BAP1 c.67+5G>C, p.?

Reasoning

Generic ACMG/AMP is applicable because no BAP1-specific VCEP or gene framework was found. PM2 Supporting is met because the variant is rare in the default all-comers population sources: gnomAD v2.1 exomes reports AF 1.99103e-05 (5/251126 alleles, zero homozygotes), and gnomAD v4.1 reports AF 1.48696e-05 (24/1614030 alleles, zero homozygotes). Both are below the supplied PM2 Supporting threshold of <=0.0001. The highest ancestry-specific frequencies also remain below that threshold, including 4.40498e-05 in gnomAD v2.1 European non-Finnish and 2.03384e-05 in gnomAD v4.1 European non-Finnish.

Rule

PM2 is Supporting when allele frequency is <=0.0001 under the supplied ClinGen SVI recommendation.

Evidence considered

• gnomAD v2.1 exomes reports 5/251126 alleles, AF 1.99103e-05, and zero homozygotes; gnomAD v4.1 reports 24/1614030 alleles, AF 1.48696e-05, and zero homozygotes.
• The highest ancestry-specific frequencies are 4.40498e-05 in gnomAD v2.1 European non-Finnish and 2.03384e-05 in gnomAD v4.1 European non-Finnish, both below 0.0001.
• The supplied ClinGen SVI recommendation downgrades PM2 to Supporting at allele frequency <=0.0001 (PMID:25741868).
• No governing BAP1-specific VCEP or gene framework was available, so the generic all-comers population source is used rather than a non-cancer subset.

Sources

Reasoning

PVS1 is not met. NM_004656.4:c.67+5G>C is an intronic +5 variant, not a nonsense, frameshift, canonical ±1,2 splice-site, or other established null-variant consequence. VariantValidator places it in exon 2i, while the available annotation gives NP_004647.1:p.? and does not establish exon skipping, NMD, a protein-length change, or loss of function. SpliceAI predicts a maximum delta of 0.161, below the generic supporting splice-impact threshold of 0.2 from Jaganathan et al. 2019 (PMID:30661751); this prediction alone does not establish a PVS1 null effect. No VCEP-specific framework is available, so the generic ClinGen SVI PVS1 framework (PMC6185798) applies.

Rule

generic ClinGen SVI PVS1 framework; no VCEP-specific specification available

Evidence considered

• NM_004656.4:c.67+5G>C is intronic and lies outside the canonical ±1,2 splice consensus; the available annotation reports NP_004647.1:p.? and no established protein consequence.
• VariantValidator identifies the affected region as exon 2i but does not provide a demonstrated transcript or protein consequence.
• SpliceAI reports DS_DL 0.161 as the maximum delta score; the generic supporting splice-impact threshold is 0.2 (Jaganathan et al. 2019, PMID:30661751).
• The generic ClinGen SVI PVS1 recommendations require evidence that a variant is a null or loss-of-function allele before applying PVS1; general ACMG guidance states that splice-site effects require RNA or protein confirmation when the consequence is not established (PMID:25741868).

Still missing

• No RNA or protein assay demonstrates exon skipping, aberrant splicing, NMD, or loss of function.
• The precise transcript consequence and protein effect remain unresolved.

Sources

Reasoning

PP3 is applicable because NM_004656.4:c.67+5G>C is an intronic splice-region variant, so SpliceAI is the sole permitted computational path. The maximum SpliceAI delta score is 0.161, which is below the generic supporting PP3 threshold of >=0.2 calibrated by Jaganathan et al. 2019 (PMID:30661751); therefore PP3 is not met. No BAP1-specific VCEP framework or pre-assigned PP3 rule is available.

Evidence considered

• The variant is intronic at c.67+5, so SpliceAI is the applicable and exclusive PP3/BP4 predictor path; REVEL is not applicable.
• SpliceAI maximum delta score is 0.161 (DS_DL 0.161), below the >=0.2 supporting PP3 threshold from the SpliceAI calibration of Jaganathan et al. 2019 (PMID:30661751).
• No applicable BAP1-specific VCEP or gene-specific computational criterion assignment was identified; generic calibration therefore applies.

