GNAS c.602G>A, p.Arg201His

Reasoning

PS3 is met at moderate strength. Taki et al. directly studied the exact protein change GNAS p.Arg201His (corresponding to NM_000516.7:c.602G>A) in conditional pancreas-specific transgenic mice. GNASR201H alone produced elevated pancreatic cAMP and pancreatic ductal/tissue abnormalities, while the model included wild-type GNAS, no-transgene, and KrasG12D comparator groups; combined GNASR201H/KrasG12D mice developed pancreatic cystic tumors in all 8 examined mice, with significantly shortened survival (P<0.001), elevated cAMP (one-way ANOVA P=0.022), activated PKA signaling, and pathology closely mimicking human IPMN. This is direct, biologically relevant in vivo evidence for a damaging/gain-of-function effect, but it is a preclinical mouse model rather than a validated and reproducible clinical diagnostic laboratory assay, so the evidence is not assigned full strong weight.

Rule

Well-established in vitro or in vivo functional studies supportive of a damaging effect on the gene or gene product. ... Functional studies that have been validated and shown to be reproducible and robust in a clinical diagnostic laboratory setting are considered the most well-established.

Evidence considered

• Taki et al. PMID:26257060, Figures 1-4 and Results: conditional pancreas-specific GNASR201H mice showed elevated cAMP, duct dilation, tubular complexes, acinar-cell loss, and fibrosis; GNASR201H/KrasG12D mice developed cystic pancreatic tumors in all 8 examined mice within 5 weeks.
• Taki et al. PMID:26257060, Figure 2 and Results: the study included wild-type, GNASR201H-only, and KrasG12D-only comparator groups; combined GNASR201H/KrasG12D mice had significantly shortened survival (P<0.001).
• Taki et al. PMID:26257060, Figure 4: pancreatic cAMP differed significantly among genotypes by one-way ANOVA (P=0.022), and tumor cells showed phosphorylated PKA substrates consistent with activated G-protein signaling.
• Strength calibration: the evidence is direct and biologically relevant but comes from a preclinical mouse model, not a validated and reproducible clinical diagnostic laboratory assay; therefore PS3 is downgraded to moderate.

Still missing

• No validated, reproducible clinical diagnostic laboratory assay for this exact variant was identified.

Sources

Reasoning

p.Arg201His lies at GNAS residue R201, a recurrent activating mutational hotspot. The Cancer Hotspots result identifies GNAS R201 as a statistically significant hotspot, and PMID:24855271 reports that all GNAS mutations in its 65-patient corticotropin-independent Cushing syndrome cohort were confined to R201 (11/65, 16.9%). No benign interpretation is supplied for the hotspot; the variant is rare in population data (gnomAD v4.1 AF 2.4797e-05; 40/1,613,098 alleles; 0 homozygotes). These data satisfy the generic PM1 hotspot route.

Rule

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

Evidence considered

• Cancer Hotspots result for GNAS R201: the queried variant lies in a statistically significant hotspot.
• Recurrent somatic mutations underlie corticotropin-independent Cushing's syndrome, Results/Fig. 1/Table S1: GNAS mutations occurred in 11/65 cases (16.9%) and all were confined to residue R201.
• gnomAD v4.1: 40/1,613,098 alleles, allele frequency 2.4797e-05, with zero homozygotes; this is rare population observation rather than established benign variation.

Sources

Reasoning

PM2 is met at supporting strength. The variant is rare but present in population databases: gnomAD v4.1 AF=2.4797e-05 (40/1,613,098 alleles; zero homozygotes), below the supplied PM2 supporting threshold of <=0.0001. The relevant non-cancer subsets are concordant: gnomAD v2.1 non-cancer exomes AF=1.26624e-05 and gnomAD v3.1 non-cancer genomes AF=1.3525e-05, both with zero homozygotes. This supports extremely low frequency rather than complete absence.

Rule

PM2 (supporting): <= 0.0001 — SVI recommendation to downgrade PM2 to Supporting (ClinGen SVI 2020) (PMID:25741868)

Evidence considered

• gnomAD v4.1: 40/1,613,098 alleles, AF=2.4797e-05, homozygotes=0; highest observed ancestry AF=6.75676e-05 in Ashkenazi Jewish individuals.
• gnomAD v2.1 non-cancer exomes: 3/236,922 alleles, AF=1.26624e-05, homozygotes=0; gnomAD v3.1 non-cancer genomes: 2/147,874 alleles, AF=1.3525e-05, homozygotes=0.
• The supplied generic PM2 calibration is <=0.0001 for supporting strength, attributed to the ClinGen SVI recommendation to downgrade PM2 to Supporting (PMID:25741868).

