MLH1c.381-1G>APathogenic

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Variant interpretation report

MLH1 c.381-1G>A, p.?

NM_000249.4:c.381-1G>A NP_000240.1:p.? View identifiers

Classification

Pathogenic

Variant description:

No variant summary was recorded for this case.

Five-step scale from benign to pathogenic. This variant is pathogenic. Select a step to read what it means.

Pathogenic.A pathogenic classification under the recorded framework.

In plain language

Gene-level background from the recorded interpretation.

About MLH1

No gene-level summary was recorded for this case.

Key facts

Evidence used: 1 very strong (pathogenic) · 1 moderate (pathogenic) · 1 supporting (pathogenic)
Expert rules: ClinGen MLH1 VCEP v2.0
Other laboratories (ClinVar): 4 Pathogenic · 1 Likely pathogenic
Pipeline record: 20 Jul 2026 · version 7.0.4

Evidence considered

View criteria

Applied evidence criteria

PVS1 (very strong): NM_000249.4:c.381-1G>A is a canonical splice acceptor variant at IVS4-1 (intron 4), disrupting the invariant AG dinucleotide. Under the InSiGHT/ClinGen MLH1 VCEP v2.0, IVS±1 variants where exon skipping disrupts the reading frame and is predicted to undergo NMD qualify for PVS1_Very_Strong. MLH1 loss of function is an established disease mechanism for Lynch syndrome. SpliceAI max delta score of 0.99 confirms a severe splicing impact. Not combined with PP3 per VCEP rules.
PM2 (supporting): NM_000249.4:c.381-1G>A is absent from gnomAD v2.1 (exomes), gnomAD v4.1 (exomes), and gnomAD-Canada v1.0 (genomes). Under the MLH1 VCEP, PM2_Supporting applies when the variant is absent or extremely rare with allele frequency <0.00002 (<1 in 50,000 alleles) in gnomAD v4.
PP4 (moderate): A patient with this MLH1 variant (reported as IVS4-1G>A) was described by Roth et al. (2016, PMID:27357288) with two synchronous primary colorectal cancers (cecum and rectosigmoid colon), both demonstrating loss of MLH1 and PMS2 protein expression by immunohistochemistry, consistent with the variant location. Under the MLH1 VCEP, 2 independent CRC tumors with loss of MMR protein expression consistent with the variant location qualifies for PP4_Moderate. Independent tumors can be from the same patient/family per VCEP guidance.

LYFE Sciences is an AI system, and it can make mistakes. Criteria may be applied incorrectly, sources may be misread, and a confident-looking classification can still be wrong. Double-check every criterion and its underlying evidence before relying on any call.

01The reasoning

How it was classified

The recorded framework is ClinGen MLH1 VCEP v2.0. The criteria and strengths shown below come from the saved assessment; this page does not recalculate the classification.

Recorded applied criteria

The applied criteria and their strengths are shown as assessed under the stated framework.

Pathogenic evidence criteria: 3 Benign evidence criteria: 0

Pathogenic criteria applied · 3

PVS1Very strong

PVS1 · very strong

NM_000249.4:c.381-1G>A is a canonical splice acceptor variant at IVS4-1 (intron 4), disrupting the invariant AG dinucleotide. Under the InSiGHT/ClinGen MLH1 VCEP v2.0, IVS±1 variants where exon skipping disrupts the reading frame and is predicted to undergo NMD qualify for PVS1_Very_Strong. MLH1 loss of function is an established disease mechanism for Lynch syndrome. SpliceAI max delta score of 0.99 confirms a severe splicing impact. Not combined with PP3 per VCEP rules.

PM2Supporting

PM2 · supporting

NM_000249.4:c.381-1G>A is absent from gnomAD v2.1 (exomes), gnomAD v4.1 (exomes), and gnomAD-Canada v1.0 (genomes). Under the MLH1 VCEP, PM2_Supporting applies when the variant is absent or extremely rare with allele frequency <0.00002 (<1 in 50,000 alleles) in gnomAD v4.

