APC c.4413A>G, p.(Ala1471=)

Reasoning

The APC InSiGHT VCEP specifies the total gnomAD v2.1.1 non-cancer dataset for PM2 and defines PM2_Supporting as allele frequency <= 0.000003 when allele count is greater than 1 or allele frequency < 0.00001 when allele count is at most 1. The variant is absent from the specified non-cancer exome dataset, equivalent to allele count 0 and allele frequency 0, so PM2_Supporting is met.

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

• The APC InSiGHT VCEP specifies the total population from gnomAD v2.1.1 non-cancer and defines PM2_Supporting as AF <= 0.000003 for allele count > 1 or AF < 0.00001 for allele count <= 1.
• The variant is absent from the gnomAD v2.1 non-cancer exome dataset, equivalent to allele count 0 and AF 0; this satisfies the allele-count-at-most-1 branch of the VCEP PM2 rule.

Sources

Reasoning

The variant is synonymous, making the splice-impact path applicable and REVEL inapplicable. SpliceAI reports a maximum delta of 0.001, which is below the generic BP4 supporting threshold of 0.1 (Jaganathan et al. 2019, PMID:30661751), indicating no predicted splice impact. This is consistent with the APC VCEP rule that synonymous or intronic variants may receive BP4 Supporting when multiple in silico splicing predictors suggest no impact; the available SpliceAI and Pangolin outputs are both near zero.

Rule

APC InSiGHT VCEP BP4 Supporting for synonymous or intronic variants with multiple no-impact splice predictions; generic SpliceAI BP4 Supporting threshold: max delta <= 0.1 (Jaganathan et al. 2019, PMID:30661751).

Evidence considered

• The APC VCEP states that BP4 Supporting applies to synonymous or intronic variants when multiple in silico splicing predictors suggest no impact on the gene or gene product.
• SpliceAI maximum delta is 0.001, below the generic BP4 supporting cutoff of 0.1 from Jaganathan et al. 2019 (PMID:30661751).
• The available Pangolin outputs are 0.012 and -0.003, concordant with the near-zero SpliceAI result and no predicted splice impact.
• The variant is synonymous, so REVEL was not used.

Sources

Reasoning

The variant is synonymous (p.(Ala1471=)), which is within BP7 scope. The APC VCEP permits BP7 Supporting for a synonymous variant when multiple splice-prediction algorithms indicate no impact; SpliceAI has a maximum delta of 0.001 and Pangolin outputs are near zero (0.012 and -0.003), providing concordant no-impact predictions. The same prediction evidence is not separately counted toward PP3.

Rule

APC InSiGHT VCEP BP7 Supporting: synonymous variant plus multiple splice-prediction algorithms indicating no splice impact.

Evidence considered

• The APC VCEP BP7 rule covers synonymous variants when multiple splice-prediction algorithms predict no impact to the splice consensus sequence or creation of a new splice site.
• SpliceAI maximum delta is 0.001, with individual delta scores from 0.000 to 0.001; Pangolin outputs are 0.012 and -0.003, concordant with no predicted splice impact.
• BP7 is assessed independently as the synonymous-variant benign criterion and does not double-count the same prediction as PP3 or BP4.

Sources

Reasoning

The variant is synonymous (NM_001127510.3:c.4413A>G; p.(Ala1471=)), so the applicable computational path is splice prediction rather than REVEL. SpliceAI max delta is 0.001, below the APC VCEP PP3 supporting requirement for multiple predictors to support a deleterious splice effect and below the generic SpliceAI PP3 supporting threshold of 0.2 (Jaganathan et al. 2019, PMID:30661751). The available splice predictions therefore do not support PP3.

Rule

APC InSiGHT VCEP PP3 Supporting: multiple in silico splicing predictors must support a deleterious effect; generic SpliceAI PP3 Supporting threshold: max delta >= 0.2 (Jaganathan et al. 2019, PMID:30661751).

Evidence considered

• The APC VCEP specifies PP3 Supporting for non-canonical splicing variants when multiple in silico splicing predictors support a deleterious effect.
• SpliceAI reports DS_AG 0.000, DS_AL 0.000, DS_DG 0.001, and DS_DL 0.001; maximum delta is 0.001, below the generic PP3 supporting cutoff of 0.2 from the SpliceAI calibration by Jaganathan et al. 2019 (PMID:30661751).
• The variant is synonymous, so REVEL is not an applicable evidence path and was not used.

Sources

Reasoning

The APC InSiGHT VCEP specifies the gnomAD v2.1.1 non-cancer dataset for BA1 and requires a gnomAD popmax filtering allele frequency of at least 0.001 (0.1%). The variant is absent from the gnomAD v2.1 non-cancer exome dataset, so it does not meet the stand-alone benign threshold.

Rule

GnomAD Popmax Filtering Allele Frequency (AF) >= 0.1% (0.001).

