POLE c.5066C>T, p.(Thr1689Ile)

Reasoning

PM2_Supporting is met because the exact POLE variant NM_006231.4:c.5066C>T (p.Thr1689Ile) is absent from gnomAD v2.1, has AC 0/AN 1,608,632 and AF 0.0 in gnomAD v4.1 with 0 homozygotes, and is absent from gnomAD-Canada v1.0. The retrieved POLE package contains no higher-priority population specification, so the generic ACMG/AMP population-absence rule is used.

Rule

Generic ACMG/AMP PM2_Supporting: apply when the variant is absent or extremely rare in population databases appropriate for the disease and ancestry context; no applicable POLE VCEP/CSPEC population rule was retrieved.

Evidence considered

• gnomAD v4.1: exact variant 12-132642284-G-A; total AC 0, total AN 1,608,632, AF 0.0, homozygotes 0; the highest-observed listed subpopulation is AFR with AC 0/AN 74,784 and AF 0.0.
• gnomAD v2.1 reports the exact variant as absent.
• gnomAD-Canada v1.0 reports the exact variant as absent.
• No relevant exact-variant population finding was present in the extracted paper record; the consulted ACMG/AMP publication provides the general population-frequency evidence framework rather than variant-specific data.

Still missing

• No disease-specific POLE maximum credible allele-frequency threshold or ancestry-complete control dataset beyond the retrieved resources was available.

Sources

Reasoning

BP4_Supporting is met by the generic missense computational fallback. p.Thr1689Ile is absent from the local POLE Supplementary Table S2 and S3 computational rules, and its REVEL score is 0.178, below the generic BP4 threshold of 0.250. The SpliceAI result is not separately used, avoiding double-counting computational evidence for this missense variant.

Rule

Generic ACMG/AMP missense computational fallback: REVEL <0.250 yields BP4_Supporting when no POLE exact-table rule applies.

Evidence considered

• The POLE local BP4 rule requires an exact Supplementary Table S2 or S3 entry classified as 'Likely benign' or 'Likely-benign' with at least four benign in silico results; p.Thr1689Ile is not listed in either table, so the framework directs generic fallback.
• The generic missense fallback assigns BP4_Supporting for REVEL <0.250; the observed REVEL score is 0.178. The use of calibrated REVEL computational thresholds is supported by Pejaver et al. (PMID:36413997).
• BayesDel score -0.338929 was not used because no verified published BayesDel threshold is available for this pipeline.

Sources

Reasoning

NM_006231.4:c.5066C>T is a missense substitution, NP_006222.2:p.(Thr1689Ile), rather than a predicted null variant. It is not a nonsense, frameshift, or canonical ±1/2 splice-site variant, so the generic PVS1 null-variant framework does not apply.

Rule

Generic ACMG/AMP PVS1 fallback: apply only to a predicted loss-of-function variant in a gene for which loss of function is an established disease mechanism; this missense substitution is not a PVS1 variant class.

Evidence considered

• The normalized ClinVar record identifies the variant as POLE c.5066C>T (p.Thr1689Ile).
• The variant-level PVS1 assessment identifies a missense consequence, no canonical splice-consensus involvement, and no eligibility for generic PVS1 because the variant is outside the nonsense, frameshift, and canonical splice null-variant buckets.
• No POLE-specific PVS1 specification was retrieved; the local POLE framework customizes only PM1, PS4, PP3, and BP4, so generic ClinGen SVI PVS1 guidance is the applicable fallback.

Sources

Reasoning

The applicable local POLE framework permits PS4_Supporting only for four specified exonuclease-domain variants (p.P286R, p.V411L, p.A456P, and p.S297F) that meet its dual COSMIC/TCGA endometrial-carcinoma recurrence requirement. POLE p.(Thr1689Ile) is not one of those exact variants, and the case evidence reports no COSMIC record.

Rule

Local POLE custom PS4: apply Supporting only to an exact established/pathogenic variant recurrent in both COSMIC and TCGA endometrial-carcinoma cohorts with combined count at least 10; no higher strength.