PP4Moderate

PP4 · moderate

A patient with this MLH1 variant (reported as IVS4-1G>A) was described by Roth et al. (2016, PMID:27357288) with two synchronous primary colorectal cancers (cecum and rectosigmoid colon), both demonstrating loss of MLH1 and PMS2 protein expression by immunohistochemistry, consistent with the variant location. Under the MLH1 VCEP, 2 independent CRC tumors with loss of MMR protein expression consistent with the variant location qualifies for PP4_Moderate. Independent tumors can be from the same patient/family per VCEP guidance.

Benign criteria applied · 0

No benign criterion marked as applied in the recorded assessment.

Recorded classification: Pathogenic

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: matched Rule3 (1 Pathogenic.Very Strong + 1 Pathogenic.Moderate + 1 Pathogenic.Supporting) with applied criteria: PVS1 very strong, PM2 supporting, PP4 moderate; maps to Pathogenic.

02The sources

Evidence at a glance

Evidence summaries and captured source records, where available. Missing entries should not be interpreted as negative findings.

gnomAD v4.1

gnomAD v2.1

Population frequency

Not recorded

gnomAD v4.1 allele counts were not recorded for this variant.

gnomAD v2.1

Absent from gnomAD v2.1.

gnomAD-Canada

Absent from gnomAD-Canada v1.0.

Applied · PM2 →

gnomAD v4.1 ↗gnomAD v2.1 ↗gnomAD-Canada ↗

SpliceAI score

0.990 maximum delta

Recorded SpliceAI maximum delta score: 0.990. Refer to the recorded PP3/BP4 rationale for interpretation.

No listed criterion applied SpliceAI ↗

ClinVar submissions

5 reported laboratory submissions

4 Pathogenic · 1 Likely pathogenic. ClinVar API review status: criteria provided, multiple submitters, no conflicts.

Context only ClinVar ↗

No screenshot for this evidence

Functional evidence

Not recorded

No standalone functional evidence summary is recorded in evidence_summary.functional.

No listed criterion applied OncoKB ↗

Somatic evidence

1 recorded samples

This variant has previously been reported in somatic cancers (COSMIC; COSV108755692, n = 1 times).

Context only COSMIC ↗

For specialists

Technical detail

ACMG/AMP criteria under ClinGen MLH1 VCEP v2.0, with the data and sources behind each one.

27 criteria with recorded statuses: 3 met · 12 not met · 0 not assessed · 12 not applicable. Open any row for the full reasoning, the rule and the evidence behind it.

Pathogenic

PVS1 PS1 PS2 PS3 PS4 PM1 PM2 PM5 PM6 PP1 PP2 PP3 PP4 PP5 PS5

Benign

BA1 BS1 BS2 BS3 BS4 BP1 BP2 BP3 BP4 BP5 BP6 BP7

Met Not met Not assessed Not applicable

NM_000249.4:c.381-1G>A is a canonical splice acceptor variant (IVS4-1) disrupting the invariant AG dinucleotide of MLH1 intron 4, meeting PVS1_Very_Strong under the InSiGHT/ClinGen MLH1 VCEP v2.0.

The variant is absent from gnomAD v2.1 and v4.1, meeting PM2_Supporting under the VCEP threshold of allele frequency <0.00002.

A patient with this variant, reported by Roth et al. (2016), had two synchronous colorectal cancers showing loss of MLH1 and PMS2 protein expression by immunohistochemistry, meeting PP4_Moderate under the VCEP rule for 2 independent tumors with MMR protein loss consistent with the variant location.

This variant was identified as a recurrent mutation in 5 African American families with Lynch syndrome by Guindalini et al. (2015), supporting its enrichment in the Lynch syndrome patient population.

ClinVar classifies this variant as Pathogenic (4 clinical laboratories) and Likely pathogenic (1 laboratory); however, the VCEP framework does not permit use of PP5 or BP6, and ClinVar classification alone is not used as an independent criterion.

Combining PVS1_Very_Strong (1) + PP4_Moderate (1) under the InSiGHT/ClinGen MLH1 VCEP v2.0 combination rules (Rule 10) yields a classification of Likely Pathogenic.

