RET c.1900T>C, p.Cys634Arg

Reasoning

RET has no VCEP-specific domain table in vcep_materials.json, so the generic ACMG/AMP PM1 definition applies. Cancer Hotspots returned a positive result for the RET C634 query and states that this variant lies in a statistically significant hotspot. This is direct hotspot evidence for residue 634; the supplied evidence does not identify a benign-variation signal that would negate the hotspot finding. The hotspot extraction is flagged as partial and should be confirmed against the captured screenshot.

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 reports domain_tables as an empty object and the case framework as generic_acmg, so no RET-specific domain list or VCEP PM1 restriction applies.
• Cancer Hotspots queried RET C634 and returned found=true with the evidence sentence: This variant lies in a statistically significant hotspot; the extractor notes that the C634 hotspot row was only partially parsed.
• The queried residue is codon 634 in RET: p.Cys634Arg.

Still missing

• Cancer Hotspots extraction is partial and its source_data is empty; confirm the hotspot row in the captured screenshot.
• No VCEP domain table is available for RET.

Sources

Reasoning

PM2 supporting is met under the supplied generic threshold of <=0.0001. gnomAD v4.1 all-comers shows total AF=2.48818e-06 and grpmax FAF=5.53e-06 from 4/1,607,602 alleles with no homozygotes. The relevant non-cancer subsets independently remain extremely rare: gnomAD v2.1 non-cancer exomes AF=4.2896e-06 (1/233,122 alleles; homozygotes=0) and gnomAD v3.1 non-cancer genomes-only AF=1.35465e-05 with grpmax FAF=2.265e-05 (2/147,640 alleles; homozygotes=0). No VCEP specifies a different source for this gene, so all-comers gnomAD v4.1 is the default.

Rule

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

Evidence considered

• gnomAD v4.1 all-comers: AF=2.48818e-06 (4/1,607,602 alleles; homozygotes=0), filtering grpmax FAF=5.53e-06.
• gnomAD v2.1 non-cancer exomes: AF=4.2896e-06 (1/233,122 alleles; homozygotes=0). gnomAD v3.1 non-cancer genomes only: AF=1.35465e-05 (2/147,640 alleles; homozygotes=0), grpmax FAF=2.265e-05.
• Generic calibration supplied for this case: PM2 <=0.0001 at supporting strength (ClinGen SVI recommendation; PMID:25741868).

Sources

Reasoning

The variant was reported in patients with MEN 2A and was absent from tested parents and relatives. Tessitore et al. reported negative testing in both parents, three sisters, and one brother and described the mutations as de novo, while Vieira et al. reported C634R only in the index case and called it a possible de novo mutation. Confirmation of paternity was reported in Tessitore et al., but confirmation of maternity was not documented; therefore the evidence supports assumed de novo status rather than PS2.

Rule

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

Evidence considered

• Tessitore et al. 1999 (PMID:10522989), Results, family screening: both parents, three sisters, and one brother were negative for the RET mutations; four informative microsatellite loci ruled out nonpaternity, and the authors concluded the mutations were de novo.
• Vieira et al. 2002 (PMID:11987030), Abstract and Family 2 clinical and molecular findings: C634R was found only in the index case; neither parent nor any of four brothers had the abnormality, suggesting a de novo RET mutation.

Still missing

• Maternity confirmation is not documented, so this remains PM6 rather than PS2.

Sources

Reasoning

The variant is a missense substitution, NM_020975.6:c.1900T>C (p.Cys634Arg), so the generic rule requires the missense REVEL path only; SpliceAI and BayesDel are not used for PP3/BP4 in this situation. REVEL is 0.972, meeting the generic PP3 strong threshold of >=0.932. The threshold is the ClinGen SVI REVEL 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: missense, NM_020975.6:c.1900T>C (p.Cys634Arg).
• Local REVEL lookup reports a score of 0.972 for this variant, which meets the generic PP3 strong cutoff >=0.932.
• The REVEL cutoff is the ClinGen SVI calibration published by Pejaver et al. 2022 (PMID:36413997).
• SpliceAI max delta 0.008 and BayesDel 0.557748 were not counted because the governing generic path for a missense variant is REVEL only, and no generic BayesDel calibration is established.

Sources

Reasoning

The governing rule is that the patient's phenotype or family history must be highly specific for a disease with a single genetic etiology. The exact-variant reports describe the MEN2A phenotype, including medullary thyroid carcinoma with pheochromocytoma and, in one report, hyperparathyroidism; MEN2A is the specific RET-associated syndrome relevant to this variant. This supports PP4 at supporting strength. The phenotype evidence is from published reports rather than a separate current-patient phenotype record.

