MSH3 c.1313C>T, p.Ala438Val

Reasoning

The variant is a missense SNV, p.(Ala438Val), so the generic missense path applies: REVEL is the applicable predictor and SpliceAI is not used for PP3/BP4 in this situation. REVEL is 0.145, which is at or below the generic BP4 supporting threshold of <= 0.29; therefore BP4 is met at supporting strength. The REVEL threshold is the supplied ClinGen SVI calibration from Pejaver et al. 2022 (PMID:36413997).

Rule

Multiple lines of computational evidence suggest no impact on gene or gene product (conservation, evolutionary, splicing impact, etc)

Evidence considered

• Variant consequence is missense SNV NM_002439.5:c.1313C>T, NP_002430.3:p.(Ala438Val), so the generic REVEL-only missense path applies.
• Local REVEL lookup reports score 0.145 for NM_002439.5:c.1313C>T.
• The supplied generic ClinGen SVI REVEL calibration assigns BP4 supporting at <= 0.29; Pejaver et al. 2022 (PMID:36413997).
• SpliceAI max delta 0.003 was not used for BP4 because the governing generic path for a missense variant is REVEL only.

Sources

Reasoning

The variant is NP_002430.3:p.(Ala438Val). The available exact-variant ClinVar record is conflicting, with Likely benign from 4 laboratories, likely benign from 1 laboratory, and Uncertain significance from 3 laboratories; no established pathogenic variant producing the same amino-acid change is documented in the supplied evidence. Therefore PS1 is not met.

Rule

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

Evidence considered

• ClinVar variation 791261, exact variant NM_002439.5(MSH3):c.1313C>T (p.Ala438Val): Likely benign from 4 laboratories, likely benign from 1 laboratory, and Uncertain significance from 3 laboratories; no expert-panel submission and no pathogenic submission.

Sources

Reasoning

The variant is p.Ala438Val in MSH3. No VCEP covers MSH3, vcep_materials.json has no domain-table entry, and the supplied evidence identifies no established MSH3 critical functional domain or mutational hotspot containing residue 438. Cancer Hotspots reports no statistically significant hotspot for MSH3 A438; the single COSMIC somatic observation does not establish a generic ACMG/AMP hotspot or critical-domain criterion. Therefore PM1 is not met.

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

• vcep_materials.json: no CSPEC and domain_tables is empty for MSH3, so no VCEP-approved critical-domain list applies.
• Cancer Hotspots query MSH3 A438: no result; evidence sentence states that the variant does not lie in a statistically significant hotspot.
• COSMIC: COSV99433070, one somatic observation; this is not sufficient by itself to establish a generic ACMG/AMP mutational hotspot or critical functional domain.

Sources

Reasoning

PM2 is not met because the variant is present in population controls at a frequency above the generic PM2 supporting threshold. Under the default all-comers population source, gnomAD v4.1 reports AF=0.000346346 (559/1613994 alleles), which exceeds the supplied threshold of <=0.0001. The variant is therefore neither absent nor extremely rare enough for generic PM2.

Rule

Absent from controls (or at extremely low frequency if recessive) in Exome Sequencing Project, 1000 Genomes or Exome Aggregation Consortium.

Evidence considered

• No VCEP specifies a non-cancer or exome-only source for this gene, so the default all-comers gnomAD source was used.
• gnomAD v4.1: AF=0.000346346 (559/1613994 alleles; 5 homozygotes).
• Generic calibration supplied in this case: PM2 supporting threshold <=0.0001 (ClinGen SVI recommendation cited in the prompt with PMID:25741868).

Sources

Reasoning

The observed variant is p.Ala438Val. The supplied same-residue comparator assessment found zero candidates at residue 438 and no expert-panel or non-expert majority pathogenic/likely pathogenic comparator. The exact-variant ClinVar record is not pathogenic. Thus no different missense change at Ala438 has been established as pathogenic, so PM5 is not met.

