MET c.3682G>A, p.Asp1228Asn

Reasoning

PM2 is met at supporting strength. The variant is reported absent from gnomAD v2.1 and gnomAD v4.1. The case has no governing VCEP requiring a different population source, so the all-comers datasets are the default; the available non-cancer records also report absence. This is consistent with the supplied generic PM2 supporting threshold of <=0.0001, since the observed allele frequency is zero in the queried datasets.

Rule

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

Evidence considered

• gnomAD v2.1 reports the variant absent.
• gnomAD v4.1 reports the variant absent.
• The available gnomAD v2.1 non-cancer and gnomAD v3.1 non-cancer records also report the variant absent; no VCEP-specific source requirement applies.
• Generic calibration supplied for PM2 is <=0.0001 and supports downgrading PM2 to Supporting, identified in the case instructions as the ClinGen SVI 2020 recommendation citing ACMG/AMP 2015 (PMID:25741868).

Sources

Reasoning

NM_000245.4:c.3682G>A is a missense variant producing p.(Asp1228Asn), so the governing generic PP3/BP4 path is REVEL only; SpliceAI and BayesDel are not used for this missense assessment. REVEL is 0.723, meeting the generic PP3 supporting threshold of >=0.644 from the ClinGen SVI REVEL calibration (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.). ... PP3 (supporting): >= 0.644 — ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet) (PMID:36413997)

Evidence considered

• Variant consequence: missense, NM_000245.4:c.3682G>A, NP_000236.2:p.(Asp1228Asn), from the normalized case evidence.
• REVEL score 0.723; generic PP3 supporting threshold >=0.644, ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet, PMID:36413997).
• SpliceAI is not counted because the generic rule selects REVEL only for a missense variant; BayesDel is not used because no generic ACMG-strength calibration is established.

Sources

Reasoning

The variant is MET p.Asp1228Asn (D1228N). PMID:28522754 reports D1228N in a patient with acquired resistance to crizotinib, but the source does not establish this variant as pathogenic, and no previously established pathogenic variant producing the same amino-acid change by a different nucleotide substitution was identified. Therefore PS1 is not met. The reported resistance-associated D1228N observation cannot be converted into PS1 solely from the paper or from the ClinVar pathogenic submission without assertion criteria.

Rule

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

Evidence considered

• PMID:28522754 explicitly reports MET D1228N in a patient with MET exon 14-skipping lung adenocarcinoma who developed acquired resistance to crizotinib, but describes it as a subclonal resistance-associated missense mutation rather than an established pathogenic germline variant.
• ClinVar variation 13884 has an OMIM pathogenic submission without assertion criteria and a single-submitter uncertain-significance submission; neither provides a validated PS1 comparator.

Still missing

• A validated pathogenic comparator producing the same p.Asp1228Asn amino-acid change by a different nucleotide change is not available.

Sources

Reasoning

The queried variant is MET p.Asp1228Asn. PMID:27694386 documents a different missense change at the same residue, D1228V, and performs functional resistance studies on D1228V, but does not determine D1228V to be pathogenic in the ACMG/AMP disease-classification sense. The paper describes both substitutions as acquired drug-resistance mutations in cancer. Thus the required different missense change at the same residue determined to be pathogenic is not present, and 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

• PMID:27694386 reports acquired MET D1228V at the same residue and evaluates its drug-resistance mechanism in modeling and cell assays, while separately describing D1228N as a previously reported acquired-resistance observation.
• The paper does not provide an ACMG/AMP pathogenic determination for D1228V.

Still missing

• A different MET missense variant at Asp1228 with a validated pathogenic determination is not available.

Sources

Reasoning

PP5 is not met under the required generic rule because the exact-variant ClinVar record has no expert-panel submission. The only exact-variant Pathogenic submission is OMIM with no assertion criteria and no expert-panel status; the other submission is Uncertain significance from a single laboratory. The exact-variant ClinVar classification therefore does not meet the required expert-panel condition for PP5.

