NRAS c.182A>T, p.(Gln61Leu)

Reasoning

The NRAS RASopathy VCEP (cspec v2.3) defines PM1 at Moderate strength as applicable only to a fixed set of critical, well-established functional domains: P-loop (AA 10-17), SW1 (AA 25-40), SW2 (AA 57-64), and SAK (AA 145-156). Codon 61 (Gln61) falls within the Switch II (SW2, AA 57-64) domain per the VCEP's domain table. The VCEP framework further specifies PM1 and PM5 may be applied in conjunction at moderate strength, and PM1 is only excluded if PM5_Strong is separately applied; because PM5 comparator evidence for this variant is not established in this case (see PM5), PM1 is applied independently at Moderate strength.

Rule

Moderate: variant residue falls within a VCEP-defined critical functional domain (P-loop 10-17, SW1 25-40, SW2 57-64, SAK 145-156) (NRAS RASopathy VCEP v2.3).

Evidence considered

• cspec (NRAS VCEP v2.3): 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).'
• Case summary normalization: NP_002515.1:p.(Gln61Leu) places the variant at residue 61, within the VCEP-defined SW2 domain boundary (AA 57-64).
• Final classification framework instructionsToUse for PM1: 'Applicable for observed analogous residue positions in HRAS, KRAS, MRAS, NRAS, RIT1, and RRAS2. PM1 and PM5 may be used in conjunction at moderate levels, however, PM1 may not be applied if PM5_Strong is applied to avoid overweighting.'

Sources

Reasoning

The NRAS RASopathy Expert Panel specifies PM2 at supporting strength when the variant is absent from gnomAD. NM_002524.5:c.182A>T (NRAS p.Gln61Leu) is reported absent from both gnomAD v2.1 and gnomAD v4.1; PM2_Supporting is met.

Rule

NRAS RASopathy Expert Panel PM2_Supporting: variant absent from controls (gnomAD).

Evidence considered

• The governing NRAS RASopathy Expert Panel specification defines PM2 at supporting strength when the variant is absent from controls in gnomAD.
• gnomAD v2.1 and gnomAD v4.1 each report NM_002524.5:c.182A>T (NRAS p.Gln61Leu) as absent.

Still missing

• The collected gnomAD records do not provide sample-number or coverage details.

Sources

Reasoning

NM_002524.5:c.182A>T (p.Gln61Leu) is a missense variant. The ClinGen RASopathy VCEP (NRAS-specific CSPEC v2.3) PP3 rule specifies: for missense variants, REVEL >= 0.7 meets PP3 at Supporting strength. The local REVEL lookup for this variant returns 0.897, which exceeds the VCEP threshold. PP3 is applied once at Supporting strength per the VCEP's own missense sub-path; the splicing sub-path is not invoked because SpliceAI shows no meaningful splice disruption (see BP4 rationale), avoiding any double counting between the two PP3 sub-paths.

Rule

REVEL >= 0.7 (missense sub-path, ClinGen RASopathy VCEP NRAS CSPEC v2.3)

Evidence considered

• VCEP (ClinGen RASopathy Expert Panel NRAS CSPEC v2.3) PP3 rule: 'For missense variants: REVEL >= 0.7. For splicing impact, predicted outcome must match disease mechanism.' Default strength Supporting.
• Local REVEL v1.3 lookup for chr1:114713908 T>A returns REVEL score = 0.897, which is >= the VCEP's 0.7 missense threshold for PP3.

Sources

Reasoning

The NRAS RASopathy Expert Panel BA1 threshold is gnomAD filtering allele frequency >=0.05%. The variant is reported absent from gnomAD v2.1 and v4.1, so it cannot meet BA1.

Rule

NRAS RASopathy Expert Panel BA1: gnomAD filtering allele frequency >=0.05%.

Evidence considered

• The governing NRAS RASopathy Expert Panel specification (version 2.3) defines BA1 as gnomAD filtering allele frequency >=0.05%.
• gnomAD v2.1 and gnomAD v4.1 each report NM_002524.5:c.182A>T (NRAS p.Gln61Leu) as absent.

Still missing

• The collected gnomAD records do not provide sample-number or coverage details; this does not affect the conclusion that an absent variant is below the BA1 frequency threshold.

Sources

Reasoning

The NRAS RASopathy Expert Panel BS1 threshold is gnomAD filtering allele frequency >=0.025%. The variant is reported absent from gnomAD v2.1 and v4.1, so it cannot meet BS1.

