CDKN2A c.369T>A, p.(His123Gln)

Reasoning

PS1 requires that the same amino-acid change, regardless of nucleotide change, has previously been established as pathogenic in another proband (Richards et al. 2015, PMID:25741868). The variant under assessment is NM_000077.5:c.369T>A (NP_000068.1:p.His123Gln). The only record of this exact protein change is ClinVar VCV000127526, whose 15 submitting laboratories return 3 Benign, 9 Likely benign/likely benign and 3 Uncertain significance assertions, with no expert-panel review (1 star; API status 'Conflicting classifications of pathogenicity'). The single publication that explicitly names the H123Q substitution, PMID:18573309, reports only an in-silico PolyPhen prediction (PSIC score difference 2.696) and structural modelling of p16INK4A; it contains no clinical classification and no affected-individual data. The ClinVar submission audit likewise yielded no same-amino-acid-change pathogenic lead for this record. No pathogenic or likely pathogenic assertion for p.His123Gln therefore exists in any consulted source, so the same-amino-acid-change premise of PS1 is not satisfied.

Rule

ACMG/AMP 2015 PS1 (generic fallback; no gene-specific VCEP/CSPEC available)

Evidence considered

• ClinVar VCV000127526 (variation 127526; NM_000077.5(CDKN2A):c.369T>A (p.His123Gln)): 15 submissions totalling 3 Benign, 9 Likely benign/likely benign, 3 Uncertain significance; no expert panel; 1 star; API review status 'criteria provided, conflicting classifications' (last updated 2026-04-13).
• ClinVar submission audit for this record: the only extracted evidence signals were in-silico PP3 statements in two submissions; no submission presented a pathogenic same-residue or same-amino-acid-change comparator.
• PMID:18573309 names H123Q (rs6413463) only as a PolyPhen-predicted deleterious nsSNP (PSIC SD 2.696) and as a structural model (1a52 H123Q); it provides no clinical pathogenicity determination for the change.
• Framework check: no ClinGen CSPEC was found (cspec.found=false), no gene VCEP directory exists (gene_vcep_dir=null) and vcep_materials.json domain_tables is empty, so generic ACMG/AMP 2015 (PMID:25741868) governs this criterion.

Sources

Reasoning

PS3 (Richards et al. 2015, PMID:25741868) requires well-established in vitro or in vivo functional assay data that demonstrate a damaging effect on the encoded protein, with documented assay controls and biological relevance to the disease mechanism; computational prediction alone cannot satisfy it. The variant under review is NM_000077.5:c.369T>A, predicted to produce NP_000068.1:p.(His123Gln), a missense substitution in p16INK4A/CDKN2A. No functional assay of this exact substitution was found in any retrieved source. The only paper that names the exact change is PMID:18573309, a purely in-silico study that predicted H123Q (rs6413463) to be damaging by PolyPhen (PSIC score difference 2.696) and carried a mutant model labelled '1a52 H123Q' forward into structural modelling; no wet-lab assay was performed, no assay controls exist, and H123Q was not among the three modelled mutants (P81L, H83Q, D84Y) for which RMSD, total-energy and binding-free-energy values were reported. Its Discussion asserts, without presenting the underlying primary data, that three PolyPhen-deleterious nsSNPs including H123Q are 'well supported by previously reported experimental mutational data'; an unsourced secondary claim of that kind cannot be credited as a well-established functional assay. Every other candidate source is a gene-level study that reports no result for p.His123Gln: homozygous CDKN2A/p16 deletions in leukaemia cohorts (PMID:9166859, PMID:7718873), CDKN2A/B-ARF tumour-suppressor loss in BCR-ABL ALL (PMID:18519632), germline multigene-panel mutation frequencies (PMID:25186627, PMID:27978560, PMID:28944238), and a structure/activity analysis of twelve tumorigenic p16INK4A mutants (PMID:9660926) whose abstract names none of them and whose full text could not be retrieved. OncoKB holds no variant-specific functional annotation for CDKN2A p.His123Q (Unknown Oncogenic Effect), and the three COSMIC records (COSV58721567) are somatic observations, not germline functional assay evidence. PS3 is therefore not met; residual uncertainty is confined to the five candidate PMIDs whose full texts were unavailable and which, at abstract level, contain no variant-specific assay.

Evidence considered

• PS3 (Richards et al. 2015, PMID:25741868) is defined as well-established in vitro or in vivo functional studies supportive of a damaging effect on the gene or gene product; prediction-tool output is PP3-type evidence and cannot satisfy PS3.
• PS3 was assessed against the exact variant only: NM_000077.5:c.369T>A, NP_000068.1:p.(His123Gln), missense in CDKN2A (p16INK4A).
• PMID:18573309 (full text read): names the exact substitution (rs6413463, nucleotide change A/T, H123Q) in Table 1 with a PolyPhen PSIC score difference of 2.696, and includes a mutant model labelled '1a52 H123Q' in its structural modelling set; no wet-lab assay, no substrate/cell controls, and no reported RMSD or interaction energy for H123Q, which was not among the three mutants (P81L, H83Q, D84Y) with RMSD > 3.00 A. The paper is in silico only.
• PMID:18573309 Discussion, paragraph 2: the authors state that three PolyPhen-deleterious nsSNPs including rs6413463 (H123Q) are 'also well supported by the above-mentioned experimental mutational data'. This is a secondary claim about third-party experiments that were not performed or reproduced in this paper and were not traceable to a primary assay in the retrieved corpus, so it was not credited as functional evidence.
• PMID:9660926 (abstract only; full text not retrievable): 'Twelve tumorigenic mutants of p16INK4A have been constructed and analyzed for their structure and activity'; the abstract names no mutant, and H123Q is not mentioned, so no variant-specific assay result can be extracted.
• PMID:9166859 (abstract): homozygous p16/MTS1 deletions in 48 pediatric ALL patients correlated with outcome - a copy-number/cohort study, not an assay of any missense allele.
• PMID:7718873 (abstract): semiquantitative multiplex PCR for homozygous p16 gene deletions in 34 CML blast-crisis, 19 ALL and 25 AML patients - gene-level deletion study, no missense functional data.
• PMID:18519632 (abstract): discussion of CDKN2A/B (INK4A/B-ARF)-mediated tumour suppression failure and targeted-therapy resistance in BCR-ABL ALL - gene-level biology, no variant-specific assay.
• PMID:25186627 (abstract): cross-sectional 25-gene panel testing in 2158 individuals referred for BRCA1/2 testing; reports germline mutation frequencies, not functional assay data for CDKN2A p.His123Gln.
• PMID:27978560 and PMID:28944238: germline panel/targeted-sequencing cohort papers; the exact variant is not named, and neither reports functional assay results for CDKN2A.
• OncoKB: 'Unknown Oncogenic Effect', is_variant_specific=false, no variant-specific reviewed functional evidence for CDKN2A H123Q; only gene-level context is provided.
• COSMIC: this variant has three recorded somatic occurrences (COSV58721567); somatic tumour recurrences are not germline functional assay evidence for PS3.
• No ClinGen CSPEC and no gene-specific criteria specification exists for CDKN2A (vcep_materials.json: no CSPEC URL, final_classification_framework found=false, source=generic_acmg_fallback), so generic ACMG/AMP 2015 standards govern PS3.