Met · 3

PVS1 Very strongNM_000249.4:c.381-1G>A is a canonical splice acceptor variant at IVS4-1 (intron 4), disrupting the invariant AG dinucleotide. Under the InSiGHT/ClinGen MLH1 VCEP v2.0, IVS±1 variants where exon skipping disrupts the reading frame and is predicted to undergo NMD qualify for PVS1_Very_Strong. MLH1 loss of function is an established disease mechanism for Lynch syndrome. SpliceAI max delta score of 0.99 confirms a severe splicing impact. Not combined with PP3 per VCEP rules. Details ▾Hide ▴

Evidence considered

Canonical splice acceptor variant c.381-1G>A (IVS4-1G>A) in MLH1
SpliceAI max delta score 0.99
predicting severe splicing aberration
MLH1 LOF is established disease mechanism for Lynch syndrome per VCEP
VCEP PVS1 rule: IVS±1 variants disrupting reading frame with predicted NMD → PVS1_Very_Strong

Sources

cspec ↗spliceai ↗pvs1_variant_assessmentpvs1_gene_context

PM2 SupportingNM_000249.4:c.381-1G>A is absent from gnomAD v2.1 (exomes), gnomAD v4.1 (exomes), and gnomAD-Canada v1.0 (genomes). Under the MLH1 VCEP, PM2_Supporting applies when the variant is absent or extremely rare with allele frequency <0.00002 (<1 in 50,000 alleles) in gnomAD v4. Details ▾Hide ▴

Evidence considered

Absent from gnomAD v2.1
Absent from gnomAD v4.1
Absent from gnomAD-Canada v1.0
Allele frequency meets VCEP PM2_Supporting threshold (<0.00002)

Sources

gnomad_v2 ↗gnomad_v4 ↗gnomad_canada ↗

PP4 ModerateA patient with this MLH1 variant (reported as IVS4-1G>A) was described by Roth et al. (2016, PMID:27357288) with two synchronous primary colorectal cancers (cecum and rectosigmoid colon), both demonstrating loss of MLH1 and PMS2 protein expression by immunohistochemistry, consistent with the variant location. Under the MLH1 VCEP, 2 independent CRC tumors with loss of MMR protein expression consistent with the variant location qualifies for PP4_Moderate. Independent tumors can be from the same patient/family per VCEP guidance. Details ▾Hide ▴

Evidence considered

Patient with 2 synchronous colon cancers showing loss of MLH1/PMS2 by IHC (PMID:27357288
Patient 2)
IHC pattern (MLH1/PMS2 loss) consistent with MLH1 variant location

Sources

PMID:27357288 ↗

Not met · 12 · evaluated, threshold not reached

PS2 No de novo occurrence data identified for NM_000249.4:c.381-1G>A in the reviewed evidence. VCEP PS2 requires confirmation of both maternity and paternity in a proband with a Lynch syndrome-spectrum tumor; no such data are available in the case materials or literature. Details ▾Hide ▴

Still missing

No de novo reports identified in ClinVar submissions or literature

PS3 No variant-specific functional assay data or calibrated functional odds for NM_000249.4:c.381-1G>A were identified. The VCEP functional assay SVI documentation spreadsheet was searched and this variant was not listed. The variant is a canonical splice site variant; splicing impact is predicted by SpliceAI (delta 0.99) and captured under PVS1, not PS3. Details ▾Hide ▴

Evidence considered

VCEP functional assay spreadsheet searched
variant not listed in calibrated assays

Still missing

No functional assay data for this specific splice variant

Sources

vcep_functional_assay_svi_documentation_mmr

PP1 No co-segregation data with Bayes Likelihood Ratio calculations were identified in the reviewed evidence. The VCEP PP1 rules require combined Bayes Likelihood Ratio >2.08 for supporting strength; no pedigrees with segregation analysis were available. Details ▾Hide ▴

Still missing

No co-segregation pedigrees or Bayes Likelihood Ratio calculations available

PP3 The VCEP PVS1 rule for canonical (±1/±2) splice variants explicitly states: 'Not to be combined with PP3.' PP3 for splice prediction is reserved for non-canonical splice variants with SpliceAI delta >= 0.2. The splicing impact of this canonical acceptor variant is already fully captured by PVS1_Very_Strong; applying PP3 would constitute double-counting of the same evidence. For missense application, the variant is not a missense change and was not found in the MLH1 HCI prior lookup table. Details ▾Hide ▴

