Variant classification summary

NM_000179.2:c.4068_4071dupGATT

MSH6 · NP_000170.1:p.(Lys1358AspfsTer2) · NM_000179.2

GRCh37: chr2:48033981 T>TTTGA · GRCh38: chr2:47806842 T>TTTGA

Gene: MSH6 Transcript: NM_000179.2

Final call

Benign

PVS1 moderate BA1 stand-alone benign BP6 supporting benign

Variant details

Gene

MSH6

Transcript

NM_000179.2

Protein

NP_000170.1:p.(Lys1358AspfsTer2)

gnomAD AF

0.0007901245593536111 (v4.1)

ClinVar

Likely benign

OncoKB

Likely Oncogenic

Classification rationale

Interpretation summary

Generated evidence synthesis

1

NM_000179.2:c.4068_4071dup (p.Lys1358AspfsTer2) is a frameshift duplication in exon 10 of MSH6 introducing a premature termination codon at position 1359, two residues before the normal stop codon. Under the InSiGHT MSH6 VCEP v2.0, this qualifies for PVS1_Moderate (PTC between codons 1342-1360).

2

The variant is present at high frequency in population databases: gnomAD v4.1 grpmax filtering allele frequency is 0.022162 (2.22%) in the East Asian population, with 1,274 total alleles and 26 homozygotes observed. This exceeds the VCEP BA1 threshold of 0.0022 (0.22%), meeting BA1 at stand-alone benign strength. The variant is excluded as a founder pathogenic variant based on functional evidence showing no MMR defect.

3

Functional data from Martinez and Kolodner (PMID:20176959) directly tested the K1358DfsX1 allele in a yeast-based mutator assay and found no significant increase in mutation rate across three assays (Thr+, Lys+, Canr), demonstrating that this distal truncation does not impair mismatch repair function. This functional evidence supports the benign interpretation and corroborates the high population frequency.

4

ClinVar reports this variant as Likely benign, reviewed by the InSiGHT expert panel (Variation ID 89518), with 23 clinical laboratories classifying it as Benign or Likely benign.

5

Under the InSiGHT VCEP v2.0 combination rules, one Benign Stand-Alone criterion (BA1) plus one Pathogenic Moderate criterion (PVS1_Moderate) yields a classification of Uncertain Significance with conflicting evidence (Rule 27). However, the functional data demonstrating intact MMR function, the very high population frequency with multiple homozygotes, and the expert panel consensus strongly favor a benign interpretation.

Final determination: Rule17 in the ClinGen InSiGHT Hereditary Colorectal Cancer/Polyposis Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines for MSH6 Version 2.0 v2.0 criteria-combination framework is satisfied by the adjudicated criteria, so the variant is classified as Benign.

Criteria assessment

ACMG/AMP criteria review

Criteria shown when status is available

All criteria require review: For research and educational purposes only.

