APC c.3927_3931del, p.Glu1309AspfsTer4

Reasoning

NM_000038.6:c.3927_3931del (c.3927_3931delAAAGA; NP_000029.2:p.(Glu1309AspfsTer4), p.(E1309Dfs*4)) is a frameshift with the frameshift start at codon 1309. The APC VCEP states that LOF is the predominant disease mechanism and its modified PVS1 decision tree assigns PVS1, without downgrade, to frameshift or premature-truncation variants between and including codons 49 and 2645. Codon 1309 lies within that interval, so PVS1 is met at very strong strength. The VCEP notes that NMD is not essential for pathogenicity in APC; no specific downgrade rule applies to this variant.

VCEP rule

APC is a gene where LOF is the predominant mechanism of disease. ... Variants causing a frameshift or premature truncation codon between and including codon 49 and 2645

Evidence considered

• APC VCEP supplementary material, PVS1 row: APC is a gene where LOF is the predominant mechanism of disease; the document identifies nonsense and frameshift variants as null variants.
• APC VCEP Figure 1(A), PVS1 decision-tree row: frameshift or premature-truncation codons between and including codons 49 and 2645 are assigned PVS1.
• Case normalization: NM_000038.6:c.3927_3931del produces NP_000029.2:p.(Glu1309AspfsTer4) / p.(E1309Dfs*4), with frameshift start at codon 1309.
• Castellsagué et al. 2010 NMD discussion in the APC VCEP supplementary material: truncated alleles show no NMD for the last exon and only partial NMD for other exons; the VCEP states NMD is not essential for APC pathogenicity.
• The exact variant is reported as the APC 5-bp deletion c.3927_3931delAAAGA at codon 1309 in 17/58 suspected de novo patients and 59/917 FAP patients in PMID:14523376.

Sources

Reasoning

The APC VCEP PS4 rule assigns Very Strong strength at "≥ 16 phenotype points". PMID:14523376 reports the exact APC c.3927_3931delAAAGA / p.(E1309Dfs*4) deletion in 59 of 917 unrelated FAP patients, including 17/58 suspected de novo cases, 17/38 proven de novo cases, and 26/505 familial cases. Under Table 1, a documented FAP diagnosis is worth 0.5 phenotype point per proband; treating the 59 reported FAP patients as documented FAP cases gives 29.5 phenotype points, and 29.5 ≥ 16? yes. As an additional case-control comparison, the affected frequency was 59/917 = 0.0643402 and the gnomAD v4 control frequency was 12/1,614,038 = 0.00000743477; the calculated carrier-count odds ratio was 9,248.98 with 95% CI 4,954.19–17,266.93. The exact variant was not found as a pre-assigned row in Table-1%262.pdf.txt, so human review is retained for the per-proband point mapping; the result relies on the paper's explicit FAP cohort description and the VCEP Table 1 documented-FAP category, not on a generic PS4 threshold.

VCEP rule

≥ 16 phenotype points. For phenotype points curation see **Table 1**. ... Other colorectal phenotype: ... OR a documented diagnosis of FAP / AFAP

Evidence considered

• PMID:14523376, Table 1 and Results: the exact APC 5-bp deletion at codon 1309 was observed in 59/917 unrelated FAP patients overall, including 17/58 suspected de novo cases (29%), 17/38 proven de novo cases (45%), and 26/505 familial cases (5%).
• Table-1%262.pdf.txt, Table 1: under Other colorectal phenotype, a documented diagnosis of FAP / AFAP is assigned 0.5 phenotype point per proband; 59 FAP probands therefore contribute 29.5 points.
• APC_specifications_supplementary-material_V2.pdf.txt, PS4 rule as represented in cspec: ≥16 phenotype points is Very Strong; explicit inequality: 29.5 ≥ 16? yes.
• gnomAD v4.1: 12/1,614,038 control alleles (AF 7.43477e-06). Comparing 59/917 affected carriers with 12/1,614,038 controls gives OR 9,248.98, 95% CI 4,954.19–17,266.93; this calculation was performed from the reported counts.
• APC_specifications_supplementary-material_V2.pdf.txt and Table-1%262.pdf.txt were searched for c.3927_3931del, p.Glu1309AspfsTer4, and p.E1309Dfs*4; the exact variant was found in the supplementary file's PM2/BS1 contextual text but not as a Table 1 variant row.