Rule

NRAS RASopathy Expert Panel BS1: gnomAD filtering allele frequency >=0.025%.

Evidence considered

• The governing NRAS RASopathy Expert Panel specification (version 2.3) defines BS1 as gnomAD filtering allele frequency >=0.025%.
• gnomAD v2.1 and gnomAD v4.1 each report NM_002524.5:c.182A>T (NRAS p.Gln61Leu) as absent.

Still missing

• The collected gnomAD records do not provide sample-number or coverage details; this does not affect the conclusion that an absent variant is below the BS1 frequency threshold.

Sources

Reasoning

The VCEP BP4 rule requires REVEL <= 0.3 for missense variants; the observed REVEL score of 0.897 is far above this threshold, so BP4 is not met via the missense sub-path. On the splicing sub-path, SpliceAI predicts no significant splice impact (max delta score = 0.01, all four delta scores <=0.01), which is consistent with no splice disruption but is not the basis the VCEP specifies for BP4 (no explicit SpliceAI threshold is defined in the NRAS CSPEC BP4 rule for missense variants, and applying it here would double count against the PP3 REVEL-based pathogenic-supporting call). BayesDel (0.356835) was also returned by prefetch but is not used: no verified published calibration threshold with a citable PMID is available for BayesDel in this pipeline, so it is treated as not_available for BP4 per standing policy, regardless of the raw score.

Rule

REVEL <= 0.3 (missense sub-path, ClinGen RASopathy VCEP NRAS CSPEC v2.3); observed REVEL = 0.897, rule not satisfied

Evidence considered

• VCEP (ClinGen RASopathy Expert Panel NRAS CSPEC v2.3) BP4 rule: 'For missense variants: REVEL <= 0.3.' Default strength Supporting.
• Local REVEL v1.3 lookup for chr1:114713908 T>A returns REVEL score = 0.897, well above the VCEP's 0.3 BP4 threshold, so BP4 is not met.
• SpliceAI Lookup for NM_002524.5:c.182A>T reports max delta score = 0.01 (DS_AG=0.0, DS_AL=0.009, DS_DG=0.003, DS_DL=0.002), indicating no significant predicted splice impact, but this is not independently sufficient under the VCEP BP4 rule for a missense variant and is not used to call BP4 met.

Still missing

• No VCEP-defined BP4 SpliceAI threshold exists for this missense variant to independently support a splicing-based benign call.
• BayesDel score (0.356835) exists in prefetch but has no verified published calibration threshold citable to a specific PMID/table, so it cannot be used for BP4.

Sources

Reasoning

The NRAS RASopathy VCEP (cspec v2.3) extends PS1 to 'observed analogous pathogenic residue positions' in HRAS, KRAS, MRAS, NRAS, RIT1, and RRAS2 (same amino acid change at the homologous codon, regardless of nucleotide change). p.Gln61Leu is a well-known oncogenic hotspot substitution (OncoKB: Oncogenic, gain-of-function) but the case bundle's dedicated PM5/PS1 comparator search (pm5_candidates.json) was scoped only to NRAS residue 61 itself and returned zero candidates; it did not query the homologous Q61 codon in HRAS, KRAS, MRAS, RIT1, or RRAS2 for an established germline pathogenic classification of the identical Gln->Leu change. No source in this bundle documents a prior ClinVar/expert-panel pathogenic classification (germline) of Gln61Leu at this or an analogous paralog residue, so the criterion cannot be affirmed from available evidence.

Rule

Same amino acid change as a previously established pathogenic variant regardless of nucleotide change; applicable for observed analogous pathogenic residue positions in HRAS, KRAS, MRAS, NRAS, RIT1, and RRAS2 (NRAS RASopathy VCEP v2.3).

Evidence considered

• cspec (NRAS VCEP v2.3): PS1 rule = 'Same amino acid change as a previously established pathogenic variant regardless of nucleotide change. Applicable for observed analogous pathogenic residue positions in HRAS, KRAS, MRAS, NRAS, RIT1, and RRAS2.'
• pm5_candidates.json comparator search (classic_same_residue_missense mode, scoped to NRAS codon 61 only) returned candidate_summary.same_residue_candidates = 0 and recommendation.level = 'no_useful_candidates_found'; it explicitly instructs not to apply PM5 (or by extension the paralog-analog form of PS1) from this artifact alone.

