canada v1.0.

Allele frequency by ancestry

three datasets · side by side

gnomAD v4.1

0.00012% · 2 / 1,614,132
0 hom

Middle Eastern 1 / 6,062	0.016%
African/African American 1 / 75,028	0.0013%

+ 8 not observed (Remaining individuals, Admixed American, European (Finnish), Amish, East Asian, South Asian, Ashkenazi Jewish, European (non-Finnish))

gnomAD v2.1

0.0008% · 2 / 251,448
0 hom

African/African American 1 / 16,254	0.0062%
European (non-Finnish) 1 / 113,732	0.00088%

+ 6 not observed (Admixed American, Ashkenazi Jewish, East Asian, European (Finnish), Remaining individuals, South Asian)

gnomAD Canada 🇨🇦

Absent · 0 / ?
0 hom

Not observed in any ancestry group.

gnomAD v4 ↗ gnomAD v2 ↗ gnomAD 🇨🇦 ↗

ClinVar

This variant has been reported in ClinVar as Uncertain significance (3 clinical laboratories) and as Likely benign (1 clinical laboratory) and as Benign (1 clinical laboratory) and as Uncertain Significance (1 clinical laboratory). (ClinVarID = 483842)

ClinVar ↗

In silico

SpliceAI predicts no significant splice impact for this variant (max delta score = 0.00). REVEL score = 0.084. BayesDel score = -0.372375. HCI prior probability for pathogenicity = 0.0003. MAPP score = 1.62. Custom PP2 score = 0.001.

SpliceAI ↗

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. MSH6, a DNA mismatch repair protein, is frequently mutated in colorectal, small bowel, and endometrial cancers.

OncoKB ↗

COSMIC

Cancer hotspots

Somatic evidence Not in COSMIC / hotspots

COSMIC

This variant does not lie in a statistically significant hotspot. This variant has not previously been reported in somatic cancers (COSMIC).

Hotspots

This variant does not lie in a statistically significant hotspot.

COSMIC ↗ Hotspots ↗

Sources & reference links

9Sources

CSpec VCEP

ClinVar

gnomAD v2.1

gnomAD v4.1

gnomAD-Canada

SpliceAI

OncoKB

COSMIC

Cancer hotspots

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

31672839 ↗ Management of patients with increased risk for familial pancreatic cancer: updated recommendations from the International Cancer of the Pancreas Screening (CAPS) Consortium. CLINVAR

15604628 ↗ Genetic cancer risk assessment and counseling: recommendations of the national society of genetic counselors. CLINVAR

23535968 ↗ Informing family members of individuals with Lynch syndrome: a guideline for clinical geneticists. CLINVAR

25356965 ↗ ACMG policy statement: updated recommendations regarding analysis and reporting of secondary findings in clinical genome-scale sequencing. CLINVAR

25452455 ↗ Hereditary colorectal cancer syndromes: American Society of Clinical Oncology Clinical Practice Guideline endorsement of the familial risk-colorectal cancer: European Society for Medical Oncology Clinical Practice Guidelines. CLINVAR

28492532 ↗ Sherloc: a comprehensive refinement of the ACMG-AMP variant classification criteria. CLINVAR