Still missing

• Table-1%262.pdf.txt was searched for c.3927_3931del, p.Glu1309AspfsTer4, and p.E1309Dfs*4; no exact variant entry was found.
• The paper reports an FAP cohort but does not provide a separate phenotype-point row for each of the 59 carriers; the 0.5-point assignment uses the VCEP Table 1 category for a documented diagnosis of FAP.
• PMID:19029688 was listed as a PS4 lead in literature_pass.json, but no source_registry key, paper_extracts entry, or full-text file is available for independent variant-level verification; it was not used in the decision.

Sources

Reasoning

The APC VCEP's PM6 conversion assigns Strong evidence to 2-3.5 de novo scores. PMID:14523376 reports the exact c.3927_3931delAAAGA variant in two parental-origin Table 1 entries and reports 17/38 proven de novo codon-1309 deletions. The study identifies the patients as FAP patients, supplying at least 0.5 phenotype point per proband under the APC Table 1 category for a documented FAP diagnosis. The parental-origin table documents two exact-variant de novo observations with traced parental origin, supporting two 1-point confirmed-relationship scores and a total of 2 de novo scores; the exact family-level relationship confirmation is not fully mapped, so human review is required.

VCEP rule

2-3.5 _de novo_ scores. For curation of _de novo_ score see **Tables 1** and **2**.

Evidence considered

• PMID:14523376, Table 1: c.3927_3931delAAAGA appears twice among 16 de novo APC mutation cases with traced parental origin.
• PMID:14523376, Results: the exact codon-1309 deletion was present in 17/38 proven de novo cases.
• Table-1%262.pdf, Table 1: a documented diagnosis of FAP is a 0.5 phenotype point per proband.
• Table-1%262.pdf, Table 2: a 0.5-point proband contributes 1 de novo score with confirmed parental relationships and 0.5 with unconfirmed relationships; two confirmed entries therefore total 2 scores.

Still missing

• PMID:14523376 does not explicitly identify the family number and complete parental-relationship confirmation status for each of the two c.3927_3931delAAAGA Table 1 entries.

Sources

Reasoning

Expert panel ClinGen InSiGHT Hereditary Colorectal Cancer/Polyposis Variant Curation Expert Panel classified as Pathogenic.

VCEP rule

Not used as recommended by Biesecker et al. 2018 (PMID: 29543229): Expert opinions should only be considered if accompanied by the primary evidence used.

Evidence considered

• APC_specifications_supplementary-material_V2.pdf.txt, PP5 comment: Not used as recommended by Biesecker et al. 2018 (PMID: 29543229): Expert opinions should only be considered if accompanied by the primary evidence used.
• ClinVar expert panel classification

Sources

Reasoning

The APC VCEP specifies BS1 Strong at GnomAD Popmax Filtering Allele Frequency (AF) ≥ 0.001%. The governing non-cancer gnomAD v2.1 exome dataset has a highest observed subpopulation frequency of 1/5600 = 0.0001785714 (0.017857%), which exceeds 0.001%; the total frequency is 2/236524 = 0.0000084558 (0.00084558%). The variant meets BS1 Strong on the population-maximum frequency path.

VCEP rule

Allele frequency ≥ 0.001% (preferably based on the Popmax Filtering Allele Frequency of the non-cancer dataset from gnomAD (v2.1.1))

Evidence considered

• APC VCEP BS1 rule: GnomAD Popmax Filtering Allele Frequency (AF) ≥ 0.001% (0.00001).
• GNOMAD_V2_1_NON_CANCER, exomes: c.3927_3931delAAAGA, highest observed subpopulation was Remaining individuals, AC 1/5600, AF 0.00017857142857142857 (0.017857%), exceeding 0.001%; total AC 2/236524, AF 8.455801525426595e-06 (0.00084558%).
• The VCEP states that the non-cancer gnomAD v2.1.1 dataset is generally recommended for determining allele frequency and that subpopulations may be used considering the highest minor allele frequency.