Rule

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

Evidence considered

• pm5_candidates.json: residue Ala438, observed alternate Val, total candidates 0, same-residue candidates 0, expert-panel pathogenic/likely pathogenic candidates 0, and non-expert majority pathogenic/likely pathogenic candidates 0.
• ClinVar variation 791261: the exact p.Ala438Val variant has conflicting classifications of Likely benign and Uncertain significance, with no pathogenic classification.

Sources

Reasoning

The variant is a missense SNV, p.(Ala438Val), so the generic missense path applies: REVEL is the applicable predictor and SpliceAI is not used for PP3/BP4 in this situation. REVEL is 0.145, below the generic PP3 supporting threshold of >= 0.644; therefore PP3 is not met. BayesDel is not used because no generic ACMG-strength calibration is established for it. The REVEL threshold is the supplied ClinGen SVI calibration from Pejaver et al. 2022 (PMID:36413997).

Rule

Multiple lines of computational evidence support a deleterious effect on the gene or gene product (conservation, evolutionary, splicing impact, etc.).

Evidence considered

• Variant consequence is missense SNV NM_002439.5:c.1313C>T, NP_002430.3:p.(Ala438Val), so the generic REVEL-only missense path applies.
• Local REVEL lookup reports score 0.145 for NM_002439.5:c.1313C>T.
• The supplied generic ClinGen SVI REVEL calibration assigns PP3 supporting at >= 0.644; Pejaver et al. 2022 (PMID:36413997).
• SpliceAI max delta 0.003 and BayesDel -0.486396 were not used for PP3 because the governing generic path for a missense variant is REVEL only; BayesDel has no established generic ACMG-strength calibration.

Sources

Reasoning

The governing rule concerns a reputable recent pathogenic report, but the generic application specified for this case permits PP5 only for an exact-variant ClinVar expert-panel classification of Pathogenic or Likely pathogenic. ClinVar variation 791261 has no expert-panel submissions; its exact-variant submissions are clinical-laboratory assertions classified as Likely benign or Uncertain significance, with conflicting classifications overall. No qualifying expert-panel pathogenic or likely pathogenic report exists, so 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 variation 791261 records expert_panels as an empty object and reports conflicting classifications from clinical laboratories: Likely benign and Uncertain significance; no expert-panel Pathogenic or Likely pathogenic assertion is present.

Sources

Reasoning

BA1 is not met. The applicable filtering frequency is the gnomAD grpmax FAF, not the African/African American popmax point estimate. gnomAD v4.1 reports grpmax FAF=0.00416115 for African/African American individuals (342/75014 observed alleles), which is below the generic stand-alone threshold of >=0.05. The qualifying population has more than 2000 observed alleles, and no frequency-based BA1 threshold is reached.

Rule

Allele frequency is above 5% in Exome Sequencing Project, 1000 Genomes or Exome Aggregation Consortium.

Evidence considered

• gnomAD v4.1: overall AF=0.000346346 (559/1613994 alleles); African/African American popmax AF=0.00455915 (342/75014 alleles), with filtering grpmax FAF=0.00416115.
• The grpmax population has 75014 observed alleles, exceeding the required 2000-allele minimum; the popmax point estimate was not used.
• Generic calibration supplied in this case: BA1 stand-alone threshold >=0.05, ACMG/AMP 2015 stand-alone benign allele frequency (PMID:25741868).

Sources

Reasoning

BS1 is not met on the supplied generic frequency threshold. The relevant filtering frequency is gnomAD v4.1 grpmax FAF=0.00416115 in African/African American individuals with 75014 observed alleles, below the generic BS1 default threshold of >=0.01. The ancestry-specific popmax point estimate of 0.00455915 was not substituted for FAF. Because the filtering frequency does not reach the default threshold, BS1 is not applied and no maximum-credible-frequency calculation is needed to support this negative determination.

Rule

Allele frequency is greater than expected for the disorder.