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 NM_000245.4(MET):c.3682G>A (p.Asp1228Asn) has 0 expert-panel submissions.
• The exact-variant OMIM submission is Pathogenic but has review status 'no assertion criteria provided', is not an expert-panel submission, and is classified as weak/literature-only in the submission audit.
• The other exact-variant submission is Uncertain significance from a single clinical laboratory, not a reputable expert-panel pathogenic classification.
• The reviewed papers do not constitute an exact-variant ClinVar expert-panel classification.

Sources

Reasoning

BA1 is not met. The variant is absent from gnomAD v2.1 and gnomAD v4.1; therefore no filtering allele frequency or grpmax FAF is available, and the allele-frequency threshold of >=0.05 is not reached. The generic BA1 rule is not overridden by the available ClinVar record because it reports conflicting historical/non-expert information rather than a reliable pathogenic consensus, and the population data do not support a stand-alone benign call.

Rule

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

Evidence considered

• gnomAD v2.1 reports the variant absent; no allele count, allele number, popmax point estimate, or grpmax FAF is reported.
• gnomAD v4.1 reports the variant absent; no allele count, allele number, popmax point estimate, or grpmax FAF is reported.
• Generic calibration supplied for BA1 is >=0.05, identified in the case instructions as the ACMG/AMP 2015 stand-alone benign allele-frequency threshold (PMID:25741868).

Sources

Reasoning

BS1 is not met on the available population data. The variant is absent from gnomAD v2.1 and v4.1, so there is no filtering allele frequency or grpmax FAF above the generic default threshold of >=0.01. Because no elevated frequency is observed, an ancestry-specific maximum-credible-frequency calculation is not required for this criterion.

Rule

Allele frequency is greater than expected for the disorder.

Evidence considered

• gnomAD v2.1 reports the variant absent; no filtering allele frequency or grpmax FAF is available.
• gnomAD v4.1 reports the variant absent; no filtering allele frequency or grpmax FAF is available.
• Generic calibration supplied for BS1 is >=0.01; the case instructions identify Whiffin et al. 2017 (PMID:28518168) as the preferred per-gene maximum-credible-frequency framework.

Sources

Reasoning

NM_000245.4:c.3682G>A is a missense variant producing p.(Asp1228Asn), so the governing generic PP3/BP4 path is REVEL only; SpliceAI cannot substitute for a gray-zone REVEL result. REVEL is 0.723, which is above the generic BP4 supporting cutoff of <=0.29, and therefore BP4 is not met. BayesDel is not used because no generic ACMG-strength calibration is established.

Rule

Multiple lines of computational evidence suggest no impact on gene or gene product (conservation, evolutionary, splicing impact, etc). ... BP4 (supporting): <= 0.29 — ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet) (PMID:36413997)

Evidence considered

• Variant consequence: missense, NM_000245.4:c.3682G>A, NP_000236.2:p.(Asp1228Asn), from the normalized case evidence.
• REVEL score 0.723; generic BP4 supporting threshold <=0.29, ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet, PMID:36413997).
• SpliceAI is not counted because the generic rule selects REVEL only for a missense variant, and a gray-zone or non-supportive REVEL result does not license substitution of the splice path.

Sources

Reasoning

BP6 is not met under the required generic rule because the exact-variant ClinVar record contains no expert-panel submission and no expert-panel Benign or Likely benign classification. The available submissions are Pathogenic without assertion criteria and Uncertain significance from a single laboratory.

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 NM_000245.4(MET):c.3682G>A (p.Asp1228Asn) has 0 expert-panel submissions.
• The exact-variant submissions are OMIM Pathogenic with no assertion criteria and Ambry Genetics Uncertain significance; neither is an expert-panel Benign or Likely benign assertion.
• No exact-variant ClinVar expert-panel benign classification is present.

