RB1 c.275_282del, p.(Ile92LysfsTer15)

Reasoning

PM2 (absent/extremely rare in population databases) applies at supporting strength per generic ACMG/AMP. No RB1-specific VCEP or non-cancer/exome-only population-source specification was identified for this case (cspec.found=false, framework_mode=generic_acmg), so the default all-comers gnomAD v2.1 and v4.1 entries were used as instructed. The variant NM_000321.2:c.275_282del is absent from gnomAD v2.1 exomes, gnomAD v4.1, the GNOMAD_V2_1_NON_CANCER and GNOMAD_V3_1_NON_CANCER subsets, and gnomAD-Canada v1.0 (all search_status=absent, found=false), consistent with absence from population databases.

Rule

Generic ACMG/AMP PM2: absent or extremely low frequency in population databases (gnomAD).

Evidence considered

• gnomAD v2.1 exome direct URL lookup (13-48916744-ATTCAAAAG-A) returned search_status=absent, found=false, evidence_sentence='Absent from gnomAD v2.1.'
• gnomAD v4.1 direct URL lookup (chr13-48342608-ATTCAAAAG-A) returned search_status=absent, found=false, evidence_sentence='Absent from gnomAD v4.1.'
• GNOMAD_V2_1_NON_CANCER and GNOMAD_V3_1_NON_CANCER subset lookups at the same coordinates also returned absent, confirming absence irrespective of which population source (all-comers vs non-cancer) is applicable.
• gnomAD-Canada v1.0 (HostSeq genomes) lookup also returned absent.

Sources

Reasoning

No ClinGen VCEP or gene-specific expert panel specification was found for RB1 (framework_mode is generic_acmg with cspec.found=false), so generic ACMG/AMP thresholds apply using the default all-comers gnomAD totals rather than any non-cancer/exome-only subset. NM_000321.2:c.275_282del (NC_000013.10:g.48916745_48916752del) is absent from gnomAD v2.1 exomes and gnomAD v4.1, as well as from the GNOMAD_V2_1_NON_CANCER and GNOMAD_V3_1_NON_CANCER subset lookups and gnomAD-Canada v1.0. An absent variant has an effective population allele frequency of 0, far below any BA1 stand-alone benign threshold (>=5% generic, or the RB1-specific >1% used elsewhere in this group), so BA1 is not met.

Rule

Generic ACMG/AMP BA1: absent from population databases, allele frequency 0 vs BA1 threshold.

Evidence considered

• gnomAD v2.1 exome direct URL lookup (13-48916744-ATTCAAAAG-A) returned search_status=absent, found=false.
• gnomAD v4.1 direct URL lookup (chr13-48342608-ATTCAAAAG-A) returned search_status=absent, found=false.
• GNOMAD_V2_1_NON_CANCER and GNOMAD_V3_1_NON_CANCER subset queries at the same coordinates also returned absent, so the population-source choice (all-comers vs non-cancer) does not change the outcome.
• No governing VCEP/CSPEC was located for RB1 in this case (cspec.found=false, framework_mode=generic_acmg), so the default all-comers gnomAD entries were used per instructions.

Sources

Reasoning

Using the same default all-comers gnomAD v2.1/v4.1 population source (no RB1 VCEP non-cancer/exome-only specification is available), the variant is absent from all queried gnomAD releases and subsets, giving an allele frequency of 0. This is well below any BS1 threshold set relative to retinoblastoma birth prevalence (roughly 1 in 15,000-20,000 live births per the RB1 literature reviewed for PVS1 gene context), so BS1 is not met.

Rule

Generic ACMG/AMP BS1: allele frequency greater than expected for disorder; not met because observed AF is 0.

Evidence considered

• gnomAD v2.1 and v4.1, plus non-cancer subsets, all report the variant as absent (search_status=absent, found=false).
• RB1-associated retinoblastoma birth prevalence cited in reviewed literature (PMID:41764457) as approximately 1 in 15,000-20,000 live births, used only as background context; frequency comparison is moot since observed AF is 0.

Sources

Reasoning

BS2 requires observation of the variant in homozygous, hemizygous, or compound heterozygous state in individuals unaffected by the fully penetrant disorder, at an age past typical retinoblastoma onset. The variant is completely absent from every population database queried (gnomAD v2.1, gnomAD v4.1, both non-cancer subsets, and gnomAD-Canada), so there is no homozygote/carrier data of any kind, healthy or affected, to evaluate. No evidence supports BS2.

Rule

Generic ACMG/AMP BS2: observed in healthy adult homozygotes/hemizygotes for a fully penetrant early-onset disorder; not met, no such observations exist.

Evidence considered

• gnomAD v2.1, v4.1, and both non-cancer subsets show zero allele count / absent status, meaning no homozygote or unaffected-carrier data exists to invoke BS2.

Still missing

• No individual-level genotype or homozygote data available since the variant is not present in any queried population database.

