MUTYH c.1103G>A, p.Gly368Asp

Reasoning

The queried variant is NP_001041639.1:p.(Gly368Asp), c.1103G>A. PMID:11818965 reports the identical Gly>Asp substitution as MYH Gly382Asp caused by c.1145G>A under an alternate MUTYH numbering convention and describes affected siblings with colorectal polyposis/carcinoma carrying that allele. PMID:16557584 likewise identifies the corresponding G382D substitution as an assumed pathogenic MUTYH missense mutation and reports it in biallelic MAP cases. These are the same amino-acid change, not merely a different change at the same residue; therefore PS1 is met.

Rule

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

Evidence considered

• PMID:11818965, Results: the paper reports MYH Gly382Asp (1145G>A), corresponding to the queried MUTYH p.Gly368Asp (c.1103G>A) under the alternate numbering convention, in affected siblings with multiple colorectal adenomas and carcinoma.
• PMID:16557584, Results: c.1145G>A/p.G382D, corresponding to the queried p.Gly368Asp, is identified as an assumed pathogenic missense mutation and occurs in biallelic MAP cases.

Sources

Reasoning

PMID:11818965 directly evaluated the allele reported as Gly382Asp (the alternate MUTYH numbering corresponding to NP_001041639.1:p.(Gly368Asp), NM_001048174.2:c.1103G>A). In an in vitro single-turnover adenine-removal assay using the equivalent E. coli MutY Gly253Asp protein, activity was reduced by approximately 86% for G:A and 85% for 8-oxoG:A substrates relative to wild type. The study also provided family observations and RT-PCR analysis showing no aberrant splicing. This is biologically relevant functional evidence with wild-type comparison and disease-relevant substrates, but it used a bacterial homolog and a single study rather than a validated, reproducible clinical diagnostic assay; therefore PS3 is applied at supporting strength, not stronger strength.

Rule

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

Evidence considered

• Al-Tassan et al. 2002 (PMID:11818965), family N and functional-assay results for the alternate Gly382Asp MUTYH numbering corresponding to the queried c.1103G>A/p.Gly368Asp allele: equivalent E. coli MutY Gly253Asp showed approximately 86% reduced activity on G:A and 85% reduced activity on 8-oxoG:A substrates versus wild type.
• Al-Tassan et al. 2002 (PMID:11818965): RT-PCR/cloning of normal colonic mucosa found no aberrant splicing for the Gly382Asp allele; the functional assay used wild-type comparison and biologically relevant adenine-removal substrates.

Still missing

• No independently replicated, clinically validated diagnostic-laboratory functional assay was identified.

Sources

Reasoning

Met. Aretz et al. evaluated 329 unrelated APC-mutation-negative polyposis patients and 116 healthy German controls. The queried variant NM_001048174.2:c.1103G>A, p.(Gly368Asp), is the same amino-acid substitution reported under the alternate MUTYH numbering c.1145G>A, p.G382D. The variant was present in 31/658 case chromosomes and 0/232 control chromosomes. This is a case-control comparison even though the paper did not print an odds ratio. Because the control cell was zero, I used a Haldane-Anscombe continuity correction: OR = (31.5 x 232.5)/(626.5 x 0.5) = 23.38; approximate 95% CI = 1.42-383.64. The exact governing comparison is OR >5.0 with a confidence interval that does not include 1.0; 23.38 > 5.0 and the CI excludes 1.0, so PS4 is met. The literature_pass PS4 index was checked; its listed papers were not already read as PS4 in packet.ai_classified_papers, which contains only PMID:20848659 classified for PS3. PMID:16557584 was verified directly because it provided the case-control counts.

Rule

The prevalence of the variant in affected individuals is significantly increased compared to the prevalence in controls. ... Note 1: 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

• Aretz et al. 2006, Results/Table II: the alternate-numbered c.1145G>A (p.G382D), corresponding to the queried c.1103G>A (p.Gly368Asp), occurred in 31/658 alleles among 329 polyposis patients; none of the assumed pathogenic variants was detected in 232 control chromosomes.
• Computed the case-control odds ratio because the paper did not print one: Haldane-Anscombe corrected OR = 23.38, approximate 95% CI = 1.42-383.64.
• The case-control direction is enrichment in affected patients, and the substituted inequality is 23.38 > 5.0? yes; the 95% CI 1.42-383.64 does not include 1.0.
• The literature_pass packet.ai_classified_papers entry for PMID:20848659 was verified as PS3 only, not PS4; PMID:16557584 was independently verified for the PS4 counts.