Sources

Reasoning

The APC VCEP PM2 Supporting rule defines rarity as AF ≤0.0003% (0.000003) when allele count is >1, or AF <0.001% (0.00001) when allele count is ≤1. In the governing non-cancer gnomAD v2.1 exome dataset, the variant has allele count 2 and AF 0.00084558%, which is above the applicable 0.0003% threshold; PM2 Supporting is not met.

VCEP rule

Rare in controls, defined by an allele frequency ≤ 0.0003% (0.000003) if the allele count is > 1 OR by an allele frequency < 0.001% (0.00001) if the allele count is ≤ 1.

Evidence considered

• APC VCEP PM2 Supporting rule: AF ≤0.0003% (0.000003) if allele count is >1, or AF <0.001% (0.00001) if allele count is ≤1.
• GNOMAD_V2_1_NON_CANCER, exomes: c.3927_3931delAAAGA, allele count 2, allele number 236524, AF 8.455801525426595e-06 = 0.00084558%, homozygotes = 0.
• The APC VCEP recommends the total population from the non-cancer gnomAD v2.1.1 dataset for PM2 frequency determination.

Sources

Reasoning

The APC VCEP specifies BA1 at GnomAD Popmax Filtering Allele Frequency (AF) ≥ 0.1% (0.001). In the governing non-cancer gnomAD v2.1 exome dataset, the highest observed subpopulation frequency is 1/5600 = 0.0001785714 (0.017857%), and the total frequency is 2/236524 = 0.0000084558 (0.00084558%); both are below 0.1%. The variant is therefore not eligible for BA1.

VCEP rule

Allele frequency ≥ 0.1% (preferably based on the Popmax Filtering Allele Frequency of the non-cancer dataset from gnomAD (v2.1.1))

Evidence considered

• APC VCEP BA1 rule: GnomAD Popmax Filtering Allele Frequency (AF) ≥ 0.1% (0.001%).
• GNOMAD_V2_1_NON_CANCER, exomes: c.3927_3931delAAAGA, total AC 2/236524, AF 8.455801525426595e-06; highest observed subpopulation was Remaining individuals, AC 1/5600, AF 0.00017857142857142857 (0.017857%).
• The APC VCEP supplementary table explicitly identifies c.3927_3931delAAAGA p.(Glu1309Aspfs*4) as the common pathogenic APC variant used in its frequency calibration; this is not a pre-assigned BA1 code for the case variant.

Sources

Reasoning

The APC VCEP assigns PS2 by de novo score using Tables 1 and 2. PMID:14523376 reports 17 proven de novo c.3927_3931delAAAGA cases and includes two exact-variant entries in its parental-origin Table 1, but the available record does not map each exact-variant entry to complete maternity and paternity confirmation together with an individually documented phenotype point. Therefore PS2 cannot be assigned without assuming case-level details.

VCEP rule

≥ 4 _de novo_ scores. For curation of _de novo_ score see **Tables 1** and **2**.

Evidence considered

• PMID:14523376, Results: the five-base-pair deletion c.3927_3931delAAAGA occurred in 17/38 proven de novo cases; Table 1 lists c.3927_3931delAAAGA twice among de novo APC mutations of maternal origin.
• PMID:14523376, Methods: de novo status was based on exclusion of the mutation from both parents, while paternity testing was performed in most but not all parental-origin cases.
• Table-1%262.pdf, Tables 1 and 2: phenotype points must be assigned per proband before converting confirmed or unconfirmed de novo observations to a de novo score.

Still missing

• APC_specifications_supplementary-material_V2.pdf.txt: the PS2 text is present, but no exact-variant case-level record establishes both maternity and paternity confirmation.
• Table-1%262.pdf.txt: the phenotype-point categories and de novo-score conversion are present, but no individual phenotype point is assigned to each exact-variant proband.

Sources

Reasoning

The APC VCEP permits PS3 at Very Strong, Strong, Moderate, or Supporting strength only for specified RNA or protein assay findings. The available case evidence identifies PMID:10213492 as a functional-study lead for the APC1309 mutant and describes a beta-catenin/Tcf transcription assay in a classifier note, but the primary paper is not available in the case files or paper extracts, so assay design, controls, quantitative result, and the VCEP-required endpoint cannot be verified. The available prefetch evidence also states that functional studies have not been reported. The exact variant was searched in APC_specifications_supplementary-material_V2.pdf; variant-specific mentions were found in the PM2/BS1 population discussion, but no pre-assigned PS3 code or variant-specific PS3 assay entry was found.

