KMT2A c.10244C>T, p.(Pro3415Leu)

Reasoning

The variant is absent from gnomAD v2.1 and gnomAD-Canada v1.0 and is extremely rare in gnomAD v4.1, with only 2 alternate alleles among 1,614,194 alleles (AF 1.23901e-06), zero homozygotes, and group maximum FAF 4.42e-06. This is compatible with PM2 for a rare KMT2A-related disorder under the generic ACMG/AMP framework. The presence of two alleles in gnomAD does not negate PM2 because the observed frequency remains extremely low and no credible disease-specific frequency excess is shown.

Rule

generic_acmg_pm2: absent_or_extremely_rare_in_population_databases

Evidence considered

• gnomAD v2.1: variant absent.
• gnomAD v4.1: total AF 1.23901e-06 (2/1,614,194 alleles), group maximum FAF 4.42e-06, highest subpopulation AF 2.66539e-05, and homozygotes = 0.
• gnomAD-Canada v1.0: variant absent.
• Population evidence is evaluated independently from ClinVar or other source classifications; no source classification was used to infer PM2.

Sources

Reasoning

No governing CSPEC/VCEP for KMT2A is available in this case (cspec.found=false; vcep_materials note='no cspec url available'; final_classification_framework falls back to generic ACMG/AMP 2015), so generic calibrated computational thresholds were applied. This is a missense variant (NP_005924.2:p.(Pro3415Leu)); consequence_class='missense', with no canonical splice consensus disruption (canonical_splice_consensus=false, variant_bucket='other'). REVEL score = 0.397, which falls in the indeterminate zone between the published ClinGen SVI calibration cutoffs (Pejaver et al. 2022, PMID:36413997): PP3_Supporting requires REVEL >=0.644, and this score does not reach that threshold, so REVEL does not support PP3. BayesDel score (-0.0423011) is available in prefetch data but this pipeline has no verified published calibration publication/table for BayesDel thresholds, so per policy it is treated as not_available/insufficiently calibrated for PP3 regardless of the numeric value. SpliceAI predicts no significant splice impact (max delta score = 0.002; DS_AG=0.001, DS_AL=0.001, DS_DG=0.002, DS_DL=0.0), which is consistent with absence of a splicing effect but does not itself provide positive evidence toward pathogenicity. With the only calibrated, citable missense predictor (REVEL) indeterminate and BayesDel excluded for lack of a citable calibration source, there is insufficient convergent calibrated computational evidence to apply PP3.

Rule

generic_acmg_pp3_pejaver_revel_calibration

Evidence considered

• REVEL score = 0.397 for NM_005933.3:c.10244C>T (NP_005924.2:p.(Pro3415Leu)), retrieved from local REVEL v1.3 lookup (chr11:118506145 C>T).
• Published ClinGen SVI REVEL calibration thresholds (Pejaver et al. 2022, PMID:36413997) require REVEL >=0.644 for PP3_Supporting; 0.397 does not meet this threshold, placing the score in the indeterminate zone.
• BayesDel score = -0.0423011 was retrieved from local BayesDel (BayesDel_170824_noAF) lookup, but no verified published threshold/publication is available to this pipeline for BayesDel, so it is treated as not_available for PP3 per policy regardless of the numeric value.
• SpliceAI Lookup (spliceailookup.broadinstitute.org) reports max delta score = 0.002 (all four delta scores <=0.002), indicating no predicted splicing impact; this does not support PP3.

Still missing

• REVEL score (0.397) falls in the indeterminate zone of the published SVI calibration and does not reach the PP3_Supporting threshold (>=0.644).
• BayesDel score is present in prefetch data but no verified published calibration publication/table is available to this pipeline, so it cannot be used to support or refute PP3.
• No gene-specific KMT2A VCEP PP3/BP4 lookup material was available to override or refine the generic REVEL/SpliceAI thresholds.

Sources

Reasoning

PP5 is not met. The exact-variant ClinVar record (Variation ID 4044414; matched by genomic HGVS) contains one Ambry Genetics single-submitter uncertain-significance assertion with criteria provided. It is not an expert-panel classification and therefore cannot trigger PP5.