Sources

Still missing

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

Reasoning

The case evidence bundle contains no proband/family clinical data whatsoever: no trio testing, no parental genotyping results, and no statement of confirmed or presumed de novo occurrence. No CSPEC/VCEP was found for RB1 (framework_mode: generic_acmg), and no gene-specific de novo scoring table is available. Available sources are limited to population databases (gnomAD v2.1/v4.1, both absent), SpliceAI, OncoKB, COSMIC, and cancer hotspot data, none of which speak to parentage or family segregation. Without any de novo confirmation data (paternity/maternity confirmed status, number of independent occurrences, phenotype consistency), PS2 cannot be assessed.

Still missing

• No parental testing results (confirmed/assumed paternity and maternity status) provided.
• No statement of de novo occurrence or number of independent de novo observations.
• No phenotype documentation for the proband to establish phenotype consistency.

Reasoning

No validated functional/splicing assay data specific to NM_000321.2:c.275_282del (p.(Ile92LysfsTer15)) was located. The five ONCOKB-sourced RB1 papers retrieved (PMID:14769601, PMID:22205104, PMID:26607597, PMID:30206110, PMID:31138663) were searched for this exact variant designation (c.275_282del, Ile92, I92Kfs) and none of them mention it; they describe protein truncation testing methodology, second-malignancy cohort outcomes, murine Rb1 knockout models, PALOMA-3 CDK4/6i resistance genomics, and generic RB pathway DNA-damage biology, none of which constitute a validated functional assay of this specific variant's protein product or splicing behavior. No RB1 VCEP-specific PS3 functional assay framework (e.g., pRb phosphorylation/binding assay thresholds) was available in this case (no CSPEC/VCEP materials found; framework_mode is generic_acmg fallback). Absent variant-specific assay data and an applicable calibrated framework, PS3 cannot be scored and is flagged for human review in case a functional study surfaces on further literature search.

Evidence considered

• Searched PMID:14769601, PMID:22205104, PMID:26607597, PMID:30206110, PMID:31138663 full texts for exact variant strings 'c.275_282del', 'Ile92', 'I92Kfs' - no matches found in any paper.
• No CSPEC/VCEP RB1 framework was available in this case (vcep_materials.json: found=false, framework_mode=generic_acmg fallback), so no gene-specific PS3 assay-validation criteria could be applied.

Still missing

• No variant-specific functional/splicing assay (e.g., minigene, pRb binding/phosphorylation, RNA analysis) identified for c.275_282del.
• No RB1 VCEP or gene-specific PS3 framework available to define assay validation/control requirements.
• Retrieved literature set is gene-level/pathway-level (PTT methodology, cohort outcomes, mouse models, CDK4/6i resistance), not variant-specific.

Sources

Reasoning

No case-control or cohort enrichment data (e.g., odds ratio, case counts vs. controls) specific to NM_000321.2:c.275_282del were located. The literature bundle contains only functional/mechanistic RB1 papers (PTT methodology, second-malignancy risk cohort, mouse Rb1 models, PALOMA-3 resistance, DNA damage studies) and none report proband counts or enrichment statistics for this exact variant. No CSPEC/VCEP RB1 framework was found to define a gene-specific PS4 threshold, so generic ACMG PS4 (statistically significant case-control enrichment, typically OR meeting a pre-specified threshold) cannot be evaluated for lack of any quantitative case observation data tied to this specific allele.

Evidence considered

• Five OncoKB-linked PMIDs retrieved (14769601, 22205104, 26607597, 30206110, 31138663) are gene-level functional/mechanistic or unrelated resistance studies, none reporting variant-specific case-control enrichment for c.275_282del.

Still missing

• No case-control cohort or proband count data specific to c.275_282del was identified in the retrieved literature.
• No RB1 VCEP/CSPEC PS4 threshold available to apply.

Reasoning

PM6 requires de novo occurrence without confirmed parentage (i.e., assumed rather than confirmed de novo). No clinical family or trio data of any kind is present in the bundle, and there is no mention of assumed de novo status for this proband. The available evidence is restricted to population frequency, splicing prediction, and somatic-oncology annotation resources; none address the mode of inheritance or parentage confirmation for an individual patient.

Still missing

• No indication of assumed de novo status (parentage not confirmed) for this proband.
• No clinical case report or family history data in the bundle.

Reasoning

PP1 requires co-segregation of the variant with disease across multiple affected family members/meioses. The bundle contains no pedigree, no family segregation data, and no meiosis count for this specific variant. The five literature_pass/OncoKB-linked PMIDs (14769601, 22205104, 26607597, 30206110, 31138663) are gene-level functional/cohort studies about RB1 broadly (protein truncation testing methodology, second-malignancy risk, mouse Rb1 models, CDK4/6 resistance genomics, DNA damage biology) and none report this specific c.275_282del variant in a segregation context; per grounding rules, general gene-level discussion does not support a variant-specific PP1 conclusion. No paper_extracts entries exist to support variant-specific segregation findings either.