Still missing

• No functional assay of CDKN2A p.(His123Gln) was identified in any retrieved source. The only exact-variant data are in-silico predictions (PMID:18573309: PolyPhen PSIC score difference 2.696 plus homology modelling of p16INK4A-CDK6), which cannot support PS3 at any strength.
• Five ClinVar-referenced candidate papers could not be retrieved in full (no files exist under publications/ for them, and Sci-Hub and Europe PMC retrieval failed from this environment): PMID:18519632, PMID:25186627, PMID:9166859, PMID:7718873, PMID:9660926. Only PMID:9660926 describes variant-level functional work (twelve tumorigenic p16INK4A mutants constructed and analysed for structure and activity), and its abstract does not name H123Q; a reviewer with full-text access should confirm whether H123Q is among those twelve mutants.
• If a qualifying assay were found, PS3 would additionally require documented positive and negative assay controls, a quantitative effect size, and demonstration that the assay interrogates the CDKN2A/p16-CDK4-RB tumour-suppressor mechanism relevant to this gene's disease mechanism.

Sources

Reasoning

PS4 requires that the prevalence of the variant in affected individuals be statistically increased over controls. The variant is CDKN2A NM_000077.5:c.369T>A (p.His123Gln), ClinVar variation 127526. No case-control or case-cohort enrichment study reporting this exact variant was identified. Literature triage surfaced only gene-level CDKN2A papers: the CDKN2A-containing cohorts (PMID:27978560, 450 early-onset colorectal cancer patients tested on a 25-gene panel, and PMID:28944238, 1231 CRC cases vs 93 unaffected controls) never name c.369T>A/p.His123Gln and report other CDKN2A missense alleles instead; the only publication that explicitly names H123Q (PMID:18573309) is an in silico PolyPhen/structural analysis with no patient or control counts. There is therefore no affected-vs-control numerator/denominator comparison to evaluate, and PS4 cannot be affirmatively supported. No CDKN2A VCEP/CSPEC specification exists, so generic ACMG/AMP 2015 applies (PMID:25741868), which sets no numeric PS4 threshold. PS4 is not met.

Rule

Generic ACMG/AMP 2015 criteria (PMID:25741868); no CDKN2A-specific VCEP/CSPEC specification available

Evidence considered

• PMID:18573309 is the only retrieved publication that explicitly names the protein change H123Q (rs6413463, nucleotide change A/T); it is an in silico PolyPhen/structure-modelling study (PSIC score difference 2.696) with no patient cohort, control group, or case-control counts, so it provides no PS4 enrichment data.
• PMID:27978560 (Ohio Colorectal Cancer Prevention Initiative) tested 450 early-onset CRC patients on a 25-gene panel including CDKN2A and reported one patient with a pathogenic CDKN2A mutation, but the specific nucleotide/protein change is not given, so this variant cannot be assigned to that case and no enrichment can be computed.
• PMID:28944238 sequenced 36 CRC susceptibility genes in 1231 CRC cases and 93 unaffected controls with CDKN2A included; the CDKN2A variants reported were p.Ala127Ser and p.Arg144Cys, not c.369T>A, so this study provides no case-control frequency for this variant.
• ClinVar variation 127526 (the exact variant) contains 16 ordinary single-submitter laboratory records and no cohort/enrichment data; it therefore carries no affected-vs-control comparison usable for PS4.
• No CDKN2A CSPEC or VCEP specification was found in the case materials or the case's VCEP directory; generic ACMG/AMP 2015 (PMID:25741868) applies and defines no numeric PS4 threshold, so PS4 rests solely on the presence/absence of case-control enrichment evidence.

Still missing

• No case-control or case-enrichment study reporting the exact variant c.369T>A (p.His123Gln) was identified; PS4 cannot be affirmatively supported without affected-versus-control counts for this variant.

Sources

Reasoning

PM1 requires the variant to lie in a mutational hotspot and/or a critical, well-established functional domain that lacks benign variation (Richards et al. 2015, PMID:25741868). vcep_materials.json contains no domain_tables entry for CDKN2A (domain_tables = {}) and no CSPEC or gene-specific framework was located (framework_mode = generic_acmg), so there is no VCEP-approved critical-domain list against which His123 can be checked and the generic rule applies. Cancer Hotspots returned no hotspot row for CDKN2A H123 ('This variant does not lie in a statistically significant hotspot'), and COSMIC records only 3 somatic occurrences. Crucially, benign variation at this residue is documented rather than absent: the identical His123Gln allele is present in gnomAD v2.1 at 116/274,982 alleles (AF 0.042%; African/African American 109/24,428 alleles, AF 0.45%) with zero homozygotes, and 12 of 15 ClinVar submitters classify the change Benign or Likely benign. Because neither hotspot status nor a benign-variation-free critical domain can be established for residue His123, PM1 is not met.

Rule

ACMG/AMP 2015 PM1 (generic fallback; no gene-specific VCEP/CSPEC or domain table available)

Evidence considered

• vcep_materials.json: domain_tables = {} (no CDKN2A entry) and no gene VCEP directory, so no approved critical-functional-domain list is available; generic ACMG/AMP PM1 was applied.
• Cancer Hotspots (cancerhotspots.org) query 'CDKN2A H123' returned search_status 'no_result' / found=false with evidence sentence 'This variant does not lie in a statistically significant hotspot.'
• COSMIC records this variant somatically 3 times (COSV58721567).
• gnomAD v2.1: 116/274,982 alleles (AF 0.000421846, 0.042%); African/African American 109/24,428 (AF 0.004462, 0.45%); 0 homozygotes; grpmax FAF 0.00387.
• ClinVar VCV000127526: 12 of 15 submissions are Benign or Likely benign/likely benign, i.e. benign missense variation is demonstrably present at this residue rather than absent.

Still missing

• No VCEP/CSPEC-approved CDKN2A critical-domain table exists in the case evidence (vcep_materials.json domain_tables empty), so domain membership for His123 could only be assessed against generic ACMG criteria rather than an authoritative gene-specific domain list.

Sources

Reasoning

PM2 (supporting) is met only when the variant is absent or extremely rare in population databases; the supplied threshold is an allele frequency of 0.0001 or below (SVI recommendation downgrading PM2 to supporting). This variant exceeds that threshold in every dataset examined. gnomAD v4.1 (all-comers) total AF is 0.000223 (359/1,610,194 alleles; 1 homozygote) with grpmax FAF 0.004159, and gnomAD v2.1 total AF is 0.000422 (116/274,982 alleles; 0 homozygotes) with grpmax FAF 0.003871; gnomAD-Canada total AF is 0.000163 (3/18,422) with faf95 grpmax 0.000348. The African/African American subpopulation frequency reaches about 0.46% in both gnomAD v4.1 (0.004557) and v2.1 (0.004462), roughly 45-fold above the 0.0001 threshold. PM2 is not met.

Evidence considered

• PM2 supporting threshold applied: AF <= 0.0001 (ClinGen SVI recommendation to downgrade PM2 to supporting, per the supplied calibration block citing PMID:25741868).
• gnomAD v4.1 (all-comers): total AF 0.000222955 (359/1,610,194 alleles; total homozygotes = 1); grpmax FAF 0.00415903; African/African American AF 0.00455684 (342/75,052).
• gnomAD v2.1 (all-comers): total AF 0.000421846 (116/274,982 alleles; total homozygotes = 0); grpmax FAF 0.00387069; African/African American AF 0.00446209 (109/24,428).
• gnomAD-Canada v1.0: total AF 0.00016285 (3/18,422); grpmax faf95 0.00034821.
• Total all-comers AF (0.000163-0.000422) and, more decisively, the African/African American AF (~0.46%) both exceed the 0.0001 PM2 threshold; the variant is not rare enough for PM2.