Still missing

• No paralog-position (HRAS/KRAS/MRAS/RIT1/RRAS2 Gln61Leu or equivalent) comparator search was performed or returned in this case's evidence bundle.
• OncoKB 'Oncogenic' is a somatic functional annotation, not a germline ACMG/AMP pathogenic classification, and cannot on its own satisfy PS1's 'previously established pathogenic variant' requirement.

Sources

Reasoning

No case-level evidence establishes that the variant is de novo in an affected individual with both maternity and paternity confirmed.

Rule

NRAS RASopathy VCEP PS2: apply SVI de novo point-based scoring; PS2 requires confirmed de novo status, with 4, 2, or 1 points corresponding to Very Strong, Strong, or Moderate strength.

Evidence considered

• The NRAS RASopathy VCEP specification directs PS2/PM6 assessment by SVI point-based scoring, gives full points for RASopathy phenotypes, and reduces points when phenotypic information is minimal.
• The consolidated case-evidence bundle contains no proband phenotype, parental genotypes, parentage confirmation, or documented de novo observation for NM_002524.5:c.182A>T.

Still missing

• Obtain the proband phenotype and parental molecular test results.
• If both parents test negative, obtain confirmation of biological maternity and paternity and phenotype specificity sufficient for RASopathy point assignment.

Sources

Reasoning

The ClinGen RASopathy VCEP v2.3 specification marks PS3 applicable for NRAS with strength capped at Moderate (>=2 different approved assays) or Supporting (1 approved assay), where 'approved assays' are restricted to a defined list (RAS Activation Assay, MEK Activation Assay, ERK Activation Assay, SHP-2 Phosphatase Activity, BRAF/RAF1 Kinase Activity, LZTR1 Stability/Localization). The case bundle contains no primary experimental data (e.g., RAS-GTP pulldown ratios, phospho-MEK/phospho-ERK quantification, or other approved-assay readouts) generated for or reporting on this specific variant, NRAS c.182A>T (p.Gln61Leu). Available functional-adjacent sources are: (1) OncoKB's curated annotation labeling the variant Oncogenic with a Gain-of-function biological-effect tag, which is a database-level curation summary rather than a described, quantifiable assay result; and (2) structural/biochemical literature on Ras Q61 catalytic mechanism and GTP hydrolysis chemistry (PMID:20194776, PMID:10574788) that discuss the RasQ61L mutant and general GTPase hydrolysis kinetics of Q61 mutants as a class, but do not report a quantitative, approved-assay-format functional study specific to this variant with defined normal/mutant range and controls. Without a described approved assay result meeting the VCEP's evidentiary bar, PS3 cannot be scored.

Rule

SVI-RASopathy-VCEP-V2 PS3: Two or more different approved assays -> Moderate; One approved assay -> Supporting.

Evidence considered

• OncoKB (oncokb) curated annotation for NRAS Q61L: biological-effect context Gain-of-function, oncogenicity label Oncogenic; this is a database curation summary, not a primary approved-assay readout, so it was not used to score PS3.
• PMID:20194776 (Buhrman et al., Allosteric modulation of Ras positions Q61 for a direct role in catalysis) discusses the RasQ61L oncogenic mutant in the context of switch II ordering and Q61 catalytic positioning, but reports structural/biochemical mechanism rather than a VCEP-approved functional assay result for this variant.
• PMID:10574788 (pre-hydrolysis state of p21(ras) in complex with GTP) addresses general GTP hydrolysis mechanism in Ras-like GTPases, not a variant-specific approved assay.
• SVI-RASopathy-VCEP-V2-Approved-Functional-Studies material (vcep_svi_rasopathy_vcep_v2_approved_functional_studies) defines the approved assay list and applicable strength tiers (Moderate for >=2 assays, Supporting for 1 assay) but the bundle contains no results from any of these listed assays for this variant.

Still missing

• No approved-assay (RAS/MEK/ERK activation, SHP-2, BRAF/RAF1 kinase, LZTR1 stability) experimental data retrieved for this specific variant in the case bundle.
• OncoKB Gain-of-function/Oncogenic label is a curated classification, not a described assay with normal-range/mutant-range controls, so it cannot be used to satisfy the VCEP's PS3 approved-assay requirement.
• Structural biology papers on Q61 catalytic mechanism (PMID:20194776, PMID:10574788) address general Q61 GTPase chemistry, not a variant-specific quantitative functional assay meeting VCEP approved-assay criteria.

