CTNNB1 c.1582C>T, p.Arg528Cys

Reasoning

PM2 is met at supporting strength under the supplied generic SVI threshold. The variant is present at an extremely low frequency in gnomAD v4.1 (10/1614008 alleles; AF 6.19576e-06; zero homozygotes), with grpmax FAF=8.75e-06, which is below the <=0.0001 PM2 threshold. Concordant low-frequency observations are present in gnomAD v2.1 all-comers exomes (1/251420; AF 3.97741e-06), gnomAD v2.1 non-cancer exomes (1/236882; AF 4.22151e-06), and gnomAD v3.1 non-cancer genomes (3/147916; AF 2.02818e-05; grpmax FAF=4.88e-06).

Rule

PM2: Absent from controls (or at extremely low frequency if recessive) in Exome Sequencing Project, 1000 Genomes or Exome Aggregation Consortium. ... PM2 (supporting): <= 0.0001 — SVI recommendation to downgrade PM2 to Supporting (ClinGen SVI 2020) (PMID:25741868)

Evidence considered

• gnomAD v4.1 reports total AF 6.19576e-06 (10/1614008 alleles), zero homozygotes, and grpmax FAF=8.75e-06.
• gnomAD v2.1 all-comers exomes reports AF 3.97741e-06 (1/251420 alleles) and zero homozygotes; the non-cancer exome subset reports AF 4.22151e-06 (1/236882 alleles) and zero homozygotes.
• gnomAD v3.1 non-cancer genomes reports AF 2.02818e-05 (3/147916 alleles), zero homozygotes, and grpmax FAF=4.88e-06.
• The generic PM2 calibration threshold supplied for this case is <=0.0001, supporting strength (ClinGen SVI recommendation; PMID:25741868).

Sources

Reasoning

The variant is a missense substitution, CTNNB1 p.Arg528Cys. Gene-level germline evidence states that most patients with CTNNB1 syndrome carry pathogenic nonsense or frameshift variants generating premature termination codons, and a cohort report describes 21 CTNNB1 loss-of-function variants in 24 affected individuals. This supports the generic BP1 pattern that disease is primarily associated with truncating variants, although missense disease associations also exist and prevent treating this as an exclusion of missense pathogenicity.

Rule

Missense variant in a gene for which primarily truncating variants are known to cause disease.

Evidence considered

• The gene-context file's PMID:41082542 abstract excerpt states: Patients with CTNNB1 syndrome display heterogeneous clinical manifestations, and most of them carry CTNNB1 pathogenic nonsense or frameshift variants that generate premature termination codons (PTC).
• The gene-context file's PMID:36153650 abstract excerpt reports 21 loss-of-function variants among 24 affected Chinese patients and states that the disorder is caused by heterozygous germline loss-of-function variants in CTNNB1.
• The available gene-context evidence also lists CTNNB1 missense disease reports; BP1 is therefore applied only at supporting strength and does not by itself classify this variant as benign.

Sources

Reasoning

The variant is a missense substitution, NM_001904.4:c.1582C>T (p.Arg528Cys), so the governing generic rule requires the missense REVEL path only; SpliceAI is not counted for PP3/BP4 in this situation. REVEL is 0.4, below the generic PP3 supporting threshold of >= 0.644, the moderate threshold of >= 0.773, and the strong threshold of >= 0.932. BayesDel is not used because no generic ACMG-strength calibration is established. Therefore PP3 is not met.

Rule

PP3 (supporting): >= 0.644 — ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet) (PMID:36413997)

Evidence considered

• NM_001904.4:c.1582C>T is an exonic missense variant producing p.Arg528Cys in exon 10.
• Local REVEL lookup reports a score of 0.4 for NC_000003.12:g.41234196C>T.
• The generic REVEL calibration gives PP3 thresholds of >= 0.644 supporting, >= 0.773 moderate, and >= 0.932 strong; these thresholds are from Pejaver et al. 2022, PMID:36413997.
• SpliceAI was not counted because the governing path for a missense variant is REVEL only; BayesDel was not used because no generic ACMG-strength calibration is established.