Sources

Reasoning

The variant is reported under the equivalent historical MUTYH numbering as Gly382Asp (1145G>A). In family N, three affected siblings with multiple colorectal adenomas and carcinoma were compound heterozygotes for MYH Tyr165Cys and Gly382Asp, establishing that the queried allele was detected in trans with another disease-associated MUTYH variant. This satisfies PM3 for the autosomal-recessive MUTYH-associated polyposis context. The phase conclusion is based on the reported compound-heterozygous family genotype, not on the ClinVar classification.

Rule

PM3: For recessive disorders, detected in trans with a pathogenic variant ... Note: This requires testing of parents (or offspring) to determine phase.

Evidence considered

• Al-Tassan et al. 2002 (PMID:11818965), family N: the queried c.1103G>A/p.(G368D) allele is reported under alternate numbering as Gly382Asp (1145G>A), and three affected siblings were compound heterozygotes for Tyr165Cys and Gly382Asp.
• The family-level compound-heterozygous observation supplies the required trans-phase evidence; no ClinVar classification was used as a substitute for phase evidence.

Sources

Reasoning

PMID:11818965 describes family N, in which three affected siblings with multiple colorectal adenomas and carcinoma were compound heterozygotes for the MYH Tyr165Cys and Gly382Asp alleles; the paper's Gly382Asp notation corresponds to the queried MUTYH c.1103G>A, p.(Gly368Asp) under the alternate numbering system. Unaffected relatives were reported as heterozygous for one variant or normal. This is co-segregation in multiple affected family members, supporting PP1 at supporting strength; no quantitative meiosis or likelihood-ratio threshold was supplied by the governing generic rule, and the evidence comes from one family with alternate numbering that warrants human confirmation.

Rule

Co-segregation with disease in multiple affected family members in a gene definitively known to cause the disease. ... May be used as stronger evidence with increasing segregation data.

Evidence considered

• Al-Tassan et al., PMID:11818965, reports three affected siblings in family N with multiple colorectal adenomas and carcinoma who were compound heterozygotes for MYH Tyr165Cys and Gly382Asp; the paper identifies Gly382Asp as a guanine-to-adenine substitution at nucleotide 1145 in exon 13.
• The paper extract maps the reported Gly382Asp substitution under the alternate numbering convention to the queried MUTYH c.1103G>A, p.(Gly368Asp), and reports unaffected relatives as heterozygous for one variant or normal.
• The generic PP1 rule requires co-segregation with disease in multiple affected family members; three affected siblings satisfy the qualitative family-member requirement.

Still missing

• Independent confirmation that the paper's alternate Gly382Asp notation is identical to the queried c.1103G>A, p.(Gly368Asp) transcript representation.
• An auditable pedigree and quantified informative meioses or segregation likelihood ratio.

Sources

Reasoning

The variant is a missense substitution, NM_001048174.2:c.1103G>A (p.Gly368Asp), so the governing generic rule selects the missense REVEL path only; SpliceAI is not used for PP3/BP4 in this situation. REVEL is 0.954, meeting the generic strong PP3 threshold of >= 0.932. BayesDel is not used because no generic ACMG-strength calibration is established. The ClinGen SVI REVEL calibration is 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.). ... Missense variant -> REVEL only. ... PP3 (strong): >= 0.932 — ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet) (PMID:36413997)

Evidence considered

• Local REVEL predictor lookup: REVEL score 0.954 for NM_001048174.2:c.1103G>A (p.Gly368Asp).
• Generic calibration applied verbatim: REVEL PP3 strong threshold >= 0.932, from ClinGen SVI REVEL calibration (Pejaver et al. 2022, American Journal of Human Genetics; PMID:36413997).
• Path selection: this is a missense variant, so REVEL is the sole PP3/BP4 predictor path; the available SpliceAI max delta 0.02 was not used for this missense criterion.

Sources

Reasoning

PM2 is not met. The supplied generic PM2 supporting threshold is allele frequency at or below 0.0001. This variant is present in gnomAD v4.1 at AF=0.00473005 (7,634/1,613,938 alleles), with 26 homozygotes; its grpmax FAF is 0.00576855. These observations are far above the PM2 threshold and are incompatible with absence or extremely low frequency in controls.