Evidence considered

• The APC InSiGHT VCEP specifies gnomAD v2.1.1 non-cancer as the population source and sets BA1 at popmax AF >= 0.001 (0.1%).
• The variant is absent from the gnomAD v2.1 non-cancer exome dataset, corresponding to an observed allele frequency of 0, which is below 0.001.

Sources

Reasoning

The APC InSiGHT VCEP specifies the gnomAD v2.1.1 non-cancer dataset for BS1 and requires a gnomAD popmax filtering allele frequency of at least 0.00001 (0.001%). The variant is absent from the gnomAD v2.1 non-cancer exome dataset, so the BS1 threshold is not met.

Rule

GnomAD Popmax Filtering Allele Frequency (AF) >= 0.001% (0.00001).

Evidence considered

• The APC InSiGHT VCEP specifies gnomAD v2.1.1 non-cancer as the population source and sets BS1 at popmax AF >= 0.00001 (0.001%).
• The variant is absent from the gnomAD v2.1 non-cancer exome dataset, corresponding to an observed allele frequency of 0, which is below 0.00001.

Sources

Reasoning

The APC InSiGHT VCEP permits the gnomAD v2.1.1 non-cancer dataset for searching for homozygous individuals and requires at least two homozygous observations for strong BS2. The variant is absent from that dataset, with no homozygous observations, and no qualifying individually characterized healthy-person evidence is provided.

Rule

>= 10 points for healthy individuals OR >= 2 times in homozygous state.

Evidence considered

• The APC InSiGHT VCEP defines strong BS2 as at least 10 healthy-individual points or at least 2 homozygous observations and allows the non-cancer dataset only to search for homozygotes.
• The variant is absent from the gnomAD v2.1 non-cancer exome dataset, providing zero observed homozygotes; no separate qualifying healthy-individual observations are available.

Sources

Reasoning

The APC Version 2.1 specification requires de novo evidence with phenotype consistency and scores based on confirmed or unconfirmed parental relationships. The available case evidence contains no proband-level de novo observation, parental genotype results, confirmation of maternity or paternity, or family-history documentation for NM_001127510.3:c.4413A>G. Therefore PS2 cannot be assessed.

Rule

APC V2.1: PS2 is scored from de novo evidence using Table 1 phenotype points and Table 2 parental-relationship status; >=4 de novo scores is Very Strong, 2-3.5 Strong, and 1-1.5 Moderate.

Evidence considered

• The APC Version 2.1 rule assigns PS2 strength from de novo scores: Very Strong for at least 4, Strong for 2–3.5, and Moderate for 1–1.5 de novo scores.
• The supplied case materials provide no patient-level parental testing, maternity or paternity confirmation, or documented de novo observation from which an APC de novo score could be calculated.

Still missing

• Documented affected-proband phenotype consistent with APC-associated polyposis
• Parental genotype results for c.4413A>G
• Confirmation of maternity and paternity
• Family history relevant to de novo assessment

Sources

Reasoning

The APC InSiGHT VCEP specifies PS3 for variant-specific RNA evidence showing a premature stop or relevant exon skipping/loss with the required reduction or absence of full-length transcript, or for protein evidence showing increased beta-catenin-regulated transcription activity and/or decreased beta-catenin binding by surface plasmon resonance. The variant is synonymous, NM_001127510.3:c.4413A>G (p.(Ala1471=)), but the reviewed case evidence and variant-level literature contain no RNA or protein assay result for this variant. SpliceAI prediction is computational evidence and cannot substitute for the required functional assay. Therefore PS3 cannot be assessed.

Rule

APC VCEP PS3: RNA assays show a premature stop codon or in-frame skipping of exon 13 or 14, with strength calibrated by the amount of full-length transcript; moderate/supporting pathways also cover specified exon loss, intronic insertion, alternative transcript over-expression, or protein assays showing increased beta-catenin-regulated transcription activity and/or decreased beta-catenin binding by surface plasmon resonance for variants in the beta-catenin binding domain.

Evidence considered

• The APC InSiGHT VCEP PS3 specification requires defined RNA assay findings for transcript loss or exon skipping, or specified protein assay findings for APC functional impairment.
• The ACMG/AMP functional-study guidance states that mRNA-level assays can evaluate effects such as mRNA stability, processing, or translation, but PMID:25741868 does not report an assay result for APC c.4413A>G.
• The reviewed variant-specific evidence identifies a synonymous APC change, p.(Ala1471=), but provides no variant-specific RNA or protein functional result.

Still missing

• A validated variant-specific RNA assay in a constitutional patient sample, including transcript quantification and relevant controls, is unavailable.
• A validated variant-specific protein assay measuring APC function or beta-catenin interaction is unavailable.

