POLD1 c.653G>A, p.(Arg218His)

Reasoning

The Ashkenazi Jewish frequency is 0.5135% in gnomAD v4.1 (149/29,016 alleles), exceeding the applicable generic non-VCEP BS1 operating threshold of 0.3%. The elevation is replicated in gnomAD v2.1 (0.4238%) and gnomAD-Canada (0.4808%), so it is not a single-dataset artifact.

Rule

Generic non-VCEP BS1: apply strong benign evidence when an appropriate population allele frequency exceeds 0.3%.

Evidence considered

• gnomAD v4.1 reports Ashkenazi Jewish AF 0.51351% (149/29,016 alleles), with an AF of 0.52157% in Ashkenazi Jewish XX individuals (74/14,188 alleles).
• The Ashkenazi Jewish frequency is concordant across independent datasets: gnomAD v2.1 0.42382% (41/9,674 alleles) and gnomAD-Canada 0.48077% (4/832 alleles).
• The generic non-VCEP BS1 operating threshold is AF greater than 0.3%; the v4.1 Ashkenazi Jewish AF exceeds it.

Still missing

• No POLD1 VCEP specification or gene-specific maximum credible allele-frequency calculation was provided; this application uses the generic non-VCEP threshold.

Sources

Reasoning

No ClinGen VCEP CSpec/lookup exists for POLD1; generic ACMG/AMP with ClinGen SVI-calibrated computational thresholds applies. As this is a missense variant, the missense in-silico sub-path (REVEL) is the governing mechanism for BP4, not the splicing sub-path. REVEL score = 0.19 is below the ClinGen SVI-calibrated BP4 supporting threshold (<=0.290) from Pejaver et al. 2022 (PMID 36413997), supporting a benign computational prediction at the supporting strength (score is above the 0.183 moderate-benign cutoff, so only supporting strength applies, not moderate). SpliceAI max delta = 0.003 is consistent with (does not contradict) this benign missense-path conclusion but is not separately counted, since splicing is not the primary plausible mechanism for a missense substitution and double-counting across sub-paths for the same criterion is avoided. BayesDel score (-0.292993) exists but has no verified published calibration threshold available to this pipeline and was not used.

Rule

generic_acmg_BP4_missense_REVEL_calibrated

Evidence considered

• REVEL score = 0.19 (local REVEL v1.3 lookup) falls below the ClinGen SVI-calibrated BP4 supporting threshold of <=0.290 established in Pejaver et al. 2022, PMID 36413997 (Table 2, REVEL supporting/moderate/strong benign thresholds); score is above the 0.183 moderate-benign cutoff, so strength assigned is supporting only.
• SpliceAI predicts no significant splice impact for this variant (max delta score = 0.003), consistent with but not independently counted toward BP4 since the missense REVEL sub-path is the governing mechanism for this substitution.
• BayesDel score -0.292993 is present in prefetch data but no verified published calibration threshold for BayesDel is available to this pipeline, so it was not used as computational evidence for BP4.
• No gene-specific ClinGen VCEP CSpec or PP3/BP4 lookup spreadsheet exists for POLD1 in this case's vcep_materials.json, so generic ACMG/AMP calibrated thresholds were applied.

Sources

Reasoning

PM2 requires absence or extreme rarity in population databases. This variant is observed at 0.5135% in the Ashkenazi Jewish gnomAD v4.1 population and exceeds the generic non-VCEP BS1 threshold; it is not sufficiently rare for PM2.

Rule

Generic ACMG/AMP PM2: absent or extremely rare in population databases; not applicable when the observed population frequency exceeds the benign-frequency threshold.

Evidence considered

• gnomAD v4.1: Ashkenazi Jewish AF 0.51351% (149/29,016 alleles); global AF 0.04872% (782/1,604,992 alleles).
• gnomAD v2.1: Ashkenazi Jewish AF 0.42382% (41/9,674 alleles).
• gnomAD-Canada: Ashkenazi Jewish AF 0.48077% (4/832 alleles).