Sources

Reasoning

PP5 is restricted here to an exact-variant ClinVar expert-panel classification of Pathogenic or Likely pathogenic. The exact ClinVar record NM_004656.4(BAP1):c.67+5G>C has 0 expert-panel submissions; its classifications are uncertain significance and likely benign from ordinary laboratory submissions. Therefore PP5 is not met.

Rule

Apply PP5 only when the exact variant has a ClinVar expert-panel Pathogenic or Likely pathogenic classification; ordinary laboratory submissions, aggregate labels, and non-expert assertions do not qualify.

Evidence considered

• The exact ClinVar variation 489641 has review status criteria provided, conflicting classifications, with 0 expert-panel submissions.
• The exact-variant submissions are Uncertain significance or Likely benign from individual clinical laboratories; ordinary submissions and aggregate labels are excluded from PP5.

Sources

Reasoning

Generic ACMG/AMP is applicable because no BAP1-specific VCEP or gene framework was found. BA1 requires an allele frequency of at least 0.05 under the supplied ACMG/AMP 2015 threshold. The highest relevant all-comers frequency is 1.99103e-05 in gnomAD v2.1 exomes, with 5/251126 alleles and zero homozygotes; gnomAD v4.1 is similarly low at 1.48696e-05 (24/1614030 alleles, zero homozygotes). Neither frequency approaches 0.05, so BA1 is not met.

Rule

BA1 requires allele frequency >=0.05 under the generic ACMG/AMP 2015 calibration.

Evidence considered

• gnomAD v2.1 exomes reports 5/251126 alleles, AF 1.99103e-05, and zero homozygotes; gnomAD v4.1 reports 24/1614030 alleles, AF 1.48696e-05, and zero homozygotes.
• The supplied generic ACMG/AMP 2015 BA1 calibration is allele frequency >=0.05 (PMID:25741868); the observed frequencies are well below this cutoff.
• No governing BAP1-specific VCEP or gene framework was available, so the generic all-comers population source is used rather than a non-cancer subset.

Sources

Reasoning

Generic ACMG/AMP is applicable because no BAP1-specific VCEP or gene framework was found. The supplied generic BS1 threshold is allele frequency >=0.01. The highest relevant all-comers frequency is 1.99103e-05 in gnomAD v2.1 exomes, and gnomAD v4.1 reports 1.48696e-05; both are substantially below 0.01. The highest observed ancestry-specific values also remain below the threshold (4.40498e-05 in gnomAD v2.1 European non-Finnish and 2.03384e-05 in gnomAD v4.1 European non-Finnish), so BS1 is not met.

Rule

BS1 requires allele frequency >=0.01 under the supplied generic SVI default.

Evidence considered

• gnomAD v2.1 exomes reports AF 1.99103e-05 overall and 4.40498e-05 in the European non-Finnish subgroup; gnomAD v4.1 reports AF 1.48696e-05 overall and 2.03384e-05 in the European non-Finnish subgroup.
• The supplied generic BS1 calibration is allele frequency >=0.01, identified as the generic SVI default with per-gene derivation preferred (Whiffin et al. 2017, PMID:28518168); all observed frequencies are below this cutoff.
• No governing BAP1-specific VCEP or gene framework was available, so the generic all-comers population source is used rather than a non-cancer subset.

Sources

Reasoning

BP4 is applicable because NM_004656.4:c.67+5G>C is an intronic splice-region variant, so SpliceAI is the sole permitted computational path. The maximum SpliceAI delta score is 0.161, which exceeds the generic supporting BP4 threshold of <=0.1 calibrated by Jaganathan et al. 2019 (PMID:30661751); therefore BP4 is not met. No BAP1-specific VCEP framework or pre-assigned BP4 rule is available.

Evidence considered

• The variant is intronic at c.67+5, so SpliceAI is the applicable and exclusive PP3/BP4 predictor path; REVEL is not applicable.
• SpliceAI maximum delta score is 0.161 (DS_DL 0.161), above the <=0.1 supporting BP4 threshold from the SpliceAI calibration of Jaganathan et al. 2019 (PMID:30661751).
• No applicable BAP1-specific VCEP or gene-specific computational criterion assignment was identified; generic calibration therefore applies.

