LYFE Sciences · Project HERA
Variant Interpretation · Classification Report
Generated: 2026-07-27
Case ID: NM_000249.3_c.1177C_A_20260727_091454
Framework: ACMG/AMP 2015
Variant classification summary

NM_000249.3:c.1177C>A

MLH1  · NP_000240.1:p.(Leu393Ile)  · NM_000249.3
GRCh37: chr3:37067266 C>A  ·  GRCh38: chr3:37025775 C>A
Gene: MLH1 Transcript: NM_000249.3
Final call
PM2 supporting BP4 supporting benign
All criteria require review: For research and educational purposes only.
Variant details
Gene
MLH1
Transcript
NM_000249.3
Protein
NP_000240.1:p.(Leu393Ile)
gnomAD AF
ClinVar
OncoKB
Unknown Oncogenic Effect
Classification rationale
Interpretation summary
Generated evidence synthesis
1
NM_000249.3:c.1177C>A (p.Leu393Ile) in MLH1 is a missense substitution absent from gnomAD population databases (PM2_Supporting met) but with a low HCI prior probability of pathogenicity of 0.0225 (BP4_Supporting met).
2
The variant is absent from ClinVar and has not been observed in any publication. No variant-specific functional data, tumor phenotype data, segregation data, or de novo observations are available.
3
Multiple in silico predictors are indeterminate or benign: SpliceAI max delta 0.02 (no splicing impact), REVEL 0.415, BayesDel -0.07.
4
Under the InSiGHT MLH1 VCEP v2.0 framework, the evidence profile (PM2_Supporting + BP4_Supporting) is insufficient to reach a Likely Pathogenic or Likely Benign classification. The variant remains a Variant of Uncertain Significance.
Final determination: Rule31 in the 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 is satisfied by the adjudicated criteria, so the variant is classified as Uncertain Significance - Conflicting Evidence.
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 N/A Missense substitution (p.Leu393Ile) does not qualify for PVS1 under the InSiGHT MLH1 VCEP v2.0 framework, which restricts PVS1 to null variants (nonsense, frameshift, canonical splice, large deletions, or initiation codon variants).
pvs1_variant_assessment
PS1 Not met No different nucleotide change encoding the same amino acid substitution (p.Leu393Ile) has been established as pathogenic by this VCEP. The variant is absent from ClinVar entirely.
clinvar vcep_vcep_pilot_variants_mmr
PS2 Not met No de novo observations have been reported for NM_000249.3:c.1177C>A in the available literature or databases.
PS3 Not met No variant-specific functional data are available for c.1177C>A (p.Leu393Ile). The variant is not listed in the InSiGHT VCEP pilot variants spreadsheet, and no calibrated functional assay results (CIMRA, CRISPR hESC, or other MMR functional assays) were identified for this substitution. OncoKB reports Unknown Oncogenic Effect with no variant-specific reviewed functional evidence.
oncokb vcep_functional_assay_svi_documentation_mmr vcep_vcep_pilot_variants_mmr
PS4 N/A PS4 is designated Not Applicable by the InSiGHT MLH1 VCEP v2.0.
cspec
PS5 N/A PS5 is not included in the InSiGHT MLH1 VCEP v2.0 criteria set.
cspec
PM1 N/A PM1 is designated Not Applicable by the InSiGHT MLH1 VCEP v2.0.
cspec
PM2 Met NM_000249.3:c.1177C>A is absent from gnomAD v2.1 (exomes), gnomAD v4.1 (exomes), and gnomAD-Canada v1.0, meeting the InSiGHT VCEP PM2_Supporting threshold of allele frequency <0.00002 (<1 in 50,000 alleles) in gnomAD v4.
gnomad_v2 gnomad_v4 gnomad_canada
PM5 Not met No missense change at amino acid residue 393 has been classified as pathogenic or likely pathogenic by this VCEP. The PM5 candidate search yielded zero same-residue comparator variants.
pm5_candidates vcep_vcep_pilot_variants_mmr
PM6 N/A PM6 is designated Not Applicable by the InSiGHT MLH1 VCEP v2.0.
cspec
PP1 Not met No co-segregation data are available for this variant in any pedigree.
PP2 N/A PP2 is designated Not Applicable by the InSiGHT MLH1 VCEP v2.0.
cspec
PP3 Not met HCI prior probability for pathogenicity is 0.0225, well below the InSiGHT VCEP PP3 thresholds (>0.81 for moderate, >0.68 for supporting). SpliceAI predicts no splicing impact (max delta score 0.02, below the 0.2 threshold). REVEL score is 0.415 (indeterminate).
hci_prior spliceai revel bayesdel
PP4 Not met No tumor data (MSI-H status, MMR protein expression, or MLH1 promoter methylation) are available for carriers of this variant.
PP5 N/A PP5 is designated Not Applicable by the InSiGHT MLH1 VCEP v2.0.
cspec
BA1 Not met The variant is absent from gnomAD v4.1, not meeting the BA1 threshold of gnomAD v4 Grpmax filtering allele frequency >= 0.001 (0.1%).
gnomad_v4
BS1 Not met The variant is absent from gnomAD v4.1, not meeting the BS1 threshold of gnomAD v4 Grpmax filtering allele frequency >= 0.0001 and < 0.001 (0.01-0.1%).
gnomad_v4
BS2 Not met No observation of this variant in trans with a known pathogenic MLH1 variant in a patient without CMMRD features has been reported.
BS3 Not met No functional data demonstrating a benign effect for c.1177C>A (p.Leu393Ile) are available. The variant has not been tested in calibrated MMR functional assays.
vcep_functional_assay_svi_documentation_mmr oncokb
BS4 Not met No lack-of-segregation data are available for this variant.
BP1 N/A BP1 is designated Not Applicable by the InSiGHT MLH1 VCEP v2.0.
cspec
BP2 N/A BP2 is designated Not Applicable by the InSiGHT MLH1 VCEP v2.0.
cspec
BP4 Met HCI prior probability for pathogenicity is 0.0225, meeting the InSiGHT VCEP BP4_Supporting threshold of <0.11. Multiple in silico predictors also suggest a benign or indeterminate effect: SpliceAI max delta 0.02 (no splicing impact), REVEL 0.415 (indeterminate), BayesDel -0.07 (indeterminate).
hci_prior spliceai revel bayesdel
BP5 Not met No tumor data demonstrating MSS status, retained MMR protein expression, or BRAF V600E/MLH1 methylation in carriers of this variant are available.
BP6 N/A BP6 is designated Not Applicable by the InSiGHT MLH1 VCEP v2.0 per ClinGen SVI VCEP Review Committee recommendation.
cspec
BP7 N/A BP7 under the InSiGHT VCEP is restricted to synonymous (silent) or intronic variants at or beyond -21/+7 splice positions. c.1177C>A is a missense substitution (p.Leu393Ile) and does not qualify.
cspec
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