Sources

Reasoning

No ClinGen VCEP CSpec found for POLD1 (case_summary.cspec.found = false; vcep_materials.json has no gene-specific PP3/BP4 lookup content); generic ACMG/AMP with ClinGen SVI-calibrated computational thresholds applies. This is a missense variant (NM_002691.4:c.653G>A, NP_002682.2:p.(Arg218His)), so the missense in-silico sub-path (REVEL) governs rather than the splicing sub-path. REVEL score = 0.19, well below the ClinGen SVI-calibrated PP3 supporting threshold (>=0.644) from Pejaver et al. 2022 (PMID 36413997). SpliceAI max delta score = 0.003 (DS_AG 0.003, DS_AL 0.002, DS_DG 0.0, DS_DL 0.0), far below any splice-impact threshold, so no splicing-based pathogenic signal exists either. BayesDel score (-0.292993) is available but this pipeline has no verified published calibration threshold for BayesDel, so it is treated as not_available per current calibration policy and is not used to support PP3. No calibrated computational tool result supports a pathogenic in-silico prediction; PP3 is not met.

Rule

generic_acmg_PP3_missense_REVEL_calibrated

Evidence considered

• REVEL score = 0.19 (local REVEL v1.3 lookup), below the ClinGen SVI-calibrated PP3 supporting threshold of >=0.644 established in Pejaver et al. 2022, PMID 36413997 (Table 2, REVEL supporting/moderate/strong pathogenic thresholds).
• SpliceAI predicts no significant splice impact for this variant (max delta score = 0.003; DS_AG=0.003, DS_AL=0.002, DS_DG=0.0, DS_DL=0.0), ruling out a splicing-based pathogenic mechanism as well.
• BayesDel score -0.292993 is present in prefetch data but no verified published calibration threshold for BayesDel is available to this pipeline, so it was not used as computational evidence for PP3 (treated as not_available).
• No gene-specific ClinGen VCEP CSpec or PP3/BP4 lookup spreadsheet exists for POLD1 in this case's vcep_materials.json, so generic ACMG/AMP calibrated thresholds were applied.

Sources

Reasoning

The exact ClinVar record has no expert-panel submissions and no Pathogenic/Likely pathogenic expert-panel classification. Therefore PP5 is not met.

Rule

Apply PP5 only to an exact-variant ClinVar expert-panel Pathogenic or Likely pathogenic assertion; ordinary laboratory submissions and aggregate labels do not qualify.

Evidence considered

• ClinVar variation 239365 is an exact match to NM_002691.4:c.653G>A (p.Arg218His); its expert_panels field is empty and the submission audit reports zero expert-panel submissions.

Still missing

• An exact-variant ClinVar expert-panel Pathogenic or Likely pathogenic assertion.

Sources

Reasoning

The highest observed population AF is 0.5216% in Ashkenazi Jewish XX individuals in gnomAD v4.1 (74/14,188 alleles); this is well below the generic ACMG/AMP BA1 stand-alone frequency threshold of 5%.

Rule

Generic ACMG/AMP BA1: stand-alone benign evidence requires an allele frequency greater than 5% in an appropriate population dataset.

Evidence considered

• gnomAD v4.1: 782/1,604,992 alleles overall (AF 0.04872%), with the highest population frequency in Ashkenazi Jewish individuals: 149/29,016 alleles (AF 0.51351%) and one homozygote; Ashkenazi Jewish XX individuals have 74/14,188 alleles (AF 0.52157%).
• gnomAD v2.1 independently shows Ashkenazi Jewish AF 0.42382% (41/9,674 alleles).
• gnomAD-Canada v1.0 shows Ashkenazi Jewish AF 0.48077% (4/832 alleles).

Sources

Reasoning

The exact ClinVar record has no expert-panel submissions and no Benign/Likely benign expert-panel classification. Therefore BP6 is not met.

Rule

Apply BP6 only to an exact-variant ClinVar expert-panel Benign or Likely benign assertion; ordinary laboratory submissions and aggregate labels do not qualify.

Evidence considered

• ClinVar variation 239365 is an exact match to NM_002691.4:c.653G>A (p.Arg218His); its expert_panels field is empty and the submission audit reports zero expert-panel submissions. Benign and Likely benign assertions are ordinary laboratory submissions.

Still missing

• An exact-variant ClinVar expert-panel Benign or Likely benign assertion.