Evidence considered

• The local POLE framework's PS4 rule is an exact-variant somatic-recurrence adaptation capped at Supporting and names only p.P286R, p.V411L, p.A456P, and p.S297F as qualifying variants.
• The evaluated variant is NM_006231.4:c.5066C>T, NP_006222.2:p.(Thr1689Ile); the case evidence reports no COSMIC result for this variant.

Sources

Reasoning

The applicable local POLE framework specifies PM1 only for exact listed exonuclease-domain hotspot or recurrent substitutions. The queried p.Thr1689Ile is not one of the specified strong, moderate, or supporting substitutions (P286R, V411L, S297F, A456P, S459F; F367S, L424I, M295R, P436R, M444K, D368Y; A465V, L424V, T278M, A428T). The case bundle also reports no statistically significant hotspot for T1689.

Rule

Local POLE PM1 rule: apply PM1 only to the exact specified exonuclease-domain hotspot/recurrent substitutions; do not generalize the rule to all POLE missense substitutions or all residues.

Evidence considered

• The local POLE framework is a literature-derived internal framework, not an official ClinGen CSPEC/VCEP specification, and limits PM1 to the exact substitutions listed in its rule payload; p.Thr1689Ile is absent from those lists.
• Cancer Hotspots returned no statistically significant hotspot for POLE T1689.

Sources

Reasoning

The variant is a single amino-acid substitution, p.(Thr1689Ile), with no in-frame insertion/deletion, stop-loss, or other protein-length alteration. PM4 is therefore not met.

Rule

Generic ACMG/AMP PM4: protein-length-changing variants, such as in-frame insertions/deletions or stop-loss variants, may qualify when not occurring in a repetitive region without known function. A missense substitution does not meet this criterion.

Evidence considered

• The normalized ClinVar record identifies POLE c.5066C>T as p.(Thr1689Ile), which replaces one residue without changing protein length.
• No POLE-specific PM4 specification was retrieved; generic ACMG/AMP PM4 semantics apply.

Sources

Reasoning

PP3 is not met. This is a missense variant (p.Thr1689Ile). It is absent from both POLE supplementary computational tables, so the local POLE PP3 rule does not apply. Under the required generic missense fallback, the REVEL score of 0.178 is below the PP3 threshold and instead meets the BP4 threshold. SpliceAI is not independently counted for this missense variant.

Rule

POLE local PP3 exact-table rule was not triggered; generic missense REVEL fallback applied.

Evidence considered

• The POLE local framework requires an exact missense-table entry with REVEL class 'likely disease causing' and no more than one benign in silico result for PP3; p.Thr1689Ile is not listed in Supplementary Table S2 or S3.
• The generic missense fallback assigns PP3 only for REVEL >0.750 and BP4 for REVEL <0.250. The observed REVEL score is 0.178. The use of calibrated REVEL computational thresholds is supported by Pejaver et al. (PMID:36413997).
• SpliceAI max delta is 0.161, but the governing generic workflow requires a missense substitution to be assessed by REVEL rather than independently combining a splice-prediction result.

Sources

Reasoning

ClinVar has one exact-variant submission classified as uncertain significance from a single clinical laboratory and no expert-panel submissions. Therefore no exact-variant Pathogenic/Likely pathogenic ClinVar expert-panel assertion is available for PP5.

Rule

Apply PP5 only for an exact-variant ClinVar expert-panel classification of Pathogenic or Likely pathogenic.

Evidence considered

• ClinVar variation 2576435 is reported as uncertain significance with review status 'criteria provided, single submitter'; the submission audit reports zero expert-panel submissions.

Sources

Reasoning

BA1 is not met. The exact variant NM_006231.4:c.5066C>T (POLE p.Thr1689Ile) has a gnomAD v4.1 allele count of 0 among 1,608,632 alleles (AF 0.00000%), far below the generic ACMG/AMP stand-alone benign threshold of >5%.