Sources

Reasoning

The available case evidence does not document a proband with a confirmed disease phenotype, absence of family history, or confirmed maternity and paternity for NM_000245.4:c.3682G>A; therefore de novo status 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 documents the variant and general ClinVar/literature records but provides no proband-level de novo report or parental testing results.
• No evidence of confirmed maternity, confirmed paternity, or absence of the variant in both biological parents was found in the supplied case materials.

Still missing

• Proband disease phenotype and diagnostic context
• Family-history assessment
• Parental testing confirming both maternity and paternity
• Evidence that the variant is absent from both biological parents

Reasoning

The available literature does not establish PS3 for NM_000245.4:c.3682G>A (NP_000236.2:p.(Asp1228Asn)). PMID:28522754 reports D1228N in a clinical progression sample and states that it had previously been reported to activate MET and cause inhibitor resistance in vitro, but the study itself performed no D1228N-specific functional assay. PMID:27694386 likewise mentions D1228N only as a prior clinical resistance observation; its functional experiments evaluated the different substitution D1228V. No validated, reproducible D1228N assay with appropriate controls and a directly interpretable damaging readout is available, so the evidence is insufficient to apply PS3 at any strength.

Rule

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

Evidence considered

• PMID:28522754, Results, 'Response to crizotinib and acquired resistance in a patient with METΔ14-Mutated NSCLC,' paragraph 2: D1228N was detected as a subclonal mutation and described as previously reported to activate MET and cause inhibitor resistance in vitro, but this paper did not perform a D1228N-specific functional assay.
• PMID:27694386, Discussion, paragraph 2: D1228N was cited as a prior clinical resistance observation, while the paper's functional and modeling experiments used MET D1228V rather than D1228N.
• No qualifying assay controls, reproducibility data, or diagnostic-laboratory validation were available for the exact p.Asp1228Asn substitution.

Still missing

• A validated, reproducible functional assay directly testing p.Asp1228Asn with appropriate positive, negative, and wild-type controls is needed.

Sources

Reasoning

The governing PS4 threshold is an odds ratio or relative risk >5.0 with a confidence interval that does not include 1.0. The reviewed variant-specific literature reports D1228N in isolated clinical cancer observations, but provides no affected-case versus unaffected/population-control counts, case frequency, matched-ancestry gnomAD frequency, odds ratio, relative risk, or confidence interval. Therefore, no case-control comparison can be computed and PS4 cannot be adjudicated. The literature_pass.json packet has an empty ai_classified_papers list, so it contains no pre-read PS4 paper requiring additional verification.

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:27694386 reports that D1228N had been reported in a patient with MET-driven NSCLC and acquired crizotinib resistance, but reports no control group or case-control statistic for D1228N.
• PMID:28522754 reports D1228N as a subclonal mutation detected at progression in one patient with MET exon 14-skipping, MET-amplified lung adenocarcinoma, but reports no unaffected/population comparison or D1228N-specific odds ratio, relative risk, or confidence interval.
• gnomAD v2.1 and v4.1 report the exact variant as absent, but no affected-case frequency or matched case-control calculation is available; absence alone does not supply the required PS4 estimate and confidence interval.
• The literature_pass.json ai_classified_papers list is empty; no paper was pre-read as PS4 in that packet.

Still missing

• Affected-case and unaffected or population-control counts or frequencies for the exact variant
• Odds ratio or relative risk with a 95% confidence interval

Sources

Reasoning

MET p.Asp1228Asn lies in the MET kinase ATP docking pocket, which PMID:28522754 describes as containing D1228N and other resistance-associated missense changes. The Cancer Hotspots result also reports a statistically significant MET D1228 hotspot, but its row was only partially parsed. The available evidence does not establish the required absence of benign variation across the relevant critical domain or hotspot region, so PM1 cannot be conclusively assigned under the generic rule.

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

• PMID:28522754 states that D1228N and other MET missense mutations are located in the ATP docking pocket.
• The Cancer Hotspots result for MET D1228 states that the variant lies in a statistically significant hotspot, but the captured row is flagged as partially parsed.
• The queried D1228N variant is absent from gnomAD v2.1 and gnomAD v4.1; this does not by itself establish absence of benign variation across the entire functional region.