Evidence considered

SpliceAI delta 0.99 confirms splicing impact
PVS1 rule prohibits combining PP3 with PVS1 for canonical splice variants
Variant not found in HCI-PRIORS-MLH1 lookup

Sources

cspec ↗spliceai ↗

BA1 NM_000249.4:c.381-1G>A is absent from gnomAD v4.1. The VCEP BA1 threshold requires grpmax filtering allele frequency >= 0.001 (0.1%), which is not met. Details ▾Hide ▴

Evidence considered

Variant absent from gnomAD v4.1

Sources

gnomad_v4 ↗

BS1 Variant is absent from gnomAD v4.1. The VCEP BS1 threshold requires grpmax filtering allele frequency >= 0.0001 and < 0.001 (0.01-0.1%), which is not met. Details ▾Hide ▴

Evidence considered

Variant absent from gnomAD v4.1

Sources

gnomad_v4 ↗

BS2 No evidence of co-occurrence in trans with a known pathogenic MLH1 variant was identified in the case materials or reviewed literature. VCEP BS2 requires confirmed phase (parental testing) demonstrating co-occurrence in trans with a known pathogenic variant in a patient with CRC after age 45 and no CMMRD features. Details ▾Hide ▴

Still missing

No data on co-occurrence in trans with pathogenic MLH1 variant

BS3 No functional assay data demonstrating a benign effect for NM_000249.4:c.381-1G>A were identified. The VCEP functional assay SVI documentation spreadsheet was searched and this variant was not listed among calibrated assays. Details ▾Hide ▴

Evidence considered

VCEP functional assay spreadsheet searched
variant not listed

Still missing

No functional data demonstrating benign effect

Sources

vcep_functional_assay_svi_documentation_mmr

BS4 No lack-of-segregation data were identified for NM_000249.4:c.381-1G>A. VCEP BS4 requires pedigrees with combined Bayes Likelihood Ratio <0.05 (strong) or between 0.05 and 0.48 (supporting); no such analysis is available. Details ▾Hide ▴

Still missing

No co-segregation or lack-of-segregation data available

BP4 BP4_Supporting requires either a missense variant with HCI prior probability <0.11 or an intronic/synonymous variant with SpliceAI delta score <= 0.1. This variant is a canonical splice site variant with SpliceAI max delta score of 0.99, far exceeding the 0.1 threshold. It was not found in the HCI-PRIORS-MLH1 lookup table. Details ▾Hide ▴

Evidence considered

SpliceAI delta = 0.99
far above BP4 threshold of <= 0.1
Variant not in HCI prior lookup table for missense HCI probability assessment

Sources

spliceai ↗vcep_hci_priors_mlh1

BP5 No evidence was identified of tumors with MSS and/or intact MMR protein expression, or BRAF V600E/MLH1 methylation with MSI-H/MLH1 loss, in patients carrying this variant. VCEP BP5 requires tumors showing evidence inconsistent with a pathogenic MMR defect. Details ▾Hide ▴

Still missing

No tumor data showing evidence inconsistent with pathogenic role

BP7 BP7 applies to synonymous or intronic variants at or beyond -21/+7 (i.e., deep intronic or distal exonic positions). NM_000249.4:c.381-1G>A is a canonical splice acceptor variant at position -1, within the splice consensus region, not at or beyond -21/+7. Additionally, the variant is not synonymous/silent. Details ▾Hide ▴

Evidence considered

Variant at position -1 of intron 4
within splice consensus region (-21 to +7)

Not applicable · 12· recorded as not applicable Show reasons ▾Hide reasons ▴

PS1 N/APS1 applies to missense variants encoding the same amino acid change as a known pathogenic variant, or to non-canonical splice variants affecting the same nucleotide as a known pathogenic splice variant. c.381-1G>A is a canonical (±1/±2) splice site variant, not a missense substitution, and the VCEP PS1 rule for splice applies only to non-canonical positions. Details ▾Hide ▴