Rule

PM2 (supporting): <= 0.0001 — SVI recommendation to downgrade PM2 to Supporting (ClinGen SVI 2020)

Evidence considered

• gnomAD v4.1 reports AF=0.00473005, 7,634/1,613,938 alleles, and 26 homozygotes; grpmax FAF=0.00576855.
• The observed gnomAD v4.1 frequency is greater than the supplied generic PM2 threshold of 0.0001, and the variant is not absent from controls.
• The MUTYH InSiGHT CSPEC is listed but has no released criterion rules, so the generic ACMG/AMP PM2 definition and supplied ClinGen SVI threshold govern.

Sources

Reasoning

Not met. ClinVar contains an exact-variant record for NM_001048174.2:c.1103G>A (p.Gly368Asp) with Pathogenic/Likely pathogenic laboratory submissions, but the audit records zero expert-panel submissions. Under the required PP5 restriction, ordinary laboratory submissions, aggregate labels, and non-expert assertions do not trigger PP5; no exact-variant ClinVar expert-panel pathogenic or likely pathogenic classification is available.

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-variant record 5294 reports Pathogenic/Likely pathogenic laboratory submissions, while the submission audit records expert_panel_submissions = 0.
• The available ClinVar classifications are not an expert-panel classification and therefore do not satisfy the required PP5 source restriction.

Sources

Reasoning

BA1 is not met. Generic ACMG/AMP BA1 requires an allele frequency of at least 0.05. The governing frequency measure is gnomAD v4.1 grpmax FAF, 0.00576855 (0.576855%), based on the European (non-Finnish) population with 1,180,038 observed alleles; this is below 0.05, and the qualifying population exceeds the 2,000-observed-allele minimum.

Rule

BA1 (stand-alone): >= 0.05 — ACMG/AMP 2015 stand-alone benign allele frequency (PMID:25741868)

Evidence considered

• gnomAD v4.1 reports grpmax FAF=0.00576855, with the highest observed population frequency in European (non-Finnish) individuals: AF=0.00588456, 6,944/1,180,038 alleles, 25 homozygotes.
• The gnomAD v4.1 European (non-Finnish) population has 1,180,038 observed alleles, exceeding the 2,000-observed-allele requirement for using the grpmax FAF.
• The MUTYH InSiGHT CSPEC is listed but has no released criterion rules, so the generic ACMG/AMP thresholds supplied for this case govern.

Sources

Reasoning

BS1 is not met. The generic default threshold is 0.01, but the required filtering measure is gnomAD grpmax FAF rather than the popmax point estimate. gnomAD v4.1 grpmax FAF is 0.00576855 (0.576855%), below 0.01; therefore no maximum-credible-frequency calculation is needed to reject the default BS1 path, despite MUTYH-associated polyposis being recessive and potentially attenuated.

Rule

Generic rules for BS1: the criterion is 'allele frequency is greater than expected for the disorder'; the 1% figure is only a default.

Evidence considered

• Generic BS1 uses allele frequency greater than expected for the disorder, with 0.01 as the supplied default threshold; the gnomAD v4.1 grpmax FAF is 0.00576855, below 0.01.
• The gnomAD v4.1 maximum observed subgroup point estimate is AF=0.00588456 in European (non-Finnish) individuals, but the point estimate is not substituted for the filtering allele frequency.
• The gnomAD v4.1 European (non-Finnish) population has 1,180,038 observed alleles, exceeding the 2,000-observed-allele requirement for the grpmax FAF.

Sources

Reasoning

The available direct functional evidence does not show no damaging effect. Instead, the assay corresponding to the queried MUTYH Gly>Asp allele reported approximately 85–86% reduced adenine-removal activity relative to wild type, supporting impaired function. No well-established benign functional or splicing result was identified, so BS3 is not met.

Rule

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

Evidence considered

• Al-Tassan et al. 2002 (PMID:11818965): the corresponding Gly>Asp MUTYH allele showed approximately 85–86% reduced adenine-removal activity compared with wild type, and no benign functional result was reported.

Sources

Reasoning

The available family observation places the queried allele in trans with Tyr165Cys, not in cis with a pathogenic variant. MUTYH-associated polyposis is described in the case as autosomal recessive, so the fully penetrant dominant-gene/disorder branch of BP2 is not applicable; the cis branch is not satisfied.

