KRAS c.57G>T, p.(Leu19Phe)

Reasoning

PM2 is met at Supporting strength under the KRAS VCEP because NM_033360.2:c.57G>T (KRAS p.Leu19Phe) is absent from gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada v1.0. The VCEP specifically requires absence from controls in gnomAD for PM2 Supporting.

Rule

The variant must be absent from controls (gnomAD).

Evidence considered

• The KRAS VCEP PM2 rule is Supporting when the variant is absent from controls in gnomAD.
• The variant is explicitly reported absent from gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada v1.0.

Still missing

• The case bundle does not provide numeric gnomAD allele counts, sample sizes, ancestry-specific counts, or confidence intervals; however, each queried population resource returned an explicit absent result.

Sources

Reasoning

NM_033360.2:c.57G>T (NP_203524.1:p.(Leu19Phe)) is a missense variant. The governing ClinGen RASopathy VCEP specification for KRAS (v2.3) defines PP3 for missense variants as REVEL >= 0.7. The local REVEL lookup returns a score of 0.797 for this variant, which meets the VCEP threshold. PP3 is applied at Supporting strength per the VCEP's defaultStrength/rule for this criterion. SpliceAI predicts no splice impact (max delta 0.011), so there is no splicing-path evidence contributing to PP3 for this variant; the missense REVEL path is the sole basis.

Rule

PP3_Supporting

Evidence considered

• ClinGen RASopathy VCEP KRAS specification v2.3 (cspec) defines PP3 for missense variants as REVEL >= 0.7, Supporting strength.
• Local REVEL predictor lookup (revel) returns a score of 0.797 for NM_033360.2:c.57G>T, which is >= 0.7, satisfying the VCEP PP3 threshold.

Sources

Reasoning

The KRAS VCEP v2.3 restricts PM1 to variants falling within four specific critical/well-established functional domains: P-loop (AA 10-17), Switch I (AA 25-40), Switch II (AA 57-64), and SAK (AA 145-156). Leu19 (codon 19) falls just outside the P-loop boundary (10-17) and outside all other listed domains, so the variant does not meet the VCEP's PM1 domain criterion. The VCEP alignment table (Alignment-with-PM1-domains.pptx) confirms the P-loop boundary is HRAS/KRAS AA 10-17, consistent with the cspec rule text.

Rule

KRAS VCEP v2.3 PM1: residue must fall in P-loop (10-17), Switch I (25-40), Switch II (57-64), or SAK (145-156); Leu19 is outside all four ranges.

Evidence considered

• KRAS VCEP v2.3 (cspec) PM1 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).'
• Alignment-with-PM1-domains.pptx table lists P-loop as HRAS/KRAS AA 10-17, confirming residue 19 (Leu19) falls outside this and all other listed domain boundaries.

Sources

Reasoning

No exact-variant ClinVar expert-panel classification was identified. ClinVar contains a Likely pathogenic clinical-laboratory submission with no assertion criteria and zero review stars; it is not eligible to trigger PP5. The KRAS VCEP also marks PP5 as not applicable.

Rule

PP5 may be applied only for an exact-variant ClinVar expert-panel Pathogenic/Likely pathogenic classification; ordinary laboratory submissions, aggregate labels, and non-expert assertions do not qualify.

Evidence considered

• ClinVar variation 376034 is reported as Likely pathogenic from a clinical-laboratory submission with no assertion criteria provided; the audit records zero expert-panel submissions and zero review stars, so this assertion cannot trigger PP5.
• The KRAS Version 2.3 VCEP marks PP5 as not applicable.

Still missing

• No exact-variant ClinVar expert-panel Pathogenic or Likely pathogenic classification.

Sources

Reasoning

BA1 is not met because the variant is reported absent from gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada v1.0; therefore no allele frequency reaches the KRAS VCEP stand-alone benign threshold of 0.05%.

Rule

GnomAD filtering allele frequency >=0.05%.