Evidence considered

• gnomAD v4.1: African/African American popmax AF=0.00455915 (342/75014 alleles), with filtering grpmax FAF=0.00416115.
• The grpmax population has 75014 observed alleles, exceeding the required 2000-allele minimum; the popmax point estimate was not used for the decision.
• Generic calibration supplied in this case: BS1 default threshold >=0.01; the prompt identifies Whiffin et al. 2017 (PMID:28518168) as the publication supporting per-gene maximum-credible-frequency derivation, but that publication is not an evidence_refs key for this case.

Sources

Reasoning

The governing rule concerns a reputable recent benign report, but the generic application specified for this case permits BP6 only for an exact-variant ClinVar expert-panel classification of Benign or Likely benign. ClinVar variation 791261 has no expert-panel submissions; the available Likely benign assertions are from individual clinical laboratories and the aggregate record is conflicting with Uncertain significance submissions. Ordinary laboratory assertions and aggregate ClinVar labels cannot trigger BP6, so 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 variation 791261 reports 0 expert-panel submissions and only single-submitter laboratory assertions, including Likely benign and Uncertain significance, with conflicting classifications overall; no qualifying expert-panel Benign or Likely benign assertion is present.

Sources

Reasoning

No documented proband disease phenotype, no family-history assessment, and no parental genotyping establishing both maternity and paternity were provided for NM_002439.5:c.1313C>T; therefore the de novo evidence required for PS2 cannot be assessed.

Rule

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

Evidence considered

• The available case evidence does not document parental testing, confirmed maternity and paternity, a de novo result, or the required proband disease and family-history context for this variant.

Still missing

• Documented disease phenotype and family history for the proband
• Parental genotyping confirming maternity and paternity
• Evidence that the variant is absent from both biological parents

Reasoning

No variant-specific validated in vitro or in vivo functional assay was identified for MSH3 NM_002439.5:c.1313C>T (NP_002430.3:p.(Ala438Val)). The available literature extracts do not report this variant or a functional result for it, and OncoKB explicitly reports no variant-specific reviewed functional evidence. ClinVar submissions and computational predictions are not functional assay evidence and cannot establish PS3. Therefore PS3 is not assessed rather than met or not met.

Rule

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

Evidence considered

• OncoKB variant review for MSH3 A438V states that no variant-specific reviewed functional evidence was identified.
• Richards et al. 2015 (PMID:25741868) was reviewed as the generic ACMG/AMP framework source; its supplied extract contains no MSH3 p.(Ala438Val) functional result.
• Karbassi et al. 2016 (PMID:26467025) describes a general pathogenicity-scoring dataset, not a functional assay result for this variant.
• Pritchard et al. 2018 (PMID:29641532) reports cohort sequencing and variant assessment, but the supplied extract contains no MSH3 p.(Ala438Val) functional assay or result.

Still missing

• A reproducible, biologically relevant functional assay of MSH3 p.(Ala438Val), with appropriate wild-type and damaging controls, is not available.

Sources

Reasoning

The governing rule requires increased prevalence in affected individuals versus controls; its note specifies RR or OR >5.0 with a confidence interval that does not include 1.0. The literature-pass packet identified five PS4 candidate papers, but packet.ai_classified_papers is empty. The available paper extracts and reviewed abstracts do not report the exact MSH3 NM_002439.5:c.1313C>T (p.Ala438Val) in affected and unaffected/control groups, nor provide counts, an OR/RR, or a confidence interval. gnomAD provides population control frequency (gnomAD v4.1 AF 0.000346346; 559/1,613,994 alleles) but no matched affected-case frequency, so the required comparison cannot be computed. The criterion is therefore not assessed rather than inferred from ClinVar classifications or gene-level disease context.

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

• gnomAD v4.1 reports NM_002439.5:c.1313C>T at AF 0.000346346 (559/1,613,994 alleles; 5 homozygotes), but this is a population frequency and not an affected-versus-control case-control comparison.
• The literature-pass PS4 index listed PMID:24493721, PMID:25394175, PMID:29641532, PMID:33516529, and PMID:39541563; reviewed variant searches and available extracts found no exact-variant affected/control counts, OR/RR, or confidence interval.
• No PS4 value was computed because the affected-case frequency needed to compare with the population control frequency was unavailable.