Rule

PP5 may be applied only when an exact-variant ClinVar expert panel classifies the variant as Pathogenic or Likely pathogenic; a single-laboratory uncertain-significance assertion does not qualify.

Evidence considered

• The exact-match ClinVar submission is classified Uncertain significance, has review status criteria provided/single submitter, and has zero expert-panel submissions.
• The ClinVar submission audit identifies the sole submitter as Ambry Genetics, marks is_expert_panel false, and records no criterion-lead submission; ordinary laboratory and aggregate ClinVar assertions are excluded from PP5 by the governing instruction.

Sources

Reasoning

The variant does not meet the generic BA1 population-frequency threshold. gnomAD v4.1 reports a total allele frequency of 1.23901e-06 (2/1,614,194 alleles), a maximum observed subpopulation AF of 2.66539e-05 in the African/African American population, and a group maximum FAF of 4.42e-06; these frequencies are far below the >1% threshold required for BA1. It is absent from gnomAD v2.1 and gnomAD-Canada v1.0.

Rule

generic_acmg_ba1: allele_frequency > 0.01

Evidence considered

• gnomAD v4.1: total AF 1.23901e-06, 2/1,614,194 alleles; highest subpopulation AF 2.66539e-05 in African/African American individuals; group maximum FAF 4.42e-06; homozygotes 0.
• gnomAD v2.1 and gnomAD-Canada v1.0: variant absent.
• Generic ACMG/AMP fallback threshold applied: BA1 requires a population allele frequency greater than 1%.

Sources

Reasoning

The available population data do not show an allele frequency greater than the expected disease-specific maximum credible frequency. The variant is present only twice in gnomAD v4.1, with total AF 1.23901e-06, group maximum FAF 4.42e-06, and highest observed subpopulation AF 2.66539e-05. These values are not consistent with an allele frequency higher than expected for a rare autosomal-dominant KMT2A disorder and do not support BS1.

Rule

generic_acmg_bs1: population_frequency_higher_than_expected_for_disorder

Evidence considered

• gnomAD v4.1: 2 alternate alleles among 1,614,194 total alleles; total AF 1.23901e-06; group maximum FAF 4.42e-06; highest subpopulation AF 2.66539e-05 in African/African American individuals.
• gnomAD v2.1 and gnomAD-Canada v1.0: variant absent.
• No disease prevalence, penetrance, or validated KMT2A-specific maximum credible allele-frequency threshold in the case bundle indicates that this frequency is excessive.

Sources

Reasoning

BP1 applies when a missense variant occurs in a gene for which primarily truncating variants are known to cause disease (i.e., missense is not a recognized disease mechanism). While the germline literature review indicates the predominant KMT2A germline disease mechanism (Wiedemann-Steiner syndrome) is haploinsufficiency via truncating variants, it also documents that missense variants are a recognized, if less common, pathogenic mechanism, particularly when clustering in specific functional domains. Because missense pathogenicity is an established mechanism for this gene (not absent), BP1 is not met.

Evidence considered

• pvs1_gene_context.json germline literature review states that most pathogenic KMT2A variants arise de novo and are typically nonsense or frameshift, but missense variants have also been reported and may have complex functional consequences, indicating missense is a recognized (non-excluded) disease mechanism for this gene, which precludes applying BP1.

Still missing

• No formal missense-constraint statistic or ClinGen VCEP-specific BP1 threshold for KMT2A is available in this case bundle to further quantify this call.

Sources

Reasoning

No governing CSPEC/VCEP for KMT2A is available in this case, so generic calibrated computational thresholds were applied. REVEL score = 0.397 is below the PP3_Supporting threshold but also above the published ClinGen SVI BP4_Supporting cutoff (REVEL <=0.290, Pejaver et al. 2022, PMID:36413997), so it falls in the indeterminate zone and does not support BP4 either. BayesDel score is present but excluded from consideration because no verified published calibration threshold/publication is available to this pipeline. SpliceAI predicts no significant splice impact (max delta score = 0.002), which is consistent with a benign splicing outcome, but this variant is a missense substitution with no canonical splice-site disruption noted (canonical_splice_consensus=false); a single negative splice-impact signal alone, without a concordant benign missense in-silico call, is insufficient to satisfy the 'multiple lines of computational evidence' standard for BP4. Because the only calibrated, citable missense predictor (REVEL) is indeterminate rather than benign-supporting, and BayesDel cannot be used, BP4 is not met.