Still missing

• No CDKN2A VCEP/CSPEC or local framework was available to supply a gene-specific PM2 frequency cutoff; the generic SVI 0.0001 threshold was used.
• No non-cancer or exome-only gnomAD subset was available for this variant; all-comers gnomAD v2.1/v4.1 totals were used.

Sources

Reasoning

PM5 requires a different missense change at the same codon (His123) that has already been determined to be pathogenic (Richards et al. 2015, PMID:25741868). The case's PM5 comparator harvest returned no candidates whatsoever (total_candidates_seen = 0, same_residue_candidates = 0), and the classic same-residue search was skipped (pm5_mode = 'unknown', eligible_for_classic_pm5_search = false). No consulted source identifies a pathogenic missense variant at CDKN2A codon 123: the ClinVar record for this variant (VCV000127526) lists no residue-123 comparator, its submission audit extracted only in-silico PP3 signals, and none of the fetched publications reports another His123 missense change. Because the criterion's prerequisite — a distinct established pathogenic missense at His123 — is absent, PM5 is not met. The absence finding is not exhaustive, however: automated residue-level harvesting was not executed and no manual ClinVar/Literature scan for alternate codon-123 changes was performed.

Rule

ACMG/AMP 2015 PM5 (generic fallback; no gene-specific VCEP/CSPEC available)

Evidence considered

• pm5_candidates.json: found=false, pm5_mode='unknown', eligible_for_classic_pm5_search=false, total_candidates_seen=0, same_residue_candidates=0, expert_panel_plp_candidates=0, candidates=[]; recommendation level 'not_applicable' with instruction not to apply PM5 from the artifact alone.
• ClinVar VCV000127526 submission list (15 SCVs) contains no variant-level record of a different pathogenic missense at codon 123.
• ClinVar submission audit: the only criteria/signals extracted were in-silico PP3 statements; no same-residue comparator lead was identified.
• No fetched full-text publication reports a pathogenic missense change at His123 other than the variant under assessment.

Still missing

• Automated same-residue PM5 candidate harvesting was not run (pm5_mode 'unknown', eligible_for_classic_pm5_search false); an exhaustive manual residue-level ClinVar/literature search for other missense changes at CDKN2A codon 123 was not performed.

Sources

Reasoning

PP2 requires a missense variant in a gene that has a low rate of benign missense variation and in which missense changes are a common mechanism of disease (Richards et al. 2015, PMID:25741868). The mechanism half of the criterion holds: missense substitution is a documented CDKN2A disease mechanism (PMID:18573309 characterises deleterious p16INK4A missense nsSNPs in malignant melanoma, and germline CDKN2A mutation carriers are managed as a high-risk melanoma/pancreatic-cancer population, PMID:25645574; PMID:31672839). The low-benign-missense-variation half is not supported by the case evidence: benign missense variation is documented in this small gene — two CDKN2A missense variants at residues flanking His123, p.Ala127Ser and p.Arg144Cys, were reported but not classified as pathogenic and were enriched in African American/admixed individuals (PMID:28944238) — and the same His123Gln allele is present in gnomAD v2.1 at AF 0.042% while 12 of 15 ClinVar submitters assert Benign or Likely benign. Because the required low rate of benign missense variation cannot be established for CDKN2A, PP2 is not met; no quantitative gene-level missense-constraint metric was available to the case.

Rule

ACMG/AMP 2015 PP2 (generic fallback; no gene-specific VCEP/CSPEC available)

Evidence considered

• CDKN2A missense mechanism: PMID:18573309 analyses 14 coding nsSNPs of p16INK4A and reports deleterious missense changes in malignant melanoma; PMID:31672839 and PMID:25645574 each classify germline CDKN2A mutation carriers as a high-risk cancer population, confirming the gene's dominant disease mechanism operates through sequence change including missense.
• Benign missense variation in CDKN2A: PMID:28944238 reported p.Ala127Ser and p.Arg144Cys as Tier 2, not classified as pathogenic, and predominantly present in African American or admixed individuals.
• Population data: the His123Gln allele is present in gnomAD v2.1 at 116/274,982 alleles (AF 0.042%, 0 homozygotes), and ClinVar VCV000127526 carries 12 Benign/Likely benign assertions from 15 submitters.
• No gene-level missense-constraint metric (mis_z / missense o/e) was present in the case evidence, so PP2 was evaluated qualitatively rather than against a rate threshold.

Still missing

• No quantitative gene-level missense-constraint metric (e.g. gnomAD mis_z or missense observed/expected) was available in the case evidence; PP2 was assessed from documented benign missense variation rather than a numeric rate threshold.

Sources

Reasoning

Scope check first: NM_000077.5:c.369T>A is a coding missense substitution, NP_000068.1:p.(His123Gln) (VEP most_severe_consequence = missense_variant; pvs1_variant_assessment consequence_class = missense; no canonical splice-consensus or splice-region involvement). For a missense variant the calibrated computational path is REVEL only; the SpliceAI result on this case says nothing about whether an amino-acid substitution is tolerated and is therefore not consulted for this criterion. The local REVEL v1.3 lookup returns a score of 0.542. Under the ClinGen SVI REVEL calibration (Pejaver et al. 2022, PMID:36413997), PP3 requires a REVEL score >= 0.644 at supporting strength (>= 0.773 moderate, >= 0.932 strong). 0.542 is below the supporting PP3 threshold, so PP3 is not met. This is a true indeterminate/gray-zone REVEL result: it does not meet PP3 and it simultaneously does not meet BP4 (BP4 supporting requires <= 0.29). BayesDel is present (0.365078) but no ACMG-strength calibration for BayesDel exists in the governing materials or in generic ACMG, so it is not_available and contributes nothing here. No gene-specific (CSPEC/VCEP) PP3/BP4 lookup table exists for CDKN2A, so the generic thresholds govern.

Rule

ClinGen SVI REVEL calibration (Pejaver et al. 2022, PMID:36413997): PP3 supporting >= 0.644; PP3 not met below that value.

Evidence considered

• Variant consequence for path selection: VEP most_severe_consequence = missense_variant and pvs1_variant_assessment consequence_class = missense for NM_000077.5:c.369T>A (p.His123Gln), so the missense/REVEL path governs PP3/BP4 and the SpliceAI path does not apply.
• Local REVEL v1.3 predictor lookup for chr9:21970990 A>T (NM_000077.5:c.369T>A) returned score = 0.542, found = true.
• ClinGen SVI REVEL calibration thresholds (Pejaver et al. 2022, Am J Hum Genet, PMID:36413997): PP3 supporting >= 0.644, PP3 moderate >= 0.773, PP3 strong >= 0.932.
• REVEL 0.542 falls in the indeterminate/gray zone between the BP4 supporting cutoff (<= 0.29) and the PP3 supporting cutoff (>= 0.644), so neither PP3 nor BP4 is met and no PP3 strength can be assigned.
• BayesDel score 0.365078 is present in the case data but no generic ACMG-strength or gene-specific calibration exists for BayesDel, so it is treated as not_available and is not used for PP3 or BP4.
• SpliceAI max delta score 0.01 for this variant was deliberately not used for PP3/BP4: the variant is missense, and a splice prediction is not evidence about the tolerance of an amino-acid substitution.
• Governing framework: no ClinGen CSPEC and no gene-specific/VCEP PP3/BP4 lookup table is available for CDKN2A (framework_mode = generic_acmg; final_classification_framework found = false), so generic ACMG/AMP computational-evidence thresholds apply.

