KRAS c.34G>T, p.Gly12Cys

Reasoning

The KRAS VCEP applies PS1 at Strong strength when the variant has the same amino acid change as a previously established pathogenic variant, regardless of nucleotide change, for analogous positions in HRAS, KRAS, MRAS, NRAS, RIT1, and RRAS2. This case is KRAS NM_004985.5:c.34G>T, p.Gly12Cys (p.G12C), and ClinVar variation 12578 records the exact p.G12C variant as Pathogenic with criteria provided, multiple submitters, and no conflicts, including three clinical laboratories. The exact same amino acid change is therefore previously established as pathogenic; the KRAS VCEP rule is satisfied.

VCEP rule

Same amino acid change as a previously established pathogenic variant regardless of nucleotide change. Applicable for observed analogous residue positions in _HRAS, KRAS, MRAS, NRAS, RIT1,_ and _RRAS2._

Evidence considered

• ClinGen RASopathy Expert Panel KRAS Version 2.3 PS1 rule: same amino acid change as a previously established pathogenic variant regardless of nucleotide change, applicable to analogous positions in KRAS and the named RASopathy genes.
• ClinVar variation 12578 for NM_004985.5:c.34G>T, p.Gly12Cys/p.G12C: Pathogenic; criteria provided, multiple submitters, no conflicts; three clinical laboratories reported Pathogenic.
• PMID:11745231, Table 2; Results, K-ras Mutation Analysis: exact codon 12 cysteine substitution reported as "GGT to TGT at codon 12, cysteine".

Sources

Reasoning

The variant-specific literature includes a KRAS RAS-activation functional study using RAF1-RBD pulldown in isogenic human cells. KRAS-G12C increased GTP-bound KRAS by up to approximately 2-fold after EGF stimulation relative to control cells, providing abnormal pathway-function evidence. The VCEP functional-study table identifies RAS Activation Assay as an approved assay class, but its KRAS-specific listed publications do not include PMID:25705018 and the VCEP file contains no pre-assigned c.34G>T, p.Gly12Cys, or p.G12C entry. Under the VCEP table, an assay not listed for the gene may support PS3 only at Supporting strength; therefore PS3 is met at Supporting, not Moderate.

VCEP rule

One approved assay. ... Assays not listed here are presumed to lack sufficient historical evidence to evaluate the functional impact of a given variant and may only be sufficient for the application of PS3_Supporting or BS3_Supporting.

Evidence considered

• SVI-RASopathy-VCEP-V2-Approved-Functional-Studies.xlsx, sheet 'RAS Activation Assay': the KRAS assay is defined as measuring bound RAS protein immunoprecipitated with RAF1 or RBD; the assay is marked approved, with proposed strength PS3_Supporting and BS3_NA for listed instances.
• SVI-RASopathy-VCEP-V2-Approved-Functional-Studies.xlsx, sheet 'READ ME': assays not listed for a gene may only be sufficient for PS3_Supporting or BS3_Supporting, and two or more unique assay types are required to upgrade PS3 to Moderate.
• PMID:25705018, Results and Discussion: in isogenic MCF10A cells, KRAS-G12C increased GTP-bound KRAS up to approximately 2-fold after EGF stimulation; the assay used RAF1-RBD pulldown and near-physiological variant expression.
• The VCEP functional-study file was searched for c.34G>T, p.Gly12Cys, and p.G12C; no variant-specific row or pre-assigned code was found.

Still missing

• No c.34G>T, p.Gly12Cys, or p.G12C entry was found in SVI-RASopathy-VCEP-V2-Approved-Functional-Studies.xlsx.txt after searching all three notations.
• The study supporting PS3 is not one of the KRAS-specific PMID sets listed in the VCEP functional-study table, so the evidence cannot be upgraded to Moderate under the VCEP rules.
• The variant-specific studies in PMID:24256730 and PMID:26841430 were reviewed but do not establish an additional VCEP-listed KRAS assay instance for strength upgrading.

Sources

Reasoning

The KRAS VCEP limits PM1 to critical and well-established functional domains in its supplementary table: P-loop amino acids 10-17, SW1 amino acids 25-40, SW2 amino acids 57-64, and SAK amino acids 145-156. The variant changes residue 12, placing p.Gly12Cys within the authoritative P-loop range 10-17. The governing PM1 domain-alignment file was searched for c.34G>T, p.Gly12Cys, and p.G12C, with no entry found; this is a file-search gap, not evidence that the approved P-loop domain is absent. The VCEP rule supports PM1 at Moderate strength.

