Variant classification summary

NM_033360.3:c.34_35delinsTGTGC

KRAS · NP_203524.1:p.(Gly12delinsCysAla) · NM_033360.3

GRCh37: chr12:25398284 CC>GCACA · GRCh38: chr12:25245350 CC>GCACA

Gene: KRAS Transcript: NM_033360.3

Final call

VUS

PM1 moderate PM2 supporting PM4 moderate

Variant details

Gene

KRAS

Transcript

NM_033360.3

Protein

NP_203524.1:p.(Gly12delinsCysAla)

gnomAD AF

ClinVar

OncoKB

Oncogenic

Classification rationale

Interpretation summary

Generated evidence synthesis

1

This variant resides at codon 12 within the P-loop domain (AA 10-17), a critical functional domain per RASopathy VCEP specifications. Codon 12 is a well-established mutational hotspot with no benign variation. The variant lies in a statistically significant hotspot.

2

The variant is absent from gnomAD v2.1, v4.1, and gnomAD-Canada, supporting rarity in the general population.

3

The variant is an in-frame deletion-insertion that alters protein length (p.Gly12delinsCysAla) in a non-repetitive region, consistent with a potential gain-of-function effect in the RAS/MAPK pathway.

4

No variant-specific functional studies, de novo observations, case reports, or segregation data were identified in the available literature or databases. OncoKB classifies this variant as Oncogenic with gain-of-function effect.

5

Insufficient evidence exists to reach a definitive classification under the RASopathy VCEP v2.3.0 framework. Met criteria: PM1 (Moderate), PM2 (Supporting), PM4 (Moderate). Key data gaps include absence of functional studies, patient phenotype data, and segregation analysis.

Final determination: No criteria-combination rule matched the adjudicated criteria in the ClinGen RASopathy Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines for KRAS Version 2.3.0 v2.3.0 framework, so the variant remains a Variant of Uncertain Significance pending human review.