Sources

Reasoning

PS1 could not be assessed because its agent did not produce valid output.

Still missing

• attempt 1: evidence_refs must contain only source_registry keys
• attempt 2: evidence_refs must contain only source_registry keys
• attempt 3: evidence_refs must contain only source_registry keys

Reasoning

Not assessed: no proband phenotype, parental genotypes, parentage confirmation, or de novo testing result is present in the case-evidence bundle.

Rule

Generic ACMG/AMP 2015 PS2: de novo occurrence with maternity and paternity confirmed in an affected patient; no applicable POLD1-specific specification was retrieved.

Evidence considered

• No POLD1-specific ClinGen VCEP specification or local gene-specific framework was supplied; generic ACMG/AMP guidance is therefore the governing framework.
• Generic ACMG/AMP PS2 requires a de novo variant with maternity and paternity confirmed in a patient with the disease and no family history; none of these case-level data are available.

Still missing

• Obtain the proband phenotype and indication for testing.
• Obtain parental test results for NM_002691.4:c.653G>A (p.Arg218His), including confirmation of maternity and paternity.
• Confirm that the proband phenotype is consistent with the relevant POLD1-associated condition and document family history.

Sources

Reasoning

No ClinGen VCEP specification is available for POLD1 (cspec.found=false; framework_mode=generic_acmg), so generic ACMG/AMP applies. No functional/biochemical assay data (e.g., polymerase activity, exonuclease/proofreading fidelity, in vitro or cell-based complementation assays) for NM_002691.4:c.653G>A (p.Arg218His) was found anywhere in the case bundle. OncoKB returned only gene-level, non-variant-specific commentary with no reviewed functional evidence for this variant. The literature set retrieved via ClinVar citation trails consists exclusively of ACMG/AMP methodology papers (PMID:25741868, PMID:28492532) and clinical practice/case-ascertainment guidelines (PMID:24493721, PMID:25394175, PMID:33516529, PMID:24905773, PMID:24929052) with no functional assay content; none report a validated assay of this variant's effect on protein function.

Still missing

• No validated functional assay (polymerase activity, exonuclease/proofreading, replication fidelity, or cell-based) for p.Arg218His was identified in the available evidence bundle.
• No gene-specific (VCEP) functional assay framework is available for POLD1 to define approved assay types or strength calibration.
• A targeted literature search beyond the ClinVar-cited PMID set (e.g., PubMed search for POLD1 proofreading/exonuclease domain functional studies) was not performed as part of this evidence bundle and could reveal assay data not captured here.

Reasoning

No case-control or other statistically valid enrichment data for the exact POLD1 c.653G>A (p.Arg218His) variant were provided. ClinVar condition labels and laboratory assertions do not establish case enrichment.

Rule

Generic ACMG/AMP fallback: PS4 requires a statistically increased prevalence of the exact variant in affected individuals compared with controls.

Evidence considered

• The exact ClinVar record contains conflicting single-laboratory assertions and condition labels, but no expert-panel assertion and no case-control counts or enrichment analysis for c.653G>A (p.Arg218His).

Still missing

• A phenotype-defined case series or case-control dataset reporting exact-variant counts in affected individuals and appropriately matched controls, with an analysable enrichment statistic.

Sources

Reasoning

PM1 could not be assessed because its agent did not produce valid output.

Still missing

• attempt 1: evidence_refs must contain only source_registry keys
• attempt 2: evidence_refs must contain only source_registry keys
• attempt 3: evidence_refs must contain only source_registry keys

Reasoning

No applicable POLD1 VCEP specification was retrieved; generic ACMG/AMP criteria apply. The evidence bundle contains no affected individual with NM_002691.4(POLD1):c.653G>A (p.Arg218His) and a pathogenic/likely pathogenic POLD1 variant, no confirmed trans phase, and no recessive POLD1 disease-context observation. PM3 is therefore not assessable.

Rule

generic ACMG/AMP PM3: in-trans observation with a pathogenic variant in a recessive disorder

Evidence considered

• No POLD1 CSPEC or other gene-specific VCEP framework was retrieved; the case framework designates generic ACMG/AMP 2015 fallback.
• The consolidated case evidence provides no affected-proband co-occurrence, companion pathogenic/likely pathogenic POLD1 variant, or phase result for p.Arg218His.
• Richards et al. state that, in a recessive disorder, a variant in trans with a known pathogenic variant can support PM3; this required observation is absent here.