Rule

Generic ACMG/AMP BA1: apply only when the allele frequency is >5% in a suitable population database; no applicable POLE VCEP/CSPEC specification was retrieved.

Evidence considered

• gnomAD v4.1 reports total AC 0, total AN 1,608,632, total AF 0.0, and 0 homozygotes for 12-132642284-G-A.
• The generic ACMG/AMP framework treats BA1 as a stand-alone benign criterion when the allele frequency exceeds 5%; this variant is absent rather than common.

Sources

Reasoning

BS1 is not met. The variant is not observed in gnomAD v4.1, gnomAD v2.1, or gnomAD-Canada v1.0, so there is no allele frequency above the disease-specific maximum credible population frequency to support a benign-frequency finding.

Rule

Generic ACMG/AMP BS1: apply when the allele frequency is greater than the maximum credible frequency for the relevant disorder; use a disease-specific threshold when available. No POLE-specific threshold was retrieved.

Evidence considered

• gnomAD v4.1 reports AC 0/AN 1,608,632 and AF 0.0, with 0 homozygotes; every listed population, including AFR, also has AC 0.
• The variant is reported absent from gnomAD v2.1 and gnomAD-Canada v1.0.
• Because no frequency excess is observed, the available population data do not support BS1; a disease-specific prevalence and penetrance calculation was not possible from this case bundle.

Still missing

• No POLE-specific maximum credible allele-frequency threshold or disease prevalence/penetrance model was available in the retrieved framework; however, the observed allele frequency is 0 and therefore does not meet BS1.

Sources

Reasoning

BS2 is not met. No healthy adult carrier or homozygous individual with this exact variant is documented in the case evidence. gnomAD reports 0 homozygotes, and population allele counts are zero; absence of carriers is not positive BS2 evidence.

Rule

Generic ACMG/AMP BS2: apply when the variant is observed in healthy adults, including homozygotes where relevant to the disease mechanism; database absence or zero homozygotes alone does not satisfy BS2.

Evidence considered

• gnomAD v4.1 reports 0 alleles and 0 homozygotes for the exact variant; gnomAD v2.1 and gnomAD-Canada v1.0 report the variant as absent.
• No source in the case bundle reports this exact variant in a healthy adult or in unaffected individuals with an informative age/phenotype assessment.

Still missing

• No phenotype-ascertained healthy-control cohort or documented healthy adult carrier was provided for this exact variant.
• The available gnomAD records provide allele and homozygote counts but do not establish a clinical healthy-adult observation for BS2.

Sources

Reasoning

p.(Thr1689Ile) is a missense substitution and does not create an in-frame protein-length change. Consequently, repeat-region context cannot trigger BP3 for this variant.

Rule

Generic ACMG/AMP BP3: an in-frame deletion/insertion in a repetitive region without a known function may qualify. This missense substitution has no in-frame length change and therefore does not meet BP3.

Evidence considered

• The normalized ClinVar record identifies POLE c.5066C>T as p.(Thr1689Ile), a one-residue substitution with no deletion or insertion.
• No POLE-specific BP3 specification was retrieved; generic ACMG/AMP BP3 semantics apply.

Sources

Reasoning

ClinVar has no expert-panel submission for this exact variant and its sole laboratory submission is uncertain significance. Therefore no exact-variant Benign/Likely benign ClinVar expert-panel assertion is available for BP6.

Rule

Apply BP6 only for an exact-variant ClinVar expert-panel classification of Benign or Likely benign.

Evidence considered

• ClinVar variation 2576435 has a single uncertain-significance laboratory submission; the submission audit reports zero expert-panel submissions.

Sources

Reasoning

The variant is POLE NP_006222.2:p.(Thr1689Ile), but the case bundle contains no verified pathogenic or likely pathogenic variant producing the same amino-acid change at Thr1689. ClinVar reports this exact variant as uncertain significance from a single submitter and does not provide same-amino-acid pathogenic evidence.

Rule

Generic PS1 requires a previously established pathogenic variant resulting in the same amino-acid change; an uncertain-significance record cannot satisfy PS1.

