NM_000435.2:c.4688C>T (p.Pro1563Leu) is a missense variant in NOTCH3. It is absent from gnomAD v2.1 (0/242,354 alleles) and ultra-rare in gnomAD v4.1 (1/1,602,340 alleles; AF=6.24e-7), meeting PM2_Supporting.1 Multiple computational predictors indicate a benign impact: REVEL score 0.08, BayesDel score -0.37, and SpliceAI max delta 0.00. This meets BP4 (supporting benign).2 The variant is a missense substitution (not a null variant), so PVS1 is not applicable. It alters a non-cysteine residue (Pro1563) in EGF-like repeat 30, where the majority of established pathogenic NOTCH3 variants affect conserved cysteine residues; PM1 is not met.3 The variant is absent from ClinVar and has no reported functional studies, de novo observations, case-control data, or family segregation data. OncoKB classifies it as 'Unknown Oncogenic Effect.'4 In the generic ACMG/AMP 2015 framework, the current evidence yields PM2_Supporting and BP4_Supporting_Benign. These criteria are insufficient for a definitive classification; the variant remains a Variant of Uncertain Significance (VUS) pending additional clinical and functional data.5
NOTCH3
Final classification
VUS
NOTCH3 c.4688C>T · p.Pro1563Leu
NOTCH3
NM_000435.2:c.4688C>T (p.Pro1563Leu) is a missense variant in NOTCH3. It is absent from gnomAD v2.1 (0/242,354 alleles) and ultra-rare in gnomAD v4.1 (1/1,602,340 alleles; AF=6.24e-7), meeting PM2_Supporting.
gene-specific framework lacked a usable explicit final combination framework, so generic ACMG/AMP 2015 final-combination rules were applied as fallback; applied criteria: PM2 supporting, BP4 supporting benign; combination = 1 supporting + 1 supporting benign, which maps to VUS.
Classification rationale
PM2
BP4
VUS
NOTCH3 c.4688C>T
PM2 + BP4
→
VUS
2
revelbayesdelspliceai ↗
3
pvs1_variant_assessmentpvs1_generic_framework ↗
5
generic_acmg_combination_rules
Gene diagram
· NM_000435.2 · variants mapped to exon structure
NOTCH3
NM_000435.2
Fetching transcript structure from UCSC…
Applied criteria · 2 applied · 19 assessed
Applied · 2
Strength
Supporting
Moderate
Strong
Very strong
✓
PM2
supporting
Pathogenic
NM_000435.2:c.4688C>T is absent from gnomAD v2.1 (0/242,354 alleles) and observed at an ultra-rare frequency in gnomAD v4.1 (1/1,602,340 alleles; AF=6.24e-7), well below the 0.1% threshold for PM2_Supporting in the generic ACMG/AMP framework.
gnomAD v2.1: 0 alleles out of 242354 (AF=0.0%)gnomAD v4.1: 1 allele out of 1
✓
BP4
supporting
Benign
Multiple lines of computational evidence support a benign impact. REVEL score is 0.08 (below 0.5 threshold), BayesDel score is -0.37 (below pathogenic threshold), and SpliceAI predicts no splicing impact (max delta = 0.00). All three in silico predictors are consistent with a benign or neutral effect.
REVEL score: 0.08 (benign-leaning)BayesDel score: -0.37 (benign-leaning)SpliceAI max delta: 0.00 (no predicted splicing impact)
Assessed · not applied
Pathogenic
PS2
No de novo observation with confirmed maternity and paternity has been identified for this variant.
PS3
No well-established functional studies demonstrating a damaging effect for the exact variant NM_000435.2:c.4688C>T (p.Pro1563Leu) were identified.
PS4
No case-control or cohort data demonstrating statistically significant enrichment of this variant in affected individuals versus controls were identified.
PM1
Pro1563 resides in EGF-like repeat 30 within the extracellular domain of NOTCH3, a region known to harbor CADASIL-causing mutations.
