Variant classification summary

NM_033632.3:c.1394G>A

FBXW7 · NP_361014.1:p.(Arg465His) · NM_033632.3

GRCh37: chr4:153249384 C>T · GRCh38: chr4:152328232 C>T

Gene: FBXW7 Transcript: NM_033632.3

Final call

Likely Pathogenic

PS3 strong PM1 moderate PM2 supporting PP3 supporting

Variant details

Gene

FBXW7

Transcript

NM_033632.3

Protein

NP_361014.1:p.(Arg465His)

gnomAD AF

0.0 (v4.1)

ClinVar

Tier I - Strong

OncoKB

Likely Oncogenic

Classification rationale

Interpretation summary

Generated evidence synthesis

1

NM_033632.3:c.1394G>A (p.Arg465His) in FBXW7 is a missense variant in the WD40 substrate-recognition domain of this E3 ubiquitin ligase tumor suppressor.

2

Two independent publications demonstrate that the R465H substitution severely impairs FBXW7 ubiquitin ligase activity: deficient in Notch1 ICD polyubiquitylation (PMID:17575125) and deficient in GRα polyubiquitylation (PMID:23228967), meeting PS3 at strong strength.

3

Arg465 is located on the surface of the WD40 β-propeller, the substrate-binding pocket critical for FBXW7 function, in a statistically significant mutational hotspot, meeting PM1 at moderate strength.

4

The variant is absent from gnomAD v2.1 (0/231,580 alleles) and v4.1 (0/1,593,060 alleles), meeting PM2 at supporting strength.

5

REVEL in silico prediction score of 0.557 supports a deleterious effect, meeting PP3 at supporting strength.

6

Applying generic ACMG/AMP 2015 combination rules: 1 strong (PS3) + 1 moderate (PM1) + 2 supporting (PM2, PP3) classifies this variant as Likely Pathogenic.

Final determination: Generic ACMG/AMP 2015 fallback rules support a Likely Pathogenic classification based on the observed combination of pathogenic criteria.