Rule

generic_acmg_bp4_pejaver_revel_calibration

Evidence considered

• REVEL score = 0.397 for NM_005933.3:c.10244C>T (NP_005924.2:p.(Pro3415Leu)), retrieved from local REVEL v1.3 lookup (chr11:118506145 C>T); published ClinGen SVI calibration (Pejaver et al. 2022, PMID:36413997) requires REVEL <=0.290 for BP4_Supporting, which 0.397 does not meet.
• BayesDel score = -0.0423011 retrieved from local BayesDel lookup, excluded from BP4 assessment because no verified published threshold/publication is available to this pipeline for BayesDel.
• SpliceAI Lookup reports max delta score = 0.002 (DS_AG=0.001, DS_AL=0.001, DS_DG=0.002, DS_DL=0.0), consistent with no predicted splicing impact, but this is not sufficient alone to satisfy BP4 for a missense variant given the indeterminate missense (REVEL) result.

Still missing

• REVEL score (0.397) is above the BP4_Supporting threshold (<=0.290) and therefore does not support a benign computational call.
• BayesDel score is present in prefetch data but excluded because no verified published calibration publication/table is available to this pipeline.
• SpliceAI alone (negative for splice impact) is not sufficient to independently satisfy BP4 for a missense variant without a concordant benign missense predictor result.

Sources

Reasoning

BP6 is not met. The exact-variant ClinVar record has only one non-expert single-laboratory uncertain-significance submission and no expert-panel Benign or Likely benign classification.

Rule

BP6 may be applied only when an exact-variant ClinVar expert panel classifies the variant as Benign or Likely benign; a non-expert uncertain-significance assertion does not qualify.

Evidence considered

• The exact-match ClinVar record reports Uncertain significance with review status criteria provided/single submitter and zero expert-panel submissions.
• The ClinVar audit identifies Ambry Genetics as the sole non-expert submitter and explicitly records is_expert_panel false; this cannot trigger BP6 under the governing instruction.

Sources

Reasoning

PS1 requires a different nucleotide substitution producing the identical amino acid change (p.Pro3415Leu) that has been established as pathogenic. The only ClinVar record for this residue is this variant itself (VCV004044414, Uncertain significance, 1 submitter). No comparator ClinVar record with the same predicted protein change (p.Pro3415Leu) arising from a different codon substitution was identified in the case bundle.

Evidence considered

• ClinVar VCV004044414 for this exact variant (NM_005933.3:c.10244C>T, p.Pro3415Leu) is classified Uncertain significance with a single submitter (Ambry Genetics, criteria provided) and lists no other reviewed pathogenic variant with the same amino-acid outcome to serve as a PS1 comparator.

Still missing

• No dedicated ClinVar/database search for alternate nucleotide changes yielding the identical p.Pro3415Leu substitution was captured in this case bundle; a targeted search is needed before PS1 can be ruled in or out.

Sources

Reasoning

No case-level evidence documents that NM_005933.3:c.10244C>T (KMT2A p.Pro3415Leu) arose de novo in an affected proband. The bundle contains no proband description, parental genotypes, confirmed maternity/paternity, or parental testing results; therefore PS2 cannot be assessed.

Rule

Generic ACMG/AMP PS2: a de novo variant in a patient with the disease and no family history, with confirmed maternity and paternity; parental testing and phenotype consistency are required for application.

Evidence considered

• Review of the consolidated case-evidence bundle found no proband-level, parental-testing, de novo, or family-history evidence relevant to PS2.
• The available bundle identifies the variant as a missense change and reports only population, database, predictor, and unrelated literature-triage information; none establishes de novo occurrence.

Still missing

• Affected proband phenotype and diagnosis are not documented in the case-evidence bundle.
• Maternal and paternal testing results for the variant are not provided.
• Confirmed maternity and paternity are not documented.
• Family history and exclusion of an inherited variant are not documented.