Sources

Reasoning

BP6 is restricted here to an exact-variant ClinVar expert-panel classification of Benign or Likely benign. The exact ClinVar record NM_004656.4(BAP1):c.67+5G>C has 0 expert-panel submissions; the Likely benign classifications are ordinary laboratory submissions. Therefore BP6 is not met.

Rule

Apply BP6 only when the exact variant has a ClinVar expert-panel Benign or Likely benign classification; ordinary laboratory submissions, aggregate labels, and non-expert assertions do not qualify.

Evidence considered

• The exact ClinVar variation 489641 has 0 expert-panel submissions.
• Two exact-variant laboratory submissions are Likely benign, but non-expert laboratory assertions and aggregate ClinVar labels are explicitly excluded from BP6.

Sources

Reasoning

No case-level evidence documents a de novo occurrence of NM_004656.4:c.67+5G>C in an affected proband. The available records contain no proband-parent genotypes, parental testing results, confirmation of maternity and paternity, or information about parental mosaicism. Therefore, confirmed de novo evidence required for PS2 is unavailable and PS2 cannot be applied.

Rule

Generic ACMG/AMP PS2 requires a confirmed de novo variant in an affected proband with confirmed parentage; absent such case-level evidence, do not apply PS2.

Evidence considered

• The case-level evidence contains no documented de novo observation, parental testing result, trio genotype, or parentage confirmation for NM_004656.4:c.67+5G>C.
• The governing framework record reports that no BAP1-specific VCEP/CSPEC specification is available; generic ACMG/AMP criteria are therefore the applicable fallback.

Still missing

• Proband and both biological parents' variant genotypes are not documented.
• Maternity and paternity confirmation is not documented.
• Parental mosaicism assessment is not documented.

Reasoning

No validated variant-specific functional assay was identified for NM_004656.4:c.67+5G>C. The available evidence includes only computational splice prediction, with SpliceAI max delta 0.161, which is not an RNA or protein functional study and cannot establish abnormal splicing or loss of BAP1 function for PS3. The ACMG/AMP guidance states that splice effects require confirmation by RNA or protein analysis, and the reviewed literature specifically notes that most predicted +3 to +5 intronic effects have not been experimentally tested. Because no assay result, assay controls, biological-relevance validation, or calibrated functional-strength rule is available, PS3 cannot be assigned.

Rule

Generic ACMG/AMP PS3 framework; no applicable BAP1-specific VCEP specification was available. Variant-specific well-established functional evidence is required; computational splice prediction alone is insufficient.

Evidence considered

• SpliceAI reports a maximum delta score of 0.161 for NM_004656.4:c.67+5G>C; this is an in-silico prediction and does not constitute functional assay evidence for PS3.
• The ACMG/AMP guidance states: "Although splice site variants are predicted to lead to a null effect, confirmation of impact requires functional analysis by either RNA or protein analysis."
• A review of variant pathogenicity assessments reports that variants predicted to affect splicing at intronic positions +3 to +5 were largely not experimentally tested, with no BAP1-specific assay reported for this variant.

Still missing

• Variant-specific RNA assay demonstrating abnormal BAP1 splicing or transcript consequence is unavailable.
• Variant-specific protein or cellular assay demonstrating impaired BAP1 function is unavailable.
• Assay controls and biological-relevance validation for this variant are unavailable.

Sources

Reasoning

No case-control enrichment analysis, affected-carrier count, control-carrier count, odds ratio, or statistically significant association was identified for the exact variant NM_004656.4:c.67+5G>C. The available literature extraction contains no exact-variant clinical observation, and the ClinVar record is not a case-control dataset. PS4 therefore cannot be assessed from the available evidence.

Rule

Generic ACMG/AMP PS4 requires a statistically significant enrichment of the variant in affected individuals compared with appropriate controls; no applicable BAP1-specific specification was available.

Evidence considered

• The case evidence contains no exact-variant case-control counts, odds ratio, likelihood ratio, or statistical enrichment result.
• The extracted literature findings for PMID:25394175 and PMID:26467025 do not report this BAP1 variant or a relevant BAP1 cohort result.

