HRAS c.173C>T, p.(Thr58Ile)

Reasoning

The exact variant NM_005343.4:c.173C>T (p.Thr58Ile) has been observed in at least 4 independent individuals with a RASopathy, including at least one reported de novo occurrence. The HRAS VCEP framework specifies PS4 Moderate at 3 or more points; the exact-variant expert-panel evidence signal explicitly documents PS4 at Moderate strength.

Rule

HRAS VCEP v2.3: PS4 Moderate for ≥3 points; exact-variant evidence documents ≥4 affected individuals with RASopathy.

Evidence considered

• The exact ClinVar variant record is NM_005343.4(HRAS):c.173C>T (p.Thr58Ile).
• The exact-variant ClinVar expert-panel evidence signal states that at least 4 independent occurrences were detected in patients with a RASopathy, including 1 de novo occurrence, and labels the evidence PS4 Moderate.
• The HRAS VCEP Version 2.3 framework defines PS4 Supporting at ≥1 point, Moderate at ≥3 points, and Strong at ≥5 points.

Still missing

• The available evidence does not provide a formal case-control enrichment calculation; PS4 is applied here using the HRAS VCEP case-count framework.

Sources

Reasoning

HRAS VCEP v2.3 PM1 is applicable at Moderate strength to variants falling within critical, well-established functional domains: P-loop (AA10-17), SW1/Switch I (AA25-40), SW2/Switch II (AA57-64), or SAK (AA145-156). Residue Thr58 falls within the SW2 (Switch II) domain (AA57-64) as defined by the VCEP alignment/domain supplement, satisfying the PM1 domain criterion.

Rule

cspec PM1: variant residue within P-loop (10-17), SW1 (25-40), SW2 (57-64), or SAK (145-156) domains -> Moderate.

Evidence considered

• cspec (HRAS 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).'
• Variant residue is Thr58 (NP_005334.1:p.(Thr58Ile)), which lies within the VCEP-defined SW2 domain range (AA57-64).
• VCEP Alignment-with-PM1-domains.pptx supplement lists P-loop (HRAS 10-17) and Switch I (HRAS 25-40) domain boundaries in its alignment table, corroborating the domain-boundary structure used to place residue 58 within SW2 per the cspec rule text (SW2 AA57-64 is stated directly in the cspec criterion text, not separately re-tabulated in the excerpted PPTX pages reviewed).

Sources

Reasoning

The exact HRAS c.173C>T (p.T58I) variant has been reported in multiple affected individuals with de novo or presumed de novo context, but the reviewed evidence does not consistently establish confirmed maternity and paternity. This supports PM6 at a conservative Supporting level under the HRAS VCEP point-based framework (0.5 point), while case-level parental records should be reviewed before any upward adjustment.

Rule

PM6: assumed de novo without confirmation of paternity and maternity; HRAS VCEP point scoring: 0.5 point Supporting, 1 point Moderate, 2 points Strong.

Evidence considered

• The HRAS VCEP specification defines PM6 as assumed de novo without confirmation of paternity and maternity and assigns 0.5 point at Supporting strength, 1 point at Moderate strength, and 2 points at Strong strength.
• PMID:18247425 reports the exact p.T58I change in a Costello syndrome patient and describes de novo HRAS mutations as the underlying disease mechanism, but does not provide explicit parental exclusion for that patient in the reviewed abstract.
• PMID:26888048 reports the exact variant with parental Sanger testing; the abstract confirms testing of the patient and both parents but does not explicitly state parental negativity or de novo status.

Still missing

• Case-level parental genotypes and testing methodology are not available for each reported de novo occurrence.
• The number of independent exact-variant presumed de novo occurrences and their phenotype detail require confirmation from the full case reports.

Sources

Reasoning

NM_005343.4:c.173C>T is a missense variant (p.Thr58Ile). The ClinGen RASopathy VCEP HRAS specification v2.3 defines PP3 at Supporting strength for missense variants with REVEL >= 0.7. The local REVEL v1.3 lookup for this exact variant (chr11:533883 G>A, GRCh37) returned a score of 0.777, which meets the >=0.7 threshold. REVEL's calibrated pathogenic threshold is drawn from the original REVEL publication (PMID 36413997, ClinGen SVI-endorsed calibration). No SpliceAI score was retrieved in this case's evidence bundle (only a lookup URL/screenshot placeholder is present, with no delta scores captured), so the splice-impact sub-path was not independently assessable; the missense REVEL path alone is sufficient to meet PP3 per the VCEP rule, and PP3 can only be applied once regardless of how many lines of evidence support it.

