NM_006231.4:c.2561+21G>A is an intronic variant in POLE located 21 bases into intron 22. SpliceAI predicts no splicing impact (max delta score = 0.00), consistent with a functionally silent intronic change.1 The variant is rare in population databases (gnomAD v2.1 AF = 0.0085%, 24/282,148 alleles; v4.1 AF = 0.0032%, 51/1,607,222 alleles), but rarity is expected for functionally silent intronic variants and is not informative for pathogenicity.2 The variant is absent from ClinVar; no germline clinical classification is available.3 It has been observed once as a somatic event in COSMIC (COSV114455825), which does not inform germline pathogenicity. The León-Castillo 2020 custom POLE framework applies exclusively to missense variants; this intronic variant falls outside its scope.4 No functional, segregation, de novo, case-control, or phenotypic data exist for this variant. No ACMG/AMP pathogenic or benign criteria are met; the variant is classified as a Variant of Uncertain Significance (VUS).
POLE
Final classification
VUS
POLE c.2561+21G>A · p.?
POLE
NM_006231.4:c.2561+21G>A is an intronic variant in POLE located 21 bases into intron 22.
No ACMG/AMP pathogenic or benign criteria are met for NM_006231.4:c.2561+21G>A. All evaluated criteria returned 'not_met' or 'not_applicable'. The León-Castillo 2020 custom POLE framework applies exclusively to missense variants, and this intronic variant falls outside its scope. Per ACMG/AMP 2015 default, when no criteria of any strength are satisfied the variant is classified as a Variant of Uncertain Significance (VUS).
Classification rationale
VUS
POLE c.2561+21G>A
Gene diagram
· NM_006231.4 · variants mapped to exon structure
POLE
NM_006231.4
Fetching transcript structure from UCSC…
Applied criteria · 0 applied · 20 assessed
Applied · 0
No criteria were applied for this variant.
Assessed · not applied
Pathogenic
PS1
No previously classified pathogenic variant at nucleotide position c.2561+21 is reported in ClinVar or the literature.
PS2
No de novo observation data are available for this variant.
PS3
No functional studies have characterized this intronic variant.
PS4
The León-Castillo 2020 custom POLE framework restricts PS4 to exact missense variants recurrent in COSMIC and TCGA endometrial carcinoma cohorts with combined EC count ≥10.
PM1
The León-Castillo 2020 custom POLE PM1 framework applies only to exact exonuclease-domain missense substitutions.
PM2
Although the variant is rare in population databases (gnomAD v2.1 AF = 0.0085%, 24/282,148 alleles; v4.1 AF = 0.0032%, 51/1,607,222 alleles), rarity of a deep intronic variant with no predicted functional consequence (SpliceAI delta = 0.00) is expected of most intronic polymorphisms and is not informative for pathogenicity.
PM6
No de novo observation data are available for this variant.
PP1
No segregation data are available for this variant.
PP3
SpliceAI predicts no splicing impact (max delta = 0.00).
PP4
No proband phenotype or clinical specificity data are available to assess whether the variant is in an individual with a phenotype specific for POLE-related disease.
PP5
This variant is absent from ClinVar; no reputable source has classified it as pathogenic.
Benign
BA1
Global allele frequency in gnomAD v2.1 is 0.0085% (24/282,148 alleles), far below the 1% BA1 threshold.
BS1
Global allele frequency in gnomAD v2.1 is 0.0085%, far below the 0.3% BS1 threshold.
BS2
No confirmed observation in a healthy adult individual with full penetrance POLE-related disorder phenotype data is available.
BS3
No functional studies demonstrating a neutral or benign effect exist for this variant.
BS4
No segregation data are available to assess lack of cosegregation with disease.
BP2
No observation of this variant in trans with a known pathogenic POLE variant has been reported.
BP4
SpliceAI predicts no splicing impact (max delta = 0.00), but BP4 requires multiple lines of computational evidence suggesting no impact.
BP5
No evidence of an alternate molecular basis for disease has been identified in this case.
BP6
This variant is absent from ClinVar; no reputable source has classified it as benign.
N/A · 8
PVS1 · PM3 · PM4 · PM5 · PP2 · BP1 · BP3 · BP7
Research & evidence
Population frequency
gnomAD v4.1
gnomAD v2.1
v4.1
This variant is present in gnomAD v4.1 (AF= 3.17318e-05; MAF= 0.00317%, 51/1607222 alleles, homozygotes = 0) and has highest observed frequency in the Admixed American population (AF= 0.000216775; MAF= 0.02168%, 13/59970 alleles, homozygotes = 0); grpmax FAF= 0.00012751.
v2.1
This variant is present in gnomAD v2.1 (AF= 8.50617e-05; MAF= 0.00851%, 24/282148 alleles, homozygotes = 0) and has highest observed frequency in the Remaining individuals population (AF= 0.000415973; MAF= 0.04160%, 3/7212 alleles, homozygotes = 0); grpmax FAF= 9.376e-05.
🇨🇦 CA
This variant is present in gnomAD-Canada v1.0 (AF= 0.00016299032924046506, 3/18406 alleles, homozygotes = 0).
Allele frequency by ancestry
three datasets · side by side
gnomAD v4.1
0.0032%
· 51 / 1,607,222
0 hom · FAF 0.013%
0 hom · FAF 0.013%
Admixed American 13 / 59,970 |
0.022% |
African/African American 9 / 74,758 |
0.012% |
Remaining individuals 6 / 62,266 |
0.0096% |
South Asian 7 / 90,954 |
0.0077% |
European (Finnish) 2 / 63,388 |
0.0032% |
European (non-Finnish) 14 / 1,174,468 |
0.0012% |
+ 4 not observed (Amish, East Asian, Middle Eastern, Ashkenazi Jewish)
gnomAD v2.1
0.0085%
· 24 / 282,148
0 hom · FAF 0.0094%
0 hom · FAF 0.0094%
Remaining individuals 3 / 7,212 |
0.042% |
African/African American 6 / 24,956 |
0.024% |
Admixed American 6 / 35,424 |
0.017% |
South Asian 5 / 30,614 |
0.016% |
East Asian 1 / 19,952 |
0.005% |
European (non-Finnish) 3 / 128,966 |
0.0023% |
+ 2 not observed (Ashkenazi Jewish, European (Finnish))
gnomAD Canada 🇨🇦
0.016%
· 3 / 18,406
0 hom · FAF 0.042%
0 hom · FAF 0.042%
Latino/Admixed American 2 / 838 |
0.24% |
Remaining individuals 1 / 1,136 |
0.088% |
+ 7 not observed (African/African American, Ashkenazi Jewish, East Asian, European (Finnish), Middle Eastern, European (non-Finnish), South Asian)
In silico
SpliceAI predicts no significant splice impact for this variant (max delta score = 0.00).
Functional
No data
No calibrated functional assay or RNA evidence was identified for this variant.
COSMIC
Somatic evidence
COSMIC
This variant has previously been reported in somatic cancers (COSMIC; COSV114455825, n = 1 times).
Hotspots
This variant does not lie in a statistically significant cancer hotspot.
Sources & reference links