Rule

BP2: 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

• Al-Tassan et al. 2002 (PMID:11818965) reports the queried allele and Tyr165Cys as compound heterozygous alleles in affected siblings, supporting trans rather than cis phase.
• The case summary identifies the MUTYH disease context as autosomal recessive; therefore BP2 can only be supported here by cis observation, which was not reported.

Sources

Reasoning

The variant is a missense substitution, so the governing generic rule selects the missense REVEL path only; SpliceAI is not used for BP4 in this situation. REVEL is 0.954, which does not meet any benign BP4 threshold, including the strong threshold of <= 0.016. The SpliceAI max delta of 0.02 is therefore not transferable to BP4 for this missense variant. BayesDel is not usable because no generic ACMG-strength calibration is established. The ClinGen SVI REVEL calibration is 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). ... Missense variant -> REVEL only. ... BP4 (strong): <= 0.016 — ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet) (PMID:36413997)

Evidence considered

• Local REVEL predictor lookup: REVEL score 0.954 for NM_001048174.2:c.1103G>A (p.Gly368Asp).
• Generic calibration applied verbatim: REVEL BP4 strong threshold <= 0.016, from ClinGen SVI REVEL calibration (Pejaver et al. 2022, American Journal of Human Genetics; PMID:36413997).
• Path selection: this is a missense variant, so REVEL is the sole PP3/BP4 predictor path; the available SpliceAI max delta 0.02 cannot substitute for a non-supportive REVEL result.

Sources

Reasoning

Not met. The exact-variant ClinVar record contains Pathogenic/Likely pathogenic laboratory assertions and no benign or likely benign expert-panel classification. Ordinary submissions, aggregate labels, and non-expert assertions cannot trigger BP6, and no qualifying exact-variant expert-panel benign source is present.

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-variant record 5294 reports Pathogenic/Likely pathogenic laboratory submissions; the audit records expert_panel_submissions = 0 and no benign expert-panel assertion.

Sources

Reasoning

No case-specific evidence documents a de novo occurrence, confirmed maternity and paternity, the patient's disease phenotype, or absence of family history. The available literature describes familial affected siblings rather than a de novo case, so 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 case evidence contains no auditable patient-level de novo report or parental genotype results.
• PMID:11818965 reports a familial series with three affected siblings carrying the variant under alternate MUTYH numbering, not a de novo occurrence.

Still missing

• Case-level documentation of the patient's disease phenotype and family history.
• Variant testing results for both biological parents with confirmed maternity and paternity.

Sources

Reasoning

The MUTYH VCEP material has no released criteria and its domain_tables entry is empty, so no authoritative approved critical-domain list is available for residue 368. The case evidence reports no statistically significant hotspot and no VCEP-named hotspot route. The available paper evidence establishes functional disease relevance for the amino-acid change but does not establish that residue 368 lies in a qualifying mutational hotspot or critical, well-established functional domain without benign variation. PM1 therefore cannot be assessed from the available evidence.

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

• The MUTYH VCEP specification is listed but has no released criteria, and vcep_materials.json contains an empty domain_tables object.
• Cancer Hotspots result: This variant does not lie in a statistically significant hotspot.
• No available source establishes residue 368 as part of a qualifying critical functional domain without benign variation.

Still missing

• Authoritative MUTYH critical-domain boundaries or a qualifying hotspot designation covering residue 368
• Evidence that the qualifying domain or hotspot lacks benign variation

Sources

Reasoning

The available evidence confirms a previously pathogenic variant with the same Gly-to-Asp amino-acid change as the query, which supports PS1 rather than PM5. No different missense substitution at MUTYH residue 368 is documented as pathogenic in the available comparator evidence, and the PM5 candidate file contains no candidates. A complete same-residue pathogenic comparator search is therefore not established, so PM5 is not assessed.

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: zero same-residue candidates were collected.
• PMID:11818965 and PMID:16557584 document the same Gly-to-Asp substitution under alternate MUTYH numbering, not a different missense substitution at the residue.

Still missing

• A documented pathogenic missense substitution other than Gly368Asp at MUTYH residue 368

Sources

Reasoning

No case-specific evidence documents an assumed de novo occurrence. The available literature describes familial affected siblings rather than a de novo case without parental confirmation, so PM6 cannot be assessed.

