Variant classification summary

NM_002524.5:c.35G>A

NRAS · NP_002515.1:p.(Gly12Asp) · NM_002524.5

GRCh37: chr1:115258747 C>T · GRCh38: chr1:114716126 C>T

Gene: NRAS Transcript: NM_002524.5

Final call

VUS

PS1 strong PS3 moderate PM1 moderate PM5 moderate PP3 supporting

Variant details

Gene

NRAS

Transcript

NM_002524.5

Protein

NP_002515.1:p.(Gly12Asp)

gnomAD AF

1.0533124818768293e-05 (v4.1)

ClinVar

Pathogenic

OncoKB

Oncogenic

Classification rationale

Interpretation summary

Generated evidence synthesis

1

NM_002524.5:c.35G>A (p.Gly12Asp) is located in the P-loop domain (AA 10-17), a critical GTP-binding domain defined by the ClinGen RASopathy VCEP as a well-established functional domain without benign variation.

2

p.Gly12Asp is the same amino acid change as well-established pathogenic variants across NRAS and other RAS paralogs. G12D is a canonical oncogenic substitution reported as Pathogenic in ClinVar by 11 clinical laboratories and observed in 1,303 somatic cancer samples in COSMIC.

3

Multiple VCEP-approved functional assays demonstrate that NRAS G12D is a gain-of-function variant. N-RasG12D exhibits increased GTP-bound Ras (RAS Activation Assay), constitutive MEK and ERK phosphorylation (MEK/ERK Activation Assays), and transforming activity in focus formation assays.

4

At codon 12, multiple different missense changes (G12V, G12C, G12S, G12A) have been established as pathogenic, satisfying PM5 at Moderate strength per the VCEP rule requiring at least one [likely] pathogenic residue change at the same codon.

5

REVEL score of 0.783 meets the VCEP PP3 threshold (≥0.7), indicating multiple lines of computational evidence support a deleterious effect. SpliceAI predicts no splice impact (max delta = 0.00).

6

This variant is present in gnomAD at extremely low frequency (v2.1: 0.0008%, 2/251,484 alleles; v4.1: 0.00105%, 17/1,613,956 alleles), with no homozygotes observed. The variant is absent from gnomAD-Canada. These frequencies are well below the VCEP BS1 (≥0.025%) and BA1 (≥0.05%) thresholds.