Rule

cspec PP3: REVEL >= 0.7 (Supporting) for missense variants

Evidence considered

• ClinGen RASopathy VCEP HRAS specification v2.3 (cspec): PP3 Supporting applies when REVEL >= 0.7 for missense variants.
• Local REVEL v1.3 lookup (source_registry key 'revel') for HRAS chr11:533883 G>A returned REVEL score = 0.777, meeting the >=0.7 VCEP threshold.
• REVEL's pathogenicity threshold calibration is documented in PMID 36413997 (ClinGen SVI-endorsed REVEL threshold recommendations).

Still missing

• SpliceAI delta scores were not captured in this case's evidence (only a lookup URL is registered); splice-impact sub-path could not be independently verified, though this does not change the PP3 outcome since REVEL already meets the VCEP threshold.

Sources

Reasoning

Expert panel ClinGen RASopathy Variant Curation Expert Panel classified as Pathogenic.

Rule

Apply PP5 at Supporting only for an exact-variant ClinVar expert-panel Pathogenic/Likely pathogenic classification; exclude ordinary laboratory submissions and aggregate labels.

Evidence considered

• ClinVar identifies the exact variant as NM_005343.4(HRAS):c.173C>T (p.Thr58Ile).
• ClinVar records an exact-match Pathogenic submission from the ClinGen RASopathy Variant Curation Expert Panel with review status reviewed by expert panel.
• The eight clinical-laboratory Pathogenic submissions and aggregate ClinVar classification were excluded from the PP5 decision.
• ClinVar expert panel classification

Sources

Reasoning

The HRAS VCEP requires the variant to be absent from gnomAD for PM2 at Supporting strength. Although it is absent from gnomAD v2.1 and gnomAD-Canada v1.0, it is present in gnomAD v4.1 at 1/1,613,288 alleles (AF 6.19852e-07), including one Middle Eastern allele; therefore the specified absence requirement is not met.

Rule

The variant must be absent from controls (gnomAD).

Evidence considered

• The ClinGen RASopathy Expert Panel HRAS version 2.3 specification defines PM2 at Supporting strength when the variant is absent from controls in gnomAD.
• The variant is absent from gnomAD v2.1 and gnomAD-Canada v1.0, but gnomAD v4.1 identifies one allele among 1,613,288 total alleles (AF 6.19852e-07; 0.00006%), with the allele observed in the Middle Eastern population.

Sources

Reasoning

The exact variant is reported in a father and son, which provides one affected-relative transmission and one informative meiosis if both individuals are considered affected. The HRAS VCEP threshold requires at least 3 informative meioses for PP1 Supporting, so the available segregation evidence is below threshold.

Rule

PP1: at least 3 informative meioses Supporting, at least 5 Moderate, and at least 7 Strong.

Evidence considered

• The HRAS VCEP specification requires at least 3 informative meioses for PP1 Supporting, at least 5 for Moderate, and at least 7 for Strong; segregation in more than one family is recommended.
• PMID:22488832 states that two additional individuals with the exact mutation were identified as father and son and that the mutation was transmitted, providing evidence for one informative meiosis rather than the required three.

Still missing

• Additional affected relatives and independently documented informative meioses are needed to reach the VCEP threshold.

Sources

Reasoning

The HRAS NM_005343.4:c.173C>T (p.Thr58Ile) variant has a gnomAD v4.1 total allele frequency of 6.19852e-07 (1/1,613,288 alleles; 0.00006%) and the highest reported subpopulation frequency is 0.000164962 (0.01650%) in the Middle Eastern population. Both are below the HRAS VCEP BA1 threshold of 0.05%.

Rule

GnomAD filtering allele frequency ≥0.05%.