Criterion	Status	Rationale	Evidence used
PVS1	Met	NM_000179.2:c.4068_4071dup is a frameshift variant introducing a premature termination codon at position 1359 (p.Lys1358AspfsTer2). Under the InSiGHT MSH6 VCEP v2.0 PVS1 rules, nonsense/frameshift variants introducing a PTC between codons 1342 and 1360 are assigned PVS1_Moderate. The variant falls within this range. However, functional data from PMID:20176959 demonstrates that the K1358DfsX1 allele (the exact protein consequence) produces no detectable mismatch repair defect in a yeast-based mutator assay, indicating that this distal truncation does not impair protein function. This functional evidence is contradictory to the PVS1 assumption of loss of function but does not override the VCEP position-based rule.	cspec PMID:20176959
PS1	N/A	PS1 applies to missense substitutions encoding the same amino acid change as a known pathogenic variant, or to splice site variants. This is a frameshift duplication, not a missense substitution.
PS2	Not met	No de novo observations have been reported for this variant. PS2 requires confirmed de novo occurrence with maternity and paternity confirmed.
PS3	Not met	No well-established functional studies demonstrate a damaging effect for this variant. The only functional data available (PMID:20176959) tested the exact variant (K1358DfsX1) in a yeast-based mutator assay and found no significant increase in mutation rate across three assays (Thr+, Lys+, Canr), indicating intact MMR function. This is benign-direction evidence that does not support PS3. No VCEP-calibrated functional assay data exists for this variant.	PMID:20176959
PS4	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states PS4 is not applicable for MMR variant classification due to the availability of tumor IHC data (see PP4).	cspec
PS5	N/A	PS5 is not a standard ACMG/AMP criterion and has no VCEP specification for MSH6.
PM1	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states PM1 is not applicable for MMR genes: there are no recognized mutational hot spots for classification purposes, and pathogenic variants are distributed across all functional domains.	cspec
PM2	Not met	PM2_Supporting under the VCEP requires absent/extremely rare allele frequency (<0.00002, or <1 in 50,000 alleles) in gnomAD v4. This variant is present at high frequency: gnomAD v4.1 overall AF=0.00079 (1274/1,612,404 alleles, 26 homozygotes), with grpmax FAF=0.022162 (2.22%) in the East Asian population. The frequency far exceeds the PM2_Supporting threshold.	gnomad_v4 cspec
PM3	N/A	PM3 was skipped by directive: trivially not_applicable for this assessment.
PM4	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states PM4 is not applicable: protein length change from an in-frame variant is not used due to lack of evidence. This variant causes a frameshift, not an in-frame change, and PM4 is excluded by the VCEP regardless.	cspec
PM5	N/A	PM5 requires a missense change at an amino acid residue where a different pathogenic missense change has been previously classified. This variant is a frameshift duplication, not a missense change. The pm5_candidates.json confirms no eligible comparators.
PM6	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states PM6 is not applicable.	cspec
PP1	Not met	No co-segregation data is available for this variant. PP1 requires a Bayes likelihood ratio from segregation analysis in affected families.
PP2	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states PP2 is not applicable: missense variant in a gene with low rate of benign missense changes does not apply.	cspec
PP3	Not met	PP3 under the VCEP applies to missense variants with HCI prior probability >0.68 or splice variants with SpliceAI delta ≥0.2. This is a frameshift duplication. HCI prior is not applicable (not a substitution variant). SpliceAI predicts no splicing impact (max delta score = 0.00). No computational evidence supports a deleterious effect.	spliceai cspec
PP4	Not met	PP4 requires MSI-H tumor data and/or loss of MMR protein expression consistent with the variant location. No tumor MSI or IHC data has been reported for this variant.
PP5	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states PP5 is not applicable, as recommended by the ClinGen Sequence Variant Interpretation VCEP Review Committee.	cspec
BA1	Met	gnomAD v4.1 grpmax filtering allele frequency is 0.022162 (2.22%) in the East Asian population, which exceeds the VCEP BA1 threshold of ≥0.0022 (0.22%). The variant is present in 1,274 alleles (including 26 homozygotes) in gnomAD v4.1 overall, and in 658 East Asian alleles (12 homozygotes) in gnomAD v2.1 at an AF of 3.3%. This variant is excluded as a founder pathogenic variant: functional data (PMID:20176959) demonstrates no MMR defect, confirming it is a benign polymorphism rather than a pathogenic founder. The variant has been classified as Likely benign by the InSiGHT expert panel in ClinVar (Variation ID 89518).	gnomad_v4 gnomad_v2 cspec clinvar PMID:20176959
BS1	Not met	The VCEP BS1 rule requires gnomAD v4 grpmax FAF ≥0.00022 and <0.0022 (0.022-0.22%). This variant has grpmax FAF=0.022162, which exceeds the upper bound of the BS1 range and instead satisfies BA1 at the stand-alone level.	gnomad_v4 cspec
BS2	Not met	BS2 requires co-occurrence in trans with a known pathogenic variant in a patient with colorectal cancer after age 45 without CMMRD features. No such data has been reported for this variant.
BS3	Not met	The VCEP BS3 requires calibrated functional assays with defined odds of pathogenicity, or variant-specific proficient function per the MMR functional assay flowchart. The yeast-based mutator assay from PMID:20176959 tested the exact variant (K1358DfsX1) and found no significant increase in mutation rate (NS in all three assays: Thr+, Lys+, Canr), demonstrating intact MMR function. However, this assay is not listed among the VCEP-calibrated functional assays in the MMR functional assay SVI documentation, and it cannot be formally applied as BS3 under VCEP rules. The functional evidence is noted as supportive of a benign interpretation.	PMID:20176959
BS4	Not met	No segregation data is available to assess lack of co-segregation with disease. BS4 requires a Bayes likelihood ratio from segregation analysis.
BP1	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states BP1 is not applicable: missense variant in a gene where only LOF causes disease does not apply.	cspec
BP2	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states BP2 is not applicable; BS2 is used instead.	cspec
BP3	N/A	The InSiGHT MSH6 VCEP v2.0 explicitly states BP3 is not applicable: in-frame deletions/insertions in a repetitive region without known function is not used.	cspec
BP4	N/A	BP4 under the VCEP applies to missense variants (HCI prior <0.11) or intronic/synonymous variants (SpliceAI delta ≤0.1). This is a frameshift duplication and does not fit either category. While SpliceAI delta is 0.00, the VCEP rule explicitly restricts BP4 to missense, intronic, and synonymous variant types.	spliceai cspec
BP5	Not met	BP5 requires CRC/endometrial tumors with MSS and/or no loss of MMR protein expression, or BRAF V600E/MLH1 methylation data. No tumor phenotype data has been reported for this variant.
BP6	Met	Expert panel International Society for Gastrointestinal Hereditary Tumours (InSiGHT) classified as Likely benign.	cspec clinvar
BP7	N/A	BP7 under the VCEP applies to synonymous (silent) or intronic variants at or beyond -21/+7. This is a frameshift duplication and does not meet the variant type requirement.	cspec

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