Variant classification summary

NM_000251.2:c.367-1G>A

MSH2 · NP_000242.1:p.? · NM_000251.2

GRCh37: chr2:47637232 G>A · GRCh38: chr2:47410093 G>A

Gene: MSH2 Transcript: NM_000251.2

Final call

Pathogenic

PVS1 very strong PM2 supporting PP5 supporting

Variant details

Gene

MSH2

Transcript

NM_000251.2

Protein

NP_000242.1:p.?

gnomAD AF

ClinVar

Likely pathogenic

OncoKB

Classification rationale

Interpretation summary

Generated evidence synthesis

1

NM_000251.2:c.367-1G>A is a canonical splice acceptor variant (IVS2-1G>A) disrupting the invariant AG dinucleotide of intron 2 in MSH2, a gene for which loss of function is an established mechanism for Lynch syndrome.

2

SpliceAI predicts strong splice disruption with a maximum delta score of 0.99 (acceptor loss 0.99), and cDNA analysis from patient lymphocytes (Wolf et al. 2005, PMID:15926618) confirmed aberrant splicing: the variant creates a new splice acceptor 1 bp downstream, resulting in deletion of a guanine (r.367delg) and a frameshift (p.122fsX173) predicted to introduce a premature termination codon subject to nonsense-mediated decay.

3

Under InSiGHT MMR VCEP v2.0, canonical splice site variants at IVS+/-1 or IVS+/-2 where exon skipping or cryptic splice site use disrupts the reading frame and is predicted to undergo NMD qualify for PVS1 at very_strong strength. The premature termination codon at position 173 is well upstream of the MSH2 NMD boundary at codon 891.

4

The variant is absent from gnomAD v2.1, v4.1, and gnomAD-Canada, meeting the VCEP PM2_Supporting threshold of <0.00002 allele frequency. It was also absent from 100 healthy control alleles in the Wolf et al. study.

5

ClinVar classifies this variant as Likely pathogenic (Variation ID 91075), reviewed by the InSiGHT expert panel, consistent with the VCEP assessment.

6

No benign or conflicting evidence was identified. BS3 is contradicted by the confirmed splicing aberration. BP4 is contradicted by the strong SpliceAI prediction. PP3 is not applied per VCEP rules prohibiting combination with PVS1 for canonical splice variants.

7

Under InSiGHT MMR VCEP v2.0 combination rules (Rule 1): 1 PVS1_VeryStrong criterion is sufficient for Pathogenic classification.

Final determination: Rule4 in the ClinGen InSiGHT Hereditary Colorectal Cancer/Polyposis Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines for MSH2 Version 2.0 v2.0 criteria-combination framework is satisfied by the adjudicated criteria, so the variant is classified as Pathogenic.