Still missing

• No ClinGen CSPEC/VCEP gene-specific PP3/BP4 calibration exists for CDKN2A; generic ClinGen SVI REVEL thresholds were used.

Sources

Reasoning

PP5 requires that the exact variant be classified as Pathogenic or Likely pathogenic by an expert panel in ClinVar. ClinVar variation 127526 (the exact variant NM_000077.5:c.369T>A, p.His123Gln) carries review status 'criteria provided, single submitter' (1-star) with 0 expert-panel submissions across 16 records; all 16 submissions are ordinary single-submitter laboratory entries (8 Likely benign, 3 Benign, 3 Uncertain significance, 1 likely benign), and the aggregate label is Likely benign. No ClinGen CDKN2A VCEP/CSPEC specification exists. Because no exact-variant expert-panel Pathogenic/Likely pathogenic classification exists - and the aggregate ClinVar label is in any case Likely benign - PP5 is not met.

Rule

Generic ACMG/AMP 2015 criteria (PMID:25741868); PP5 applied only for an exact-variant ClinVar expert-panel Pathogenic/Likely pathogenic classification

Evidence considered

• ClinVar variation 127526 for the exact variant has review status 'criteria provided, single submitter' (1 star) and expert_panel_submissions = 0 out of 16 submissions; no ClinVar expert panel has asserted a classification for this variant.
• The ClinVar aggregate label for variation 127526 is Likely benign (8 clinical laboratories), Benign (3), Uncertain significance (3) and likely benign (1) - an aggregate of ordinary laboratory submissions, which must not trigger PP5.
• PMID:25741868 (ACMG/AMP 2015) defines PP5 and its requirement for a recognised expert-panel assertion, which is the standard applied here; no CDKN2A expert-panel specification exists to modify it.

Sources

Reasoning

BA1 (stand-alone benign) requires an allele frequency of at least 0.05 in a population database. No CDKN2A CSPEC/VCEP or local gene-specific framework exists for this case, so the generic ACMG/AMP 2015 threshold applies and the all-comers gnomAD v2.1/v4.1 totals are the default population source. gnomAD v4.1 (all-comers) reports a total AF of 0.000223 (359/1,610,194 alleles; 1 homozygote) with a maximum population frequency of 0.004557 (342/75,052) in African/African American and a grpmax filtering AF of 0.004159. gnomAD v2.1 reports a total AF of 0.000422 (116/274,982 alleles; 0 homozygotes) with African/African American AF 0.004462 (109/24,428) and grpmax FAF 0.003871. gnomAD-Canada v1.0 (HostSeq, 10,487 genomes) reports a total AF of 0.000163 (3/18,422) with a grpmax African AF of 0.00196 (faf95 0.000348). The single highest population frequency observed anywhere is about 0.46%, far below the 5% BA1 stand-alone threshold. BA1 is not met.

Evidence considered

• No CDKN2A CSPEC/VCEP or local gene-specific framework is present (framework_complete=false, generic_acmg fallback), so the generic ACMG/AMP 2015 stand-alone benign threshold (AF >= 0.05) applies.
• gnomAD v4.1 (all-comers): total AF 0.000222955 (359/1,610,194 alleles; total homozygotes = 1); maximum population African/African American AF 0.00455684 (342/75,052; 1 homozygote); grpmax FAF 0.00415903.
• gnomAD v2.1 (all-comers, exome+genome): total AF 0.000421846 (116/274,982 alleles; total homozygotes = 0); African/African American AF 0.00446209 (109/24,428; 0 homozygotes); grpmax FAF 0.00387069.
• gnomAD-Canada v1.0 (HostSeq genomes, n = 10,487): total AF 0.00016285 (3/18,422 alleles); grpmax African AF 0.00196078 (2/1,020; faf95 0.00034821).
• The highest frequency in any population in any dataset (~0.46%) is roughly 11-fold below the 5% BA1 threshold; BA1 is not satisfied.

Still missing

• No CDKN2A VCEP/CSPEC or local gene-specific framework was available to override the generic 0.05 BA1 stand-alone threshold.
• No non-cancer or exome-only gnomAD subset was available for this variant; all-comers gnomAD v2.1/v4.1 totals were used.

Sources

Reasoning

BS1 (strong benign) applies at an allele frequency of at least 0.01 (generic SVI default of 0.01; per-gene derivation is unavailable because no CDKN2A VCEP/CSPEC or local framework exists for this case, and the all-comers gnomAD totals remain the default population source). The highest frequencies observed are gnomAD v4.1 African/African American AF 0.004557 (342/75,052; grpmax FAF 0.004159) and gnomAD v2.1 African/African American AF 0.004462 (109/24,428; grpmax FAF 0.003871). Every other subpopulation is at or below ~0.0002 (for example, gnomAD v4.1 non-Finnish European AF 0.0000017, Admixed American 0.0001), and gnomAD-Canada's grpmax African AF is 0.00196. No population reaches the 1% threshold; the maximal ancestry-specific value, African/African American at ~0.46%, is less than half of it. BS1 is not met.

Evidence considered

• No CDKN2A CSPEC/VCEP or local framework exists, so the generic BS1 strong-benign threshold of AF >= 0.01 was applied; the generic default derives from Whiffin et al. 2017 (PMID:28518168), which recommends per-gene derivation where data allow.
• gnomAD v4.1 (all-comers): African/African American AF 0.00455684 (342/75,052; grpmax FAF 0.00415903); total AF 0.000222955 (359/1,610,194).
• gnomAD v2.1 (all-comers): African/African American AF 0.00446209 (109/24,428; grpmax FAF 0.00387069); total AF 0.000421846 (116/274,982).
• gnomAD-Canada v1.0: grpmax African AF 0.00196078 (2/1,020; faf95 0.00034821); total AF 0.00016285 (3/18,422).
• The maximal ancestry-specific frequency in any dataset (~0.46%, African/African American) is below half of the 1% BS1 threshold; BS1 is not satisfied.

Still missing

• No CDKN2A VCEP/CSPEC or local gene-specific framework was available to supply a per-gene BS1 threshold; the generic SVI default of 0.01 was used.
• A data-derived BS1 threshold from disease prevalence/penetrance could not be computed; the generic default stands.

Sources

Reasoning

BS2 is met when a variant is observed in a healthy adult individual in the context of a disorder for which full penetrance is expected at an early age (for a dominant disorder, this typically means observing homozygotes or otherwise implausible healthy carriers). gnomAD v4.1 (all-comers) contains exactly one homozygote among 359 total alleles (total AF 0.000223); that homozygote sits in the African/African American subpopulation (1/75,052, AF 0.004557). gnomAD v2.1 contains 116 alleles and 0 homozygotes, and gnomAD-Canada contains 3 alleles and 0 homozygotes. The disorder context here is CDKN2A-associated hereditary melanoma (familial atypical multiple mole melanoma) and melanoma-pancreatic cancer syndrome, an autosomal dominant condition with reduced (incomplete) and adult-onset penetrance rather than full penetrance at an early age. The penetrance premise on which BS2 depends is therefore not satisfied, and a single reference-population homozygote, without linked phenotype or age, does not establish the criterion. BS2 is not met.