Still missing

• A complete, auditable assessment of benign variation across the MET ATP docking pocket or D1228 hotspot region is unavailable.
• The Cancer Hotspots D1228 row requires verification because the captured extraction was partial.

Sources

Reasoning

PM3 cannot be assessed because the case contains no affected-proband observation, parental or offspring testing, documented phase, or identified pathogenic variant in trans. The available variant-specific literature describes MET p.Asp1228Asn in acquired resistance to crizotinib in MET-driven lung cancer, not germline allelic configuration. The case materials also do not establish a recessive MET disorder context for applying PM3.

Rule

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

Evidence considered

• PMID:27694386: the paper mentions MET D1228N in a prior patient with MET-driven NSCLC and acquired crizotinib resistance but reports no germline segregation, parental testing, or phase information.
• PMID:28522754: the paper reports D1228N as a subclonal mutation detected during acquired resistance in a MET exon 14-skipping lung adenocarcinoma case and provides no allelic phase or pathogenic-in-trans observation.
• Case evidence contains no affected-proband segregation record, parent/offspring testing result, cis/trans assignment, or pathogenic partner allele.

Still missing

• Determine whether the variant is being interpreted for a recessive disorder.
• Obtain parental or offspring testing to establish phase.
• Identify and classify any pathogenic allele in trans.

Sources

Reasoning

The available case evidence does not document an assumed de novo occurrence in a proband or testing showing the variant is absent from either parent; therefore PM6 cannot be assessed.

Rule

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

Evidence considered

• The supplied materials contain no clinical case report or family study documenting this variant as assumed de novo.
• No parental genotype information is provided.

Still missing

• Proband-level clinical observation
• Evidence supporting de novo origin
• Parental genotypes or explicit lack of parental confirmation

Reasoning

The available case evidence provides no affected relatives, family genotypes, or meioses showing co-segregation of NM_000245.4:c.3682G>A with disease; therefore PP1 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 supplied case materials do not report a family pedigree, affected relatives, relative genotypes, or segregation counts for this variant.
• The available literature extracts concern unrelated variant-specific or gene-level observations and do not provide segregation evidence for this variant.

Still missing

• Affected family members and their clinical diagnoses
• Genotypes of informative relatives
• Number of informative meioses
• Assessment of phenotype consistency across segregating relatives

Reasoning

The generic PP2 rule requires both a low rate of benign missense variation in the gene and missense variants as a common disease mechanism. The case evidence does not provide validated gene-level data establishing either requirement for MET in the applicable germline disease context. Variant-specific absence from gnomAD and oncogenic somatic literature cannot substitute for both gene-level PP2 requirements.

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 evidence quantifying benign missense variation in MET is unavailable.
• Validated evidence that missense variants are a common mechanism of the relevant disease is unavailable.

Reasoning

The case materials identify the variant and gene but do not provide the patient's phenotype, family history, or a documented highly specific clinical presentation linked to a disease with a single genetic etiology. PP4 therefore cannot be adjudicated from the available case evidence.

Rule

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

Evidence considered

• The case summary provides the MET c.3682G>A (p.Asp1228Asn) variant and generic disease contexts but no patient-level phenotype or family-history assessment.

Still missing

• Patient phenotype and family history
• Disease-specific evidence that the presentation is highly specific for a single genetic etiology

Reasoning

BS2 cannot be assessed from the available evidence. No documented observation of this variant in a healthy adult individual, no homozygote or hemizygote observation, and no disease-specific penetrance or age-of-onset framework are provided. Absence from gnomAD is population-frequency evidence for PM2, not evidence that the variant was observed in a healthy adult.

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

• The supplied gnomAD records report the variant as absent in v2.1 and v4.1; they do not provide a healthy-adult observation for BS2.
• No source in the supplied population evidence documents a homozygous, heterozygous, or hemizygous healthy-adult carrier of this variant.