Rule

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

Evidence considered

• Tessitore et al. (PMID:10522989) reports the exact RET p.Cys634Arg variant in a patient with MEN2A, medullary thyroid carcinoma, and pheochromocytoma.
• Vieira et al. (PMID:11987030) reports the exact C634R variant in an MEN2A index case with medullary thyroid carcinoma, pheochromocytoma, and hyperparathyroidism; the abstract states that MEN2A patients are at risk for early medullary thyroid carcinoma.

Sources

Reasoning

The queried variant is RET NM_020975.6:c.1900T>C, p.Cys634Arg. The evidence establishes this same nucleotide and amino-acid change as pathogenic, including a published MEN2A case and multiple ClinVar submissions, but does not establish a previously pathogenic p.Cys634Arg variant caused by a different nucleotide change. Therefore the same-amino-acid/different-nucleotide requirement for PS1 is not met.

Rule

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

Evidence considered

• Variant normalization: NM_020975.6:c.1900T>C produces NP_066124.1:p.(Cys634Arg).
• Tessitore et al. (PMID:10522989) reports the queried c.1900T>C transition as producing cysteine-to-arginine substitution at codon 634; this is the same nucleotide change, not an alternate nucleotide change.
• ClinVar variation 13917 lists the exact variant NM_020975.6(RET):c.1900T>C (p.Cys634Arg) with Pathogenic/Likely pathogenic submissions, but does not provide a distinct nucleotide change yielding the same amino-acid substitution.

Still missing

• No previously established pathogenic p.Cys634Arg variant caused by a different nucleotide change was identified in the supplied evidence.

Sources

Reasoning

The variant was reported in a patient with MEN 2A and was absent from tested relatives in the Tessitore case. However, the paper states that four informative microsatellite loci ruled out nonpaternity but does not establish maternity; confirmation of paternity alone is explicitly insufficient for PS2. A second family report also described the variant as possible de novo because it was absent from both parents and four siblings, without documented maternity and paternity confirmation.

Rule

De novo (both maternity and paternity confirmed) in a patient with the disease and no family history. ... Confirmation of paternity only is insufficient.

Evidence considered

• Tessitore et al. 1999 (PMID:10522989): in a 26-year-old woman with MEN 2A carrying RET p.Cys634Arg plus Ala640Gly, both parents, three sisters, and one brother lacked both mutations; four informative microsatellite loci ruled out nonpaternity, but the report did not document maternity confirmation.
• Vieira et al. 2002 (PMID:11987030): Family 2 had RET C634R only in the index case; neither parent nor four brothers carried it, and the authors called it a possible de novo mutation without documented maternity and paternity confirmation.

Still missing

• Documented confirmation of both maternity and paternity is absent.

Sources

Reasoning

The governing rule requires a reputable recent report of the variant as pathogenic when the evidence is unavailable to the laboratory for independent evaluation. ClinVar contains exact-variant Pathogenic/Likely pathogenic submissions, but the case audit records zero expert-panel submissions; the available assertions are ordinary laboratory, OMIM/literature-only, or aggregate classifications. Therefore the required exact-variant expert-panel classification is absent and PP5 is not met. PMID:10522989 is a case report describing the variant as disease-associated, not an expert-panel classification.

Rule

Apply them only for an exact-variant ClinVar expert-panel classification: Pathogenic/Likely pathogenic supports PP5; Benign/Likely benign supports BP6. Ordinary laboratory submissions, aggregate ClinVar labels, and non-expert assertions must never trigger either criterion.

Evidence considered

• ClinVar exact match NM_020975.6(RET):c.1900T>C (p.Cys634Arg) is classified Pathogenic/Likely pathogenic, but the submission audit reports expert_panel_submissions = 0 and identifies the records as laboratory, OMIM/literature-only, or other non-expert assertions.
• Tessitore et al. (PMID:10522989) is an exact-variant case report and does not constitute an expert-panel classification.

Sources

Reasoning

BA1 is not met. The applicable gnomAD v4.1 filtering frequency is grpmax FAF=5.53e-06, based on 59,968 observed alleles, which is far below the supplied generic stand-alone threshold of 0.05. The gnomAD v4.1 total frequency is 2.48818e-06 (4/1,607,602 alleles; homozygotes=0); gnomAD v2.1 all-comers is 1.21195e-05 (3/247,534 alleles; homozygotes=0), and gnomAD v2.1 non-cancer exomes is 4.2896e-06 (1/233,122 alleles; homozygotes=0). The higher Admixed American point estimate is not substituted for grpmax FAF.