Evidence considered

• BS2 as defined by ACMG/AMP 2015 requires the variant to be observed in a healthy adult for a disorder with full penetrance expected at an early age.
• gnomAD v4.1 (all-comers): total homozygotes = 1 (total AF 0.000222955, 359/1,610,194); the homozygote is in African/African American (homozygote_count = 1; AF 0.00455684; 342/75,052).
• gnomAD v2.1 (all-comers): total homozygotes = 0 (116/274,982 alleles); all subpopulations homozygote_count = 0.
• gnomAD-Canada v1.0: homozygote_count = 0 (3/18,422 alleles).
• CDKN2A germline disease (familial melanoma / melanoma-pancreatic cancer syndrome) is autosomal dominant with reduced, adult-onset penetrance, so the full-penetrance-at-early-age premise of BS2 does not hold; a homozygous observation in a reference cohort is not the BS2 scenario for such a gene.

Still missing

• The single gnomAD v4.1 homozygote carries no linked phenotype, age, or ancestry-quality annotation, so BS2 rests on CDKN2A's known reduced, adult-onset penetrance rather than direct genotype-phenotype evidence.
• No CDKN2A VCEP/CSPEC or local framework was available to specify a gene-specific BS2 application (e.g. explicit penetrance assumptions).

Sources

Reasoning

BS3 (Richards et al. 2015, PMID:25741868) requires well-established functional assay data that demonstrate no damaging effect on protein function. The variant is the missense substitution NM_000077.5:c.369T>A (NP_000068.1:p.(His123Gln)) in CDKN2A/p16INK4A. No functional assay of this substitution was found in any retrieved source, so there is no assay result demonstrating preserved p16INK4A function (e.g. retained CDK4/CDK6 inhibition, preserved cell-cycle arrest, or normal protein stability). The only exact-variant evidence, PMID:18573309, is in-silico and points in the opposite direction - PolyPhen classified H123Q as damaging with a PSIC score difference of 2.696 - and computational prediction is not functional-assay evidence for BS3 any more than it is for PS3. The remaining sources are gene-level deletion, cohort or guideline studies (PMID:9166859, PMID:7718873, PMID:18519632, PMID:25186627, PMID:27978560, PMID:28944238, PMID:25645574, PMID:31672839) that provide no assay of this allele, and the one variant-level functional study identified (PMID:9660926, twelve tumorigenic p16 mutants analysed for structure and activity) does not name H123Q at abstract level and could not be retrieved in full. OncoKB holds no variant-specific functional evidence for this variant, and the somatic COSMIC records (COSV58721567) are not germline functional data. BS3 is therefore not met.

Evidence considered

• BS3 (Richards et al. 2015, PMID:25741868) is defined as well-established in vitro or in vivo functional studies showing no damaging effect on the gene or gene product; the evidence must be an assay of the exact variant.
• Variant assessed: NM_000077.5:c.369T>A, NP_000068.1:p.(His123Gln), missense in CDKN2A.
• PMID:18573309 (full text read): the only source naming the exact substitution H123Q (rs6413463); it reports in-silico PolyPhen prediction of a damaging effect (PSIC score difference 2.696) and a homology model, not an assay of function, so it provides no evidence of a non-damaging effect.
• PMID:27978560 reports a pathogenic CDKN2A mutation in 1 of 450 early-onset colorectal cancer patients but gives no nucleotide/protein change and no functional assay, so it neither names nor assays p.His123Gln.
• PMID:28944238 (full text read): CDKN2A missense variants seen in the CRC cohort were p.Ala127Ser and p.Arg144Cys only; p.His123Gln is not mentioned.
• PMID:29533785 (full text read): functional-genomic viability platform covering 1049 somatic mutations in 95 genes; CDKN2A is not among the genes tested and the variant is never named, so it supplies no assay result either way.
• PMID:25645574 and PMID:31672839: clinical practice/consensus guidelines that name CDKN2A only at gene level (pancreatic cancer risk and surveillance); no variant-level functional data.
• PMID:9166859, PMID:7718873, PMID:18519632: leukaemia deletion/cohort and mechanism studies performed at gene/locus level, not assays of the p.His123Gln allele.
• OncoKB: no variant-specific reviewed functional evidence ('Unknown Oncogenic Effect'), so it cannot support preserved function.
• No ClinGen CSPEC or gene-specific specification exists for CDKN2A, so generic ACMG/AMP 2015 standards govern BS3.

Still missing

• No functional assay of CDKN2A p.(His123Gln) showing preserved protein function was identified; BS3 therefore cannot be met on the retrieved evidence.
• Candidate papers that might contain p16INK4A assay data could not be retrieved in full (PMID:9660926, PMID:9166859, PMID:7718873, PMID:18519632, PMID:25186627); PMID:9660926 is the only one describing variant-level functional work and its abstract does not name H123Q.
• A BS3-grade assay would need documented positive and negative controls, a quantitative null effect size versus wild-type, and relevance to the CDKN2A/p16-CDK4-RB tumour-suppressor mechanism before any strength could be assigned.

Sources

Reasoning

BP1 applies only to a missense variant in a gene for which disease is caused primarily by truncating variants (Richards et al. 2015, PMID:25741868). CDKN2A does not meet that premise: missense substitutions in p16INK4A are an established pathogenic mechanism for melanoma and other CDKN2A-associated cancers (PMID:18573309 analyses deleterious CDKN2A/p16INK4A missense nsSNPs in malignant melanoma and cites supporting experimental mutational data), and CDKN2A missense variants are identified directly in cancer-susceptibility sequencing cohorts (PMID:28944238). Because both truncating and missense changes are recognised causes of CDKN2A disease — the gene is not truncating-predominant — BP1, whose only route to benign support is a truncating-restricted disease mechanism, is not met for this missense variant.

Rule

ACMG/AMP 2015 BP1 (generic fallback; no gene-specific VCEP/CSPEC available)

Evidence considered

• PMID:18573309 characterises deleterious p16INK4A missense nsSNPs in malignant melanoma and cites previously reported experimental mutational data for three of them, establishing missense substitution as a CDKN2A disease mechanism.
• PMID:28944238 identified CDKN2A missense variants (p.Ala127Ser, p.Arg144Cys) in a cancer-susceptibility sequencing cohort, showing missense changes are observed in the CDKN2A disease context rather than being tolerated.
• No consulted source supports truncating variants as the primary or sole CDKN2A disease mechanism, so the truncating-predominance premise required by BP1 cannot be established.

Sources

Reasoning

Scope check first: NM_000077.5:c.369T>A is a coding missense substitution, NP_000068.1:p.(His123Gln) (VEP most_severe_consequence = missense_variant), so BP4 is evaluated on the missense/REVEL path only. The local REVEL v1.3 lookup gives 0.542. Under the ClinGen SVI REVEL calibration (Pejaver et al. 2022, PMID:36413997), BP4 requires REVEL <= 0.29 at supporting strength (<= 0.183 moderate, <= 0.016 strong). 0.542 is well above every BP4 cutoff, so BP4 is not met. The gray-zone REVEL result does not license reaching across to the SpliceAI path: the SpliceAI max delta of 0.01 (which would otherwise appear to satisfy the <= 0.1 SpliceAI BP4 cutoff) is not BP4 support for a missense variant, because a splice prediction was never calibrated to assess whether an amino-acid substitution is tolerated. BayesDel (0.365078) has no ACMG-strength or gene-specific calibration and is therefore not_available for BP4. No CDKN2A CSPEC/VCEP lookup table pre-assigns BP4, so the generic threshold governs.