Still missing

• A reliable report of the variant in a healthy adult, with zygosity, ancestry, phenotype ascertainment, and relevant disease penetrance/age-of-onset context, is required to assess BS2.

Reasoning

No study in the available evidence demonstrates that NM_000245.4:c.3682G>A (NP_000236.2:p.(Asp1228Asn)) has no damaging effect on MET protein function or splicing. PMID:28522754 reports the variant in a clinical resistance context without testing its function, and PMID:27694386 discusses D1228N clinically but functionally evaluates D1228V. Therefore, there is no well-established benign functional result, controlled normal-function assay, or splicing assay supporting BS3.

Rule

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

Evidence considered

• PMID:28522754, Results, paragraph 2: the study reports D1228N in a patient but provides no D1228N-specific functional or splicing assay.
• PMID:27694386, Discussion, paragraph 2: the paper mentions D1228N as a clinical observation, while its own functional experiments concern D1228V, not D1228N.
• No controlled normal-function, normal-splicing, reproducibility, or clinical-laboratory validation evidence was available for the exact p.Asp1228Asn substitution.

Still missing

• A validated assay directly demonstrating preserved MET function or normal splicing for p.Asp1228Asn is needed.

Sources

Reasoning

The available case evidence does not identify affected relatives who were tested for the variant and does not document non-segregation; therefore lack of segregation cannot be assessed.

Rule

Lack of segregation in affected members of a family.

Evidence considered

• No family study in the supplied materials reports affected relatives carrying a non-segregating genotype or otherwise documents lack of segregation.
• Because no discordant affected relatives are documented, the BS4 phenocopy and multiple-variant caveats cannot be evaluated.

Still missing

• Affected relatives evaluated for the disease
• Genotypes of those affected relatives
• Phenotype and age information for tested relatives
• Evaluation for phenocopies or multiple pathogenic variants if non-segregation is observed

Reasoning

The generic BP1 rule requires a gene for which primarily truncating variants are known to cause disease. The supplied evidence does not establish that MET-associated disease is primarily caused by truncating variants, nor does it provide a validated disease-specific comparison of truncating versus missense mechanisms. Somatic MET kinase-domain resistance literature is not sufficient to satisfy or exclude this germline gene-level criterion.

Rule

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

Still missing

• A validated gene-level disease mechanism showing primarily truncating MET variants cause disease is unavailable.

Reasoning

BP2 cannot be assessed because the case contains no affected-proband observation, documented cis or trans phase, or pathogenic variant paired with NM_000245.4:c.3682G>A. The available variant-specific literature concerns acquired somatic resistance and does not establish either a trans observation in a fully penetrant dominant disorder or a cis observation in any inheritance pattern.

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:27694386: D1228N is discussed as an acquired-resistance observation in MET-driven NSCLC; no cis/trans phase or paired pathogenic allele is reported.
• PMID:28522754: D1228N is described among subclonal resistance mutations in a lung adenocarcinoma case; no cis/trans phase or pathogenic allele relationship is reported.
• Case evidence contains no affected-proband allelic observation, phase result, or documented pathogenic variant in cis or trans.

Still missing

• Obtain testing or specimen-level data establishing cis or trans phase with a pathogenic variant.
• For a trans observation, establish whether the relevant disorder is fully penetrant and dominant.

Sources

Reasoning

No case evidence documents the exact variant in a patient with an alternate molecular basis for the disease. The available variant-specific reports describe MET D1228N in cancer contexts but do not establish an alternate molecular diagnosis for a disease case, so BP5 cannot be adjudicated.

Rule

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

Evidence considered

• PMID:27694386 describes a prior patient with MET-driven NSCLC and acquired resistance, but does not document an alternate molecular basis for the relevant disease.
• PMID:28522754 describes D1228N as a subclonal progression mutation in a patient with MET exon 14-skipping and MET amplification, but this is not a documented BP5 case of an alternate molecular basis for the disease under evaluation.

Still missing

• A patient-level report showing the exact variant and an independently established alternate molecular basis for the disease

Sources