VCEP rule

Applicable only to critical and well-established functional domains available in the supplementary table (P-loop [AA 10-17], SW1 [AA 25-40], SW2 [AA 57-64], SAK [AA 145-156]). Not applicable to specific amino acid residues (see PM5).

Evidence considered

• ClinGen RASopathy Expert Panel KRAS Version 2.3 PM1 rule: approved domains are P-loop AA 10-17, SW1 AA 25-40, SW2 AA 57-64, and SAK AA 145-156; p.Gly12Cys is at residue 12 and is within P-loop AA 10-17.
• Alignment-with-PM1-domains.pptx.pdf.txt was searched for c.34G>T, p.Gly12Cys, and p.G12C; no variant-specific entry was found.
• PMID:25705018, Results and Discussion: KRAS-G12C at codon 12 increased GTP-bound KRAS up to 2-fold; this hotspot observation was reviewed but was not used as an independent PM1 route because the KRAS VCEP limits PM1 to the listed approved domains.

Still missing

• No c.34G>T, p.Gly12Cys, or p.G12C entry was found in Alignment-with-PM1-domains.pptx.pdf.txt after searching all three notations; PM1 is determined from the VCEP's explicit P-loop range in cspec.

Sources

Reasoning

The governing KRAS VCEP permits PM2 at Supporting strength when the variant is absent from controls in gnomAD. No alternate allele was observed in gnomAD v2.1 exomes (0/249,272; AF 0%; 0 homozygotes), and the variant was absent from gnomAD v4.1. No VCEP rule requires a non-cancer subset or a separate ancestry threshold, so the available all-comers gnomAD results are used.

VCEP rule

The variant must be absent from controls (gnomAD).

Evidence considered

• ClinGen RASopathy Expert Panel KRAS specification v2.3, PM2 criterion: The variant must be absent from controls (gnomAD); Supporting is the applicable PM2 strength.
• gnomAD v2.1 exome result for NM_004985.5:c.34G>T / p.(Gly12Cys): alternate allele count 0, allele number 249,272, AF 0.0%, homozygotes 0; all reported subpopulation counts are also 0.
• gnomAD v4.1 result: variant absent.

Sources

Reasoning

The VCEP applies PP3 to missense variants using REVEL ≥ 0.7, with a separate splicing-impact path requiring the predicted outcome to match the disease mechanism. This variant is a missense change, NM_004985.5:c.34G>T (p.Gly12Cys), and the governing missense predictor is available with REVEL 0.853, which meets the VCEP cutoff. The splice prediction is not used to create a second PP3 assignment.

VCEP rule

For missense variants: REVEL ≥ 0.7. For splicing impact, predicted outcome must match disease mechanism.

Evidence considered

• VCEP CSPEC PP3 rule: For missense variants, REVEL ≥ 0.7; the case variant is NM_004985.5:c.34G>T (p.Gly12Cys), classified as missense.
• Local REVEL predictor lookup: REVEL score 0.853 for NM_004985.5:c.34G>T, exceeding the VCEP cutoff of 0.7.

Sources

Reasoning

The governing KRAS VCEP requires a gnomAD filtering allele frequency of at least 0.05% for BA1. The variant has 0 alternate alleles among 249,272 gnomAD v2.1 exome alleles (AF 0%; 0 homozygotes) and is absent from gnomAD v4.1, so the threshold is not met.

VCEP rule

GnomAD filtering allele frequency ≥0.05%.

Evidence considered

• ClinGen RASopathy Expert Panel KRAS specification v2.3, BA1 criterion: GnomAD filtering allele frequency ≥0.05%.
• gnomAD v2.1 exome result for NM_004985.5:c.34G>T / p.(Gly12Cys): alternate allele count 0, allele number 249,272, AF 0.0%, homozygotes 0; highest observed subpopulation AF also 0.0% (African/African American, 0/16,042 alleles).
• gnomAD v4.1 result: variant absent.

Sources

Reasoning

The governing KRAS VCEP requires a gnomAD filtering allele frequency of at least 0.025% for BS1. The variant has 0 alternate alleles among 249,272 gnomAD v2.1 exome alleles (AF 0%; 0 homozygotes) and is absent from gnomAD v4.1, so the threshold is not met.