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	Not applicable per ClinGen RASopathy VCEP v2.3.0 specifications; variant is an in-frame deletion-insertion, not a null variant (nonsense, frameshift, or canonical splice site).	cspec
PS1	Not met	PS1 requires the same amino acid change as a previously established pathogenic variant. This variant produces p.Gly12delinsCysAla, an in-frame insertion at codon 12, which does not match any known pathogenic missense change at this codon. No prior pathogenic variant with this amino acid change has been identified.	cspec
PS2	Not assessed	No de novo observations for this specific variant were identified in the available literature or ClinVar submissions. PS2 requires confirmed de novo occurrence in a patient with RASopathy phenotype.
PS3	Not assessed	No variant-specific functional studies for NM_033360.3:c.34_35delinsTGTGC were identified. OncoKB classifies this variant as Oncogenic, but this is a curated knowledgebase label rather than direct functional assay evidence. The only available publication (PMID:24642870) characterized different KRAS insertion variants (c.30_31insGGA, c.33_34insGGAGCT), not this specific variant. VCEP-approved assays (RAS Activation, MEK Activation, ERK Activation) have not been applied to this variant.	oncokb
PS4	Not assessed	No patient case data or case-control studies involving this variant were identified. The variant is absent from ClinVar with no submissions from clinical laboratories. PS4 point-based scoring requires proband counts per VCEP specifications.	clinvar
PS5	N/A	Not applicable for this VCEP per ClinGen RASopathy VCEP v2.3.0; the VCEP Review Committee recommends against use of PP5.	cspec
PM1	Met	The variant affects codon 12 within the P-loop domain (amino acids 10-17), a critical and well-established functional domain per the RASopathy VCEP supplementary table. Codon 12 is a known mutational hotspot with no benign variation in this region. Hotspot analysis confirms this residue is statistically significant.	cspec vcep_alignment_with_pm1_domains_pptx
PM2	Met	This variant is absent from all population databases including gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada v1.0. Per RASopathy VCEP v2.3.0, PM2 is applied at Supporting strength when the variant is absent from gnomAD controls.	gnomad_v2 gnomad_v4 gnomad_canada cspec
PM4	Met	This is an in-frame deletion-insertion (c.34_35delinsTGTGC) resulting in a protein length change (p.Gly12delinsCysAla). KRAS has no known repetitive regions per the RASopathy VCEP. PM4 is applied at Moderate strength per VCEP rule: 'No known repetitive areas in gene. Use as described.'	cspec
PM5	N/A	VCEP PM5 rule applies to missense changes at the same codon where a different pathogenic missense change has been established. This variant is an in-frame deletion-insertion, not a missense change. Per PM5 candidate analysis, the variant class is not missense-like and is not eligible for classic same-residue PM5.	cspec pm5_candidates
PM6	Not assessed	No assumed de novo observations for this variant were identified. PM6 point-based scoring requires de novo proband data per the SVI recommendations cited by the RASopathy VCEP.
PP1	Not assessed	No co-segregation data for this variant was identified in the available literature. PP1 requires segregation analysis in affected family members; ≥3 informative meioses for Supporting, ≥5 for Moderate, ≥7 for Strong per VCEP.
PP2	N/A	Not applicable per RASopathy VCEP v2.3.0. The missense z-score for KRAS is <3.09 in gnomAD, so PP2 cannot be applied.	cspec
PP3	Not met	VCEP PP3 applies to missense variants with REVEL ≥0.7 or splicing impact matching disease mechanism. This is an in-frame indel, not a missense variant; REVEL and BayesDel scores are not available. SpliceAI predicts no significant splice impact (max delta score = 0.01). Neither computational criterion is satisfied.	spliceai cspec
PP4	N/A	Not applicable per RASopathy VCEP v2.3.0. PP4 is superseded by PS4 and is not separately evaluated in this framework.	cspec
PP5	N/A	Not applicable for this VCEP per ClinGen RASopathy VCEP v2.3.0; the VCEP Review Committee recommends against use of PP5.	cspec
BA1	Not met	RASopathy VCEP BA1 threshold is gnomAD filtering allele frequency ≥0.05%. This variant is absent from gnomAD v2.1, v4.1, and gnomAD-Canada (allele frequency = 0). BA1 is not met.	gnomad_v2 gnomad_v4 gnomad_canada cspec
BS1	Not met	RASopathy VCEP BS1 threshold is gnomAD filtering allele frequency ≥0.025%. This variant is absent from all gnomAD populations (allele frequency = 0). BS1 is not met.	gnomad_v2 gnomad_v4 gnomad_canada cspec
BS2	Not assessed	No data regarding observation of this variant in healthy adult individuals was available. BS2 point-based scoring requires phenotypic information about heterozygous or homozygous carriers.
BS3	N/A	Not applicable per RASopathy VCEP v2.3.0. BS3 (well-established functional studies showing no damaging effect) is marked as Not applicable by the VCEP.	cspec
BS4	Not assessed	No segregation data was available to assess lack of segregation in affected family members. BS4 requires only one informative meiosis showing lack of segregation per VCEP.
BP1	N/A	RASopathy VCEP BP1 applies to truncating variants (nonsense, frameshift, canonical splice sites, initiation codon, exon/gene deletion) in genes where RASopathies have a gain-of-function mechanism. This variant is an in-frame deletion-insertion, not a truncating variant.	cspec
BP2	Not assessed	No data regarding observation of this variant in trans with a pathogenic variant or cis with a pathogenic variant was available. BP2 point-based scoring requires evidence of an alternative molecular cause in the same gene.
BP3	N/A	Not applicable per RASopathy VCEP v2.3.0. No known benign repetitive areas exist in RASopathy genes, and BP3 is specific to in-frame deletions/insertions in repetitive regions.	cspec
BP4	N/A	RASopathy VCEP BP4 applies to missense variants with REVEL ≤0.3 or splicing variants with no predicted impact matching disease mechanism. This is an in-frame indel; REVEL is not available and BP4 is not applicable to non-missense, non-splice variants.	cspec
BP5	Not assessed	No data regarding an alternative molecular cause of RASopathy was available. BP5 point-based scoring requires evidence of an alternative molecular cause in a different gene with phenotype consistent with RASopathy severity.
BP6	N/A	Not applicable for this VCEP per ClinGen RASopathy VCEP v2.3.0; the VCEP Review Committee recommends against use of BP6.	cspec
BP7	N/A	BP7 applies to synonymous (silent) variants with no predicted splice impact. This variant is an in-frame deletion-insertion at codon 12, not a synonymous variant.	cspec

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