Reasoning

No functional/biochemical assay data (e.g. methyltransferase activity, cell-based reporter, in vitro binding, or other validated functional study) addressing NM_005933.3:c.10244C>T (p.Pro3415Leu) in KMT2A was found anywhere in the case evidence bundle. No ClinGen KMT2A VCEP specification is present (cspec.found = false; framework_mode = generic_acmg), so no gene-specific PS3 functional assay framework or approved assay list is available. The literature associated with this case (ClinVar-linked PMIDs) consists entirely of newborn screening policy/guideline papers with no mention of this variant or of any KMT2A functional assay; none are functional studies.

Still missing

• No published or unpublished functional assay data for this variant were located.
• No KMT2A-specific VCEP functional evidence framework (assay validation criteria, control ranges) is available to calibrate strength even if a study were found.

Reasoning

No affected individuals carrying the exact NM_005933.3:c.10244C>T (NP_005924.2:p.Pro3415Leu) variant, case-control enrichment analysis, or statistically supported excess of the variant in affected cases is documented in the available evidence bundle.

Rule

Generic ACMG/AMP PS4 requires statistically significant enrichment of the variant in affected individuals compared with controls or a well-established excess of affected individuals carrying the variant; no qualifying exact-variant evidence is available.

Evidence considered

• The case-evidence bundle contains no exact-variant case series, affected-carrier count, case-control comparison, or enrichment statistic suitable for PS4.
• The only exact-variant ClinVar submission is an uncertain-significance single-laboratory assertion and does not provide PS4 case-control evidence.

Still missing

• Exact-variant case observations with affected status and phenotype details are unavailable.
• No case-control counts, statistical enrichment estimate, or comparison cohort is available.

Reasoning

PM1 requires the variant to fall in a well-established mutational hotspot or critical/well-characterized functional domain (e.g. an active site) without benign variation. No hotspot database evidence was found (cancerhotspots.org returned no significant hotspot at KMT2A P3415), and no domain-boundary annotation (UniProt/InterPro) for residue 3415 was retrieved in this case bundle. KMT2A's well-characterized functional domains (AT-hooks, CXXC zinc finger, PHD fingers, bromodomain, FYRN/FYRC, SET domain) are described narratively in the germline literature review but exact residue coordinates for KMT2A p.3415 relative to those domains are not confirmed by a cited source in this bundle, so domain membership cannot be verified without fabricating coordinates.

Evidence considered

• cancerhotspots.org query for 'KMT2A P3415' returned no_result / no statistically significant hotspot row for this residue.
• OncoKB gene-level page for KMT2A/P3415L returned no variant-specific reviewed functional or oncogenic-effect annotation (classification: Unknown Oncogenic Effect, not variant specific), providing no domain/hotspot support for PM1.

Still missing

• No UniProt/InterPro domain-boundary lookup for KMT2A residue 3415 was performed or captured in this case bundle.
• cancerhotspots.org search for KMT2A P3415 returned no significant hotspot (screenshot only, no positive supporting source).

Sources

Reasoning

No case-level affected-proband observations, second pathogenic variant, phase information, or segregation data are provided for NM_005933.3:c.10244C>T (NP_005924.2:p.Pro3415Leu). The bundle indicates no applicable KMT2A-specific VCEP/CSPEC, so the generic ACMG/AMP framework is used. PM3 cannot be assessed without evidence that this variant is in trans with a pathogenic or likely pathogenic allele in an affected individual with a recessive disorder; no such evidence is available.

Evidence considered

• The consolidated case-evidence bundle contains no affected-proband, compound-heterozygous, phase, or segregation observations for this variant.
• The bundle reports no applicable KMT2A-specific VCEP/CSPEC or gene-specific framework; generic ACMG/AMP rules therefore apply.

Still missing

• Affected-proband or affected-family observations involving this variant
• A second pathogenic or likely pathogenic variant in the same individual
• Validated cis/trans phase information
• Inheritance model and disease-specific applicability for PM3

Reasoning

PM5 requires a previously classified pathogenic/likely pathogenic missense variant at the same residue with a different amino-acid substitution. The dedicated PM5 candidate-collection artifact for this case explicitly did not run automated same-residue candidate harvesting ('eligible_for_classic_pm5_search': false) because classic PM5 semantics could not be safely confirmed, and returned zero candidates.