Evidence considered

• ClinVar identifies the exact variant NM_006231.4:c.5066C>T (p.Thr1689Ile) as uncertain significance with one single-submitter germline submission; this is not evidence of a pathogenic same-amino-acid change.

Still missing

• A verified pathogenic or likely pathogenic comparator with the same amino-acid change at Thr1689 is not available in the case evidence.

Sources

Reasoning

No proband-level de novo assertion, parental genotypes, maternity/paternity confirmation, or parental testing results are documented in the case-evidence bundle. PS2 therefore cannot be assessed.

Rule

Generic ACMG/AMP PS2 requires a confirmed de novo occurrence in the affected proband with maternity and paternity confirmed; strength may depend on phenotype specificity and parental testing completeness.

Evidence considered

• The consolidated case-evidence bundle contains no pedigree, proband/parent genotype results, trio testing report, or confirmed de novo observation for NM_006231.4:c.5066C>T (NP_006222.2:p.(Thr1689Ile)).
• ClinVar contains one germline single-submitter record classified as uncertain significance with no condition specified; it does not provide usable de novo or parental-testing evidence for this criterion.

Still missing

• Documented variant presence in the proband
• Variant-negative testing of both biological parents
• Confirmed maternity and paternity
• Phenotype and de novo-event details

Reasoning

No variant-specific validated functional assay evidence was identified for NM_006231.4:c.5066C>T (NP_006222.2:p.(Thr1689Ile)). The available ClinVar-linked publication is a general ACMG/AMP guideline and does not report this POLE variant or functional results. The local POLE framework does not specify PS3, so generic ACMG/AMP functional-evidence requirements apply; those requirements are not met by the evidence bundle because no assay result, appropriate controls, or biologically relevant functional readout is available.

Evidence considered

• The case bundle states that no official POLE CSPEC/VCEP specification was found; the local custom POLE framework customizes PM1, PS4, PP3, and BP4 only and provides no PS3 rule, so generic ACMG/AMP functional-evidence guidance is used.
• The paper extract for PMID:25741868 states that the paper does not explicitly mention POLE c.5066C>T or p.Thr1689Ile and provides no POLE-specific functional data; it contains only general ACMG/AMP guidance on evaluating functional studies.
• OncoKB reports no variant-specific reviewed functional evidence for POLE p.Thr1689Ile; its available information is gene-level context rather than an assay result.

Still missing

• A validated functional assay directly testing p.Thr1689Ile is unavailable.
• No assay controls, replication, quantitative result, or biologically relevant readout for this variant are available.

Sources

Reasoning

The variant is NM_006231.4:c.5066C>T (NP_006222.2:p.(Thr1689Ile)) in POLE. PM3 cannot be assessed because the case-evidence bundle provides no affected-proband observations, no confirmed biallelic disease, no pathogenic or likely pathogenic comparator variant, and no phase information establishing that this variant is in trans with such a variant. POLE inheritance mode is also not established in the supplied case evidence.

Rule

Generic ACMG/AMP PM3: moderate evidence for a recessive disorder when the variant is observed in trans with a pathogenic or likely pathogenic variant in an affected individual; phase and disease relevance must be established, and strength may be adjusted according to applicable specification.

Evidence considered

• The consolidated case-evidence bundle contains no family, pedigree, affected-proband, genotype-phase, or second-variant observations that satisfy the PM3 evidence requirements.
• The general ACMG/AMP standards were consulted for the criterion framework, but PMID:25741868 contains no POLE-specific clinical or segregation data and therefore does not provide positive PM3 evidence.

Still missing

• Affected-proband genotype and phenotype data are not supplied.
• No pathogenic or likely pathogenic variant in the same gene is reported for a relevant affected individual.
• No parental testing, segregation, or other phasing evidence is supplied to establish trans configuration.
• The applicable POLE disease inheritance model is not established in the supplied evidence.

Sources

Reasoning

No verified same-residue comparator variants at Thr1689 were identified in the case evidence. The PM5 candidate artifact contains zero candidates and explicitly states that classic PM5 semantics could not be safely confirmed.