Still missing

• An exact-variant case cohort and matched unaffected/control cohort with carrier counts or frequencies are needed to calculate RR/OR and its 95% confidence interval.

Sources

Reasoning

PM3 cannot be assessed because no affected-proband observation documents NM_002439.5:c.1313C>T in trans with a pathogenic or likely pathogenic MSH3 variant, and no parental or offspring testing establishes phase. The available ClinVar record contains no cis/trans, co-occurrence, segregation, or phase data for this variant.

Rule

For recessive disorders, detected in trans with a pathogenic variant Note: This requires testing of parents (or offspring) to determine phase.

Evidence considered

• ClinVar variation 791261 for NM_002439.5(MSH3):c.1313C>T (p.Ala438Val) reports conflicting classifications but provides no affected-proband, second pathogenic-variant, parental, offspring, cis/trans, or phase observation relevant to PM3.

Still missing

• Affected-proband observations with a second pathogenic or likely pathogenic MSH3 variant are not documented.
• Parental or offspring testing establishing trans phase is not documented.

Sources

Reasoning

No documented assumed-de-novo observation or parental testing result was provided for NM_002439.5:c.1313C>T. The available evidence therefore does not establish PM6.

Rule

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

Evidence considered

• The available case evidence contains no documented parental testing, de novo observation, or family data for NM_002439.5:c.1313C>T.

Still missing

• A documented de novo observation in the proband
• Parental genotyping or a clear statement that maternity and paternity were unconfirmed

Reasoning

No affected relatives, informative meioses, variant genotypes, or phenotype-consistent segregation results were documented for NM_002439.5:c.1313C>T. Consequently, co-segregation cannot be assessed.

Rule

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

Evidence considered

• The available case evidence does not provide a pedigree, affected-family-member genotypes, phenotype concordance, or segregation counts for this variant.

Still missing

• Pedigree identifying affected family members
• Variant genotypes for affected and unaffected relatives
• Phenotype assessments and informative meioses

Reasoning

PP2 requires both a low rate of benign missense variation in the gene and missense variants as a common mechanism of disease. The bundle provides population frequency for this single MSH3 variant and literature-derived support for MSH3 loss of function, but it does not provide a gene-wide benign-missense-rate assessment or establish that missense variation is a common MSH3 disease mechanism. The required two-part PP2 evidence is therefore insufficient.

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

• gnomAD v4.1 provides frequency for the individual p.Ala438Val variant (AF 0.000346346; highest population AF 0.00455915), not a gene-wide benign-missense-rate analysis.
• pvs1_gene_context.json supports MSH3 loss of function as a disease mechanism, but does not establish the PP2 requirement that missense variants are a common disease mechanism.

Still missing

• A validated gene-wide assessment showing a low rate of benign MSH3 missense variation.
• Evidence that missense variants are a common mechanism of MSH3-related disease.

Sources

Reasoning

The governing rule requires a patient's phenotype or family history that is highly specific for a disease with a single genetic etiology. The supplied case evidence contains no patient phenotype, family-history description, or disease-specific clinical assessment for this variant; ClinVar condition labels and gene-level cancer context are not a patient phenotype. Therefore PP4 cannot be assessed.

Rule

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

Still missing

• A documented patient phenotype and relevant family history, linked to a disease with a sufficiently specific single genetic etiology, are needed.

Reasoning

BS2 is not assessed. The variant is observed in gnomAD homozygotes (1 in v2.1 and 5 in v4.1), but the population records do not establish that these individuals were healthy adults. They also do not establish that the relevant MSH3-associated disorder has full penetrance expected at an early age, so the homozygote observations cannot be converted into BS2 under the generic rule.