Still missing

• A validated case-control dataset or quantitative enrichment analysis for NM_004656.4:c.67+5G>C is needed.
• Exact-variant affected and control carrier counts are unavailable.

Sources

Reasoning

No case-level evidence documents that NM_004656.4:c.67+5G>C arose de novo without parental testing. There is no report of a clinically affected proband with an apparently de novo variant, no parental genotype information, and no assessment of parentage or parental mosaicism. Because the minimum observation for PM6 is not documented, PM6 cannot be applied.

Rule

Generic ACMG/AMP PM6 requires an apparently de novo variant in an affected individual when parental testing is unavailable; no such case-level observation is available here.

Evidence considered

• The available case evidence provides no affected-proband report, family description, or apparently de novo statement for NM_004656.4:c.67+5G>C.
• The available literature extractions do not mention BAP1 or this exact variant in a segregation or de novo context.

Still missing

• A case report or clinical record identifying an affected proband with this exact variant is not documented.
• Parental testing or an explicit apparently de novo statement is not documented.
• Parentage and parental mosaicism information is not documented.

Reasoning

No family segregation data are available for NM_004656.4:c.67+5G>C. The records do not identify affected relatives carrying the variant, unaffected relatives lacking it, the number of informative meioses, phenotype status of relatives, or any statistically interpretable segregation series. Without informative affected-relative transmission, PP1 cannot be applied at supporting or increased strength.

Rule

Generic ACMG/AMP PP1 requires co-segregation of the variant with disease in multiple affected family members; strength cannot be assigned without documented informative meioses and phenotype/genotype data.

Evidence considered

• The case evidence contains no pedigree, familial genotype data, affected-relative transmission, or informative meioses for NM_004656.4:c.67+5G>C.
• No phenotype-consistent family series is documented that could support PP1.

Still missing

• No affected relatives with genotype and phenotype data are documented.
• No informative meioses or segregation count is documented.
• No family pedigree or phenotype-consistency assessment is documented.

Reasoning

No phenotype, clinical presentation, or phenotype-specific individual-level record is available for a carrier of NM_004656.4:c.67+5G>C. Although BAP1 tumor predisposition syndrome is represented in the ClinVar condition labels, those labels do not establish that the exact variant occurs in an individual with a highly specific phenotype. PP4 cannot be assessed.

Rule

Generic ACMG/AMP PP4 requires a phenotype or family history highly specific for a disease with a single-gene etiology; no applicable BAP1-specific specification was available.

Evidence considered

• ClinVar lists BAP1-related tumor predisposition syndrome among condition labels, but provides no exact-variant carrier phenotype or phenotype-specific clinical observation.
• The extracted literature findings contain no exact-variant clinical phenotype for NM_004656.4:c.67+5G>C.

Still missing

• A clinical record for an exact-variant carrier with a disease-specific phenotype is needed.
• The phenotype must be sufficiently specific for BAP1-related disease rather than inferred from the ClinVar condition label.

Sources

Reasoning

BS2 requires the variant to be observed in healthy individuals in a manner inconsistent with the disease model. The available population data report zero homozygotes in gnomAD v2.1 and v4.1, but do not provide phenotype-confirmed unaffected carrier observations, penetrance-adjusted analysis, or a disease-prevalence-based BS2 rule. Therefore the absence of homozygotes does not establish BS2, and the criterion cannot be assessed from the available evidence.

Rule

BS2 requires observation in healthy individuals inconsistent with the disease model; population allele counts alone are insufficient without phenotype or penetrance context.

Evidence considered

• gnomAD v2.1 reports zero homozygotes and gnomAD v4.1 reports zero homozygotes for the variant.
• The available population records do not identify individual carrier phenotypes, establish unaffected status, or provide penetrance and disease-prevalence calibration for applying BS2.

Still missing

• No phenotype-confirmed unaffected carriers or disease-model-specific BS2 threshold were available.