Sources

Reasoning

The governing PP5 definition requires a reputable source to report the exact variant as pathogenic while the underlying evidence is unavailable for independent evaluation. The exact-match ClinVar record is Uncertain significance from one clinical laboratory with no expert-panel submission, and the reviewed literature does not report this exact variant as pathogenic. PP5 is not met.

Rule

Reputable source recently reports variant as pathogenic, but the evidence is not available to the laboratory to perform an independent evaluation.

Evidence considered

• ClinVar exact-match record NM_001904.4(CTNNB1):c.1582C>T (p.Arg528Cys): Uncertain significance, criteria provided, single submitter; expert-panel submissions = 0.
• PMID:28191889 full-text extraction: the exact variant is not mentioned and is not reported as pathogenic.

Sources

Reasoning

BA1 is not met. The governing frequency metric is the gnomAD v4.1 grpmax FAF, 8.75e-06, which is far below the generic stand-alone threshold of 0.05. The highest point estimate is the Amish AF of 0.0010989 (1/910 alleles), but this subgroup has fewer than the required 2000 observed alleles and its point estimate must not override the filtering FAF. The gnomAD v2.1 all-comers and non-cancer exome frequencies are also far below 5% (3.97741e-06 and 4.22151e-06, respectively).

Rule

BA1: Allele frequency is above 5% in Exome Sequencing Project, 1000 Genomes or Exome Aggregation Consortium. ... BA1 (stand-alone): >= 0.05 — ACMG/AMP 2015 stand-alone benign allele frequency (PMID:25741868)

Evidence considered

• gnomAD v4.1 reports total AF 6.19576e-06 and grpmax FAF=8.75e-06; the highest observed subgroup point estimate is Amish AF 0.0010989 from 1/910 alleles.
• gnomAD v2.1 reports all-comers exome AF 3.97741e-06 (1/251420 alleles); its non-cancer exome subset reports AF 4.22151e-06 (1/236882 alleles).
• The generic BA1 calibration threshold supplied for this case is >=0.05 (ACMG/AMP 2015, PMID:25741868); filtering FAF rather than a small-subgroup popmax point estimate is required by the governing population guidance.

Sources

Reasoning

BS1 is not met on the available population data. The appropriate filtering metric, gnomAD v4.1 grpmax FAF, is 8.75e-06, below the generic 0.01 threshold; the non-cancer v3.1 genome-only grpmax FAF is even lower at 4.88e-06. The Amish point estimate of 0.0010989 is based on only 1/910 alleles, below the required 2000 observed-allele population-size floor, so it cannot establish BS1. The rare, early-onset autosomal-dominant CTNNB1 disorder context also provides no basis to replace the supplied threshold with a disease-specific maximum-credible-frequency calculation.

Rule

BS1: Allele frequency is greater than expected for the disorder. ... BS1 (strong): >= 0.01 — Generic SVI default; per-gene derivation preferred (Whiffin et al. 2017) (PMID:28518168)

Evidence considered

• gnomAD v4.1 reports grpmax FAF=8.75e-06, total AF 6.19576e-06, and Amish AF 0.0010989 from 1/910 alleles.
• The non-cancer genome-only gnomAD v3.1 result reports grpmax FAF=4.88e-06 and total AF 2.02818e-05 (3/147916 alleles).
• The generic BS1 calibration threshold supplied for this case is >=0.01; Whiffin et al. 2017 (PMID:28518168) is the cited maximum-credible-frequency framework, but the available evidence does not provide a validated disease-specific prevalence, penetrance, or allelic-heterogeneity calculation that would support a different threshold.
• The population guidance requires at least 2000 observed alleles for the population used to support frequency-based BS1; the Amish subgroup has 910 observed alleles.