Rule

ClinGen SVI REVEL calibration (Pejaver et al. 2022, PMID:36413997): BP4 supporting <= 0.29; BP4 not met above that value.

Evidence considered

• Variant consequence for path selection: VEP most_severe_consequence = missense_variant and pvs1_variant_assessment consequence_class = missense for NM_000077.5:c.369T>A (p.His123Gln), so the missense/REVEL path governs PP3/BP4 and the SpliceAI path does not apply.
• Local REVEL v1.3 predictor lookup for chr9:21970990 A>T (NM_000077.5:c.369T>A) returned score = 0.542, found = true.
• ClinGen SVI REVEL calibration thresholds (Pejaver et al. 2022, Am J Hum Genet, PMID:36413997): BP4 supporting <= 0.29, BP4 moderate <= 0.183, BP4 strong <= 0.016.
• REVEL 0.542 exceeds the supporting BP4 cutoff of 0.29, so BP4 is not met at any strength and the same gray-zone result that fails PP3 also fails BP4.
• SpliceAI max delta score 0.01 was deliberately not substituted for the REVEL path: the variant is missense, and a splice prediction is not valid BP4 support for a tolerated amino-acid substitution.
• BayesDel score 0.365078 is present but has no ACMG-strength or gene-specific calibration, so it is treated as not_available and is not used for BP4.
• Governing framework: no ClinGen CSPEC and no gene-specific/VCEP PP3/BP4 lookup table is available for CDKN2A (framework_mode = generic_acmg; final_classification_framework found = false), so generic ACMG/AMP computational-evidence thresholds apply, and no pre-assigned BP4 code exists.

Still missing

• No ClinGen CSPEC/VCEP gene-specific PP3/BP4 calibration exists for CDKN2A; generic ClinGen SVI REVEL thresholds were used.

Sources

Reasoning

BP6 requires that the exact variant be classified as Benign or Likely benign by an expert panel in ClinVar. ClinVar variation 127526 (exact variant NM_000077.5:c.369T>A, p.His123Gln) has review status 'criteria provided, single submitter' (1-star) with 0 expert-panel submissions across 16 records; its aggregate 'Likely benign' label is derived solely from ordinary single-submitter laboratory submissions (8 Likely benign, 3 Benign, 3 Uncertain significance, 1 likely benign). Per the rule applied here, ordinary laboratory submissions, aggregate ClinVar labels and non-expert assertions must never trigger BP6, and no ClinGen CDKN2A VCEP/CSPEC specification exists. BP6 is not met.

Rule

Generic ACMG/AMP 2015 criteria (PMID:25741868); BP6 applied only for an exact-variant ClinVar expert-panel Benign/Likely benign classification

Evidence considered

• ClinVar variation 127526 for the exact variant has review status 'criteria provided, single submitter' (1 star) and 0 expert-panel submissions across 16 records, so no expert-panel Benign/Likely benign classification exists.
• The ClinVar 'Likely benign' aggregate for variation 127526 is an aggregate of ordinary laboratory submissions (8 Likely benign, 3 Benign, 3 Uncertain significance, 1 likely benign), which is explicitly excluded from triggering BP6.
• PMID:25741868 (ACMG/AMP 2015) defines BP6; the governing rule restricts it to an exact-variant expert-panel Benign/Likely benign classification.

Sources

Reasoning

PS2 requires the variant to have arisen de novo with both maternity and paternity confirmed in a patient who has the disease and no family history of it. CDKN2A c.369T>A has no proband-level evidence of any kind: no source reports a tested individual's phenotype, parental genotypes, or a de novo occurrence. The ClinVar record (variation 127526) aggregates 16 submissions (14 usable, 5 with criterion leads) from clinical germline testing laboratories, but all condition labels are 'not provided', no expert-panel submission exists, and the only criterion signal any submitter asserted is PP3 (in-silico support); none asserted PS2. The one publication that names the variant (PMID:18573309) is an in-silico structural analysis of p16INK4A containing no patient, family, or genotype data. PS2 is applicable in principle to a germline variant in a cancer-predisposition gene such as CDKN2A, but with no trio or parental testing available the criterion cannot be evaluated in either direction.

Rule

PS2 (Strong): De novo (both maternity and paternity confirmed) in a patient with the disease and no family history.

Evidence considered

• All case sources were searched for proband-level information: no tested individual's phenotype, no parental genotypes, and no de novo occurrence of c.369T>A is reported anywhere.
• ClinVar (variation 127526) aggregates 16 submissions, 14 usable for classification and 5 usable for criterion leads, all 'criteria provided, single submitter'; none is an expert-panel submission, all condition labels are 'not provided', and the only criterion asserted by any submitter is PP3 (GeneDx in-silico statement). No submitter reported a de novo finding or parental testing.
• The ClinVar submission audit contains no PS2 signal; validated PMID trails behind those submissions concern panel/genotyping studies and in-silico analyses rather than trio or de novo observations.
• The only full text naming the variant (PMID:18573309) is an in-silico (PolyPhen/structural modelling) study of p16INK4A with no patient, family, or segregation data.
• CDKN2A germline loss of function is an established disease mechanism with a validated germline disease context, so PS2 is applicable in principle and is recorded as not assessed rather than not applicable; the gap is proband and parental genotype data.

Still missing

• Proband identity and phenotype (disease consistent with CDKN2A/FAMMM-spectrum disease) for the tested individual
• Parental genotypes (confirmed maternity and paternity) testing the variant in both parents
• Documentation of family history of CDKN2A-spectrum disease

Sources

Reasoning

PM3 requires the variant to be detected in trans with a pathogenic variant, a configuration that applies to genes with a recessive/compound-heterozygous disease mechanism. The required observation is entirely absent from every source available for this case rather than affirmatively excluded, so PM3 is not assessed. ClinVar variation 127526 (NM_000077.5:c.369T>A, p.His123Gln) aggregates submissions across the conditions Familial melanoma, Hereditary cancer-predisposing syndrome and Melanoma-pancreatic cancer syndrome; neither the overall record nor the five criterion-lead submissions with validated PMID trails reports a second CDKN2A variant, and no submission provides allele-phase information. The submission audit for this exact variant (16 submissions, 14 usable) contains no PM3 statement and no genotype or phasing data; the only criterion signal any submitter asserted is PP3 (in silico), and origin fields are limited to 'germline' or 'unknown'. No proband-level material exists: the case records no phenotype, family history or inheritance mode, and the one publication that names the variant (PMID:18573309) is an in silico structural analysis of p16INK4A with no patient, family or genotype data. The gene context also argues against a recessive reading: the retrieved literature describes CDKN2A p16 mutation carriers as high-risk individuals for surveillance irrespective of family history (PMID:31672839), consistent with the dominant, high-penetrance conditions recorded in ClinVar, and no source consulted describes a recessive or biallelic germline CDKN2A disorder. No CDKN2A CSPEC/VCEP or local gene-specific framework is available for this case, so the generic ACMG/AMP 2015 definition of PM3 (PMID:25741868) was applied, including its note that PM3 may be upgraded to strong only when there are multiple observations of the variant in trans with other pathogenic variants.