Sources

Reasoning

PM6 is met at supporting strength. The exact Arg201His variant was observed in an affected patient, and the primary report describes a mosaic postzygotic mutation, supporting an assumed de novo origin without confirmed parental testing. Because the evidence concerns somatic mosaicism rather than a conventionally inherited germline variant, human review is warranted before clinical use.

Rule

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

Evidence considered

• Schwindinger et al. reported the exact Arg201His variant in a patient with McCune-Albright syndrome and detected it at different levels across leukocytes and skin tissues.
• The authors concluded that their findings support a postzygotic somatic mutation causing mosaic McCune-Albright syndrome; no parental testing was reported.

Still missing

• Maternity and paternity were not confirmed.
• The report did not test parental samples.
• The evidence establishes postzygotic mosaicism but does not characterize the variant as a conventional germline de novo event.

Sources

Reasoning

The variant is a missense SNV, so the governing generic rule selects the REVEL path only; SpliceAI and BayesDel are not used for PP3 in this situation. REVEL is 0.969, which meets the generic ClinGen SVI strong PP3 threshold of >= 0.932. The threshold is from the supplied ClinGen SVI REVEL calibration by Pejaver et al. 2022 (PMID:36413997).

Rule

Missense variant -> REVEL only. ... PP3 (strong): >= 0.932 — ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet) (PMID:36413997)

Evidence considered

• The variant is annotated as a missense_variant, NP_000507.1:p.(Arg201His), so the generic missense path requires REVEL only; the available SpliceAI score is not counted for PP3.
• The local REVEL predictor lookup reports a score of 0.969.
• REVEL 0.969 meets the supplied generic ClinGen SVI strong PP3 cutoff of >= 0.932; threshold calibration: Pejaver et al. 2022, Am J Hum Genet, PMID:36413997.

Sources

Reasoning

PS2 is not met. The exact variant was identified in a patient with McCune-Albright syndrome, but the directly reviewed report describes it as a postzygotic somatic mosaic mutation and does not document confirmed maternity, confirmed paternity, or a formal parental study. Therefore, the required combination of both parental confirmations and absence of family history is not established.

Rule

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

Evidence considered

• Schwindinger et al. identified the exact Arg201His change corresponding to NM_000516.7:c.602G>A in one patient with McCune-Albright syndrome.
• The report states that the mutation was present in a mosaic distribution and supports a postzygotic somatic mutation; it does not report maternity confirmation, paternity confirmation, or parental testing.

Still missing

• No documented maternity confirmation.
• No documented paternity confirmation.
• No formal parental testing or pedigree-based exclusion of inherited transmission.

Sources

Reasoning

Under the specified application, PP5 requires an exact-variant ClinVar expert-panel classification of Pathogenic or Likely pathogenic. ClinVar variation 15934 is classified Pathogenic overall, but the submission audit reports zero expert-panel submissions; the available assertions are ordinary laboratory, literature-only, or other non-expert submissions. PP5 is therefore not met.

Rule

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

Evidence considered

• ClinVar exact variant NM_000516.7(GNAS):c.602G>A (p.Arg201His), variation 15934: overall classification Pathogenic, with expert_panel_submissions = 0.
• The available exact-match assertions are not expert-panel classifications and therefore do not satisfy the specified PP5 trigger.

Sources

Reasoning

BA1 is not met. The highest relevant all-comers frequency is 2.4797e-05 (40/1,613,098 alleles) in gnomAD v4.1, and the highest observed ancestry frequency is 6.75676e-05 in Ashkenazi Jewish individuals; both are far below the generic stand-alone threshold of 0.05. The gnomAD v2.1 and non-cancer subsets are likewise far below 5%, with no evidence of a population-frequency threshold artifact that would support BA1.

Rule

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

Evidence considered

• gnomAD v4.1: 40/1,613,098 alleles, AF=2.4797e-05, homozygotes=0; highest observed Ashkenazi Jewish AF=6.75676e-05 (2/29,600 alleles).
• gnomAD v2.1: 4/251,454 alleles, AF=1.59075e-05, homozygotes=0; non-cancer exomes: 3/236,922 alleles, AF=1.26624e-05, homozygotes=0.
• Generic ACMG/AMP stand-alone population threshold supplied for BA1: >=0.05, attributed to ACMG/AMP 2015 (PMID:25741868).