Sources

Reasoning

The variant is a missense substitution, so BP4 must be assessed through the REVEL path only; its SpliceAI result cannot substitute for an indeterminate REVEL result. REVEL is 0.4, above the generic BP4 supporting threshold of <= 0.29, the moderate threshold of <= 0.183, and the strong threshold of <= 0.016. The score is therefore gray-zone/indeterminate for BP4, and the governing rule states that an indeterminate applicable-path result does not meet BP4. BayesDel is not used because no generic ACMG-strength calibration is established.

Rule

An indeterminate result on the applicable path means PP3/BP4 are simply not met. It does not license reaching across to the other path's score - a missense variant whose REVEL sits in the gray zone gets no BP4, however clean its SpliceAI delta.

Evidence considered

• NM_001904.4:c.1582C>T is an exonic missense variant producing p.Arg528Cys in exon 10.
• Local REVEL lookup reports a score of 0.4 for NC_000003.12:g.41234196C>T.
• The generic REVEL calibration gives BP4 thresholds of <= 0.29 supporting, <= 0.183 moderate, and <= 0.016 strong; these thresholds are from Pejaver et al. 2022, PMID:36413997.
• SpliceAI was not used for BP4 because missense variants use the REVEL path only; BayesDel was not used because no generic ACMG-strength calibration is established.

Sources

Reasoning

The governing BP6 definition requires a reputable source to report the exact variant as benign while the evidence is unavailable for independent evaluation. The exact-match ClinVar record is Uncertain significance from one clinical laboratory, not benign or likely benign, with no expert-panel submission; the reviewed paper does not report this exact variant as benign. BP6 is not met.

Rule

Reputable source recently reports variant as benign, but the evidence is not available to the laboratory to perform an independent evaluation.

Evidence considered

• ClinVar exact-match record NM_001904.4(CTNNB1):c.1582C>T (p.Arg528Cys): Uncertain significance, criteria provided, single submitter; expert-panel submissions = 0.
• PMID:28191889 full-text extraction: the exact variant is not mentioned and is not reported as benign.

Sources

Reasoning

The variant is CTNNB1 p.Arg528Cys. The available exact-variant ClinVar record is a single-submitter uncertain-significance record, and the reviewed paper extract does not mention p.Arg528Cys. No previously established pathogenic variant producing the same Arg528Cys amino-acid change, including an alternate nucleotide change, is documented in the supplied evidence. Therefore PS1 cannot be assessed from the available evidence.

Rule

Same amino acid change as a previously established pathogenic variant regardless of nucleotide change.

Evidence considered

• ClinVar variation 1984487 is NM_001904.4(CTNNB1):c.1582C>T (p.Arg528Cys), classified as uncertain significance with one clinical-laboratory submission; this is not an established pathogenic comparator.
• PMID:28191889 paper extract: the paper does not mention CTNNB1 c.1582C>T or p.(R528C).

Still missing

• A verified pathogenic alternate-nucleotide change producing p.Arg528Cys is needed.

Sources

Reasoning

No case-level evidence establishes that NM_001904.4:c.1582C>T (p.Arg528Cys) is de novo, that both maternity and paternity were confirmed, that the proband has the disease, or that there is no family history. The reviewed exact-variant literature entry does not mention this variant and provides no parental or proband segregation data.

Rule

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

Evidence considered

• The available exact-variant literature extraction states that CTNNB1 c.1582C>T/p.Arg528Cys is not explicitly mentioned and reports no variant-specific clinical, segregation, or parental findings.
• No case-level proband phenotype, family history, maternity confirmation, paternity confirmation, or parental genotype results are available in the reviewed evidence.

Still missing

• Documented proband disease phenotype
• Parental testing confirming maternity and paternity
• Evidence that the variant is absent in both biological parents
• Documented absence of relevant family history

Sources

Reasoning

No variant-specific functional study demonstrating a damaging effect on CTNNB1 p.Arg528Cys was identified. PMID:28191889 includes CTNNB1 in a broader loss-of-function neurodevelopmental-disorder gene list, but its extracted text reports no p.Arg528Cys-specific assay, result, controls, or reproducibility data. The available CTNNB1 functional-literature lead does not establish that this exact variant was tested. Therefore, the evidence is insufficient to assess PS3 rather than supporting or refuting it.