Evidence considered

• The KRAS VCEP specifies BA1 at a gnomAD filtering allele frequency of at least 0.05%.
• The variant NM_033360.2:c.57G>T (KRAS p.Leu19Phe) is reported absent from gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada v1.0; an absent variant cannot meet a 0.05% frequency threshold.

Still missing

• The case bundle provides absence calls rather than numeric allele counts and allele frequencies, but absence is sufficient to exclude the BA1 frequency threshold.

Sources

Reasoning

BS1 is not met because the variant is reported absent from gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada v1.0; therefore no allele frequency reaches the KRAS VCEP BS1 threshold of 0.025%.

Rule

GnomAD filtering allele frequency >=0.025%.

Evidence considered

• The KRAS VCEP specifies BS1 at a gnomAD filtering allele frequency of at least 0.025%.
• The variant NM_033360.2:c.57G>T (KRAS p.Leu19Phe) is reported absent from gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada v1.0; an absent variant cannot meet a 0.025% frequency threshold.

Still missing

• The case bundle provides absence calls rather than numeric allele counts and allele frequencies, but absence is sufficient to exclude the BS1 frequency threshold.

Sources

Reasoning

The VCEP specification defines BP4 for missense variants as REVEL <= 0.3. The REVEL score for this variant is 0.797, well above the BP4 threshold, so BP4 is not met. SpliceAI also does not provide grounds for BP4 here since this criterion group's BP4 pathway per the VCEP is REVEL-based for missense variants, and the observed REVEL score contradicts a benign computational prediction.

Rule

BP4_not_met

Evidence considered

• ClinGen RASopathy VCEP KRAS specification v2.3 (cspec) defines BP4 for missense variants as REVEL <= 0.3, Supporting strength.
• Local REVEL predictor lookup (revel) score of 0.797 exceeds the BP4 threshold of <=0.3, so the benign computational criterion is not met.

Sources

Reasoning

No exact-variant ClinVar expert-panel Benign or Likely benign classification was identified. The only relevant ClinVar germline assertion is a non-expert clinical-laboratory Likely pathogenic submission without assertion criteria; it cannot trigger BP6. The KRAS VCEP also marks BP6 as not applicable.

Rule

BP6 may be applied only for an exact-variant ClinVar expert-panel Benign/Likely benign classification; ordinary laboratory submissions, aggregate labels, and non-expert assertions do not qualify.

Evidence considered

• ClinVar variation 376034 has no expert-panel submission; the audit records one clinical-laboratory Likely pathogenic submission with no assertion criteria and zero review stars, which cannot trigger BP6.
• The KRAS Version 2.3 VCEP marks BP6 as not applicable.

Still missing

• No exact-variant ClinVar expert-panel Benign or Likely benign classification.

Sources

Reasoning

PS1 (KRAS VCEP v2.3) requires an alternate nucleotide change producing the identical amino acid change (p.Leu19Phe) that has already been established as pathogenic/likely pathogenic. The automated comparator search (pm5_candidates.json) attempted to harvest same-residue ClinVar comparators but failed with an HTTP 429 rate-limit error before returning results, so no comparator (same or different nucleotide change at codon 19) could be confirmed or excluded. No other established pathogenic p.Leu19Phe entry was identified in the reviewed evidence.

Rule

KRAS VCEP v2.3 PS1: same amino acid change as a previously established [likely] pathogenic variant regardless of nucleotide change, including analogous residue positions in HRAS/KRAS/MRAS/NRAS/RIT1/RRAS2.

Evidence considered

• pm5_candidates.json: found=false, candidate_summary.same_residue_candidates=0, errors=['HTTP Error 429: Too Many Requests']; recommendation explicitly states not to apply PM5/PS1-type logic from this artifact alone.
• KRAS VCEP v2.3 (cspec) PS1 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.'