PM6
No de novo observation of this variant without confirmation of maternity and paternity was identified.
PP1
No family cosegregation data were identified for this variant.
PP2
NOTCH3 is not a gene with a low rate of benign missense variation.
PP3
Multiple lines of computational evidence do not support a deleterious effect.
PP4
No patient phenotype or family history data are available for this case.
PP5
No reputable source (ClinVar, published literature) reports this variant as pathogenic with accessible evidence.
Benign
BA1
The variant has an allele frequency of 6.24e-7 in gnomAD v4.1 (1/1,602,340 alleles), far below the BA1 threshold of >1% (or >5% in some frameworks).
BS1
The variant allele frequency (6.24e-7 in gnomAD v4.1; absent in v2.1) is far below the 0.3% threshold for BS1.
BS2
A single allele is observed in gnomAD v4.1, but CADASIL has adult-onset with variable expressivity and the phenotype of the gnomAD individual is unknown.
BS3
No well-established functional studies demonstrating no damaging effect for the exact variant NM_000435.2:c.4688C>T were identified.
BS4
No family segregation data (either positive or negative) were identified for this variant.
BP1
CADASIL is primarily caused by missense variants in NOTCH3, not exclusively by truncating variants.
BP2
No data on co-occurrence of this variant in trans with a known pathogenic NOTCH3 variant were identified.
BP5
No data are available on whether this variant has been observed in a case where an alternate molecular basis for disease has been identified.
BP6
No reputable source reports this variant as benign.
N/A · 4
PVS1 · PS1 · PM5 · BP7
Research & evidence
Population frequency
gnomAD v4.1
gnomAD v2.1
v4.1
This variant is present in gnomAD v4.1 (AF= 6.24087e-07; MAF= 0.00006%, 1/1602340 alleles, homozygotes = 0) and has highest observed frequency in the African/African American population (AF= 1.33751e-05; MAF= 0.00134%, 1/74766 alleles, homozygotes = 0).
v2.1
This variant is present in gnomAD v2.1 (AF= 0; MAF= 0.00000%, 0/242354 alleles, homozygotes = 0) and has highest observed frequency in the African/African American population (AF= 0; MAF= 0.00000%, 0/15004 alleles, homozygotes = 0).
🇨🇦 CA
This variant is absent from gnomAD-Canada.
Allele frequency by ancestry
three datasets · side by side
gnomAD v4.1
6.2e-05%
· 1 / 1,602,340
0 hom
0 hom
African/African American 1 / 74,766 |
0.0013% |
+ 9 not observed (Remaining individuals, Admixed American, European (Finnish), Amish, East Asian, Middle Eastern, South Asian, Ashkenazi Jewish, European (non-Finnish))
gnomAD v2.1
Absent
· 0 / 242,354
0 hom
0 hom
Not observed in any ancestry group.
+ 8 not observed (African/African American, Admixed American, Ashkenazi Jewish, East Asian, European (Finnish), European (non-Finnish), Remaining individuals, South Asian)
gnomAD Canada 🇨🇦
Absent
· 0 / ?
0 hom
0 hom
Not observed in any ancestry group.
In silico
SpliceAI predicts no significant splice impact for this variant (max delta score = 0.00). REVEL score = 0.08. BayesDel score = -0.370843.
Functional
Unknown Oncogenic Effect
OncoKB did not identify variant-specific reviewed functional evidence for this variant; gene-level curated context is available for reviewer follow-up. NOTCH3 encodes a Type I transmembrane protein of the Notch family. Missense and nonsense mutations in NOTCH3 have been identified in various cancers.
COSMIC
Cancer hotspots
Somatic evidence
Not in COSMIC / hotspots
COSMIC
This variant does not lie in a statistically significant hotspot. This variant has not previously been reported in somatic cancers (COSMIC).
Hotspots
This variant does not lie in a statistically significant hotspot.
Sources & reference links