Rule

PS3: Well-established in vitro or in vivo functional studies supportive of a damaging effect on the gene or gene product.

Evidence considered

• PMID:28191889, Online Methods, Clinical recontact and phenotyping / Patient workups: the paper lists CTNNB1 among genes with loss-of-function mutations but does not report the exact c.1582C>T or p.Arg528Cys variant or a variant-specific functional assay.
• OncoKB reports no variant-specific reviewed functional evidence for CTNNB1 p.Arg528Cys.

Still missing

• A variant-specific in vitro or in vivo functional study with appropriate controls, biologically relevant readout, and reproducible results is needed.

Sources

Reasoning

The governing PS4 definition requires significantly increased prevalence in affected individuals compared with controls, with an OR or RR >5.0 and a confidence interval excluding 1.0. The reviewed literature packet contains no variant-specific PS4 paper, and the only full-text paper reviewed does not mention CTNNB1 c.1582C>T/p.(R528C) or provide variant-specific affected-case and control counts. gnomAD v4.1 reports 10/1,614,008 alleles (AF 6.195756e-06), but no affected-case frequency is available, so an OR and 95% CI cannot be computed and PS4 cannot be adjudicated.

Rule

The prevalence of the variant in affected individuals is significantly increased compared to the prevalence in controls. ... Relative risk (RR) or odds ratio (OR), as obtained from case-control studies, is >5.0 and the confidence interval around the estimate of RR or OR does not include 1.0.

Evidence considered

• PMID:28191889 full-text extraction: CTNNB1 is listed only among genes in a broader loss-of-function neurodevelopmental-disorder analysis; the paper does not report this exact variant or variant-specific case/control counts.
• gnomAD v4.1: NM_001904.4:c.1582C>T is present at 10/1,614,008 alleles (AF 6.195756e-06), but no affected-case frequency is provided, so no case-control OR/RR or 95% CI can be calculated.

Still missing

• Variant-specific affected-case count or frequency and a matched control count or frequency are required to calculate an OR/RR and 95% CI.

Sources

Reasoning

The variant is CTNNB1 p.Arg528Cys. No CTNNB1 VCEP domain table is present, no statistically significant hotspot was identified for CTNNB1 R528, and COSMIC reports the exact variant five times in somatic cancers but does not establish a generic ACMG critical functional domain or hotspot route. The supplied evidence does not establish that residue 528 lies in a well-established critical functional domain without benign variation, so PM1 cannot be assessed.

Rule

Located in a mutational hot spot and/or critical and well-established functional domain (e.g. active site of an enzyme) without benign variation.

Evidence considered

• Cancer Hotspots query CTNNB1 R528 returned no result and stated that the variant does not lie in a statistically significant hotspot.
• COSMIC recorded the exact variant as COSV62722256 with a somatic count of 5; this is somatic occurrence evidence, not by itself proof of a generic ACMG critical domain or hotspot route.

Still missing

• A verified CTNNB1 critical-domain or generic mutational-hotspot source covering residue 528, together with absence of benign variation, is needed.

Sources

Reasoning

The variant is CTNNB1 p.Arg528Cys. The supplied evidence contains no verified pathogenic missense comparator at residue Arg528. The exact variant is a ClinVar uncertain-significance record, and the reviewed literature extract does not mention it. Consequently, the required same-residue pathogenic alternate missense observation is not available for PM5.

Rule

Novel missense change at an amino acid residue where a different missense change determined to be pathogenic has been seen before.

Evidence considered

• ClinVar variation 1984487 classifies p.Arg528Cys as uncertain significance, not pathogenic.
• The PM5 candidate summary reports zero collected same-residue candidates and instructs that PM5 cannot be applied from that artifact alone.
• PMID:28191889 paper extract does not mention CTNNB1 p.R528C.

Still missing

• A verified pathogenic different missense variant at residue Arg528 is needed.