Sources

Reasoning

BS1 is not met. The all-comers gnomAD v4.1 AF is 2.4797e-05 and the maximum population AF is 6.75676e-05, both far below the supplied generic default BS1 threshold of 0.01. The non-cancer subsets are also lower (gnomAD v2.1 non-cancer exomes AF=1.26624e-05; gnomAD v3.1 non-cancer genomes AF=1.3525e-05). No prevalence, penetrance, or allelic-heterogeneity calculation indicates that this frequency is greater than expected for the relevant GNAS disorder; consequently, BS1 cannot be applied from population frequency.

Rule

BS1 (strong): >= 0.01 — Generic SVI default; per-gene derivation preferred (Whiffin et al. 2017) (PMID:28518168)

Evidence considered

• gnomAD v4.1 all-comers: AF=2.4797e-05; highest observed ancestry AF=6.75676e-05 in Ashkenazi Jewish individuals; homozygotes=0.
• gnomAD v2.1 non-cancer exomes: AF=1.26624e-05 (3/236,922 alleles); gnomAD v3.1 non-cancer genomes: AF=1.3525e-05 (2/147,874 alleles); homozygotes=0 in both.
• The supplied generic BS1 calibration is >=0.01, with per-gene maximum-credible-frequency derivation preferred (Whiffin et al. 2017, PMID:28518168); the observed frequencies do not approach that default.

Sources

Reasoning

The variant is missense, but the supplied GNAS evidence describes recurrent activating missense disease mechanisms rather than a gene in which primarily truncating variants are known to cause disease. PMID:26257060 demonstrates constitutive activation from GNAS R201H, PMID:28868010 describes R201H as an activating mutation with constitutive cAMP signaling, and PMID:24855271 reports recurrent GNAS missense mutations at R201. Thus the BP1 gene-mechanism condition is not met.

Rule

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

Evidence considered

• PMID:26257060: GNAS R201H causes constitutive Gs-alpha activation and elevated cAMP in transgenic models.
• PMID:28868010: R201H is described as an activating mutation causing increased Gnas/XL-alpha-s activity, increased cAMP accumulation, and constitutive cAMP signaling.
• PMID:24855271: all observed GNAS mutations in the reported cohort were missense changes confined to R201.

Sources

Reasoning

The variant is a missense SNV, so the governing generic rule selects the REVEL path only; the low SpliceAI score cannot substitute for REVEL and BayesDel has no generic ACMG-strength calibration. REVEL is 0.969, which does not meet the strongest available generic BP4 supporting threshold of <= 0.29. Therefore BP4 is not met.

Rule

Missense variant -> REVEL only. ... BP4 (supporting): <= 0.29 — ClinGen SVI REVEL calibration (Pejaver et al. 2022, Am J Hum Genet) (PMID:36413997)

Evidence considered

• The variant is annotated as a missense_variant, NP_000507.1:p.(Arg201His), so the generic missense path requires REVEL only; the available SpliceAI score of 0.012 is not counted for BP4.
• The local REVEL predictor lookup reports a score of 0.969.
• REVEL 0.969 exceeds the supplied generic ClinGen SVI BP4 supporting cutoff of <= 0.29; threshold calibration: Pejaver et al. 2022, Am J Hum Genet, PMID:36413997.

Sources

Reasoning

BP5 requires the variant to be found in a case with an alternate molecular basis for disease. The reviewed exact-variant appendiceal adenocarcinoma report describes genomic profiling but does not establish an alternate molecular cause, and the GNAS R201H/KrasG12D mouse study is an experimental model rather than a same-case alternate molecular basis. No qualifying BP5 evidence is established.

Rule

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

Evidence considered

• PMID:28868010, Clinical Benefit from Trametinib in a Patient with Appendiceal Adenocarcinoma with a GNAS R201H Mutation: the exact-variant case report used comprehensive genomic profiling but does not establish an alternate molecular basis for the disease.
• PMID:26257060, GNAS(R201H) and Kras(G12D) cooperate to promote murine pancreatic tumorigenesis: the comparison is a conditional mouse model and is not evidence of an alternate molecular basis in the current human case.

Still missing

• Document an alternate pathogenic molecular finding in the same case and show that it independently provides the disease basis.

Sources

Reasoning

Under the specified application, BP6 requires an exact-variant ClinVar expert-panel classification of Benign or Likely benign. ClinVar variation 15934 is classified Pathogenic overall and the submission audit reports zero expert-panel submissions; no exact-variant expert-panel benign or likely-benign assertion is present. BP6 is therefore not met.