Rule

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

Evidence considered

• The case evidence contains no auditable patient-level assumed-de-novo report or parental genotype results.
• PMID:11818965 reports a familial series with three affected siblings carrying the variant under alternate MUTYH numbering, not an assumed de novo occurrence.

Still missing

• Case-level documentation that the variant is absent from available parental testing or is otherwise assumed de novo.
• The patient's disease phenotype and family-history assessment.

Sources

Reasoning

The available case evidence documents pathogenic MUTYH missense variants, including the queried Gly-to-Asp change, but it does not provide the gene-level evidence required to establish both a low rate of benign missense variation and missense variants as a common disease mechanism. PP2 is therefore not assessed.

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.

Still missing

• Gene-level benign missense variation rate
• Evidence that missense variation is a common MUTYH disease mechanism

Reasoning

Not assessed. The available case materials do not provide the individual patient's phenotype or family history. PMID:16557584 describes phenotypes in a study cohort of MUTYH-associated polyposis patients, including attenuated or atypical polyposis, but cohort-level phenotype data cannot establish that this patient's phenotype or family history is highly specific for a single genetic etiology.

Rule

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

Evidence considered

• Aretz et al. 2006 describes cohort phenotypes but does not identify the phenotype of this case; it reports that most cohort patients had attenuated or atypical polyposis, which is not sufficient for case-specific PP4.

Still missing

• Individual patient's clinical phenotype and family history, including adenoma burden, age at onset, colorectal cancer history, and relevant extraintestinal findings, are unavailable.

Sources

Reasoning

BS2 is not assessed. gnomAD v4.1 reports 26 homozygotes overall and 25 European (non-Finnish) homozygotes, but the database record does not document that these are healthy adults, their ages, clinical phenotypes, or absence of MUTYH-associated polyposis. Homozygote counts alone therefore do not satisfy the generic BS2 requirement.

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 v4.1 reports 26 homozygotes overall and 25 homozygotes among European (non-Finnish) individuals.
• The gnomAD evidence does not provide individual-level age, health status, phenotype, or clinical follow-up for those homozygotes.
• The MUTYH InSiGHT CSPEC is listed but has no released criterion rules, so the generic ACMG/AMP BS2 definition governs.

Still missing

• No individual-level documentation establishes that the reported homozygotes are healthy adults without MUTYH-associated polyposis.
• No age, penetrance, or clinical follow-up data are available for the gnomAD homozygotes.

Sources

Reasoning

No affected family member is documented as lacking the variant. The available family report instead describes three affected siblings carrying the relevant allele and unaffected relatives who were heterozygous for one variant or normal; this does not establish lack of segregation in affected members. The generic caveat also requires consideration of phenocopies and more than one pathogenic variant.

Rule

Lack of segregation in affected members of a family.

Evidence considered

• PMID:11818965 reports three affected siblings carrying the relevant Gly>Asp allele under alternate numbering and does not report an affected relative who lacks that allele.
• The paper extract reports unaffected relatives as heterozygous for one variant or normal, which is not evidence of non-segregation among affected members.
• The generic BS4 caveat requires consideration of phenocopies and families with more than one pathogenic variant before interpreting an apparent lack of segregation.

Still missing

• Auditable pedigree with genotype and phenotype results for affected relatives.
• Documentation of any affected relatives who lack the variant.
• Assessment for phenocopies and more than one pathogenic variant in the family.

Sources

Reasoning

MUTYH disease-associated missense variants are documented in the available literature, but the case evidence does not establish that MUTYH disease is caused primarily by truncating variants. The required gene-level variant-spectrum comparison is unavailable, so BP1 is not assessed.

Rule

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

Still missing

• Gene-level evidence that truncating variants are the primary known disease-causing variant type

Reasoning

Not assessed. The reviewed evidence does not document that this variant was found in a case with an alternate molecular basis for disease. The available literature concerns MUTYH-associated polyposis and related variant observations, but it does not establish an alternate molecular diagnosis for this case.

Rule

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

Evidence considered

• Aretz et al. 2006 reports MUTYH mutation findings in polyposis patients but does not establish an alternate molecular basis for the present case.

Still missing

• Case-level molecular workup and an independently established alternate molecular diagnosis are not available.

Sources