VCEP rule

GnomAD filtering allele frequency ≥0.025%.

Evidence considered

• ClinGen RASopathy Expert Panel KRAS specification v2.3, BS1 criterion: GnomAD filtering allele frequency ≥0.025%.
• gnomAD v2.1 exome result for NM_004985.5:c.34G>T / p.(Gly12Cys): alternate allele count 0, allele number 249,272, AF 0.0%, homozygotes 0; highest observed subpopulation AF also 0.0% (African/African American, 0/16,042 alleles).
• gnomAD v4.1 result: variant absent.

Sources

Reasoning

The VCEP applies BP4 to missense variants using REVEL ≤ 0.3. This variant is a missense change, NM_004985.5:c.34G>T (p.Gly12Cys), so BP4 is within scope. The governing predictor value is REVEL 0.853, which is above the VCEP benign cutoff and therefore does not satisfy BP4. The available splice prediction is not substituted for the VCEP's named missense predictor and is not counted as separate computational evidence.

VCEP rule

For missense variants: REVEL ≤0.3.

Evidence considered

• VCEP CSPEC BP4 rule: For missense variants, REVEL ≤ 0.3; the case variant is NM_004985.5:c.34G>T (p.Gly12Cys), classified as missense.
• Local REVEL predictor lookup: REVEL score 0.853 for NM_004985.5:c.34G>T, above the VCEP cutoff of 0.3.

Sources

Reasoning

The VCEP assigns PS2 by de novo evidence points: 4 points for Very Strong, 2 points for Strong, and 1 point for Moderate. The available case evidence does not document a proband with confirmed de novo NM_004985.5:c.34G>T (p.Gly12Cys), confirmed parental relationships, parental testing results, or phenotype consistency sufficient to assign any PS2 points.

VCEP rule

4 Points; 2 Points; 1 Point.

Evidence considered

• CSPEC criteria entry for PS2: Very Strong = 4 points; Strong = 2 points; Moderate = 1 point.
• No proband-level de novo, parental-testing, parentage, or qualifying phenotype evidence was identified in the available case materials.

Still missing

• Documented proband phenotype consistent with the KRAS-associated disorder
• Confirmed parental relationships and parental genotypes for NM_004985.5:c.34G>T
• Evidence that the variant is absent in both tested parents

Sources

Reasoning

The KRAS VCEP makes PS4 applicable through point-based scoring for autosomal-dominant RASopathy cases, with full points for RASopathy phenotypes and reduced points for minimal clinical information. Its governing rule states: "The prevalence of the variant in affected individuals is significantly increased compared to the prevalence in controls. Note 1: Relative risk (RR) or odds ratio (OR), as obtained from case-control studies, is >5.0 and the confidence interval around the estimate of RR or OR does not include 1.0. See manuscript for detailed guidance. Note 2: In instances of very rare variants where case-control studies may not reach statistical significance, the prior observation of the variant in multiple unrelated patients with the same phenotype, and its absence in controls, may be used as moderate level of evidence." The checked PMID:11745231 paper reports KRAS G12C in 25 of 106 primary lung adenocarcinoma tumors and reports KRAS mutations overall in smokers versus nonsmokers (43% versus 0%), but it is a somatic lung-cancer cohort rather than a RASopathy case-control cohort and provides no unaffected-control count for KRAS G12C. Therefore an OR and 95% CI cannot be computed for the relevant RASopathy comparison, and no VCEP PS4 point score is available. The literature_pass packet's ai_classified_papers contains no paper already read as PS4.

VCEP rule

Strength adjustment using point-based scoring for autosomal dominant cases with phenotypic specifications: full points awarded with RASopathy phenotypes, reduced points applicable to cases with minimal phenotypic information (i.e. prenatal cases, cases with a clinical order of a RASopathy panel without clinical information, and cases with limited clinical information in other global tests (such as WES)). ... The prevalence of the variant in affected individuals is significantly increased compared to the prevalence in controls. Note 1: Relative risk (RR) or odds ratio (OR), as obtained from case-control studies, is >5.0 and the confidence interval around the estimate of RR or OR does not include 1.0. See manuscript for detailed guidance. Note 2: In instances of very rare variants where case-control studies may not reach statistical significance, the prior observation of the variant in multiple unrelated patients with the same phenotype, and its absence in controls, may be used as moderate level of evidence. ... ≥5 points. ... ≥3 points. ... ≥1 points.