Sources

Reasoning

The NRAS RASopathy VCEP specification permits PS4 at Supporting (≥1 point), Moderate (≥3 points), or Strong (≥5 points), but this case bundle contains no germline RASopathy case series or case-control dataset for p.(Gln61Leu) with the proband counts, phenotype attribution, and control comparison needed to assign points.

Rule

ClinGen RASopathy Expert Panel NRAS v2.3: PS4 Supporting at ≥1 point, Moderate at ≥3 points, Strong at ≥5 points.

Evidence considered

• The governing ClinGen RASopathy Expert Panel specification for NRAS sets PS4 thresholds of ≥1 point (Supporting), ≥3 points (Moderate), and ≥5 points (Strong).
• The supplied evidence establishes only somatic cancer recurrence and non-expert ClinVar submissions; neither supplies a germline RASopathy prevalence comparison or VCEP PS4 point total.

Still missing

• Variant-specific germline RASopathy proband data and the VCEP point calculation are needed.
• A controlled enrichment analysis, or a case series with sufficiently documented phenotypes and independent observations, is needed.

Sources

Reasoning

The NRAS RASopathy VCEP (cspec v2.3) PM5 rule requires >=1 other [likely] pathogenic amino acid change at the same codon (Moderate) or >=2 different [likely] pathogenic residue changes at the same codon observed in >=5 probands (Strong). The case-generated same-residue comparator search (pm5_candidates.json, mode classic_same_residue_missense, explicitly scoped to NRAS codon 61) reviewed 7 total candidates but identified zero same-residue comparator variants (same_residue_candidates = 0, expert_panel_plp_candidates = 0, non_expert_majority_plp_candidates = 0), yielding recommendation level 'no_useful_candidates_found' and an explicit instruction not to apply PM5 from that artifact alone. No other source in this bundle documents an established [likely] pathogenic germline classification of a different amino acid substitution at NRAS codon 61 (e.g., Gln61Arg/Lys/His/Pro/Glu) with proband counts, so the rule's evidentiary threshold cannot be confirmed or excluded from available data.

Rule

Moderate: 1 established [likely] pathogenic residue change at the same codon; Strong: >=2 different [likely] pathogenic residue changes at the same codon in >=5 probands (NRAS RASopathy VCEP v2.3).

Evidence considered

• cspec (NRAS VCEP v2.3): PM5 default strength Moderate; Strong requires >=2 different [likely] pathogenic residue changes at the same codon observed in >=5 probands; Moderate requires 1 [likely] pathogenic residue change at the same codon.
• pm5_candidates.json: pm5_mode='classic_same_residue_missense', eligible_for_classic_pm5_search=true, candidate_summary.total_candidates_seen=7, same_residue_candidates=0, recommendation.summary='No clearly useful same-residue comparator variants were identified.'

Still missing

• The pm5_candidates.json search surfaced 0 usable same-residue comparators despite 7 total candidates seen; the underlying comparator records (classification, proband counts) were not retained/exposed in this bundle for direct review.
• No germline-focused ClinVar/expert-panel search for other codon-61 substitutions (e.g., Q61R, Q61K, Q61H) with established [likely] pathogenic status and proband counts is present in this case's evidence bundle.

Sources

Reasoning

No affected individual is reported with the variant absent from parents but without confirmed parentage; therefore assumed de novo evidence cannot be scored.

Rule

NRAS RASopathy VCEP PM6: apply SVI de novo point-based scoring to assumed de novo observations without confirmed maternity and paternity; 2, 1, and 0.5 points correspond to Strong, Moderate, and Supporting strength.

Evidence considered

• The NRAS RASopathy VCEP specification directs PM6 assessment by SVI de novo point-based scoring; PM6 is for assumed de novo observations without maternity and paternity confirmation and permits 2, 1, or 0.5 points for Strong, Moderate, or Supporting strength.
• The consolidated case-evidence bundle contains no proband phenotype, parental genotypes, or reported assumed de novo observation for NM_002524.5:c.182A>T.

Still missing

• Obtain the proband phenotype and parental molecular test results.
• Document whether parentage is unconfirmed, so that an observed de novo event can be assigned to PS2 rather than PM6 when appropriate.

Sources

Reasoning

No family segregation data or informative meioses are available for this variant.

Rule

NRAS RASopathy VCEP PP1: Supporting at ≥3 informative meioses, Moderate at ≥5, Strong at ≥7; segregation in more than one family is recommended.