Rule

Generic PM5 requires a novel missense change at a residue where another missense change is established as pathogenic; the comparator must be independently evaluated rather than inferred from a final classification alone.

Evidence considered

• The case PM5 candidate collection reports no same-residue candidates, no expert-panel pathogenic/likely pathogenic candidates, and no non-expert majority pathogenic/likely pathogenic candidates; it recommends not applying PM5 from that artifact.

Still missing

• A verified pathogenic or likely pathogenic missense variant at POLE residue Thr1689 is not available.

Sources

Reasoning

No reliable de novo observation without confirmed maternity and paternity is reported in the case-evidence bundle. PM6 cannot be assessed.

Rule

Generic ACMG/AMP PM6 is used for a presumed de novo occurrence when maternity and paternity have not been confirmed; the observation still requires credible case-level evidence and a consistent phenotype.

Evidence considered

• The consolidated case-evidence bundle provides no case report, family study, trio result, or parental genotype information establishing an unconfirmed de novo event for this variant.
• The available ClinVar submission cites only a general ACMG/AMP guideline and contains no variant-specific de novo evidence.

Still missing

• A documented de novo variant in the proband
• Evidence that maternity and paternity were not confirmed
• Phenotype consistency and clinical context

Reasoning

No affected or unaffected relatives, parental testing, informative meioses, or genotype-phenotype segregation data are available. PP1 cannot be assessed.

Rule

Generic ACMG/AMP PP1 requires co-segregation of the variant with disease in affected family members; strength should be calibrated to the number of informative meioses and phenotype specificity.

Evidence considered

• The consolidated case-evidence bundle contains no segregation study, pedigree, family genotype table, affected-relative report, or meiosis count for NM_006231.4:c.5066C>T.
• The variant-specific ClinVar record does not include family segregation data or a disease condition.

Still missing

• Family pedigree and relationship structure
• Genotypes of affected and unaffected relatives
• Number of informative meioses
• Phenotype definitions and clinical consistency

Reasoning

The bundle does not provide a validated POLE-specific PP2 specification or adequate gene-wide data demonstrating that pathogenic missense variation is a predominant mechanism with a low benign missense rate. The local POLE framework customizes PM1, PS4, PP3, and BP4, not PP2. Gene-level statements about POLE function and selected somatic hotspot mutations are insufficient to apply PP2.

Rule

Generic PP2 requires a gene in which missense variation is a common disease mechanism and benign missense variation is uncommon; gene function or isolated hotspots alone do not establish this criterion.

Evidence considered

• The local framework lists PM1, PS4, PP3, and BP4 as customized criteria and provides no PP2 rule.
• OncoKB provides only gene-level POLE biological context and no variant-specific or validated PP2 evidence.

Still missing

• A validated POLE missense-mechanism/benign-missense-rate assessment or applicable gene-specific PP2 rule is unavailable.

Sources

Reasoning

No proband phenotype, tumor phenotype, or evidence that the observed phenotype is highly specific for a POLE-related disorder was supplied. PP4 cannot be assessed.

Rule

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

Evidence considered

• The consolidated case evidence contains variant-level, population, and database information but no individual phenotype or phenotype-specific diagnostic evidence.

Still missing

• Provide the individual's phenotype and clinical context, with sufficient information to assess phenotype specificity and alternative etiologies.

Reasoning

No variant-specific validated benign functional assay evidence was identified for NM_006231.4:c.5066C>T (NP_006222.2:p.(Thr1689Ile)). The available ClinVar-linked publication is a general ACMG/AMP guideline and does not report this POLE variant or functional results. The local POLE framework does not specify BS3, so generic ACMG/AMP functional-evidence requirements apply; no well-established assay demonstrating normal function with suitable controls and a biologically relevant readout is available.