Evidence considered

• ClinGen KRAS VCEP v2.3 PS4 entry: PS4 is adjusted using point-based scoring for autosomal-dominant cases with phenotypic specifications; full points are for RASopathy phenotypes and reduced points are for minimal clinical information.
• PMID:11745231, Table 2 and Results, K-ras Mutation Analysis: KRAS codon 12 cysteine (GGT to TGT at codon 12) was found in 25 of 106 primary lung adenocarcinoma tumors; the cohort had 40 KRAS-mutant tumors, and KRAS mutations overall occurred in 43% of smokers versus 0% of nonsmokers.
• No unaffected or population-control count for the exact KRAS G12C variant was reported in the verified paper extraction, so no case-control OR or 95% CI was computed.
• The literature_pass ai_classified_papers entries were checked; none was classified as PS4.

Still missing

• A RASopathy-specific case-control comparison or multiple unrelated RASopathy patients with the same phenotype and appropriate control data is needed.
• The VCEP phenotypic point assignment for this case is not available.

Sources

Reasoning

The KRAS VCEP PM5 rule requires at least one [likely] pathogenic residue change at the same codon for Moderate strength, or at least two different [likely] pathogenic residue changes at the same codon observed in at least five probands for Strong strength. The available comparator collection reports zero same-residue candidates, and its harvesting step ended with an HTTP 429 error. The reviewed paper extracts describe other KRAS codon 12 substitutions and functional observations but do not establish a qualifying [likely] pathogenic comparator under this VCEP rule. PM5 is therefore not assessed rather than inferred.

VCEP rule

1 \[likely\] pathogenic residue change at the same codon.

Evidence considered

• ClinGen RASopathy Expert Panel KRAS Version 2.3 PM5 rule: one [likely] pathogenic residue change at the same codon supports Moderate; at least two different [likely] pathogenic residue changes at the same codon observed in at least five probands support Strong.
• PM5 candidate collection for NM_004985.5:c.34G>T: eligible classic same-residue missense search, zero same-residue candidates, zero expert-panel or non-expert-majority pathogenic/likely pathogenic candidates, and HTTP Error 429 during harvesting.
• PMID:25705018 was reviewed for codon 12 substitutions and functional data, but it does not by itself establish a qualifying [likely] pathogenic comparator for PM5.

Still missing

• A validated ClinVar or VCEP-supported comparator search for KRAS codon 12 alternate missense changes and their pathogenicity evidence is required; the available PM5 candidate collection found zero same-residue candidates after an HTTP 429 error.

Sources

Reasoning

The VCEP assigns PM6 according to de novo evidence points: 2 points for Strong, 1 point for Moderate, and 0.5 points for Supporting. The available case evidence does not document a presumed de novo occurrence, parental testing, confirmed parentage, or other information that would support assigning PM6 points.

VCEP rule

2 Points; 1 Point; 0.5 Points.

Evidence considered

• CSPEC criteria entry for PM6: Strong = 2 points; Moderate = 1 point; Supporting = 0.5 points.
• No presumed de novo, parental-testing, parentage, or qualifying family evidence was identified in the available case materials.

Still missing

• Documented presumed de novo proband observation
• Confirmed parental relationships and parental genotypes
• Evidence supporting the applicable PM6 point level

Sources

Reasoning

The VCEP assigns PP1 from informative meioses: at least 7 for Strong, at least 5 for Moderate, and at least 3 for Supporting. The available case evidence provides no documented informative meioses or affected-relative segregation data, so no PP1 strength can be assigned.

VCEP rule

≥7 informative meioses; ≥5 informative meioses; ≥3 informative meioses.

Evidence considered

• CSPEC criteria entry for PP1: Strong requires ≥7 informative meioses; Moderate requires ≥5; Supporting requires ≥3.
• No informative meioses or affected-relative segregation data were identified in the available case materials.

Still missing

• Documented affected relatives carrying the variant
• Documented informative meioses
• Phenotype and genotype data for each segregating relative

Sources

Reasoning

The governing KRAS VCEP makes BS2 applicable through a phenotypic point system based on healthy homozygous or heterozygous individuals, with applicable strengths of -4 points or -1 point. Population data show 0 homozygotes, but do not document a healthy adult carrier or provide the required phenotype-based point assignment; therefore BS2 cannot be evaluated.