Evidence considered

• ClinGen InSiGHT APC specification PS3 table: the VCEP strength rules require defined RNA assay findings or a specified protein-assay result; no available evidence verifies those endpoints for this variant.
• The literature packet identifies PMID:10213492, 'Dominant negative effect of the APC1309 mutation: a possible explanation for genotype-phenotype correlations in familial adenomatous polyposis,' as a functional-study lead and reports a beta-catenin/Tcf transcription assay, but this is an unverified classifier lead without the primary article or extract.
• Prefetch evidence states: 'although functional studies have not been reported,' so the available record does not establish a qualifying functional study.

Still missing

• Primary full text for PMID:10213492 is not present in the case publications directory and has no paper_extracts.json entry; the reported beta-catenin/Tcf assay therefore cannot be independently verified.
• No assay controls, quantitative functional result, RNA transcript measurement, or protein assay endpoint sufficient to apply a VCEP PS3 strength is available.
• Searched APC_specifications_supplementary-material_V2.pdf for c.3927_3931del, p.Glu1309AspfsTer4, and p.E1309Dfs*4; variant-specific mentions were found, but no PS3 pre-assigned code or variant-specific functional-assay row was found.

Sources

Reasoning

The APC VCEP requires segregation in affected relatives and calibrates PP1 by the number of meioses. The available exact-variant evidence reports de novo cases and parental-origin haplotypes but does not report affected relatives carrying the variant, genotype-positive phenotype-positive relatives, obligate carriers with phenotype, or a count of segregating meioses. PP1 therefore cannot be assessed.

VCEP rule

Variant segregates in 3-4 meioses in ≥ 1 family.

Evidence considered

• APC_specifications_supplementary-material_V2.pdf, PP1: only genotype- and phenotype-positive individuals and obligate carriers with phenotype are counted.
• APC_specifications_supplementary-material_V2.pdf, PP1: the governing calibration is based on segregating meioses, but no such meioses are documented for this exact variant.

Still missing

• APC_specifications_supplementary-material_V2.pdf.txt: no exact-variant family segregation data or meiosis count was located.
• Table-1%262.pdf.txt: no exact-variant affected-relative phenotype and genotype records are provided for PP1.

Sources

Reasoning

The APC VCEP BS2 Strong rule requires either ≥10 healthy-individual points or ≥2 observations in the homozygous state. The governing non-cancer gnomAD v2.1 exome record reports 0 homozygotes, so the homozygous path is not met. However, no age, colonoscopy, adenoma, indication, or phenotype data are available to determine healthy-individual points; the criterion cannot be fully assessed.

VCEP rule

≥ 10 points for healthy individuals **OR** ≥ 2 times in homozygous state. ... A **healthy individual** worth 1 point is defined by: ... Age ≥ 50 years  + Less than 5 adenomatous polyps in a colonoscopy  + Absence of features in Table 1 ... **OR** ... Age ≥ 50 years  + Colorectal cancer/polyposis was not the indication for testing ... A **healthy individual** worth 0.5 points is defined by keywords including control, non-cancer, normal, unaffected population.

Evidence considered

• APC VCEP BS2 Strong rule requires ≥10 points for healthy individuals OR ≥2 observations in the homozygous state.
• GNOMAD_V2_1_NON_CANCER, exomes: AC 2/236524 and homozygotes = 0 for c.3927_3931delAAAGA.
• The APC VCEP cautions that non-cancer gnomAD v2.1.1 cannot be used for heterozygous healthy individuals because phenotype information is limited, but can be used to search for homozygous individuals.

Still missing

• No qualifying healthy-individual records with age and phenotype/colonoscopy or testing-indication data were available to calculate BS2 points.

Sources

Reasoning

The APC VCEP BS3 rule requires either an RNA assay showing no mRNA aberration for a synonymous or intronic variant, with the specified expression or NMD control, or a protein assay showing retention of beta-catenin-regulated transcription activity comparable to wild type for variants within APC codons 959-2129. This frameshift variant affects codon 1309 and is within that domain, but no qualifying benign RNA or protein assay result is available. The absence of a qualifying benign assay is insufficient to call BS3 not_met; the criterion is not_assessed.