Sources

Reasoning

No validated variant-specific functional assay was identified showing normal BAP1 splicing or preserved BAP1 activity for NM_004656.4:c.67+5G>C. SpliceAI max delta 0.161 is a computational prediction and cannot establish a normal transcript or protein phenotype, particularly for an intronic +5 variant. The reviewed ACMG/AMP guidance requires RNA or protein analysis to confirm splice effects, while the reviewed scoring literature indicates that most predicted +3 to +5 intronic splice effects lack experimental testing. Without a validated benign assay, appropriate controls, and demonstrated biological relevance, BS3 cannot be assigned.

Rule

Generic ACMG/AMP BS3 framework; no applicable BAP1-specific VCEP specification was available. Variant-specific well-established functional evidence demonstrating no damaging effect is required; computational splice prediction alone is insufficient.

Evidence considered

• SpliceAI reports a maximum delta score of 0.161 for NM_004656.4:c.67+5G>C; this computational result cannot demonstrate normal splicing or preserved protein function for BS3.
• The ACMG/AMP guidance states that the impact of splice-site variants requires confirmation by functional analysis of RNA or protein, which is not available here.
• The reviewed scoring literature states that most predicted splice-affecting variants in the +3 to +5 intronic range have not been experimentally tested, so prediction alone cannot support a benign functional criterion.

Still missing

• Variant-specific RNA assay demonstrating normal BAP1 splicing is unavailable.
• Variant-specific protein or cellular assay demonstrating preserved BAP1 function is unavailable.
• Assay controls and biological-relevance validation supporting a benign functional conclusion are unavailable.

Sources

Reasoning

No non-segregation study is documented for NM_004656.4:c.67+5G>C. The available records do not provide affected relatives who lack the variant, unaffected relatives who carry it, a pedigree with reliable phenotyping, or an informative family structure sufficient to demonstrate failure of segregation. BS4 therefore cannot be applied.

Rule

Generic ACMG/AMP BS4 requires well-established lack of segregation with disease in affected family members; no informative non-segregation evidence is available here.

Evidence considered

• The case evidence contains no affected relative reported to lack NM_004656.4:c.67+5G>C and no other informative genotype-phenotype discordance.
• No family-based non-segregation analysis is documented.

Still missing

• No affected relatives with negative variant testing are documented.
• No unaffected carriers or other discordant phenotype/genotype observations are documented.
• No reliable family phenotyping and genotype-informed pedigree are documented.

Reasoning

No affected-proband genotype, family testing, read-backed phasing, or other reliable evidence is provided showing NM_004656.4:c.67+5G>C in cis with a pathogenic BAP1 variant. The available ClinVar record reports conflicting classifications for the assessed variant but provides no phase or co-occurrence observation, and the reviewed papers do not report this variant or relevant allelic data. BP2 therefore cannot be assessed.

Rule

Generic ACMG/AMP BP2 for a dominant disorder requires the variant to be observed in cis with a pathogenic variant; phase and the pathogenicity of the partner variant must be established.

Evidence considered

• ClinVar variation 489641 records NM_004656.4(BAP1):c.67+5G>C with conflicting submissions, but no phase, cis/trans, affected-proband, or co-occurrence data are provided.
• The available paper extracts do not mention BAP1 c.67+5G>C and provide no relevant allelic observations.
• No family segregation, parental testing, read-backed phasing, or second pathogenic BAP1 variant is documented in the case evidence.

Still missing

• A phased genotype or family-based test showing this variant in cis with a pathogenic BAP1 variant is needed to evaluate BP2.
• Affected-proband and inheritance information for any co-occurring BAP1 variant is unavailable.

Sources

Reasoning

No affected individual carrying NM_004656.4:c.67+5G>C is documented, and no alternative pathogenic variant or molecular diagnosis is reported in an affected individual. The available ClinVar submissions do not provide the individual-level clinical and molecular context required to evaluate BP5. BP5 cannot be assessed.

Rule

Generic ACMG/AMP BP5 requires an alternate molecular basis for the affected individual's phenotype; no applicable BAP1-specific specification was available.

Evidence considered

• The exact ClinVar record contains laboratory classifications but no affected-individual case description pairing this variant with an alternative molecular diagnosis.

Still missing

• An affected exact-variant carrier with documented phenotype and an established alternative molecular diagnosis is needed.

Sources