Rule

PM3 (Moderate): For recessive disorders, detected in trans with a pathogenic variant.

Evidence considered

• ClinVar variation 127526 (NM_000077.5(CDKN2A):c.369T>A, p.His123Gln) spans the conditions Familial melanoma, Hereditary cancer-predisposing syndrome, Melanoma-pancreatic cancer syndrome, 'not provided' and 'not specified'; no submission and no criterion-lead submission with a validated PMID trail reports a second CDKN2A variant or any allele-phase information. The only criterion signal extracted from any submitter's assertion is PP3 (in silico support); no submitter asserted PM3.
• The ClinVar submission audit for this exact variant records 16 submissions (14 usable for classification, 5 usable for criterion leads, 0 expert-panel submissions); it contains no PM3 statement, no co-occurring variant, and no phase or genotype data, with origin recorded only as 'germline' or 'unknown'.
• No proband-level evidence exists for this variant: no phenotype, family history, inheritance mode, parental genotype or segregation data are recorded, and the only publication that names the variant (PMID:18573309) is an in silico structural/functional analysis of p16INK4A (H123Q, rs6413463) containing no patient, family or genotype data.
• CDKN2A germline predisposition is described in the retrieved literature as a dominant carrier syndrome: PMID:31672839 classifies CDKN2A p16 (FAMMM) mutation carriers as high-risk individuals recommended for pancreatic surveillance irrespective of family history, matching the ClinVar conditions; no source consulted describes a recessive or biallelic germline CDKN2A disorder, which is the setting PM3's trans-with-a-pathogenic-variant configuration addresses.
• ACMG/AMP 2015 (PMID:25741868) defines PM3 as detection of the variant in trans with a pathogenic variant, and states it may be upgraded to strong with multiple observations of the variant in trans (on opposite chromosomes) with other pathogenic variants; no trans observation exists for c.369T>A.

Still missing

• No phased genotype, parental/family genotype, or second CDKN2A variant is available for c.369T>A in any case source; a proband-level genotype with phasing would be required to evaluate PM3.
• No CDKN2A CSPEC/VCEP or local gene-specific framework exists for this case, so the generic ACMG/AMP 2015 PM3 definition was used; a gene-specific specification could alter how PM3 is handled for an incompletely penetrant dominant cancer-predisposition gene.

Sources

Reasoning

PM6 covers a variant assumed to be de novo without confirmation of paternity and maternity. No source states that c.369T>A arose de novo in any individual, so even the assumption-based criterion cannot be applied. The only origin information available is that several clinical submissions in ClinVar record the variant's origin as germline, which documents germline sample testing but says nothing about whether the variant was inherited or newly arisen, and no parental genotypes are reported. As with PS2, CDKN2A germline disease context makes the criterion applicable in principle, but the underlying observation (a de novo occurrence) is entirely absent rather than merely unconfirmed.

Rule

PM6 (Moderate): Assumed de novo, but without confirmation of paternity and maternity.

Evidence considered

• Searched all case sources for any statement of a de novo occurrence: none exists for c.369T>A.
• ClinVar submission audit for variation 127526 records germline origin for several clinical submissions (e.g. PreventionGenetics, GeneDx), which confirms germline testing but provides no statement about de novo status and no parental results; no submitter asserted PM6 or PS2.
• No publication among those fetched (including the only one naming the variant, PMID:18573309) reports a de novo case or any family-based genotyping.
• PM6 is not converted to a benign/negative finding by this absence: with no de novo observation reported at all, the criterion is unevaluable.

Still missing

• Any report (published or in a clinical submission) of c.369T>A occurring de novo in an affected individual
• Parental testing status, even unconfirmed, for a proband carrying the variant
• Proband phenotype to establish that a de novo occurrence would be disease-relevant

Sources

Reasoning

PP1 requires co-segregation of the variant with disease in multiple affected members of a single family. No pedigree, relative genotypes, or co-segregation analysis for c.369T>A exists in any source: the case carries no family-level data, no paper in the literature pass was tagged for segregation, and no ClinVar submission reports family studies. The variant is present in the general population (gnomAD v2.1 AF 0.000422, 116/274982 alleles; popmax African/African American AF 0.00446) and in gnomAD v4.1 (AF 0.000223, 359/1610194, popmax AFR AF 0.00456), which is relevant to the population-frequency criteria but is not segregation evidence and was not treated as such. Because there are no genotyped affected and unaffected relatives, PP1 can be neither supported nor excluded.

Rule

PP1 (Supporting): Co-segregation with disease in multiple affected family members in a gene definitively known to cause the disease.

Evidence considered

• Searched every case source for pedigree or co-segregation content (family, affected relative, meiosis, carrier, inherited): no family-member genotypes or segregation analysis are reported for c.369T>A.
• No paper in the literature pass was tagged for segregation/PP1; the criteria index covers PP3, PP5, PS3 and PS4 only, and neither of the two germline cohort studies reviewed (PMID:27978560, PMID:28944238) reports family-based analysis of this variant (PMID:28944238 names only CDKN2A p.Ala127Ser and p.Arg144Cys; PMID:27978560 does not identify the CDKN2A variant at all).
• ClinVar variation 127526 has no expert-panel submission and none of the 16 submissions reports segregation data; the criterion-lead submissions cite panel-testing and in-silico literature.
• Population data (gnomAD v2.1 AF 0.000422, popmax AFR 0.00446; gnomAD v4.1 AF 0.000223, popmax AFR 0.00456) were noted only as context and were not used to substitute for segregation evidence.

Still missing

• A family pedigree linking genotype to phenotype for c.369T>A
• Genotypes of multiple affected relatives (and unaffected relatives) to test co-segregation
• Family history of CDKN2A-spectrum disease for the carrier(s) identified in clinical testing

Sources

Reasoning

PP4 requires that the patient's phenotype or family history be highly specific for a disease with a single genetic etiology. The case record contains no proband phenotype, no HPO terms, no clinical indication, and no family history: the review manifest, case summary, evidence bundle and ClinVar submission audit describe only the variant (NM_000077.5:c.369T>A, p.His123Gln) and the databases queried for it, with no patient-level clinical description. CDKN2A's disease associations (familial atypical multiple mole melanoma / familial melanoma, pancreatic cancer, as summarised at gene level in the gene context record and in the CDKN2A phenotype guidelines PMID:25645574 and PMID:31672839) are gene-level only and cannot substitute for patient-level phenotype specificity. With no clinical phenotype to compare against the disease spectrum, PP4 cannot be evaluated and is not assessed.

Rule

Generic ACMG/AMP 2015 criteria (PMID:25741868); no CDKN2A-specific VCEP/CSPEC specification available

Evidence considered

• The case record is a variant-level interpretation: review_manifest.json, case_summary.json, evidence.json and clinvar_submission_audit.json describe the variant, gene and queried databases but contain no proband phenotype, HPO terms, indication for testing, or family history.
• PMID:25645574 (ACG hereditary GI cancer guideline) names CDKN2A only as a gene to include when testing for suspected familial pancreatic cancer; it provides disease-level context but no patient-level phenotype for this variant.
• PMID:31672839 (International CAPS Consortium) treats germline CDKN2A carriers as high-risk for pancreatic surveillance; again gene-level management context only, with the exact variant not named.
• Gene-level CDKN2A context (p16/p14ARF tumour suppressor; familial melanoma and pancreatic cancer predisposition) does not establish patient-level phenotype specificity, which is what PP4 requires.