Rule

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

Evidence considered

• gnomAD v4.1 all-comers: total AF=2.48818e-06 (4/1,607,602 alleles; homozygotes=0); filtering grpmax FAF=5.53e-06 from 2/59,968 Admixed American alleles.
• gnomAD v2.1 all-comers exomes: AF=1.21195e-05 (3/247,534 alleles; homozygotes=0). gnomAD v2.1 non-cancer exomes: AF=4.2896e-06 (1/233,122 alleles; homozygotes=0).
• Generic calibration supplied for this case: BA1 >= 0.05, ACMG/AMP 2015 (PMID:25741868).

Sources

Reasoning

BS1 is not met on population frequency. The required filtering allele frequency is gnomAD v4.1 grpmax FAF=5.53e-06, far below the supplied generic default threshold of 0.01; therefore the higher Admixed American point estimate of 3.33511e-05 does not establish BS1. The relevant non-cancer subsets are also below the default: gnomAD v2.1 non-cancer exomes AF=4.2896e-06 and gnomAD v3.1 non-cancer genomes-only grpmax FAF=2.265e-05. Because the filtering FAF is not elevated, a maximum-credible-frequency calculation is not needed to withhold BS1.

Rule

Allele frequency is greater than expected for the disorder.

Evidence considered

• gnomAD v4.1: filtering grpmax FAF=5.53e-06; total AF=2.48818e-06 (4/1,607,602 alleles), while the Admixed American point estimate is 3.33511e-05 from 2/59,968 alleles.
• gnomAD v2.1 non-cancer exomes: AF=4.2896e-06 (1/233,122 alleles; homozygotes=0). gnomAD v3.1 non-cancer genomes only: AF=1.35465e-05 (2/147,640 alleles; homozygotes=0), grpmax FAF=2.265e-05.
• Generic calibration supplied for this case: BS1 >= 0.01; Whiffin et al. 2017 maximum-credible-frequency guidance (PMID:28518168) is not needed because the filtering FAF is far below the default.

Sources

Reasoning

The available evidence supports RET activating missense disease biology rather than a gene in which disease is primarily caused by truncating variants. OncoKB annotates RET p.C634R with a gain-of-function effect, and functional studies describe RET-C634R as a classical oncogenic or MEN2A-associated control. Thus the premise of BP1 is not supported for this variant's RET disease context.

Rule

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

Evidence considered

• OncoKB identifies RET p.C634R as Oncogenic and assigns a Gain-of-function biological-effect context.
• Functional literature uses RET-C634R as a classical MEN2A-associated or oncogenic control in signaling and transformation assays.
• These data are inconsistent with applying a benign criterion premised on a gene for which primarily truncating variants are known to cause disease.

Sources

Reasoning

The variant is a missense substitution, so BP4 must be assessed through the generic REVEL path only; SpliceAI cannot substitute for REVEL in a missense variant. REVEL is 0.972, which is not <=0.29, the generic BP4 supporting threshold, so BP4 is not met. BayesDel is not usable 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)

Evidence considered

• Variant consequence: missense, NM_020975.6:c.1900T>C (p.Cys634Arg).
• Local REVEL lookup reports a score of 0.972, which does not meet the generic BP4 supporting cutoff <=0.29.
• SpliceAI max delta 0.008 was not used because the governing generic path for a missense variant is REVEL only; BayesDel 0.557748 was not used because no generic ACMG-strength calibration is established.
• The REVEL cutoff is the ClinGen SVI calibration published by Pejaver et al. 2022 (PMID:36413997).

Sources

Reasoning

The governing rule requires an exact-variant ClinVar expert-panel classification as Benign/Likely benign for BP6 under the supplied generic rule. The exact ClinVar record has no expert-panel submissions and no benign or likely benign expert-panel classification; available submissions are Pathogenic/Likely pathogenic or non-expert laboratory/literature assertions. BP6 is therefore not met.

Rule

Apply them only for an exact-variant ClinVar expert-panel classification: Pathogenic/Likely pathogenic supports PP5; Benign/Likely benign supports BP6. Ordinary laboratory submissions, aggregate ClinVar labels, and non-expert assertions must never trigger either criterion.

Evidence considered

• ClinVar exact match NM_020975.6(RET):c.1900T>C (p.Cys634Arg) reports Pathogenic/Likely pathogenic submissions, expert_panel_submissions = 0, and no expert-panel Benign/Likely benign assertion.