Still missing

• Automated ClinVar same-residue comparator search failed (HTTP 429 Too Many Requests) before returning results; a manual ClinVar search for other nucleotide changes resulting in p.Leu19Phe (e.g. KRAS[GENE] AND L19[VARNAME]) has not been completed.

Sources

Reasoning

No germline proband with documented parental testing is provided. The only directly relevant publication reviewed describes c.57G>T/p.Leu19Phe as a tumor-specific somatic mutation in three colorectal carcinomas, with the mutation absent from patient-matched normal tissue; this does not establish a de novo germline event.

Rule

KRAS VCEP v2.3 PS2: Very Strong = 4 points; Strong = 2 points; Moderate = 1 point.

Evidence considered

• The KRAS VCEP Version 2.3 framework specifies PS2 using point-based de novo evidence, but the case bundle contains no germline proband or parental testing data.
• Akagi et al. identified the c.57G>T/p.Leu19Phe change in three primary colon carcinomas and reported that the band shift was absent from patient-matched normal tissue, indicating a tumor-specific somatic mutation; this is not evidence for germline de novo occurrence.

Still missing

• A germline proband with a confirmed phenotype consistent with KRAS-associated RASopathy is not documented.
• Parental testing results and confirmation of absence in both parents are not provided.
• The available exact-variant publication concerns somatic tumors rather than a germline de novo case.

Sources

Reasoning

The KRAS RASopathy VCEP (cspec v2.3) defines PS3 with strengths Moderate (≥2 different approved assays) or Supporting (1 approved assay), citing an approved-functional-studies list (RAS Activation Assay, MEK/ERK Activation Assays, etc.) with variant-level validation controls. No source in this case bundle reports a validated functional assay (RAS-GTP loading, MEK/ERK phosphorylation, or other VCEP-approved assay) performed specifically on KRAS p.(Leu19Phe)/c.57G>T. OncoKB and COSMIC/hotspot annotations describe curated oncogenicity and mutational recurrence, not a wet-lab functional assay result, and none of the retrieved full-text publications (PMID:17150185, PMID:20147967, PMID:28572459, PMID:22918138, PMID:23619274, PMID:34131312) mention this specific variant on full-text or abstract search. Without a variant-specific approved-assay result, PS3 cannot be applied under the VCEP rule.

Evidence considered

• SVI-RASopathy-VCEP-V2-Approved-Functional-Studies.xlsx lists approved assays (RAS Activation Assay, MEK Activation Assay, ERK Activation Assay, and gene-specific assays for PTPN11/BRAF/RAF1/LZTR1) with validation control variant tables, but no entry for KRAS L19F/c.57G>T was found in the accessible summary content.
• Full-text search of publications/17150185.txt, 20147967.txt, 28572459.txt, 22918138.txt, 23619274.txt, and 34131312.txt for 'L19', 'Leu19', '19F', 'codon 19' returned zero matches; none of these sources discuss this specific variant.
• evidence.json shows OncoKB provides a curated Gain-of-function/Oncogenic label for KRAS L19F citing PMID:17150185, PMID:20147967, PMID:28572459 as background literature, but these are curation citations, not a described functional assay methodology/result for this variant, so they do not satisfy the VCEP PS3 approved-assay requirement.

Still missing

• No VCEP-approved functional assay (RAS activation, MEK/ERK phosphorylation, or gene-specific assay) identified that specifically tested KRAS p.(Leu19Phe)/c.57G>T
• None of the case's retrieved full-text/abstract sources mention the exact variant despite functional/oncogenicity theming

Sources

Reasoning

The KRAS VCEP specifies PS4 for autosomal-dominant RASopathy cases using phenotype-adjusted point scoring: strong at >=5 points, moderate at >=3 points, and supporting at >=1 point. The available exact-variant publication describes three somatic colorectal carcinomas, not germline RASopathy patients, and provides no RASopathy case-control enrichment or germline RASopathy phenotype dataset. Therefore the available evidence cannot be scored under the governing PS4 specification.