Still missing

• No proband phenotype, HPO terms, clinical indication, or family history is present anywhere in the case record; PP4 requires patient-level clinical information that is absent here.
• If a patient phenotype (e.g. familial melanoma / FAMMM or pancreatic cancer spectrum) is supplied by the referring clinician, PP4 should be re-evaluated against its specificity for CDKN2A.

Sources

Reasoning

BS4 requires a demonstrated lack of segregation of the variant with disease in affected members of a family. No family study, pedigree, or relative genotype data exist for c.369T>A, so non-segregation cannot be shown. Two potentially tempting non-segregation-adjacent observations were deliberately not used to satisfy BS4: the ClinVar aggregate classification for the variant is Likely benign across multiple submitters, but a source's final classification is not segregation evidence and cannot be borrowed; and gnomAD v4.1 contains a homozygote (AF 0.000223, 359/1610194 alleles; popmax AFR AF 0.00456, 342/75052 with 1 homozygote), which bears on population-frequency criteria rather than on within-family segregation. With no genotyped affected and unaffected relatives, BS4 is unevaluable in either direction.

Rule

BS4 (Strong): Lack of segregation in affected members of a family.

Evidence considered

• Searched all case sources for family studies capable of demonstrating non-segregation: none were found, and no ClinVar submitter reported segregation or non-segregation data.
• The ClinVar aggregate classification of variation 127526 (Likely benign/Benign across submitters) was reviewed but explicitly not used to infer BS4, because BS4 requires within-family genotype evidence rather than another source's classification.
• gnomAD v4.1 reports the variant at AF 0.000223 (359/1610194 alleles) with one homozygote and popmax African/African American AF 0.00456 (342/75052, 1 homozygote); this is population-frequency context relevant to population criteria, not evidence of non-segregation.
• No fetched publication (including the only one naming the variant, PMID:18573309) reports family genotyping that could support or refute segregation.

Still missing

• Genotypes of affected and unaffected relatives in at least one family informative for non-segregation
• Pedigree documentation separating variant carriers from affected non-carriers
• Phenotype definition per family to determine whether absence of co-segregation would be interpretable

Sources

Reasoning

BP2 requires the variant to be observed in trans with a pathogenic variant for a fully penetrant dominant gene/disorder, or in cis with a pathogenic variant in any inheritance pattern. Neither configuration is documented anywhere in this case, and the criterion therefore cannot be evaluated in either direction. The ClinVar record (variation 127526) and its submission audit contain no co-occurring CDKN2A variant, no phase information and no BP2 assertion; the only criterion signal any submitter asserted is PP3 (in silico). None of the retrieved publications places c.369T>A, or any CDKN2A variant, in cis or in trans with a pathogenic variant: the only paper naming the variant (PMID:18573309) is an in silico structural analysis without genotypes, and the cohort study that reports a pathogenic CDKN2A mutation in one patient (PMID:27978560) reports a single mutation with no second variant and no phasing. Population genotype data do not substitute for the required observation: gnomAD v4.1 records 359/1,610,194 alleles (AF 0.000223) including one homozygote (popmax African/African American 342/75,052, AF 0.004557, 1 homozygote) and gnomAD v2.1 records 116/274,982 alleles with no homozygote; a population homozygote is not a pathogenic variant on the opposite or same allele and is not BP2 evidence, such homozygote observations being the remit of the population-frequency criteria. No CDKN2A CSPEC/VCEP or local gene-specific framework is available, so the generic ACMG/AMP 2015 BP2 definition (PMID:25741868) was applied. With no cis/trans partner variant identified, BP2 is not assessed rather than affirmatively excluded.

Rule

BP2 (Supporting): Observed in trans with a pathogenic variant for a fully penetrant dominant gene/disorder, or observed in cis with a pathogenic variant in any inheritance pattern.

Evidence considered

• ACMG/AMP 2015 (PMID:25741868) defines BP2 as observed in trans with a pathogenic variant for a fully penetrant dominant gene/disorder, or observed in cis with a pathogenic variant in any inheritance pattern.
• No source reports a co-occurring pathogenic CDKN2A variant in cis or in trans with c.369T>A: the ClinVar record (variation 127526, conditions Familial melanoma / Hereditary cancer-predisposing syndrome / Melanoma-pancreatic cancer syndrome) and its submission audit (16 submissions, 14 usable, 0 expert-panel) contain no co-occurring variant, no phasing, and no BP2 assertion; the only criterion signal any submitter asserted is PP3 (in silico).
• The only publication naming the variant (PMID:18573309) is an in silico study of H123Q with no patient or family genotypes; the cohort study (PMID:27978560) reporting a pathogenic CDKN2A mutation in 1 of 450 early-onset colorectal cancer patients reports a single mutation with no second CDKN2A variant and no allele-phase data.
• gnomAD v4.1 genotype counts for this variant are 359/1,610,194 alleles (AF 0.000223) with 1 homozygote, popmax African/African American 342/75,052 (AF 0.004557, 1 homozygote); gnomAD v2.1 is 116/274,982 alleles (AF 0.000422) with 0 homozygotes. These document population carriers only, not a pathogenic variant on the opposite or same allele, so they do not satisfy BP2.

Still missing

• No phase-resolved genotype (cis/trans) and no second CDKN2A variant were available for this variant in any source, so neither BP2 configuration could be tested.
• PMID:28492532 (Sherloc, a refinement of the ACMG/AMP criteria) appears in the ClinVar citation trail for this variant, but no full text or extraction was retrieved, so any refinement of BP2 usage could not be checked.
• No CDKN2A CSPEC/VCEP or local gene-specific framework exists for this case, so the generic ACMG/AMP 2015 BP2 wording was applied.

Sources

Reasoning

BP5 applies when a variant is found in a case in which there is an alternate molecular basis for the disease (e.g. a second pathogenic variant or a distinct genetic diagnosis explaining the phenotype). The case record contains no case-level genotype or phenotype data at all - no proband, no co-inherited/alternative pathogenic variant, and no alternate clinical diagnosis - and no retrieved publication reports c.369T>A in a patient with a competing molecular explanation. Without a described case carrying this variant, there is nothing in which an alternate molecular basis could be observed, so BP5 cannot be evaluated and is not assessed.

Rule

Generic ACMG/AMP 2015 criteria (PMID:25741868); no CDKN2A-specific VCEP/CSPEC specification available

Evidence considered

• No case or pedigree is described in the case record; the case summary, evidence bundle and ClinVar submission audit for variation 127526 contain only variant, gene and database-level information, with no alternate molecular cause reported.
• No retrieved publication reports the exact variant c.369T>A (p.His123Gln) in a patient; the only variant-naming paper (PMID:18573309) is an in silico study with no patient, so no case exists in which an alternate molecular basis could be documented.

Still missing

• No case-level genotype/phenotype data (proband, co-occurring variant, or alternate diagnosis) are present in the case record; BP5 requires a specific case in which this variant co-occurs with an alternate explanation for disease.