Evidence considered

• The NRAS RASopathy VCEP specification recommends segregation in more than one family and sets PP1 thresholds at at least 3 informative meioses for Supporting, at least 5 for Moderate, and at least 7 for Strong.
• The consolidated case-evidence bundle contains no pedigree, relative genotypes, relative phenotypes, or informative-meiosis count for NM_002524.5:c.182A>T.

Still missing

• Obtain pedigree, phenotypes, and variant testing for affected and unaffected relatives.
• Document informative meioses across more than one family when available.

Sources

Reasoning

The NRAS RASopathy Expert Panel uses phenotype-specific point-based BS2 scoring: healthy homozygous or heterozygous individuals may contribute points, with reduced points for apparently unaffected heterozygotes, and BS2_Supporting and BS2 strengths require -1 and -4 points, respectively. No healthy/unaffected individual observations, zygosity data, ages, phenotypes, or point assignments are provided for this variant.

Rule

NRAS RASopathy Expert Panel BS2: phenotype-specific point scoring; -1 point for Supporting and -4 points for Strong.

Evidence considered

• The governing NRAS RASopathy Expert Panel specification directs BS2 strength adjustment by phenotypic point-based scoring and specifies healthy homozygous or heterozygous individuals, reduced points for apparently unaffected heterozygotes, -1 point for supporting strength, and -4 points for strong strength.
• No qualifying healthy/unaffected individual evidence is present in the consolidated case-evidence bundle.

Still missing

• Variant-specific observations in healthy or apparently unaffected individuals, including zygosity, age, phenotype assessment, and sufficient information to calculate the RASopathy Expert Panel BS2 point total.

Sources

Reasoning

No informative non-segregation observation in an affected family member is available.

Rule

NRAS RASopathy VCEP BS4: Strong evidence requires lack of segregation in affected family members and one informative meiosis; assess phenocopies and alternative molecular causes.

Evidence considered

• The NRAS RASopathy VCEP specification defines BS4 as lack of segregation in affected family members, at Strong strength with one informative meiosis, and cautions that phenocopies or multiple pathogenic variants can confound apparent non-segregation.
• The consolidated case-evidence bundle contains no affected-relative genotype/phenotype comparison or informative non-segregation meiosis for NM_002524.5:c.182A>T.

Still missing

• Obtain pedigree, affected-relative phenotypes, and variant testing to identify a true informative non-segregation event.
• Evaluate phenocopies and alternative molecular causes before applying BS4.

Sources

Reasoning

BP2 is applicable in the governing NRAS framework but cannot be assessed from the available evidence. No affected-proband observation documents a second alternative molecular cause in NRAS, no pathogenic co-variant is reported, and no cis/trans phase evidence is provided.

Rule

ClinGen RASopathy Expert Panel NRAS v2.3: BP2 points require an alternative molecular cause of RASopathy in the same gene (and/or in conjunction with BP5) with phenotype severity consistent with the expected RASopathy; supporting, moderate, and strong require at least -1, -2, and -4 points, respectively.

Evidence considered

• The governing NRAS v2.3 RASopathy Expert Panel specification permits BP2 using point-based evidence for an alternative molecular cause of RASopathy in the same gene, with phenotype severity considered.
• The consolidated case-evidence bundle contains no report of a second NRAS variant, cis/trans phase, or an alternative molecular diagnosis in an affected proband.

Still missing

• Affected-proband genotype data identifying any second relevant variant in NRAS or an alternative molecular diagnosis.
• Validated phase data (cis or trans) for c.182A>T relative to any relevant co-variant.
• Phenotype information sufficient to determine whether any alternative molecular cause explains the expected RASopathy severity.

Sources

Reasoning

The NRAS RASopathy VCEP specification permits BP5 at Supporting (≥−1 point), Moderate (≥−2 points), or Strong (≥−4 points), but the supplied case evidence contains no documented alternate molecular diagnosis that fully explains the relevant phenotype and no VCEP point calculation.

Rule

ClinGen RASopathy Expert Panel NRAS v2.3: BP5 Supporting at ≥−1 point, Moderate at ≥−2 points, Strong at ≥−4 points.

Evidence considered

• The governing ClinGen RASopathy Expert Panel specification sets BP5 thresholds of ≥−1 point (Supporting), ≥−2 points (Moderate), and ≥−4 points (Strong).
• No alternate molecular diagnosis or corresponding VCEP BP5 point evidence is present in the supplied bundle.

Still missing

• A documented independent molecular diagnosis and phenotypic attribution sufficient for a negative BP5 point calculation are needed.

Sources