Evidence considered

• The ClinGen RASopathy Expert Panel HRAS version 2.3 specification defines BA1 as applicable at a gnomAD filtering allele frequency of at least 0.05%.
• gnomAD v4.1 reports total AF 6.19852e-07 (1/1,613,288 alleles; 0.00006%) and a highest observed Middle Eastern AF of 0.000164962 (0.01650%), with 0 homozygotes; these frequencies do not reach 0.05%.

Sources

Reasoning

The variant's gnomAD v4.1 total allele frequency is 6.19852e-07 (0.00006%), and its highest observed subpopulation frequency is 0.000164962 (0.01650%) in the Middle Eastern population. Both are below the HRAS VCEP BS1 threshold of 0.025%.

Rule

GnomAD filtering allele frequency ≥0.025%.

Evidence considered

• The ClinGen RASopathy Expert Panel HRAS version 2.3 specification defines BS1 at a gnomAD filtering allele frequency of at least 0.025%.
• gnomAD v4.1 reports total AF 6.19852e-07 (0.00006%) and a highest observed Middle Eastern AF of 0.000164962 (0.01650%); neither reaches 0.025%.

Sources

Reasoning

BS4 requires lack of segregation in an affected family and the HRAS VCEP specifies one informative meiosis. The available exact-variant family report instead documents transmission from an affected father to an affected son; no affected non-carrier or other non-segregating family member is reported. BS4 is therefore not supported.

Rule

BS4: lack of segregation in affected members of a family; one informative meiosis is sufficient for Strong strength.

Evidence considered

• The HRAS VCEP specification defines BS4 as lack of segregation in affected family members and requires only one informative meiosis for Strong strength.
• PMID:22488832 reports father-son transmission of the exact HRAS c.173C>T (p.T58I) variant, which is segregation rather than lack of segregation; the abstract does not report an affected non-carrier.

Still missing

• No family-level phenotype and genotype table is available documenting an affected non-carrier or other informative non-segregating meiosis.

Sources

Reasoning

The ClinGen RASopathy VCEP HRAS specification v2.3 defines BP4 at Supporting strength for missense variants with REVEL <= 0.3. The observed REVEL score for this variant is 0.777, well above the BP4 threshold, so BP4 is not met. A BayesDel score (0.433392) was also retrieved for this variant, but per this pipeline's calibration policy no verified published threshold/publication for BayesDel is available, so it was not used to inform BP4 (or PP3) and is treated as not_available for computational evidence purposes.

Rule

cspec BP4: REVEL <= 0.3 (Supporting) for missense variants

Evidence considered

• ClinGen RASopathy VCEP HRAS specification v2.3 (cspec): BP4 Supporting applies when REVEL <= 0.3 for missense variants.
• Local REVEL v1.3 lookup (source_registry key 'revel') returned REVEL score = 0.777, far above the <=0.3 BP4 threshold, so BP4 criterion is not satisfied.
• A BayesDel score of 0.433392 was retrieved (source_registry key 'bayesdel') but was not applied to BP4/PP3 because no verified published calibration threshold/publication for BayesDel is available to this pipeline.

Sources

Reasoning

The HRAS VCEP framework requires confirmed de novo status with point-based scoring. The available literature documents the exact HRAS c.173C>T (p.T58I) variant in affected individuals and parental testing in one report, but the reviewed abstract does not explicitly document that both parents were negative for the variant; another report describes a father-son transmission. Therefore, confirmed de novo evidence sufficient to assign PS2 cannot be established from the available auditable material.

Rule

PS2: confirmed de novo with maternity and paternity confirmed; HRAS VCEP point scoring: 4 points Very Strong, 2 points Strong, 1 point Moderate.

Evidence considered

• The HRAS VCEP v2.3 specification states that PS2 requires de novo occurrence with both maternity and paternity confirmed and assigns 1, 2, or 4 points for Moderate, Strong, or Very Strong evidence, respectively.
• PMID:18247425 reports an exact p.T58I patient but its abstract states generally that Costello syndrome is caused by de novo HRAS mutations; it does not provide explicit negative parental testing for the p.T58I case.
• PMID:26888048 reports the exact HRAS c.173C>T (p.T58I) variant and states that the mutation was verified by Sanger sequencing in the patient and her parents, but the reviewed abstract does not state that both parents were negative.
• PMID:22488832 documents father-son transmission of the exact variant and therefore shows that at least one reported occurrence was inherited rather than de novo.