Rule

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

Evidence considered

• ClinVar exact variant NM_000516.7(GNAS):c.602G>A (p.Arg201His), variation 15934: overall classification Pathogenic, with expert_panel_submissions = 0.
• No exact-variant expert-panel benign or likely-benign assertion is present; ordinary laboratory submissions and aggregate labels are excluded from BP6.

Sources

Reasoning

The variant is NM_000516.7:c.602G>A (p.Arg201His). PMID:1594625 directly reports the same nucleotide and amino-acid change, but PS1 requires a previously established pathogenic variant producing the same amino-acid change regardless of nucleotide change. The available evidence does not document an alternate nucleotide change producing p.Arg201His, so the required same-amino-acid comparator is not established.

Rule

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

Evidence considered

• PMID:1594625, Results and Discussion: a G-to-A transition in GNAS exon 8 changes Arg201 to His (p.Arg201His), confirming the case variant itself rather than an alternate nucleotide representation.

Still missing

• A validated alternate-nucleotide p.Arg201His comparator is not documented.

Sources

Reasoning

The ACMG/AMP PS4 definition requires significantly increased prevalence in affected individuals compared with controls, with OR or RR >5.0 and a confidence interval excluding 1.0. The exact variant p.Arg201His is reported in affected patients, and gnomAD provides population-control observations (40/1,613,098 alleles in v4.1; 4/251,454 in v2.1), but no affected-case frequency or denominator paired with a control count is available in the reviewed evidence. The literature_pass.json PS4 index lists PMID:10571700, PMID:12970318, PMID:1944469, PMID:21835143, PMID:23536913, PMID:25157968, PMID:25719192, PMID:29991465, and PMID:3720010; their supplied abstract/title records do not provide exact-variant case-control counts, and none has a paper_extracts entry for verification. The required comparison is OR > 5.0? not evaluable because no OR can be calculated, and the 95% CI cannot be calculated; therefore PS4 cannot be adjudicated.

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

• gnomAD v4.1: 40/1,613,098 alleles, AF 2.4797e-05; this is a population-control frequency without a paired affected-case frequency.
• gnomAD v2.1: 4/251,454 alleles, AF 1.59075e-05; this is a population-control frequency without a paired affected-case frequency.
• literature_pass.json PS4 index: PMID:10571700, PMID:12970318, PMID:1944469, PMID:21835143, PMID:23536913, PMID:25157968, PMID:25719192, PMID:29991465, and PMID:3720010 were checked against their supplied title/abstract records; no exact-variant case-control counts or denominators were supplied.
• The exact-variant report in PMID:1594625 describes one affected patient and mosaic detection, but no control comparison or affected-cohort denominator; this source is not cited in evidence_refs because it is not a source_registry key.

Still missing

• Obtain exact-variant affected-case counts and denominators from the PS4-indexed papers.
• Obtain matched-ancestry control counts or a justified control frequency for the same variant.
• Calculate OR and 95% CI and determine whether OR >5.0 and the CI excludes 1.0.

Sources

Reasoning

The target is p.Arg201His. PMID:24855271 reports a different missense change at the same residue, p.Arg201Cys, in affected patients, and the ClinVar evidence trail lists a report of p.Arg201Gly in McCune-Albright syndrome. However, the supplied evidence does not establish either comparator as a previously determined pathogenic variant under the required PM5 standard; the R201C paper provides cohort occurrence but no pathogenic determination, and the R201G full text is unavailable in the fetched 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

• PMID:24855271: the cohort contained GNAS p.Arg201Cys and reported all GNAS mutations at R201, but the extracted paper evidence does not determine p.Arg201Cys to be pathogenic.
• ClinVar exact-variant evidence trail: it cites PMID:10571700, titled 'A novel GNAS1 mutation, R201G, in McCune-Albright syndrome,' but the fetched evidence does not provide a validated pathogenic determination or full-text finding for p.Arg201Gly.

Still missing

• A same-residue alternate missense variant with an explicit established pathogenic determination is not documented.

Sources

Reasoning

PP1 is not assessed. The available variant-specific report concerns a single affected patient with somatic mosaicism and provides no multiple affected family members, informative meioses, or cosegregation analysis.

Rule

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

Evidence considered

• The primary report presents molecular analysis of one affected patient and does not provide a family segregation study or affected relatives carrying the variant.

Still missing

• No pedigree with affected relatives is available.
• No familial genotyping or informative meioses are reported.