Evidence considered

• Reviewed literature_pass.json high_priority_papers (PMIDs 14769601, 22205104, 26607597, 30206110, 31138663): all are gene-level RB1 functional/cohort studies without mention of this specific variant or any family segregation data for it.

Still missing

• No pedigree or number of affected family members co-segregating with the variant.
• No meiosis count for segregation analysis.
• None of the five linked PMIDs mention this specific variant (c.275_282del) in a segregation context.

Reasoning

PP4 requires that the patient's phenotype or family history be highly specific for a disease with a single genetic etiology. No individual patient/proband clinical presentation, phenotype description, or family history tied to this specific variant is present anywhere in the case bundle (case_summary, evidence.json, literature_pass.json, paper_extracts.json are all empty of proband-level phenotype data for this allele). Without a described phenotype to compare against RB1-associated retinoblastoma specificity, PP4 cannot be scored.

Still missing

• No individual patient phenotype or family history information tied to this specific variant was available in the case bundle.

Reasoning

No functional or splicing assay data showing a benign/no-effect result for NM_000321.2:c.275_282del (p.(Ile92LysfsTer15)) was located. The variant produces an early frameshift (p.Ile92Lysfs*15) predicted to trigger nonsense-mediated decay or truncate the pRb pocket domain, and OncoKB's curated biological-effect annotation is 'Likely Loss-of-function' (not a validated wet-lab assay, and not usable as direct BS3 evidence per this evaluation's rule that a source's own classification cannot be used to infer a criterion). None of the five retrieved RB1 papers (PMID:14769601, PMID:22205104, PMID:26607597, PMID:30206110, PMID:31138663) report this specific variant or any assay result consistent with normal/benign protein function for it. No RB1 VCEP-specific BS3 framework was available (generic_acmg fallback). Given the absence of any variant-specific functional evidence in either direction, BS3 is not assessed rather than defaulted to not_met, pending further targeted literature search.

Evidence considered

• Searched PMID:14769601, PMID:22205104, PMID:26607597, PMID:30206110, PMID:31138663 full texts for exact variant strings 'c.275_282del', 'Ile92', 'I92Kfs' - no matches found in any paper.
• No CSPEC/VCEP RB1 framework was available in this case (vcep_materials.json: found=false, framework_mode=generic_acmg fallback), so no gene-specific BS3 assay-validation criteria could be applied.

Still missing

• No functional assay result showing normal/benign activity identified for c.275_282del.
• No RB1 VCEP or gene-specific BS3 framework available to define assay validation/control requirements.
• OncoKB's 'Likely Loss-of-function' annotation is a curated classification label, not a wet-lab assay result, and cannot substitute for direct functional evidence.

Sources

Reasoning

BS4 requires documented non-segregation of the variant with disease in an affected family. No family or pedigree data of any kind is present in the case bundle for this variant, so non-segregation cannot be evaluated in either direction.

Still missing

• No family/pedigree data present to assess segregation or non-segregation.

Reasoning

No ClinGen RB1 VCEP specification is available (cspec.found=false), so generic ACMG/AMP BP2 would apply if data existed. BP2 requires observing the variant in trans with a pathogenic variant in a fully penetrant dominant disorder, or in cis with a pathogenic variant under any inheritance pattern. The consolidated case bundle contains no patient/pedigree information, no reported second variant, and no phase data of any kind (case_summary.json, evidence.json, and review_manifest.json each lack a genotype/family section). ClinVar search returned no result for this variant (absent from ClinVar), so no submitter-level genotype context is available there either. Without any observed second variant or phase information, this criterion cannot be evaluated one way or the other.

Evidence considered

• ClinVar search for RB1 c.275_282del returned no_result (variant absent from ClinVar), so no submitter genotype/zygosity context is available to assess phase.
• No proband, pedigree, or second-variant information appears anywhere in case_summary.json, evidence.json, or review_manifest.json.

Still missing

• No case-level genotype or pedigree data provided to determine whether this variant occurs in cis or trans with any other pathogenic variant.
• ClinVar has no record for this variant, so no submitter-reported zygosity/phase context is available.
• Would need proband trio/segregation data or an internal lab report documenting a second variant and its phase to assess BP2.

Sources

Reasoning

BP5 (observation in a case with an alternate molecular basis for disease) requires case-level data identifying another causal variant/etiology in a patient carrying this allele. No such case-level data, alternate diagnosis, or co-occurring causal variant information for this specific variant is present in the bundle. No RB1 VCEP/CSPEC framework was found defining a gene-specific BP5 rule or likelihood-ratio-style threshold, so generic BP5 cannot be evaluated for lack of any qualifying case observation.

Still missing

• No patient case observation or alternate-etiology data available for this variant.
• No RB1 VCEP/CSPEC BP5 rule or threshold identified.