Sources

Reasoning

No case-level evidence establishes an assumed de novo occurrence of NM_001904.4:c.1582C>T (p.Arg528Cys). The available exact-variant literature extraction does not mention this variant and provides no proband phenotype, family history, parental genotype, or parentage information.

Rule

Assumed de novo, but without confirmation of paternity and maternity.

Evidence considered

• The available exact-variant literature extraction states that CTNNB1 c.1582C>T/p.Arg528Cys is not explicitly mentioned and reports no variant-specific clinical, segregation, or parental findings.
• No case-level de novo observation, parental testing result, or family-history description is available in the reviewed evidence.

Still missing

• Evidence that the variant is de novo or presumed de novo in the proband
• Documented proband disease phenotype
• Parental genotype results or an explicit absence of parental testing
• Documented family history

Sources

Reasoning

No affected family members, informative meioses, or variant-positive affected relatives are documented for NM_001904.4:c.1582C>T (p.Arg528Cys). The reviewed exact-variant literature entry does not mention the variant and provides no segregation data.

Rule

Co-segregation with disease in multiple affected family members in a gene definitively known to cause the disease.

Evidence considered

• The available exact-variant literature extraction states that CTNNB1 c.1582C>T/p.Arg528Cys is not explicitly mentioned and reports no variant-specific clinical or segregation findings.
• No family pedigree, affected-relative genotypes, affected-relative phenotypes, or informative meioses are available in the reviewed evidence.

Still missing

• Documented affected relatives
• Genotypes and phenotypes of informative family members
• Number of informative meioses
• Evidence that CTNNB1 is definitively associated with the relevant familial disease in this family

Sources

Reasoning

The available gene-level evidence supports CTNNB1 loss of function as a germline disease mechanism and describes pathogenic nonsense or frameshift variants, while another supplied abstract title concerns CTNNB1 missense variants. However, the evidence does not establish both components required for PP2: a low rate of benign missense variation and missense variants as a common disease mechanism. No validated gene-wide benign-missense-rate metric or quantified missense-mechanism evidence is supplied.

Rule

Missense variant in a gene that has a low rate of benign missense variation and where missense variants are a common mechanism of disease.

Evidence considered

• The gene-context file states that CTNNB1 loss of function is a supported germline disease mechanism and cites literature describing mostly pathogenic nonsense or frameshift variants; it does not provide the low-benign-missense-rate component required for PP2.
• The same gene-context file lists a paper titled Missense variants in CTNNB1 can be associated with vitreoretinopathy, but this does not establish that missense variants are a common mechanism or quantify benign missense variation.

Still missing

• A validated CTNNB1 benign-missense-rate assessment and evidence that missense variants are a common disease mechanism are needed.

Sources

Reasoning

The governing PP4 definition requires a patient phenotype or family history highly specific for a disease with a single genetic etiology. The supplied case evidence does not provide a clinical phenotype or family history for the individual carrying NM_001904.4:c.1582C>T (p.Arg528Cys), and the reviewed paper does not report this exact variant or a variant-specific patient phenotype. PP4 therefore cannot be adjudicated.

Rule

Patient’s phenotype or family history is highly specific for a disease with a single genetic etiology.

Evidence considered

• PMID:28191889 full-text extraction: the paper does not mention CTNNB1 c.1582C>T/p.(R528C) or provide a phenotype for a carrier of this exact variant.
• No case-specific phenotype or family-history description was available in the supplied case evidence.

Still missing

• A documented patient phenotype or family history and its specificity for a single-genetic-etiology disease are required.

Sources

Reasoning

BS2 cannot be assessed from the available evidence. gnomAD shows zero homozygotes for this variant, but the available population records do not document an observed healthy adult individual with the required genotype and phenotype assessment. The single heterozygous observation in the Amish subgroup is a population allele count, not evidence that the carrier was a healthy adult with full penetrance expected at an early age.

Rule

BS2: Observed in a healthy adult individual for a recessive (homozygous), dominant (heterozygous), or X-linked (hemizygous) disorder, with full penetrance expected at an early age.