Sources

Reasoning

The available evidence shows recurrent activating GNAS missense changes at R201, including p.Arg201His, but does not establish the two gene-level requirements for PP2: a low rate of benign missense variation across GNAS and missense variants as a common disease mechanism for the gene. The single-variant gnomAD frequency and hotspot literature cannot supply those gene-wide 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.

Evidence considered

• PMID:24855271: recurrent somatic GNAS missense mutations were confined to R201 in 11/65 affected patients.
• PMID:26257060 and PMID:28868010: R201H is described as an activating GNAS missense alteration with constitutive Gs-alpha/cAMP signaling.
• gnomAD v4.1 provides the frequency of this variant, not a gene-wide benign-missense rate.

Still missing

• Gene-wide evidence for low benign missense variation and a common missense disease mechanism is not supplied.

Sources

Reasoning

PP4 requires the patient's phenotype or family history to be highly specific for a disease with a single genetic etiology. The supplied evidence describes McCune-Albright syndrome in published patients and its characteristic triad, but it does not provide the current patient's phenotype or family history. The criterion therefore cannot be assessed.

Rule

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

Evidence considered

• PMID:15126527: the abstract describes McCune-Albright syndrome as characterized by polyostotic fibrous dysplasia, café-au-lait pigmentation, and peripheral precocious puberty, but does not document the current patient's phenotype or family history.
• A published exact-variant report describes p.Arg201His in a patient with McCune-Albright syndrome, but published disease context cannot substitute for the current patient's phenotype or family history.

Still missing

• Obtain the current patient's clinical phenotype and family history.
• Establish that the phenotype is highly specific for a disease with a single genetic etiology.

Sources

Reasoning

BS2 is not assessed. The population datasets document allele counts and zero homozygotes but do not document a specific healthy adult carrier, the carrier's age, phenotype evaluation, or ascertainment. In addition, GNAS activating disease is commonly mosaic and clinically variable, so the required assumption of full penetrance at an early age is not established for this variant-disease context. Population presence alone therefore cannot satisfy BS2.

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 40/1,613,098 alleles and zero homozygotes; gnomAD v2.1 reports 4/251,454 alleles and zero homozygotes.
• The available population records provide no individual-level clinical annotation establishing a healthy adult carrier or confirming age-appropriate absence of the relevant phenotype.
• The case evidence describes GNAS p.Arg201His as an activating variant associated with mosaic/variable clinical contexts; this does not establish full penetrance at an early age for BS2.

Still missing

• A documented healthy adult individual carrying the variant in the genotype relevant to the disorder.
• Evidence that full penetrance at an early age is expected for the specific GNAS disorder and variant context.

Sources

Reasoning

No well-established functional study showing that GNAS p.Arg201His has no damaging effect on protein function or splicing was identified. The available variant-specific in vivo evidence instead supports constitutive activation and downstream signaling, but that positive evidence cannot be used to satisfy BS3.

Rule

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

Evidence considered

• Taki et al. PMID:26257060 directly studied GNAS R201H in vivo and reported constitutive Gsα activation, elevated cAMP, activated PKA signaling, and pancreatic phenotypes; it did not provide evidence of a no-damaging effect on protein function or splicing.

Still missing

• A validated functional assay demonstrating no damaging effect on GNAS protein function or splicing is absent.

Sources

Reasoning

BS4 is not assessed. No affected family members were evaluated for the exact variant, so there is no informative non-segregation result. The single-patient mosaic report cannot establish lack of segregation in an affected family.

Rule

Lack of segregation in affected members of a family.

Evidence considered

• The reviewed paper describes one patient with a mosaic postzygotic mutation and contains no affected-relative testing or family non-segregation analysis.

Still missing

• No affected family members were genotyped.
• No pedigree-based non-segregation analysis is available.
• Potential phenocopy and multiple-variant explanations cannot be evaluated without family data.

Sources

Reasoning

BP2 requires an observed phase relationship: the evaluated variant in trans with a pathogenic variant for a fully penetrant dominant gene/disorder, or in cis with a pathogenic variant in any inheritance pattern. The available case evidence reports the evaluated GNAS variant as mosaic and heterozygous but provides no second pathogenic variant and no parental, offspring, or other phase data establishing either trans or cis.

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

• PMID:1594625, Results: Arg201His was reported in one allele with mosaic tissue distribution, but the paper does not report a second pathogenic variant or cis/trans phase testing.
• Case evidence reviewed for this criterion contains no affected-proband observation with a pathogenic allele in trans and no observation of the evaluated variant in cis with a pathogenic allele.

Still missing

• A second pathogenic variant and definitive phase information from parental, offspring, or equivalent testing are not available.

Sources