Sources

Reasoning

The exact RET p.Cys634Arg variant was included in functional experiments only as a positive or comparator control. PMID:32293499 states that C634R was used as the positive control of an oncogenic RET mutation in RET phosphorylation and transformation assays, but does not report separate C634R results beyond control use. PMID:21690267 reports that RET-C634R had greater transforming activity than RET-K666E (P<0.01) and used C634R as a comparator, but does not provide a direct variant-specific comparison with wild-type RET establishing the assay result for C634R. These findings are biologically supportive but do not establish a well-established, reproducible, robust variant-specific functional result sufficient to assign PS3 under the supplied generic rule; no validated clinical diagnostic assay evidence was identified.

Rule

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

Evidence considered

• PMID:32293499: In transiently transfected HEK293T cells, C634R was used as the positive control for an oncogenic RET mutation; the paper did not report separate C634R functional results beyond control use.
• PMID:21690267: RET-C634R was used as a MEN2A-associated functional comparator in NIH3T3 focus-formation and HEK293T signaling assays; RET-K666E showed significantly lower transforming activity than RET-C634R (P<0.01), but no direct C634R-versus-wild-type result was reported.
• The available evidence supports biological plausibility of an activating effect but does not document the variant-specific, validated, reproducible and robust functional evidence required for PS3.

Still missing

• A direct, variant-specific functional assay result for RET p.Cys634Arg with wild-type and appropriate assay controls is needed.
• Reproducibility or validation in a clinical diagnostic laboratory setting is not documented.

Sources

Reasoning

The governing ACMG/AMP rule requires a case-control prevalence comparison with RR or OR >5.0 and a confidence interval that excludes 1.0. The reviewed exact-variant reports describe affected individuals with RET p.Cys634Arg, but provide no unaffected/population control count or case-control estimate. gnomAD provides a population frequency (4/1,607,602 alleles in v4.1) but no affected-case frequency from a matched study, so an OR and 95% CI cannot be calculated. The literature_pass PS4 leads were checked against the available exact-variant extracts/abstract evidence; they identify case observations or family reports, not qualifying case-control enrichment.

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

• Tessitore et al. (PMID:10522989) reports one affected MEN 2A patient carrying RET c.1900T>C (p.Cys634Arg) with a second RET mutation, but no unaffected/population control comparison.
• Vieira et al. (PMID:11987030) reports RET C634R in an MEN2A family index case and family testing, but no case-control counts or RR/OR estimate.
• gnomAD v4.1 reports 4/1,607,602 alleles (AF 2.48818e-06); this is a control-population frequency only and cannot supply the affected-case count needed for PS4.

Still missing

• A case-control study or matched affected-case and control counts for RET c.1900T>C are needed to calculate OR and its 95% CI.

Sources

Reasoning

The queried variant changes RET Cys634 to Arg. The supplied evidence includes reports that codon 634 is a recurrent RET disease-associated codon, but it does not identify a specific different missense substitution at residue 634 with an independently established pathogenic classification. The PM5 candidate artifact contains zero same-residue candidates and explicitly cannot confirm classic PM5 semantics, so PM5 cannot be assessed from the available evidence.

Rule

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

Evidence considered

• The PM5 candidate collection reports zero total candidates and zero same-residue candidates and instructs that PM5 must not be applied from that artifact alone.
• The abstract record for Hedayati et al. (PMID:9230192) discusses RET cysteine mutations at several codons and says codon 634 mutations correlate with MEN2A, but the supplied extract does not name a distinct pathogenic missense substitution at residue 634.

Still missing

• A named alternate missense variant at RET residue 634 with an independently established pathogenic classification is required.

Sources

Reasoning

No qualifying co-segregation evidence is available. The exact-variant reports describe isolated index cases with negative testing in unaffected relatives; the Tessitore patient had no children, and no multiple affected family members carrying the variant were reported. The available segregation-study lead was not accompanied by an auditable extracted source record and cannot establish PP1.

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

• Tessitore et al. 1999 (PMID:10522989): the patient had no children, and the report states that cosegregation could not be proven; tested relatives were clinically unaffected and mutation-negative.
• Vieira et al. 2002 (PMID:11987030): C634R was present only in the index case of Family 2, with parents and four brothers negative; this is not co-segregation in multiple affected family members.

Still missing

• Affected relatives with documented RET C634R status and clinical phenotype are not reported.
• No auditable family-level meioses or co-segregation count is available.

Sources

Reasoning

RET missense activation is supported by the variant-specific OncoKB biological-effect annotation and functional literature, but the supplied evidence does not establish the required gene-level low rate of benign missense variation. The single-variant gnomAD frequencies cannot substitute for a gene-level benign-missense-rate assessment. Because both components of PP2 cannot be evaluated from the available evidence, PP2 is 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.