Still missing

• Obtain the original case-level parental genotypes and explicit maternity and paternity confirmation for each claimed de novo occurrence.
• Resolve whether the reported de novo occurrences correspond to the exact HRAS c.173C>T (p.T58I) variant rather than other RASopathy variants.

Sources

Reasoning

ClinGen RASopathy VCEP v2.3 (HRAS) marks PS3 as applicable, requiring one or more of the VCEP-approved functional assays (RAS Activation Assay, MEK Activation Assay, ERK Activation Assay, SHP-2 Phosphatase Activity, BRAF/RAF1 Kinase Activity, LZTR1 Stability) performed on the variant itself: Supporting for one approved assay, Moderate for two or more. The bundle contains no functional study of HRAS p.Thr58Ile (NM_005343.4:c.173C>T) using any of those approved assays. The only biochemical functional data found (PMID:16474405, Schubbert et al. 2006) characterizes the analogous T58I substitution in a different gene, K-Ras (KRAS), not HRAS, using intrinsic GTPase hydrolysis and GAP(p120/neurofibromin)-stimulated GTP-hydrolysis assays plus a hematopoietic colony assay -- none of which are on the VCEP-approved assay list, and the study is on a paralogous gene rather than the variant under review. PMID:16921267 is a review/perspective article discussing RAS pathway biochemistry generally and does not report new HRAS T58I-specific assay data. No direct evidence meeting the VCEP PS3 rule (approved assay, variant-specific, HRAS) was located.

Rule

ClinGen RASopathy VCEP v2.3 PS3: Moderate if >=2 different approved assays; Supporting if one approved assay.

Evidence considered

• ClinGen RASopathy VCEP v2.3 PS3 rule: Moderate for two or more different approved assays, Supporting for one approved assay; approved assay list from SVI-RASopathy-VCEP-V2-Approved-Functional-Studies.xlsx includes RAS Activation Assay, MEK Activation Assay, ERK Activation Assay, SHP-2 Phosphatase Activity (PTPN11-specific), BRAF Kinase Activity (BRAF-specific), RAF1 Kinase Activity (RAF1-specific), LZTR1 Stability/Localization (LZTR1-specific).
• PMID:16474405 (Schubbert et al., Nature Genetics 2006) reports that T58I K-Ras (not HRAS) has reduced intrinsic GTPase activity relative to wild-type, is less responsive to GAP-related domains of p120 GAP and neurofibromin than wild-type, and was studied in a hematopoietic progenitor colony assay -- these assay types are not on the VCEP PS3-approved list and the variant tested is in KRAS, a different (paralogous) gene, not the HRAS variant under review.
• PMID:16921267 (Kratz et al., Cell Cycle 2006) is a review/perspective summarizing RAS pathway biochemistry and germline RAS mutations broadly; it does not present new HRAS T58I-specific functional assay results.

Still missing

• No HRAS-specific functional assay (RAS/MEK/ERK activation or other VCEP-approved assay) for p.Thr58Ile was found in the case bundle or reviewed publications.
• PMID:16474405 functional data pertains to the analogous KRAS p.Thr58Ile variant, not HRAS, and uses assay types (intrinsic/GAP-stimulated GTPase hydrolysis, colony assay) not on the VCEP-approved list, so it cannot be used to satisfy the HRAS-specific PS3 rule.
• If a VCEP-approved assay result for HRAS p.Thr58Ile exists in primary literature not included in this bundle, it should be retrieved and re-assessed.

Sources

Reasoning

HRAS VCEP v2.3 PM5 requires evidence of one or more different [likely] pathogenic missense residue changes at the same codon (Thr58) to another amino acid, distinct from the Ile substitution seen here. The dedicated pm5_candidates.json comparator search for this case explicitly failed (HTTP 429 rate-limit error during ClinVar querying), returning zero same-residue candidates and zero total candidates evaluated; the evidence bundle contains no other same-codon alternate-amino-acid variant data (e.g., Thr58Ala, Thr58Ser, Thr58Asn) with an established [likely] pathogenic classification.