Rule

KRAS VCEP Version 2.3 PS4: phenotype-adjusted point scoring; strong >=5 points, moderate >=3 points, supporting >=1 point.

Evidence considered

• The KRAS Version 2.3 VCEP specifies PS4 as applicable to autosomal-dominant RASopathy cases with phenotype-adjusted strengths of >=5 points (strong), >=3 points (moderate), and >=1 point (supporting).
• PMID:17150185 reports the exact codon-19 G>T/L19F change in three primary colon carcinomas from a 690-tumor somatic colorectal-cancer cohort; this is not germline RASopathy case-control evidence and cannot be converted into a VCEP PS4 score.

Still missing

• No germline RASopathy case series or case-control study for NM_033360.2:c.57G>T/p.Leu19Phe with phenotype-specific point assignments.
• No validated RASopathy phenotype information for the patient or affected cohort in this case bundle.

Sources

Reasoning

The KRAS VCEP v2.3 PM5 rule requires ≥1 different established [likely] pathogenic residue change at the same codon (codon 19), with strength escalating to Strong if ≥2 different pathogenic residue changes are each observed in ≥5 probands. The automated same-residue comparator search (pm5_candidates.json) targeting codon 19/Leu19 failed with an HTTP 429 rate-limit error and returned zero candidates, so it cannot be determined whether any other established pathogenic amino-acid substitution exists at codon 19.

Rule

KRAS VCEP v2.3 PM5: Moderate for 1 different [likely]pathogenic residue change at the same codon; Strong for ≥2 different such changes each seen in ≥5 probands.

Evidence considered

• pm5_candidates.json: pm5_mode='classic_same_residue_missense', eligible_for_classic_pm5_search=true, same_residue_candidates=0, errors=['HTTP Error 429: Too Many Requests']; recommendation states 'do not apply PM5 from this artifact alone.'
• KRAS VCEP v2.3 (cspec) PM5 rule: Moderate for 1 different pathogenic residue change at the same codon; Strong for ≥2 different pathogenic residue changes each observed in ≥5 probands.

Still missing

• Automated same-codon comparator search for codon 19 failed (HTTP 429); need a manual ClinVar/literature search for other pathogenic/likely pathogenic amino-acid changes at KRAS codon 19 to adjudicate PM5.

Sources

Reasoning

No presumed de novo germline occurrence without parental testing is documented. Available evidence is limited to an exact-variant somatic colon-cancer report and cannot support PM6.

Rule

KRAS VCEP v2.3 PM6: Strong = 2 points; Moderate = 1 point; Supporting = 0.5 points.

Evidence considered

• The KRAS VCEP Version 2.3 framework specifies PM6 with point-based presumed de novo evidence, but the case bundle provides no qualifying germline case.
• The reviewed exact-variant publication reports a tumor-specific somatic mutation in colorectal carcinomas, with absence from matched normal tissue; it does not describe presumed de novo germline inheritance.

Still missing

• No germline affected proband is documented.
• No parental genotypes or information showing that parental testing was unavailable are provided.
• No phenotype and inheritance assessment for a KRAS-associated RASopathy case is provided.

Sources

Reasoning

No familial segregation data or informative meioses are available. The exact-variant literature reviewed is a somatic tumor study and provides no affected relatives, unaffected relatives, parental testing, or segregation analysis.

Rule

KRAS VCEP v2.3 PP1: Supporting = ≥3 informative meioses; Moderate = ≥5; Strong = ≥7.

Evidence considered

• The KRAS VCEP Version 2.3 framework specifies PP1 as Supporting for at least 3 informative meioses, Moderate for at least 5, and Strong for at least 7; none are documented in this case.
• The reviewed exact-variant paper studied three unrelated colorectal carcinomas and did not report familial segregation or informative meioses.

Still missing

• No family history or relatives with phenotype and genotype are documented.
• No informative meioses are available to count.
• No phenotype consistency assessment across segregating relatives is provided.