Evidence considered

• OncoKB labels RET p.C634R as Oncogenic with a Gain-of-function biological-effect context.
• Functional studies used RET-C634R as an oncogenic or MEN2A-associated control, supporting a missense activation mechanism but not quantifying the gene-wide rate of benign missense variation.
• gnomAD v4.1 reports the queried variant at AF 2.48818e-06 (4/1,607,602 alleles); this is a frequency for the queried variant, not a gene-level benign-missense-rate metric.

Still missing

• A gene-level assessment showing a low rate of benign RET missense variation is missing.

Sources

Reasoning

BS2 cannot be assessed from the available population data. gnomAD reports heterozygous alleles but no homozygotes (v4.1: 4 total alleles; v2.1: 3; v3.1 non-cancer: 2); these aggregate records do not establish that any carrier was a healthy adult or provide age, phenotype, ascertainment, or follow-up. The healthy-adult requirement for an early-onset, fully penetrant dominant disorder therefore remains unverified.

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 4/1,607,602 total alleles and 0 homozygotes; gnomAD v2.1 reports 3/247,534 alleles and 0 homozygotes; gnomAD v3.1 non-cancer reports 2/147,640 alleles and 0 homozygotes.
• The available population summaries provide allele and homozygote counts but not carrier-level age, health status, phenotype, or follow-up.

Still missing

• Individual carrier age, health status, ascertainment, and clinical follow-up are unavailable.
• A source explicitly documenting an observed carrier as a healthy adult is needed to apply BS2.

Sources

Reasoning

No reviewed functional study demonstrates that RET p.Cys634Arg has no damaging effect on protein function or splicing. The available papers use C634R as an oncogenic positive or comparator control and therefore do not provide benign functional evidence. There is also no validated clinical diagnostic laboratory assay showing preserved RET function or normal splicing for this variant.

Rule

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

Still missing

• A well-established assay demonstrating no damaging effect on RET protein function or splicing is needed.
• Assay validation, controls, and reproducibility for a benign functional result are not documented.

Reasoning

BS4 requires lack of segregation in affected family members, but the available reports describe unaffected relatives who tested negative rather than affected relatives who lack the variant. No affected family member with a documented negative RET C634R result was identified, so the BS4 requirement is not evaluable.

Rule

Lack of segregation in affected members of a family.

Evidence considered

• Tessitore et al. 1999 (PMID:10522989): no clinical MEN 2A symptoms were present in tested relatives, and all tested relatives were mutation-negative; this does not constitute non-segregation in affected members.
• Vieira et al. 2002 (PMID:11987030): neither parent nor four brothers carried C634R, but the report does not identify these relatives as affected.

Still missing

• No affected relative with documented absence of RET C634R is reported.

Sources

Reasoning

BP2 can apply when a variant is observed in cis with a pathogenic variant in any inheritance pattern. PMID:10522989 documents the queried RET p.Cys634Arg and an Ala640Gly RET variant on the same allele, with both detected in germline and tumor DNA. However, the available evidence does not establish Ala640Gly as pathogenic, and no qualifying trans observation in a fully penetrant dominant disorder is documented. The cis observation therefore cannot be used to assess BP2 without pathogenicity evidence for the second variant.

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

• A novel case of multiple endocrine neoplasia type 2A associated with two de novo mutations of the RET protooncogene, PMID:10522989, reports RET p.Cys634Arg and Ala640Gly on the same RET allele; both were detected in germline and tumor DNA.
• The source reports the cis phase but does not provide a pathogenicity classification for Ala640Gly, so the required cis pathogenic-variant condition is unresolved.

Still missing

• A reliable pathogenic or likely pathogenic classification for the cis RET Ala640Gly variant is not provided.
• No qualifying observation of the queried variant in trans with a pathogenic variant in a fully penetrant dominant disorder is documented.

Sources

Reasoning

The governing rule requires the variant to be found in a case with an alternate molecular basis for disease. The reviewed exact-variant reports document RET-associated MEN2A and, in PMID:10522989, a second RET mutation on the same allele, not an alternate molecular basis independent of the queried variant. No case-specific evidence in the available packet demonstrates that an unrelated molecular diagnosis explains the relevant disease phenotype. BP5 therefore cannot be assessed as met.

Rule

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

Evidence considered

• Tessitore et al. (PMID:10522989) reports RET p.Cys634Arg together with RET Ala640Gly on the same allele in a MEN2A case; this is not an independent alternate molecular basis under BP5.
• No reviewed exact-variant source documents a separate non-RET molecular diagnosis that explains the relevant disease phenotype.

Still missing

• A documented case with an independent alternate molecular diagnosis explaining the disease phenotype is needed for BP5.

Sources