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 NRAS 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	NM_002524.5:c.35G>A is a missense variant (p.Gly12Asp). The ClinGen RASopathy VCEP v2.3.0 specifies PVS1 as not applicable; this variant does not fall into null-variant buckets (nonsense, frameshift, or canonical splice site).	cspec
PS1	Met	p.Gly12Asp (G12D) is the same amino acid change as a previously established pathogenic variant regardless of nucleotide change. G12D is a well-characterized oncogenic substitution in NRAS, KRAS, and HRAS, observed in thousands of somatic cancers (COSMIC n=1303) and reported as Pathogenic in ClinVar by 11 clinical laboratories. The VCEP explicitly authorizes PS1 at Strong for analogous pathogenic residue positions across HRAS, KRAS, MRAS, NRAS, RIT1, and RRAS2.	cspec clinvar oncokb
PS2	Not assessed	No de novo occurrence data with confirmed maternity and paternity is available in the evidence packet for this specific variant. The VCEP PS2 point-based scoring requires confirmed de novo status with RASopathy phenotypes.
PS3	Met	Well-established in vitro and in vivo functional studies demonstrate a gain-of-function effect for NRAS G12D. The VCEP-approved RAS Activation Assay, MEK Activation Assay, and ERK Activation Assay are all applicable to NRAS. N-RasG12D shows increased GTP-bound Ras (RAF-RBD pulldown), constitutive MEK and ERK phosphorylation in multiple cell types, and transforming activity in focus formation assays. Mouse models with endogenous NrasG12D expression develop myeloproliferative neoplasms. G12D is validated as a pathogenic/likely pathogenic control in VCEP-approved ERK Activation Assay for HRAS, supporting functional analogy across RAS paralogs. At least two different approved assay types (RAS Activation and MEK/ERK Activation) demonstrate damaging effect.	cspec PMID:19075190 PMID:18372904 PMID:23687087 oncokb
PS4	Not assessed	The VCEP PS4 requires point-based scoring using probands with RASopathy phenotypes. While this variant is observed in numerous somatic cancer cases and ClinVar submissions, the evidence packet does not contain specific germline RASopathy proband counts meeting VCEP phenotypic specifications for PS4 scoring.
PS5	N/A	The ClinGen RASopathy VCEP explicitly disables PP5 as not for use, and PS5 is not defined in the VCEP criteria. Applying the VCEP's intent consistently, PS5 is not applicable under this framework.	cspec
PM1	Met	Gly12 is located within the P-loop domain (amino acids 10-17), a critical and well-established functional domain defined in the VCEP supplementary table. The P-loop is essential for GTP binding and RAS protein function. This domain has no benign variation and is a known mutational hotspot.	cspec oncokb
PM2	Not met	The VCEP PM2 rule states the variant must be absent from controls in gnomAD. This variant is present in gnomAD v2.1 at AF=7.95e-06 (2/251,484 alleles) and in gnomAD v4.1 at AF=1.05e-05 (17/1,613,956 alleles). Although at extremely low frequency, the variant is not absent, so PM2_Supporting is not met under the strict VCEP definition.	cspec gnomad_v2 gnomad_v4
PM5	Met	At codon 12 of NRAS, multiple different missense changes have been established as pathogenic: G12V, G12C, G12S, and G12A are all recognized oncogenic substitutions. The VCEP PM5 Moderate rule is satisfied when ≥1 [likely] pathogenic residue change exists at the same codon. G12D represents a novel residue change at a codon with established pathogenic variants.	cspec clinvar
PM6	Not assessed	No assumed de novo events are documented in the evidence packet for this variant. The VCEP PM6 requires point-based scoring with phenotypic specifications, and no data meeting these criteria were identified.
PP1	Not assessed	No co-segregation data in affected family members is available. The VCEP PP1 requires ≥3 informative meioses for Supporting, ≥5 for Moderate, and ≥7 for Strong strength.
PP2	N/A	The VCEP declares PP2 not applicable because the NRAS missense z-score in gnomAD is less than 3.09, indicating the gene does not have a low rate of benign missense variation.	cspec
PP3	Met	REVEL score is 0.783, which meets the VCEP threshold of ≥0.7 for PP3_Supporting for missense variants. SpliceAI predicts no splice impact (max delta = 0.00), consistent with the variant's primary effect being at the protein level. The REVEL score indicates multiple lines of computational evidence support a deleterious effect.	cspec revel spliceai
PP4	N/A	The VCEP declares PP4 not applicable; see PS4 for phenotype-based evidence.	cspec
PP5	N/A	The VCEP explicitly declares PP5 not for use as recommended by the ClinGen Sequence Variant Interpretation VCEP Review Committee.	cspec
BA1	Not met	The VCEP BA1 threshold is gnomAD filtering allele frequency ≥0.05%. This variant has AF=0.0008% in gnomAD v2.1 and AF=0.00105% in v4.1, both well below the BA1 threshold.	cspec gnomad_v2 gnomad_v4
BS1	Not met	The VCEP BS1 threshold is gnomAD filtering allele frequency ≥0.025%. This variant has AF=0.0008% in gnomAD v2.1 and AF=0.00105% in v4.1, both well below the BS1 threshold.	cspec gnomad_v2 gnomad_v4
BS2	Not assessed	No data on healthy adult individuals carrying this variant with full penetrance expected at an early age are available. The VCEP BS2 uses point-based scoring based on phenotypic specifications.
BS3	N/A	The VCEP declares BS3 not applicable for NRAS. Well-established functional studies for NRAS G12D uniformly show a damaging/gain-of-function effect.	cspec
BS4	Not assessed	No lack of segregation data in affected family members is available. The VCEP BS4 requires only one informative meiosis but no such data are present.
BP1	Not met	The VCEP BP1 rule applies only to truncating variants (nonsense, frameshift, canonical splice site, initiation codon, entire gene or multi-exon deletion) in genes without established LOF correlation to disease. NM_002524.5:c.35G>A is a missense variant and does not qualify.	cspec
BP2	Not assessed	No data on an alternative molecular cause of a RASopathy in the same gene or on co-occurrence with a different causative variant are available. The VCEP BP2 uses point-based scoring for alternative molecular causes.
BP4	Not met	The VCEP BP4 threshold for missense variants is REVEL ≤0.3. The REVEL score for this variant is 0.783, which exceeds the BP4 threshold and instead meets PP3.	cspec revel
BP5	Not assessed	No data on an alternative molecular cause of disease in a different gene are available. The VCEP BP5 uses point-based scoring for phenotypes inconsistent with RASopathy and fully explained by a different causative variant.
BP6	N/A	The VCEP explicitly declares BP6 not for use as recommended by the ClinGen Sequence Variant Interpretation VCEP Review Committee.	cspec
BP7	Not met	BP7 applies only to synonymous (silent) variants. NM_002524.5:c.35G>A is a missense variant (p.Gly12Asp), not a synonymous change.	cspec

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