Evidence considered

• ClinGen InSiGHT APC specification BS3 table: qualifying evidence is a normal RNA result for a synonymous/intronic variant with the stated controls, or retention of beta-catenin-regulated transcription activity comparable to wild type in the beta-catenin binding domain.
• The variant consequence is a frameshift at codon 1309, but the case evidence contains no qualifying benign RNA or protein assay result for it.

Still missing

• No VCEP-qualifying benign functional assay, assay controls, or quantitative wild-type comparison is available for this variant.
• Searched APC_specifications_supplementary-material_V2.pdf for c.3927_3931del, p.Glu1309AspfsTer4, and p.E1309Dfs*4; variant-specific mentions were found, but no BS3 pre-assigned code or variant-specific benign functional-assay row was found.

Sources

Reasoning

The APC VCEP requires an affected member without the variant and a qualifying phenotype point from Table 1. No affected noncarrier is reported for the exact c.3927_3931del variant, and the available de novo study provides no family non-segregation observation. BS4 cannot be assessed.

VCEP rule

Affected member without the variant must score at least 0.5 phenotype points (see **Table 1**).

Evidence considered

• APC_specifications_supplementary-material_V2.pdf, BS4: the criterion requires an affected member without the variant with at least 0.5 phenotype point, or two such members each with at least 0.5 point for Strong evidence.
• Table-1%262.pdf, Table 1: phenotype points are assigned to affected individuals by APC-specific clinical features; no affected noncarrier is documented here.

Still missing

• APC_specifications_supplementary-material_V2.pdf.txt: no exact-variant affected noncarrier or non-segregation observation was located.
• Table-1%262.pdf.txt: no affected noncarrier phenotype point is available for this variant.

Sources

Reasoning

The APC VCEP BP2 rule requires the variant to be observed in trans with a (Likely) Pathogenic APC variant, or at least three times in unknown phase with different (Likely) Pathogenic APC variants. The available case evidence documents no paired APC variant observations, phase information, or qualifying co-occurrence count for NM_000038.6:c.3927_3931del. Accordingly, BP2 cannot be evaluated.

VCEP rule

Observed _in trans_ with a (Likely) Pathogenic _APC_ variant **OR** ≥ 3 times in an unknown phase with different (Likely) Pathogenic _APC_ variants.

Evidence considered

• APC VCEP CSPEC BP2 rule: observed in trans with a (Likely) Pathogenic APC variant, or at least 3 times in unknown phase with different (Likely) Pathogenic APC variants.
• Case evidence contains no documented paired APC variant, phase determination, or qualifying unknown-phase co-occurrence count for NM_000038.6:c.3927_3931del.

Still missing

• Documented co-occurrence with a different (Likely) Pathogenic APC variant in the same affected individual
• Phase determination or at least three qualifying unknown-phase observations

Sources

Reasoning

The APC VCEP BP5 rule is conditional on an alternate genetic basis of the colorectal polyposis phenotype. The case evidence documents APC/FAP observations but does not document testing results for a VCEP-listed alternate pathogenic polyposis-gene basis, including POLD1, POLE, MUTYH, NTHL1, MSH3, or the specified childhood/adolescent biallelic mismatch-repair genes. Without that result, BP5 cannot be assessed.

VCEP rule

Can be used, if a (Likely) Pathogenic variant in another adenomatous polyposis gene was identified (heterozygous variants in POLD1 or POLE; biallelic variants in MUTYH, NTHL1 or MSH3; in patients with onset in childhood / adolescence: biallelic variants in MLH1, MSH2, MSH6 or PMS2). This rule is only applicable when a colorectal polyposis phenotype is present.

Evidence considered

• vcep_materials.json cspec BP5: the criterion can be used when a (Likely) Pathogenic variant in another adenomatous polyposis gene is identified and the colorectal polyposis phenotype is present.
• The reviewed case evidence contains no documented alternate pathogenic variant result in the VCEP-listed genes.

Still missing

• Alternate pathogenic-variant testing/results for the VCEP-listed adenomatous polyposis genes are unavailable.

Sources