Sources

Reasoning

BS2 cannot be assessed from the case bundle. No healthy adult homozygotes, heterozygotes, or hemizygotes carrying this variant are documented, and there is no evidence about age, phenotype, penetrance, or disease prevalence in carriers.

Rule

Strength adjustment using point-based scoring based on phenotypic specifications; healthy homozygote or heterozygote observations require consideration of penetrance and age.

Evidence considered

• The KRAS VCEP defines BS2 using point-based scoring for healthy homozygous or heterozygous individuals, but the bundle contains no such observations for this variant.

Still missing

• No healthy-carrier observations are provided.
• No carrier genotype, zygosity, age, phenotype, penetrance, or disease-prevalence data are provided.

Reasoning

No informative meiosis demonstrates that the variant does not segregate with disease. The available exact-variant publication is a somatic tumor study and contains no familial non-segregation analysis.

Rule

KRAS VCEP v2.3 BS4: Strong requires only one informative meiosis.

Evidence considered

• The KRAS VCEP Version 2.3 framework requires at least one informative meiosis for BS4; no such meiosis is present in the case bundle.
• Akagi et al. reported a tumor-specific somatic mutation absent from matched normal tissue, not non-segregation among relatives in a germline family.

Still missing

• No affected and unaffected relatives with genotype and phenotype data are documented.
• No informative meiosis or family-based non-segregation observation is available.
• The available somatic tumor evidence cannot establish benign non-segregation in a germline disease family.

Sources

Reasoning

The KRAS VCEP permits BP2 when there is an alternative molecular cause of a RASopathy in the same gene, with phenotype consistent with the expected severity, and assigns strength by the specified negative point score. The case evidence bundle contains no documented second KRAS variant, pathogenic variant in cis or trans, parental/offspring phase result, or affected-proband family observation establishing an alternative molecular cause. Therefore no BP2 points or strength can be assigned.

Rule

Points are awarded for an alternative molecular cause of a RASopathy in the same gene (and/or in conjunction with BP5) and the phenotype is consistent with expected severity of the RASopathy. Supporting: ≥ (-1) Point; Moderate: ≥ (-2) Points; Strong: ≥ (-4) Points.

Evidence considered

• The KRAS VCEP BP2 instruction states that points are awarded for an alternative molecular cause of a RASopathy in the same gene, with phenotype consistent with expected severity; its strength rules are Supporting at at least -1 point, Moderate at at least -2 points, and Strong at at least -4 points.
• The available case evidence documents KRAS NM_033360.2:c.57G>T (p.Leu19Phe) but does not document a second KRAS variant, cis/trans phase, or an affected-proband allelic observation relevant to BP2.

Still missing

• Documented alternative pathogenic or likely pathogenic KRAS variant in the same individual or family
• Definitive phase information establishing cis or trans configuration
• Phenotypic documentation showing consistency with the expected RASopathy severity

Sources

Reasoning

The KRAS VCEP BP5 rule requires an alternative molecular cause in a different gene and a phenotype consistent with the expected RASopathy severity, or a phenotype inconsistent with RASopathy fully explained by another causative variant, with point-based strength adjustment. The case bundle provides no patient phenotype, alternate causative variant, or qualifying molecular explanation. Somatic colorectal-cancer observations do not establish the VCEP BP5 conditions.

Rule

KRAS VCEP Version 2.3 BP5: alternate molecular cause/fully explanatory alternate variant plus qualifying phenotype, with supporting at >=(-1) point, moderate at >=(-2) points, and strong at >=(-4) points.

Evidence considered

• The KRAS Version 2.3 VCEP defines BP5 using an alternative molecular cause in a different gene and a phenotype consistent with expected RASopathy severity, or an inconsistent phenotype fully explained by another causative variant; no such evidence is present in the case bundle.

Still missing

• No documented patient phenotype or RASopathy severity assessment.
• No documented alternate molecular cause in a different gene or fully explanatory alternate variant.
• No VCEP BP5 point assignment.

Sources