Rule

cspec PM5: >=2 different [likely] pathogenic residue changes at same codon in >=5 probands -> Strong; 1 [likely] pathogenic residue change at same codon -> Moderate.

Evidence considered

• pm5_candidates.json: 'found': false; 'eligible_for_classic_pm5_search': true; 'candidate_summary': {'total_candidates_seen': 2, 'same_residue_candidates': 0, ...}; 'errors': ['HTTP Error 429: Too Many Requests']; explicit instruction not to apply PM5 from this artifact alone.

Still missing

• pm5_candidates.json PM5 comparator search failed with 'HTTP Error 429: Too Many Requests' and returned 0 same-residue candidates; a repeat ClinVar/literature search for other Thr58 substitutions (e.g., p.Thr58Ala, p.Thr58Ser, p.Thr58Asn, p.Thr58Arg) with pathogenic/likely pathogenic classification is needed before PM5 can be adjudicated.

Sources

Reasoning

No evidence was provided that the variant was observed in a healthy adult individual. gnomAD v4.1 reports 0 homozygotes, but population absence of homozygotes is not evidence that a heterozygous carrier was clinically unaffected and does not satisfy the HRAS VCEP BS2 requirement.

Rule

Strength adjustment using point-based scoring based on phenotypic specifications; based on healthy homozygote or heterozygote individuals, with reduced points for apparently unaffected heterozygous individuals.

Evidence considered

• The ClinGen RASopathy Expert Panel HRAS version 2.3 specification bases BS2 on healthy homozygote or heterozygote individuals and allows strength adjustment using phenotypic specifications.
• gnomAD v4.1 reports 0 homozygotes and one allele overall, but does not establish that the carrier was a healthy adult or provide penetrance and phenotype follow-up data.

Still missing

• No documented clinically healthy adult heterozygote or homozygote for this variant was identified.
• No penetrance, age, phenotype, or clinical follow-up information is available for any population carrier.

Sources

Reasoning

The HRAS VCEP permits BP2 when an alternative molecular cause of RASopathy is present in the same gene, with point-based strength adjustment; its underlying allelic requirement is observation in trans with a pathogenic variant in a fully penetrant dominant disorder or in cis with a pathogenic variant. The case bundle documents HRAS c.173C>T (p.Thr58Ile) in affected individuals, including reported de novo observations, but does not document a second pathogenic HRAS variant, cis/trans phase, or an alternative same-gene molecular cause. Therefore BP2 cannot be applied.

Rule

Strong: ≥ (-4) points; Moderate: ≥ (-2) points; Supporting: ≥ (-1) point.

Evidence considered

• The HRAS VCEP Version 2.3 BP2 instruction awards points for an alternative molecular cause of RASopathy in the same gene and requires the relevant cis/trans allelic observation.
• ClinVar evidence in the case bundle reports the target HRAS variant as observed de novo in affected individuals and reports segregation with disease in one affected relative, but does not report a second pathogenic HRAS variant or cis/trans phase for BP2.
• The available case-level evidence concerns the target variant and affected-proband observations; it does not establish an alternate same-gene molecular cause.

Still missing

• No documented second pathogenic or likely pathogenic HRAS variant in the same individual.
• No cis/trans phase determination for the target variant relative to another pathogenic variant.
• No case-level evidence of an alternative same-gene molecular cause with phenotype consistent with the expected RASopathy severity.

Sources

Reasoning

No reliable evidence in the case bundle establishes an alternative molecular basis for the reported phenotype in individuals carrying this exact variant. The available exact-variant evidence concerns HRAS-related RASopathy and does not establish a competing diagnosis.

Rule

Use BP5 only when an alternative molecular basis explains the patient's phenotype; no such evidence is available.

Evidence considered

• The case bundle provides exact-variant HRAS-related RASopathy evidence but no documented alternative molecular basis for the phenotype.

Still missing

• No documented alternative molecular diagnosis or genotype for the affected individuals was provided.
• No case-level evidence was provided showing that the phenotype is explained by another pathogenic variant or mechanism.