Still missing

• Affected-proband observations of p.Arg218His with a pathogenic/likely pathogenic POLD1 variant in a recessive POLD1 disorder.
• Parental or read-based phase establishing that p.Arg218His is in trans with the pathogenic/likely pathogenic POLD1 variant.

Sources

Reasoning

PM5 could not be assessed because its agent did not produce valid output.

Still missing

• attempt 1: evidence_refs must contain only source_registry keys
• attempt 2: evidence_refs must contain only source_registry keys
• attempt 3: evidence_refs must contain only source_registry keys

Reasoning

Not assessed: the bundle contains no reported de novo observation, parental testing result, or parentage information for the proband.

Rule

Generic ACMG/AMP 2015 PM6: assumed de novo occurrence without confirmation of maternity and paternity; no applicable POLD1-specific specification was retrieved.

Evidence considered

• No POLD1-specific ClinGen VCEP specification or local gene-specific framework was supplied; generic ACMG/AMP guidance is therefore the governing framework.
• Generic ACMG/AMP PM6 applies to an assumed de novo observation when maternity and paternity are not confirmed; the prerequisite observation that the variant is absent from the parents is not available.

Still missing

• Obtain parental test results for NM_002691.4:c.653G>A (p.Arg218His).
• If the variant is absent from both parents, document whether maternity and paternity were confirmed.
• Obtain the proband phenotype and relevant family history to assess phenotype consistency.

Sources

Reasoning

Not assessed: no pedigree, affected-relative genotypes, phenotypes, or informative meioses are provided for evaluating co-segregation.

Rule

Generic ACMG/AMP 2015 PP1: co-segregation with disease in multiple affected family members; no applicable POLD1-specific specification was retrieved.

Evidence considered

• No POLD1-specific ClinGen VCEP specification or local gene-specific framework was supplied; generic ACMG/AMP guidance is therefore the governing framework.
• Generic ACMG/AMP PP1 requires co-segregation with disease in multiple affected family members for a gene definitively known to cause the disease; no family segregation data are available.

Still missing

• Obtain a pedigree with phenotypes and ages of onset for informative relatives.
• Obtain variant genotypes for affected and informative unaffected relatives.
• Document the number of informative meioses and confirm phenotype consistency in genotyped affected relatives.

Sources

Reasoning

PP2 could not be assessed because its agent did not produce valid output.

Still missing

• attempt 1: evidence_refs must contain only source_registry keys
• attempt 2: evidence_refs must contain only source_registry keys
• attempt 3: evidence_refs must contain only source_registry keys

Reasoning

No proband phenotype, tumour phenotype, family history, or phenotype-specific testing context was provided for this case. The listed ClinVar conditions are submission labels rather than phenotype evidence for an individual carrying the exact variant.

Rule

Generic ACMG/AMP fallback: PP4 requires a patient phenotype or family history highly specific for a disease with a single genetic etiology.

Evidence considered

• ClinVar lists colorectal cancer susceptibility, endometrial carcinoma, hereditary cancer-predisposing syndrome, and POLD1-related disorder among record-level condition labels, but supplies no patient-level phenotype supporting PP4.

Still missing

• Phenotype and testing context for the individual carrying c.653G>A (p.Arg218His), with evidence that the presentation is highly specific for a POLD1-related disorder and that relevant alternative causes have been evaluated.

Sources

Reasoning

gnomAD v4.1 reports two homozygotes overall, including one Ashkenazi Jewish homozygote, but the supplied population records do not provide individual phenotype, age, cancer status, or an established disease-specific penetrance framework. Homozygosity in a general population dataset alone is therefore insufficient for BS2 in the context of an adult-onset, variably penetrant cancer-predisposition phenotype.

Rule

Generic ACMG/AMP BS2 requires observation in healthy adult individual(s) for a disorder expected to be fully penetrant at an early age; the available aggregate population data do not establish those conditions.