Evidence considered

• pm5_candidates.json reports 0 total candidates and 0 same-residue candidates, with recommendation level 'not_applicable' and explicit instruction not to apply PM5 from that artifact alone because classic same-residue PM5 semantics could not be confirmed safely for this variant.

Still missing

• pm5_candidates.json contains no same-residue comparator variants; a manual ClinVar search for other amino-acid substitutions at KMT2A codon 3415 (p.Pro3415X, X != Leu) with pathogenic/likely pathogenic classification has not been completed for this case.

Sources

Reasoning

No case-level evidence indicates that the variant is suspected to be de novo without confirmed parental relationships or parental testing. The available materials do not describe a proband, parents, or a de novo observation, so PM6 cannot be assessed.

Rule

Generic ACMG/AMP PM6: assumed de novo, without confirmed maternity and paternity; this requires a documented affected proband and an apparent de novo observation but lacks the confirmation required for PS2.

Evidence considered

• The consolidated case-evidence bundle contains no documented suspected de novo observation for this variant.
• The literature-pass entries are unrelated newborn-screening policy papers and provide no variant-specific parental or de novo evidence.

Still missing

• No affected proband or clinical presentation is documented.
• No report of an apparent de novo occurrence is provided.
• No parental genotypes or relationship testing are provided.
• No family history is provided.

Reasoning

No affected or unaffected relatives, variant genotypes, pedigree, or informative meioses are reported. Consequently, no cosegregation evidence is available to support PP1 or to determine a strength based on the number and informativeness of meioses.

Rule

Generic ACMG/AMP PP1: cosegregation with disease in multiple affected family members in a gene definitively known to cause the disease; strength requires an appropriate number of informative meioses and consideration of phenotype specificity.

Evidence considered

• The consolidated case-evidence bundle contains no family segregation data, affected-relative observations, or informative meioses for NM_005933.3:c.10244C>T.
• Population frequency, ClinVar, somatic cancer, and computational findings do not constitute cosegregation evidence and were not used for PP1.

Still missing

• No pedigree or family structure is provided.
• No affected-relative phenotypes are documented.
• No relative genotypes are provided.
• No informative meiosis count or segregation analysis is available.

Reasoning

PP2 requires the gene to have a low rate of benign missense variation and missense to be a common mechanism of disease. No CSPEC/VCEP was found for KMT2A (framework_mode: generic_acmg, cspec.found: false), and no missense constraint metric (e.g. gnomAD missense Z-score or o/e ratio) was retrieved in this case bundle to quantify benign missense tolerance. The available germline literature indicates the dominant KMT2A disease mechanism for Wiedemann-Steiner syndrome is haploinsufficiency, with truncating (nonsense/frameshift) variants predominating and missense variants representing a minority that can also be pathogenic when they cluster in specific functional domains; this mixed picture cannot be resolved to a confident PP2 call without quantitative constraint data.

Evidence considered

• pvs1_gene_context.json germline literature review (targeted PubMed/EuropePMC search) found KMT2A germline disease (Wiedemann-Steiner syndrome) is caused primarily by loss-of-function variants, with haploinsufficiency considered the primary pathogenic mechanism; missense variants have also been reported and may have complex functional consequences, but no missense constraint statistic was captured to support or refute a 'low rate of benign missense' claim for PP2.

Still missing

• No gnomAD/ExAC missense constraint (Z-score, o/e) for KMT2A was retrieved in this case bundle.
• No official ClinGen KMT2A VCEP specification for PP2 applicability was found.

Sources

Reasoning

The available records do not document the phenotype of an individual carrying this exact variant, nor do they establish that such a phenotype is highly specific for a KMT2A-related disorder.

Rule

Generic ACMG/AMP PP4 requires the patient's phenotype or family history to be highly specific for a disease with a single-gene etiology; no qualifying patient-specific phenotype is available.