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	Canonical splice acceptor variant (IVS2-1G>A, NM_000251.2:c.367-1G>A) disrupting the invariant AG dinucleotide. cDNA analysis from patient blood (Wolf et al., PMID:15926618) confirmed the variant generates a new splice acceptor site 1 bp downstream, resulting in deletion of a guanine (r.367delg) and a frameshift predicted to produce a premature termination codon (p.122fsX173) subject to nonsense-mediated decay. Under InSiGHT MMR VCEP v2.0 PVS1 rules, IVS+/-1 or IVS+/-2 variants where exon skipping or cryptic splice site use disrupts the reading frame and is predicted to undergo NMD qualify for PVS1 at very_strong strength. Confirmed by patient-derived mRNA assay; no additional independent confirmation required as this is a canonical splice site variant.	PMID:15926618 spliceai vcep_pvs1_decisiontree_mmr pvs1_gene_context
PS1	N/A	PS1 under InSiGHT MMR VCEP v2.0 applies to (a) missense substitutions encoding the same amino acid change as a known pathogenic variant, or (b) variants affecting the same non-canonical splice nucleotide as a confirmed pathogenic splice variant. This variant is a canonical splice site change (IVS-1), not a missense or non-canonical splice variant. PS1 is not applicable.
PS2	Not met	No de novo occurrence data identified in the literature for NM_000251.2:c.367-1G>A. Under VCEP PS2 rules, de novo points require confirmed maternity and paternity in a proband with MMR-deficient LS spectrum tumor. No such reports found.
PS3	Not met	Under InSiGHT MMR VCEP v2.0, PS3 requires calibrated functional assay data with defined odds of pathogenicity (as documented in Functional-assay-SVI-documentation-MMR.xlsx) or MMR function defect per the functional assay flowchart. The cDNA splicing analysis by Wolf et al. (PMID:15926618) confirms a splicing aberration at the mRNA level, but this is not a calibrated protein-level functional assay and does not meet VCEP PS3 calibrated assay requirements. The splicing evidence is appropriately captured under PVS1. No calibrated functional assay data available for this variant.	vcep_functional_assay_svi_documentation_mmr PMID:15926618
PS4	N/A	PS4 is not applicable under the InSiGHT MMR VCEP v2.0 framework for MSH2.
PS5	N/A	PS5 is not defined in the InSiGHT MMR VCEP v2.0 criteria set for MSH2.
PM1	N/A	PM1 is designated as Not Applicable by the InSiGHT MMR VCEP v2.0 for MSH2. Domain-level criteria are not applied under this VCEP framework.
PM2	Met	Absent from gnomAD v2.1 (exomes), gnomAD v4.1, and gnomAD-Canada v1.0. Under InSiGHT MMR VCEP v2.0, PM2_Supporting applies when allele frequency is <0.00002 (<1 in 50,000 alleles) in gnomAD v4. The variant is completely absent from all population databases, satisfying this threshold.	gnomad_v2 gnomad_v4 PMID:15926618
PM5	N/A	PM5 under InSiGHT MMR VCEP v2.0 applies only to missense changes at an amino acid residue where a different missense change was classified as Pathogenic or Likely Pathogenic. NM_000251.2:c.367-1G>A is a canonical splice site variant, not a missense change. PM5 is not applicable.
PM6	N/A	PM6 is designated as Not Applicable by the InSiGHT MMR VCEP v2.0. De novo evidence for MMR genes is assessed through the PS2 points system instead.
PP1	Not met	No co-segregation data available for NM_000251.2:c.367-1G>A. Under VCEP PP1 rules, co-segregation requires combined Bayes Likelihood Ratio calculations from pedigrees. No such data identified in the literature.
PP2	N/A	PP2 is designated as Not Applicable by the InSiGHT MMR VCEP v2.0.
PP3	N/A	Under InSiGHT MMR VCEP v2.0, PP3 applies to (a) missense variants with HCI prior probability >0.68, or (b) predicted splice defects for non-canonical splice nucleotides using SpliceAI delta score >=0.2. NM_000251.2:c.367-1G>A is a canonical splice site variant (IVS-1), not a missense or non-canonical splice variant. The VCEP PVS1 rules explicitly state that PP3 must not be combined with PVS1 for IVS+/-1 or IVS+/-2 variants. The splice prediction evidence is captured under PVS1.	spliceai vcep_hci_priors_msh2
PP4	Not met	Under InSiGHT MMR VCEP v2.0, PP4 requires CRC/endometrial MSI-H tumors with consistent loss of MMR protein expression. No patient phenotype, tumor MSI, or IHC data were available in the case materials. Wolf et al. (PMID:15926618) reported one tumor with MSI-H and one with MSS in the single family carrying this variant, which does not meet PP4 Supporting threshold (1 CRC/endometrial MSI-H tumor).	PMID:15926618
PP5	Met	Expert panel International Society for Gastrointestinal Hereditary Tumours (InSiGHT) classified as Likely pathogenic.	clinvar
BA1	Not met	Under InSiGHT MMR VCEP v2.0, BA1 requires gnomAD v4 Grpmax filtering allele frequency >=0.001 (0.1%). NM_000251.2:c.367-1G>A is absent from gnomAD v4.1 and does not meet the BA1 threshold.	gnomad_v4
BS1	Not met	Under InSiGHT MMR VCEP v2.0, BS1 requires gnomAD v4 Grpmax filtering allele frequency >=0.0001 and <0.001 (0.01-0.1%). NM_000251.2:c.367-1G>A is absent from gnomAD v4.1 and does not meet the BS1 threshold.	gnomad_v4
BS2	Not met	Under InSiGHT MMR VCEP v2.0, BS2 requires co-occurrence in trans with a known pathogenic variant in the same gene in a CRC patient after age 45 without CMMRD features. No such co-occurrence data identified for NM_000251.2:c.367-1G>A.
BS3	Not met	Under InSiGHT MMR VCEP v2.0, BS3 requires calibrated functional assays demonstrating functional odds for pathogenicity <=0.05, or synonymous/intronic variants with no associated mRNA aberration by NMD-inhibited lab assays. The cDNA analysis by Wolf et al. (PMID:15926618) demonstrates the opposite: a confirmed splicing aberration producing a frameshift (r.367delg → p.122fsX173). No calibrated functional assay data supporting normal function exists. BS3 is not met.	PMID:15926618 vcep_functional_assay_svi_documentation_mmr
BS4	Not met	Under InSiGHT MMR VCEP v2.0, BS4 requires formal co-segregation analysis demonstrating lack of co-segregation with disease (Bayes Likelihood Ratio <0.05 for strong, >0.05 and <=0.48 for supporting). No segregation data available for NM_000251.2:c.367-1G>A.
BP1	N/A	BP1 is designated as Not Applicable by the InSiGHT MMR VCEP v2.0.
BP2	N/A	BP2 is designated as Not Applicable by the InSiGHT MMR VCEP v2.0; BS2 is used instead.
BP3	N/A	In-frame deletions/insertions in repetitive regions without known function; not applicable to a canonical splice site substitution.
BP4	Not met	Under InSiGHT MMR VCEP v2.0, BP4 Supporting applies to (a) missense variants with HCI prior probability <0.11, or (b) intronic/synonymous variants with SpliceAI delta score <=0.1. NM_000251.2:c.367-1G>A is an intronic variant but SpliceAI predicts strong splice impact (max delta 0.99, acceptor loss 0.99), far exceeding the 0.1 threshold. HCI prior is not applicable for non-missense variants. BP4 is not met.	spliceai bayesdel
BP5	Not met	Under InSiGHT MMR VCEP v2.0, BP5 requires tumor data demonstrating MSS and/or no loss of MMR protein expression consistent with the variant gene, or BRAF V600E/MLH1 methylation in appropriate tumor context. No tumor phenotype data available for independent BP5 assessment.
BP6	N/A	BP6 is designated as Not Applicable by the InSiGHT MMR VCEP v2.0. The VCEP explicitly states this criterion is not for use as recommended by the ClinGen Sequence Variant Interpretation VCEP Review Committee.
BP7	Not met	Under InSiGHT MMR VCEP v2.0, BP7 applies to synonymous or intronic variants at or beyond position -21 (3' splice site) or +7 (5' splice site) from the exon boundary. NM_000251.2:c.367-1G>A is at position -1 of the splice acceptor site of intron 2, which is within the canonical splice consensus region. The -1 position is a universally conserved splice dinucleotide, not a deep intronic variant eligible for BP7.

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