Evidence considered

• gnomAD v4.1 reports 2 homozygotes among 1,604,992 alleles overall; the Ashkenazi Jewish subset contains 1 homozygote among 29,016 alleles.

Still missing

• Phenotypic and age data for the reported homozygotes.
• A POLD1 disease-specific penetrance and BS2 specification, if available.

Sources

Reasoning

No ClinGen VCEP specification is available for POLD1 (cspec.found=false; framework_mode=generic_acmg), so generic ACMG/AMP applies. No functional assay data demonstrating normal/wild-type-equivalent activity for NM_002691.4:c.653G>A (p.Arg218His) was found in the case bundle. OncoKB provides only gene-level, non-variant-specific context (no reviewed functional evidence, classification 'Unknown Oncogenic Effect', is_variant_specific=false). None of the literature retrieved via ClinVar citation trails contain functional assay results for this variant.

Still missing

• No functional assay data (of any kind, showing normal or abnormal function) for p.Arg218His was identified in the available evidence bundle.
• No gene-specific (VCEP) functional assay framework is available for POLD1 to define approved assay types or strength calibration for benign evidence.
• A targeted literature search beyond the ClinVar-cited PMID set was not performed as part of this evidence bundle and could reveal assay data not captured here.

Reasoning

Not assessed: no family phenotype-genotype data or documented non-segregation observation is present.

Rule

Generic ACMG/AMP 2015 BS4: lack of segregation in affected family members; no applicable POLD1-specific specification was retrieved.

Evidence considered

• No POLD1-specific ClinGen VCEP specification or local gene-specific framework was supplied; generic ACMG/AMP guidance is therefore the governing framework.
• Generic ACMG/AMP BS4 requires lack of segregation in affected family members; no informative family data are available.

Still missing

• Obtain a pedigree and phenotype information for relatives carrying and not carrying the variant.
• Obtain genotypes for affected relatives to determine whether the variant fails to segregate with disease.
• Assess phenocopies, age-related penetrance, and the relevant POLD1-associated phenotype before interpreting any non-segregation.

Sources

Reasoning

BP1 could not be assessed because its agent did not produce valid output.

Still missing

• attempt 1: evidence_refs must contain only source_registry keys
• attempt 2: evidence_refs must contain only source_registry keys
• attempt 3: evidence_refs must contain only source_registry keys

Reasoning

No applicable POLD1 VCEP specification was retrieved; generic ACMG/AMP criteria apply. The evidence bundle contains no observation of p.Arg218His in cis with a pathogenic/likely pathogenic POLD1 variant in a recessive POLD1 disorder and no observation of p.Arg218His in trans with a pathogenic/likely pathogenic POLD1 variant in a dominant POLD1 disorder. BP2 is therefore not assessable.

Rule

generic ACMG/AMP BP2: cis/trans co-occurrence with a pathogenic variant interpreted in the relevant inheritance model

Evidence considered

• No POLD1 CSPEC or other gene-specific VCEP framework was retrieved; the case framework designates generic ACMG/AMP 2015 fallback.
• The consolidated case evidence provides no co-occurring pathogenic/likely pathogenic POLD1 variant and no cis/trans phase result for p.Arg218His.
• Richards et al. describe BP2 from a cis finding in a recessive disorder or a trans finding in a dominant disorder with a pathogenic variant; neither required observation is available.

Still missing

• A documented co-occurring pathogenic/likely pathogenic POLD1 variant in the same individual.
• Phase evidence and disease-model context showing either cis configuration for a recessive disorder or trans configuration for a dominant disorder.

Sources

Reasoning

No individual-level evidence was provided showing that a carrier's phenotype is fully explained by an alternative molecular diagnosis. Record-level ClinVar condition labels and laboratory classifications cannot establish BP5.

Rule

Generic ACMG/AMP fallback: BP5 requires a variant in a gene for a disease with a primarily single genetic etiology in a case with an alternate molecular basis for disease.

Evidence considered

• The ClinVar record contains condition labels and conflicting single-laboratory submissions but no individual-level alternative molecular diagnosis or phenotype attribution.

Still missing

• Individual-level phenotype and molecular testing results documenting an alternative diagnosis that fully explains the observed presentation.

Sources