Evidence considered

• The bundle provides gene-level disease context but no clinical description linked to NM_005933.3:c.10244C>T or p.Pro3415Leu.
• General descriptions of KMT2A-related disorders cannot establish PP4 for this individual variant without an affected-carrier phenotype.

Still missing

• Clinical phenotype and diagnostic workup for an exact-variant carrier are unavailable.
• No phenotype-specificity assessment or validated disease-specific phenotype definition is available for this variant.

Reasoning

BS2 requires observation of the variant in healthy adults, typically in homozygous or hemizygous state for a disorder expected to be fully penetrant. gnomAD v4.1 reports zero homozygotes, and the case bundle contains no individual-level clinical information establishing unaffected adult homozygous or hemizygous carriers. Therefore BS2 cannot be applied from the available evidence.

Rule

generic_acmg_bs2: observed_in_healthy_adult_homozygote_or_hemizygote

Evidence considered

• gnomAD v4.1 reports homozygotes = 0 for the variant; this does not establish the BS2 observation requirement.
• The case bundle provides no healthy-adult carrier-level phenotype, age, zygosity, or penetrance data.

Still missing

• No verified healthy-adult homozygous or hemizygous carrier observation is available.
• No individual-level phenotype and age information is available to evaluate unaffected status or penetrance.

Sources

Reasoning

No functional/biochemical assay data showing normal/wild-type-like function for NM_005933.3:c.10244C>T (p.Pro3415Leu) in KMT2A was found in the case evidence bundle. No ClinGen KMT2A VCEP specification exists for this gene (cspec.found = false), so there is no gene-specific BS3 assay framework to apply. The associated literature (ClinVar-linked PMIDs) is limited to general newborn screening policy papers with no functional relevance to this variant.

Still missing

• No published or unpublished functional assay data for this variant were located.
• No KMT2A-specific VCEP functional evidence framework is available to calibrate strength even if a study were found.

Reasoning

No family members or variant-positive relatives are described, and no unaffected relatives tested negative for the variant are documented. Therefore lack of segregation cannot be evaluated and BS4 cannot be assessed.

Rule

Generic ACMG/AMP BS4: lack of segregation in affected members of a family; application requires informative family testing and phenotype assessment, while accounting for reduced penetrance, variable expressivity, and phenocopies.

Evidence considered

• The consolidated case-evidence bundle provides no non-segregation observations or variant testing results from unaffected relatives.
• No source in the bundle establishes a family-based mismatch between the variant and the disease phenotype.

Still missing

• No pedigree or family history is available.
• No clinically unaffected relatives with reliable phenotyping are identified.
• No genotype results for relatives are provided.
• No assessment of whether non-segregation is informative or whether reduced penetrance/age-dependent expression could explain it is available.

Reasoning

No case-level evidence is available showing NM_005933.3:c.10244C>T in trans with a pathogenic variant in an affected individual with a dominant disorder, or in cis with a pathogenic variant. The case bundle provides no phase-resolved family or proband data, so BP2 cannot be assessed.

Evidence considered

• The consolidated case-evidence bundle contains no phase, co-occurring pathogenic-variant, affected-proband, or segregation data relevant to BP2.
• The single ClinVar submission is an uncertain-significance classification and provides no validated phase or allelic evidence.

Still missing

• Phase-resolved genotype data for this variant and any additional pathogenic variant
• Affected-proband or family observations establishing the relevant inheritance context
• Confirmation that any comparator allele is pathogenic or likely pathogenic

Reasoning

No affected individual carrying the exact variant is documented, and no credible alternative molecular explanation for a relevant phenotype is available; consequently BP5 cannot be evaluated from the supplied evidence.

Rule

Generic ACMG/AMP BP5 requires an affected individual with the variant and a convincing alternative molecular cause explaining the phenotype; no qualifying case-level evidence is available.

Evidence considered

• The evidence bundle contains no case-level phenotype record or molecular workup showing an alternative cause of disease in an exact-variant carrier.
• The single somatic COSMIC observation is not a germline clinical case with an alternative molecular diagnosis and is insufficient for BP5.

Still missing

• An exact-variant affected individual and the phenotype requiring explanation are not documented.
• No confirmed alternative pathogenic variant or other molecular cause in an affected individual is available.