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_033632.3:c.1394G>A is a missense substitution (p.Arg465His), not a null variant (nonsense, frameshift, or canonical ±1,2 splice site). The generic PVS1 framework per PMC6185798 does not apply to missense variants.	pvs1_generic_framework
PS1	Not met	No evidence of a different pathogenic amino acid change at codon 465 with established pathogenicity. The pm5_candidates search found no same-residue comparator variants with expert panel pathogenic classification in ClinVar.	pm5_candidates
PS2	Not assessed	No de novo data (with confirmed paternity and maternity) is available for NM_033632.3:c.1394G>A in a germline context. The variant has been observed somatically in T-ALL patients but de novo germline status was not assessed.
PS3	Met	Two independent publications directly tested the R465H variant in functional assays and demonstrated unequivocal loss of E3 ubiquitin ligase function. PMID:17575125 showed R465H is severely deficient in Notch1 ICD polyubiquitylation and fails to suppress Notch1 signaling in a luciferase reporter assay. PMID:23228967 independently demonstrated R465H is severely deficient in polyubiquitylating GRα. Two independent studies testing the exact variant with unequivocal functional defect meets the strong strength threshold.	PMID:17575125 PMID:23228967
PS4	Not assessed	No case-control or cohort data for this variant in a germline disease context. The variant has been observed in 3/26 pediatric T-ALL patients (PMID:17575125) and in COSMIC (311 somatic entries), but no germline prevalence or case-control data are available for FBXW7-related neurodevelopmental syndrome.
PS5	N/A	PS5 is not a recognized criterion in the standard ACMG/AMP 2015 framework (PMID:25741868). De novo evidence is assessed under PS2 (with confirmed parentage) or PM6 (without confirmed parentage). No separate PS5 criterion is applicable.
PM1	Met	Arg465 is located on the surface of the WD40 β-propeller domain, the substrate-recognition pocket of FBXW7 critical for E3 ubiquitin ligase function. PMID:17575125 explicitly identifies Arg465 as a residue on the β-propeller surface essential for substrate recognition. The residue is in a statistically significant hotspot per cancerhotspots.org. The WD40 domain is a well-characterized functional domain without benign variation at structurally critical arginine positions.	PMID:17575125 oncokb
PM2	Met	NM_033632.3:c.1394G>A is absent from gnomAD v2.1 (0/231,580 alleles) and gnomAD v4.1 (0/1,593,060 alleles), yielding an allele frequency of 0% in all populations. This is well below the PM2 threshold of <0.1% for a rare variant absent from population databases.	gnomad_v2 gnomad_v4 gnomad_canada
PM5	N/A	No same-residue comparator variant with established pathogenic classification could be identified. The automated pm5_candidates search found no eligible comparator variants at Arg465. While R465C has been reported as a somatic mutation in the CCRF-CEM cell line (PMID:17575125), it lacks a ClinVar pathogenic classification with expert panel support and does not satisfy PM5 requirements.	pm5_candidates
PM6	Not assessed	No de novo data (without confirmed parentage) is available for this variant. The variant has been observed as a somatic mutation in T-ALL (PMID:17575125) but no germline de novo observations have been reported.
PP1	Not assessed	No segregation data are available for this variant. No family studies have been reported.
PP2	Not assessed	PP2 requires a gene with a low rate of benign missense variation where missense variants are a common disease mechanism. While FBXW7 missense variants in the WD40 domain are known to cause disease (both somatic and germline), gene-level constraint metrics (Z-score, missense depletion) were not available in this case to confirm a low rate of benign missense variation.
PP3	Met	REVEL score of 0.557 exceeds the 0.5 threshold predictive of a damaging effect. BayesDel score of 0.112528 is below the typical 0.27 threshold and HCI prior is unavailable. SpliceAI predicts no splice impact (max delta=0.00). Despite mixed in silico results, the REVEL score provides one line of computational support for a deleterious effect.	revel bayesdel spliceai
PP4	Not assessed	No specific phenotype or clinical data for the proband are available to assess whether the patient's phenotype is specific for FBXW7-related disease.
PP5	Not met	ClinVar variation 4530533 is classified as 'Tier I - Strong' with review status 'criteria provided, single submitter' (1 star). The user's PP5 rule requires ClinVar 3-star expert panel review to apply PP5 at supporting strength. A 1-star single-submitter classification does not meet this threshold.	clinvar
BA1	Not met	Variant is absent from gnomAD v2.1 (0/231,580 alleles) and v4.1 (0/1,593,060 alleles). Allele frequency of 0% is far below the BA1 threshold of >1%.	gnomad_v2 gnomad_v4
BS1	Not met	Variant is absent from gnomAD (AF=0%), well below the BS1 threshold of >0.3% allele frequency.	gnomad_v2 gnomad_v4
BS2	Not met	No data showing this variant observed in healthy adults with full penetrance expected at an early age. The variant is absent from population databases and has only been reported in somatic cancer contexts.
BS3	Not met	Functional studies in PMID:17575125 and PMID:23228967 demonstrate that R465H impairs FBXW7 E3 ubiquitin ligase function, showing loss of function rather than normal function. These studies support a deleterious effect, not a benign one. No studies demonstrate normal protein function for this variant.	PMID:17575125 PMID:23228967
BS4	Not assessed	No segregation data available to evaluate lack of cosegregation with disease.
BP1	Not met	BP1 applies when a missense variant occurs in a gene where primarily truncating variants cause disease. FBXW7-related neurodevelopmental syndrome and FBXW7-associated cancers are caused by both missense and truncating variants in the WD40 domain. Missense variants at critical arginine residues (Arg465, Arg479, Arg505) are a well-established disease mechanism in FBXW7 (PMID:17575125, PMID:35395208).	PMID:17575125
BP2	Not met	No data available showing this variant observed in trans with a pathogenic variant. The variant has not been observed in population databases, making it impossible to assess co-occurrence in trans.
BP4	Not met	REVEL score of 0.557 predicts a deleterious effect, not a benign one. While BayesDel is below threshold (0.113), the REVEL score provides computational evidence of pathogenicity, not benignity. Multiple in silico predictors do not agree on a benign effect.	revel bayesdel
BP5	Not met	BP5 is not applicable — the variant has been observed in somatic cancers (COSMIC: 311 entries, PMID:17575125: 3/26 T-ALL patients) and is classified as Likely Oncogenic by OncoKB. This is not a case where the variant is found in a case with an alternate molecular basis for disease.	oncokb
BP6	Not met	ClinVar classification is pathogenic ('Tier I - Strong') by a single submitter (1 star). The user's BP6 rule requires a 3-star expert panel benign or likely benign classification to apply BP6 at supporting strength. The current ClinVar classification supports pathogenicity, not benignity, and is only 1 star.	clinvar
BP7	Not met	BP7 applies to synonymous variants predicted to have no impact on splicing. NM_033632.3:c.1394G>A is a missense variant (p.Arg465His), not a synonymous variant. Although SpliceAI delta is 0.00, BP7 does not apply to missense variants.	spliceai
BP3	N/A	Skipped — BP3 applies to in-frame deletions/insertions in repetitive regions; this is a substitution variant.
PM3	N/A	Skipped — PM3 applies to recessive disorders with a pathogenic variant in trans; FBXW7-related neurodevelopmental syndrome is autosomal dominant and this variant has not been observed in trans.
PM4	N/A	Skipped — PM4 applies to protein length-altering variants (in-frame deletions/insertions, stop-loss); this is a missense substitution.

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