Sources

Reasoning

The APC InSiGHT VCEP specifies PS4 as applicable and assigns strength from phenotype points: >= 16 points is Very Strong, 4-15.5 is Strong, 2-3.5 is Moderate, and 1-1.5 is Supporting. The available case evidence contains no proband phenotype, family-history details, phenotype-point assignments, case-control enrichment estimate, odds ratio, relative risk, or confidence interval for this exact variant. The reviewed literature extracts do not report NM_001127510.3:c.4413A>G or NP_001120982.1:p.(A1471=). Therefore PS4 cannot be assessed rather than inferred from the gene-level description of FAP.

Rule

APC VCEP PS4: >= 16 phenotype points = Very Strong; 4-15.5 = Strong; 2-3.5 = Moderate; 1-1.5 = Supporting. For phenotype points, use APC Table 1.

Evidence considered

• The APC InSiGHT VCEP source text states: ">= 16 phenotype points" for PS4 Very Strong, "4-15.5 phenotype points" for Strong, "2-3.5 phenotype points" for Moderate, and "1-1.5 phenotype point" for Supporting.
• The APC phenotype table assigns 1 point for highly specific phenotype findings and 0.5 points for phenotype consistent with APC, but no patient-level phenotype points are supplied for this exact variant.
• The exact variant is not mentioned in the reviewed extracts for PMID:11598466, PMID:24310308, PMID:25452455, or PMID:34012068; these sources therefore provide no exact-variant prevalence or case-control evidence.

Still missing

• Exact-variant proband phenotype and family-history data sufficient to assign APC Table 1 phenotype points.
• If using case-control evidence, an exact-variant odds ratio or relative risk with a confidence interval and control comparison.

Sources

Reasoning

The APC Version 2.1 specification requires an assumed de novo observation without confirmed paternity and maternity, with strength determined by de novo scores. The available case evidence contains no proband-level de novo observation, parental genotype results, or phenotype documentation for NM_001127510.3:c.4413A>G. An assumed de novo score cannot be established, so PM6 cannot be assessed.

Rule

APC V2.1: PM6 is scored from assumed de novo evidence without confirmed maternity and paternity; 2-3.5 de novo scores is Strong, 1-1.5 Moderate, and 0.5 Supporting.

Evidence considered

• The APC Version 2.1 rule assigns PM6 strength from de novo scores: Strong for 2–3.5, Moderate for 1–1.5, and Supporting for 0.5 de novo scores.
• No case-level evidence documents an assumed de novo occurrence or supplies the parental and phenotype information needed to calculate the score.

Still missing

• Documented affected-proband phenotype consistent with APC-associated polyposis
• Evidence that the variant is absent from tested parents or otherwise assumed de novo
• Parental relationship and genotype details sufficient to calculate a de novo score

Sources

Reasoning

The APC Version 2.1 specification requires genotype-positive, phenotype-positive affected relatives or obligate carriers with phenotype and counts evidence by meioses. The available case evidence contains no pedigrees, affected relatives, genotype results, phenotype scores, or meiosis count for NM_001127510.3:c.4413A>G. PP1 therefore cannot be assessed.

Rule

APC V2.1: PP1 Supporting requires 3-4 segregating meioses in at least 1 family, Moderate requires 5-6 meioses in at least 1 family, and Strong requires at least 7 meioses in at least 2 families.

Evidence considered

• The APC Version 2.1 rule requires at least 3-4 segregating meioses for Supporting, 5-6 meioses in at least one family for Moderate, or at least 7 meioses in at least 2 families for Strong.
• No affected relatives, genotype-positive relatives, phenotype-positive obligate carriers, family pedigrees, or informative meioses are reported in the supplied case evidence.

Still missing

• Pedigree and family-level clinical information
• Genotype results for affected relatives or obligate carriers
• Phenotype documentation for genotype-positive relatives
• Count of informative segregating meioses and number of families

Sources

Reasoning

The APC InSiGHT VCEP specifies BS3 Strong when an RNA assay of a synonymous or intronic variant in a constitutional patient sample shows no mRNA aberration together with demonstrated biallelic expression and/or nonsense-mediated decay inhibition. BS3 Supporting may use such an RNA result without those controls, or a protein assay showing beta-catenin-regulated transcription comparable to wild type for variants in the beta-catenin binding domain. Although this synonymous variant lies at codon 1471, no variant-specific RNA assay, allelic-expression analysis, nonsense-mediated decay experiment, or protein functional assay was found in the reviewed evidence. SpliceAI's max delta score of 0.001 is computational prediction, not functional evidence, so BS3 cannot be assessed.

Rule

APC VCEP BS3: Strong requires an RNA assay of a synonymous or intronic variant in a constitutional patient sample showing no mRNA aberration plus demonstrated biallelic expression and/or nonsense-mediated decay inhibition. Supporting allows the same RNA result without those controls, or a protein assay showing beta-catenin-regulated transcription comparable to wild type for variants within the beta-catenin binding domain.