Rule

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 v2.1 reports 1 homozygote among 159/282616 total alleles.
• gnomAD v4.1 reports 5 homozygotes among 559/1613994 total alleles, including 2 in the African/African American population.
• The available frequency records do not identify the homozygotes as healthy adults or provide age, phenotype, or penetrance information.

Still missing

• Confirm whether the observed homozygotes were healthy adults at an age appropriate for the relevant MSH3-associated disorder.
• Establish that the relevant disorder is expected to show full penetrance at an early age before applying BS2.

Sources

Reasoning

No variant-specific validated in vitro or in vivo functional assay was identified for MSH3 NM_002439.5:c.1313C>T (NP_002430.3:p.(Ala438Val)). The available literature extracts do not report this variant or a functional result for it, and OncoKB explicitly reports no variant-specific reviewed functional evidence. ClinVar submissions and computational predictions are not functional assay evidence and cannot establish BS3. Therefore BS3 is not assessed rather than met or not met.

Rule

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

Evidence considered

• OncoKB variant review for MSH3 A438V states that no variant-specific reviewed functional evidence was identified.
• Richards et al. 2015 (PMID:25741868) was reviewed as the generic ACMG/AMP framework source; its supplied extract contains no MSH3 p.(Ala438Val) functional result.
• Karbassi et al. 2016 (PMID:26467025) describes a general pathogenicity-scoring dataset, not a functional assay result for this variant.
• Pritchard et al. 2018 (PMID:29641532) reports cohort sequencing and variant assessment, but the supplied extract contains no MSH3 p.(Ala438Val) functional assay or result.

Still missing

• A reproducible, biologically relevant functional assay of MSH3 p.(Ala438Val), with appropriate wild-type and benign controls, is not available.

Sources

Reasoning

No affected family members with documented absence of NM_002439.5:c.1313C>T were identified, and no family segregation study or phenotype adjudication was provided. Lack of segregation therefore cannot be assessed.

Rule

Lack of segregation in affected members of a family.

Evidence considered

• The available case evidence contains no documented affected relatives who lack the variant and no family-level non-segregation analysis.

Still missing

• Affected relatives with confirmed disease phenotypes
• Reliable variant testing results for those affected relatives
• Assessment for phenocopies or multiple pathogenic variants if non-segregation is observed

Reasoning

BP1 requires evidence that disease in the gene is caused primarily by truncating variants. The supplied gene-level context supports MSH3 loss of function as a disease mechanism, but it does not quantify the proportion of disease-causing variants that are truncating or establish that truncating variants are the primary known mechanism relative to missense variants. The specific BP1 premise is therefore not established.

Rule

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

Evidence considered

• pvs1_gene_context.json: MSH3 loss of function is supported as a disease mechanism, but the file does not establish that disease-causing variants are primarily truncating.

Still missing

• A gene-level variant-spectrum analysis demonstrating that MSH3 disease is primarily caused by truncating variants.

Sources

Reasoning

BP2 cannot be assessed because no observation documents NM_002439.5:c.1313C>T in cis with a pathogenic variant. The available ClinVar record contains no cis/trans or co-occurrence data; therefore, the cis branch of BP2 is unresolved, and no evidence was found for the dominant trans branch.

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

• ClinVar variation 791261 for NM_002439.5(MSH3):c.1313C>T (p.Ala438Val) reports conflicting classifications but provides no cis-phase, trans-phase, co-occurrence, or inheritance observation relevant to BP2.

Still missing

• Cis-phase testing with a pathogenic variant is not documented.
• A fully penetrant dominant disorder context and trans observation are not documented.

Sources

Reasoning

The governing rule requires the variant to be found in a case with an alternate molecular basis for disease. The available case evidence does not document an individual case carrying this variant together with an independently established alternate molecular diagnosis or causal variant. Population frequency, ClinVar laboratory submissions, and general cancer literature do not satisfy BP5.

Rule

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

Still missing

• A case report or clinical record documenting the variant and a confirmed alternate molecular basis for the patient's disease is needed.