Evidence considered

• gnomAD v4.1 reports 10/1614008 total alleles and zero homozygotes; the Amish subgroup has 1/910 alleles and zero homozygotes.
• gnomAD v2.1 reports 1/251420 exome alleles and zero homozygotes; the non-cancer exome subset reports 1/236882 alleles and zero homozygotes.
• These population counts do not provide individual-level confirmation that a carrier was a healthy adult or that the relevant phenotype was absent.

Still missing

• A documented healthy adult individual with the relevant genotype and adequate phenotype and age-of-onset information is needed.

Sources

Reasoning

No variant-specific functional study demonstrating no damaging effect on CTNNB1 p.Arg528Cys was identified. PMID:28191889 includes CTNNB1 in a broader loss-of-function neurodevelopmental-disorder gene list, but its extracted text reports no p.Arg528Cys-specific assay, benign functional result, controls, or reproducibility data. The absence of a qualifying assay does not establish a normal protein-function or splicing result, so BS3 cannot be applied.

Rule

BS3: Well-established in vitro or in vivo functional studies show no damaging effect on protein function or splicing.

Evidence considered

• PMID:28191889, Online Methods, Clinical recontact and phenotyping / Patient workups: the paper lists CTNNB1 among genes with loss-of-function mutations but does not report the exact c.1582C>T or p.Arg528Cys variant or a variant-specific functional assay.
• OncoKB reports no variant-specific reviewed functional evidence for CTNNB1 p.Arg528Cys.

Still missing

• A variant-specific well-established functional study showing preserved protein function or splicing, with appropriate controls and reproducible results, is needed.

Sources

Reasoning

No affected family members carrying or lacking NM_001904.4:c.1582C>T (p.Arg528Cys) are documented, so lack of segregation cannot be evaluated. The available exact-variant literature extraction does not mention the variant and provides no family data; therefore phenocopy and multiple-variant explanations cannot be assessed.

Rule

Lack of segregation in affected members of a family.

Evidence considered

• The available exact-variant literature extraction states that CTNNB1 c.1582C>T/p.Arg528Cys is not explicitly mentioned and reports no variant-specific clinical or segregation findings.
• No family pedigree or affected-relative genotype and phenotype data are available to evaluate non-segregation or the phenocopy caveat.

Still missing

• Documented affected family members
• Genotypes of affected family members
• Phenotypes and diagnostic certainty of affected family members
• Assessment of phenocopies or additional pathogenic variants

Sources

Reasoning

BP2 requires either a pathogenic variant in trans in a fully penetrant dominant disorder or a pathogenic variant in cis. No second pathogenic variant, cis/trans phase, affected-proband observation, or parent/offspring phasing result is documented for NM_001904.4:c.1582C>T (p.Arg528Cys). PMID:28191889 was reviewed, but its extracted text only lists CTNNB1 among genes studied and provides no variant-specific clinical or segregation findings.

Rule

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

• PMID:28191889, Online Methods, Clinical recontact and phenotyping / Patient workups: the paper lists CTNNB1 among genes with LGD mutations but does not report variant-specific clinical, segregation, or phase findings for c.1582C>T or p.Arg528Cys.

Still missing

• A confirmed pathogenic partner variant and its cis/trans relationship to NM_001904.4:c.1582C>T are not documented.
• Parent or offspring testing establishing phase is not documented.

Sources

Reasoning

The governing BP5 definition requires the variant to be found in a case with an alternate molecular basis for disease. The supplied evidence does not document a case carrying NM_001904.4:c.1582C>T (p.Arg528Cys) together with an independently established alternate molecular diagnosis. The literature packet and ClinVar record provide no such case evidence, so BP5 cannot be adjudicated.

Rule

Variant found in a case with an alternate molecular basis for disease.

Evidence considered

• No supplied case or literature source documents the exact variant in a patient with an alternate molecular diagnosis.

Still missing

• A case report or cohort record must document this exact variant and an independently established alternate molecular basis for the patient's disease.