Evidence considered

• The APC InSiGHT VCEP BS3 Strong rule requires a normal constitutional-sample RNA result plus biallelic expression or nonsense-mediated decay inhibition; its Supporting rule allows a less controlled RNA result or a specified beta-catenin functional protein assay.
• The reviewed variant-specific evidence contains no RNA assay, allelic-expression study, nonsense-mediated decay experiment, or protein assay for APC c.4413A>G.
• SpliceAI reports a maximum delta score of 0.001, which is an in-silico prediction and does not establish the functional-study evidence required for BS3.

Still missing

• A constitutional-sample RNA assay demonstrating no mRNA aberration is unavailable.
• Biallelic expression or nonsense-mediated decay inhibition data needed for BS3 Strong are unavailable.
• A variant-specific protein assay demonstrating beta-catenin-regulated transcription comparable to wild type is unavailable.

Sources

Reasoning

The APC Version 2.1 specification requires an affected member without the variant who meets a phenotype-point threshold: at least 1 point for Strong, or at least 0.5 point for Supporting. The available case evidence contains no affected non-carrier, phenotype score, family genotype data, or information excluding phenocopy or an alternate pathogenic variant. BS4 cannot be assessed.

Rule

APC V2.1: BS4 Supporting requires an affected member without the variant scoring at least 0.5 phenotype point; Strong requires one such member scoring at least 1 point or at least two each scoring at least 0.5 point.

Evidence considered

• The APC Version 2.1 rule assigns BS4 Supporting when an affected member without the variant has at least 0.5 phenotype point, and Strong when one has at least 1 point or two each have at least 0.5 point.
• No affected non-carrier, phenotype-point score, or family-level genotype information is documented, and no evidence addresses phenocopy or an alternate pathogenic variant.

Still missing

• Affected family member documented to lack c.4413A>G
• Phenotype-point assessment for the affected non-carrier
• Family genotype data sufficient to exclude phenocopy or an alternate pathogenic variant

Sources

Reasoning

The APC VCEP permits BP2 at Supporting strength when the assessed variant is observed in trans with a (Likely) Pathogenic APC variant, or at least three times with different (Likely) Pathogenic APC variants in unknown phase. The available case evidence documents NM_001127510.3:c.4413A>G (p.Ala1471=) but provides no affected-proband observations, second APC variant, parental testing, phase determination, or three qualifying unknown-phase co-occurrences. BP2 therefore cannot be assessed.

Rule

BP2 Supporting: observed in trans with a (Likely) Pathogenic APC variant, or at least 3 times in unknown phase with different (Likely) Pathogenic APC variants.

Evidence considered

• The APC VCEP BP2 rule requires observation in trans with a (Likely) Pathogenic APC variant or at least three observations in unknown phase with different (Likely) Pathogenic APC variants.
• The available case materials identify the assessed synonymous APC variant but do not provide affected-proband co-occurrence data, a second pathogenic APC variant, or phase information.

Still missing

• Case-level observations identifying a second APC variant in affected probands are unavailable.
• Parental or other phasing data establishing trans or cis configuration are unavailable.
• No evidence of at least three qualifying unknown-phase co-occurrences with different (Likely) Pathogenic APC variants is available.

Sources

Reasoning

The APC InSiGHT VCEP permits BP5 at Supporting strength only for an alternate genetic basis of a colorectal polyposis phenotype, specifically a qualifying (Likely) Pathogenic variant in another adenomatous polyposis gene, and only when a colorectal polyposis phenotype is present. The available evidence does not establish a colorectal polyposis phenotype for this case and does not identify a qualifying alternate-gene (Likely) Pathogenic variant. Therefore BP5 is not assessed; neither the absence of reported alternate testing nor a generic ClinVar label is sufficient to call it not_met.

Rule

APC VCEP BP5 Supporting: a (Likely) Pathogenic variant in another specified adenomatous polyposis gene, with the specified inheritance pattern, and a colorectal polyposis phenotype.

Evidence considered

• The APC VCEP BP5 rule states that it is only applicable for an alternate genetic basis of the colorectal polyposis phenotype.
• The APC VCEP instructions specify qualifying alternate genes and inheritance patterns: heterozygous POLD1 or POLE; biallelic MUTYH, NTHL1, or MSH3; and, for childhood/adolescent onset, biallelic MLH1, MSH2, MSH6, or PMS2.
• No qualifying alternate-gene variant or case-specific colorectal polyposis phenotype is present in the reviewed evidence.

Still missing

• Documented colorectal polyposis phenotype in the individual or family.
• A qualifying (Likely) Pathogenic variant in another adenomatous polyposis gene, with inheritance/zygosity meeting the APC VCEP rule.

Sources