VCEP rule

Strength adjustment using point-based scoring based on phenotypic specifications. Phenotypic specifications: based on healthy homozygote or heterozygote individuals, reduced points for apparently unaffected heterozygous individuals, applicable to parent or sibling samples during clinical family evaluations. ... \-4 Points. ... \-1 Point.

Evidence considered

• ClinGen RASopathy Expert Panel KRAS specification v2.3, BS2 instructions: strength adjustment uses point-based scoring based on phenotypic specifications, including healthy homozygote or heterozygote individuals; applicable strengths are -4 points and -1 point.
• gnomAD v2.1 reports 0 homozygotes among 249,272 exome alleles; this is absence of an observed homozygote, not documentation that an individual carrying the variant was a healthy adult.
• gnomAD v4.1 reports the variant absent and provides no healthy-carrier phenotype observation.

Still missing

• A documented healthy adult homozygote or heterozygote and the VCEP phenotypic point assignment are required to assess BS2.

Sources

Reasoning

The VCEP requires only one informative meiosis for BS4. The available case evidence does not document an unaffected relative who was tested and did not carry the variant, or any other informative non-segregation meiosis; therefore BS4 cannot be assessed.

VCEP rule

Requires only one informative meiosis.

Evidence considered

• CSPEC criteria entry for BS4: Strong requires only one informative meiosis.
• No informative non-segregation meiosis or tested unaffected-relative genotype was identified in the available case materials.

Still missing

• Documented unaffected relative with relevant phenotype information
• Genotype result for the unaffected relative
• One confirmed informative non-segregation meiosis

Sources

Reasoning

The KRAS VCEP Version 2.3 criteria entry provides BP2 point thresholds (supporting at ≥ (-1) point, moderate at ≥ (-2) points, and strong at ≥ (-4) points), but the available case evidence contains no documented phase or allelic observation establishing a BP2 point value. No cis/trans relationship with a pathogenic or benign variant, affected-proband observation, or inheritance result is available to evaluate the VCEP's point requirement. BP2 is therefore not assessed rather than inferred from the variant's ClinVar classification or from generic ACMG/AMP text.

VCEP rule

Supporting: ≥ (-1) Point; Moderate: ≥ (-2) Points; Strong: ≥ (-4) Points.

Evidence considered

• ClinGen RASopathy Expert Panel KRAS Version 2.3 CSPEC BP2 entry: Supporting ≥ (-1) Point, Moderate ≥ (-2) Points, Strong ≥ (-4) Points; no case-level phase, cis/trans, or allelic point assignment is documented.

Still missing

• Documented phase and allelic context with a pathogenic or benign variant sufficient to assign the VCEP BP2 point value are unavailable.

Sources

Reasoning

The KRAS VCEP defines BP5 as point-based evidence for an alternative molecular cause of a RASopathy in a different gene, or a phenotype inconsistent with RASopathy fully explained by another causative variant. The governing strength thresholds are Strong: "≥ (-4) Points."; Moderate: "≥ (-2) Points."; Supporting: "≥ (-1) Point." No alternative causative variant, phenotype-specific point assignment, or qualifying WES result is present in the available evidence. Consequently, the BP5 point total cannot be determined and no threshold comparison is performed.

VCEP rule

Points are awarded for an alternative molecular cause of a RASopathy in a different gene (and/or in conjunction with BP2) and the phenotype is consistent with expected severity of the RASopathy. Points are also awarded for phenotypes inconsistent with a RASopathy and fully explained by a different causative variant (e.g. WES testing). ... ≥ (-4) Points. ... ≥ (-2) Points. ... ≥ (-1) Point.

Evidence considered

• ClinGen KRAS VCEP v2.3 BP5 instructions: points are awarded for an alternative molecular cause of a RASopathy in a different gene, or for phenotypes inconsistent with RASopathy fully explained by a different causative variant.
• No qualifying alternative molecular cause or VCEP-assigned BP5 points were identified in the case evidence.

Still missing

• Evidence of an alternative molecular cause in a different gene or a fully explanatory non-RASopathy phenotype is needed.
• The VCEP BP5 phenotypic point assignment is not available.

Sources