Evidence considered

• The case bundle states that no official POLE CSPEC/VCEP specification was found; the local custom POLE framework customizes PM1, PS4, PP3, and BP4 only and provides no BS3 rule, so generic ACMG/AMP functional-evidence guidance is used.
• The paper extract for PMID:25741868 states that the paper does not explicitly mention POLE c.5066C>T or p.Thr1689Ile and provides no POLE-specific functional data; it contains only general ACMG/AMP guidance on evaluating functional studies.
• OncoKB reports no variant-specific reviewed functional evidence for POLE p.Thr1689Ile; its available information is gene-level context rather than an assay result.

Still missing

• A validated functional assay demonstrating normal function for p.Thr1689Ile is unavailable.
• No assay controls, replication, quantitative result, or biologically relevant normal-function readout are available.

Sources

Reasoning

No tested unaffected relatives who lack the variant, and no informative non-segregation analysis, are documented. BS4 cannot be assessed.

Rule

Generic ACMG/AMP BS4 requires lack of segregation in affected family members or presence of the variant in appropriately assessed unaffected relatives, with adequate informative family data.

Evidence considered

• The consolidated case-evidence bundle contains no family study documenting unaffected relatives who were tested and found not to carry the variant.
• Absence of family information is not evidence of non-segregation; the available ClinVar and population records do not address BS4.

Still missing

• Genotypes of clinically unaffected relatives
• Reliable age and phenotype assessment of those relatives
• Informative family structure and segregation analysis

Reasoning

The bundle does not provide a validated POLE-specific BP1 rule or sufficient evidence that POLE disease is predominantly caused by truncating variants with missense variation generally tolerated. The available POLE material documents selected pathogenic missense hotspots in a somatic endometrial-carcinoma context and cannot establish a gene-wide benign missense pattern for this germline variant.

Rule

Generic BP1 requires a gene in which truncating variants are the established disease mechanism and missense variants are generally benign; this cannot be inferred from the absence of evidence for one variant or from selected somatic hotspots.

Evidence considered

• The local framework contains no BP1 specification and expressly limits its POLE-specific customizations to other criteria.
• The available POLE literature package concerns selected recurrent somatic exonuclease-domain missense variants and advises conservative interpretation of non-hotspot variants; it does not establish BP1 for POLE gene-wide germline missense variation.

Still missing

• A validated POLE loss-of-function predominance assessment together with evidence that missense variants are generally benign is unavailable.

Sources

Reasoning

The variant is NM_006231.4:c.5066C>T (NP_006222.2:p.(Thr1689Ile)) in POLE. BP2 cannot be assessed because the case-evidence bundle provides no observation of this variant in trans with a pathogenic or likely pathogenic variant, no phase information, and no disease-specific inheritance context supporting application of the benign allelic criterion. Absence of such data is insufficient to conclude that BP2 is not met.

Rule

Generic ACMG/AMP BP2: supporting benign evidence when the variant is observed in trans with a pathogenic variant for a fully penetrant dominant gene/disorder, or in cis with a pathogenic variant for a recessive disorder, provided the relevant inheritance model and phase are established.

Evidence considered

• The consolidated case-evidence bundle contains no phase, pedigree, parental, affected-proband, or allelic observations supporting BP2.
• The general ACMG/AMP standards were consulted for the criterion framework, but PMID:25741868 contains no POLE-specific clinical or segregation data and therefore does not provide positive BP2 evidence.

Still missing

• No confirmed trans configuration with a pathogenic or likely pathogenic variant is supplied.
• No affected or unaffected individual genotype data are supplied for allelic interpretation.
• No parental testing or other phasing evidence is supplied.
• The relevant POLE inheritance model and disease context are not established in the supplied evidence.

Sources

Reasoning

No individual clinical phenotype or alternate molecular diagnosis was supplied; the required evidence for BP5 is unavailable.

Rule

Generic ACMG/AMP fallback: BP5 requires an alternate molecular basis for disease in a case with the variant under assessment.

Evidence considered

• The consolidated case evidence provides no proband-level alternate molecular diagnosis or phenotype-explanation assessment.

Still missing

• Provide the individual's phenotype and results establishing an alternate molecular basis that fully explains that phenotype.