ESR1 c.1138G>C, p.Glu380Gln

Reasoning

PMID:8114756 directly tested the exact ESR1 p.(Glu380Gln) substitution corresponding to NM_000125.4:c.1138G>C in transiently transfected estrogen-receptor-deficient CHO cells. The study included wild-type ER controls and multiple functionally relevant assays: estradiol binding, transcriptional activation, DNA binding, and hydroxytamoxifen suppression. E380Q retained similar estradiol-binding affinity to wild type (Kd 0.38 nM versus 0.50 nM), but required approximately two to three times less estradiol for activation, had 8-fold higher activity without added estrogen, showed enhanced DNA binding, and was less suppressible by hydroxytamoxifen. These results support an altered, constitutively active estrogen-receptor function relevant to a damaging gain-of-function effect. The study is biologically relevant and has appropriate wild-type comparison, but the available evidence is from one transient overexpression study without demonstrated independent replication or validation in a clinical diagnostic laboratory; therefore PS3 is met at supporting rather than strong strength.

Rule

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

Evidence considered

• Pakdel et al. (PMID:8114756) directly tested ESR1 E380Q, corresponding to NM_000125.4:c.1138G>C / NP_000116.2:p.(Glu380Gln), using site-directed mutagenesis and transient expression in estrogen-receptor-deficient CHO cells.
• The study used wild-type ER as a comparator and performed estradiol-binding Scatchard assays, Western blotting, estrogen-responsive CAT reporter transactivation assays, promoter-interference and gel-shift DNA-binding assays, and hydroxytamoxifen suppression experiments.
• PMID:8114756 reports E380Q estradiol-binding affinity similar to wild type (Kd 0.38 nM versus 0.50 nM), approximately two- to threefold lower estradiol requirement for activation, 8-fold higher activity without added estrogen, enhanced DNA binding, and reduced hydroxytamoxifen suppression.
• PMID:8114756, Abstract: "Mutation of the one charged amino acid nearest to C381 gave a mutant receptor (E380Q) requiring two to three times less estradiol (E2) than wild type (WT) ER to achieve maximal activity and having activity in the absence of added estrogen that was 8-fold higher than that of WT receptor."

Still missing

• Independent replication of the exact E380Q functional findings is not documented in the reviewed evidence.
• Validation and reproducibility in a clinical diagnostic laboratory setting are not documented.

Sources

Reasoning

PM2 is met at supporting strength. NM_000125.4:c.1138G>C (p.Glu380Gln) is absent from gnomAD v2.1 and gnomAD v4.1, with the available non-cancer subset results also reporting absence. This is consistent with an allele frequency of 0 and satisfies the supplied generic PM2 supporting threshold of <=0.0001; no VCEP-specific population source overrides the generic 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 v2.1 (exome) reports NM_000125.4:c.1138G>C absent.
• gnomAD v4.1 reports NM_000125.4:c.1138G>C absent.
• The available non-cancer subset results report the variant absent from gnomAD v2.1 non-cancer and gnomAD v3.1 non-cancer.
• The generic PM2 calibration supplied for this case is <=0.0001 at supporting strength (ClinGen SVI recommendation, PMID:25741868).

Still missing

• The available gnomAD records provide absence results but no allele-count, coverage, or grpmax FAF fields.

Sources

Reasoning

The variant is a missense substitution, NM_000125.4:c.1138G>C (p.Glu380Gln), so the generic missense PP3 path is REVEL only. REVEL is 0.85, meeting the supplied PP3 moderate threshold of >=0.773 (ClinGen SVI REVEL calibration, Pejaver et al. 2022, PMID:36413997). The SpliceAI result is not counted because splice prediction is not the governing path for a missense variant, and BayesDel has no established generic ACMG-strength calibration.

Rule

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

Evidence considered

• Local REVEL predictor lookup for NM_000125.4:c.1138G>C (p.Glu380Gln): REVEL = 0.85.
• ClinGen SVI REVEL calibration (Pejaver et al. 2022, PMID:36413997): PP3 moderate threshold >=0.773; the observed score of 0.85 meets this threshold.
• Path selection: this is a missense variant, so only REVEL was used for PP3/BP4; the available SpliceAI score was not counted, and BayesDel was not used because no generic ACMG-strength calibration is established.

Sources

Reasoning

Generic ACMG/AMP PS4 requires a significantly increased prevalence in affected individuals compared with controls, with an RR or OR >5.0 and a confidence interval that excludes 1.0. The literature_pass.json packet identified PMID:22918138 and PMID:34131312 as PS4 candidates, but direct full-text extraction verified that neither mentions ESR1 c.1138G>C or p.(E380Q), and neither reports affected-case/control counts. No case-control comparison, counts, OR, or 95% CI is available for this exact variant; therefore PS4 is not met.

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

• literature_pass.json listed PMID:22918138 and PMID:34131312 under the PS4 index; their full-text extractions were checked rather than accepted from classifier triage.
• PMID:22918138 full text does not mention ESR1, c.1138G>C, p.(E380Q), or an equivalent notation and reports no variant-specific case observation or population-frequency information.
• PMID:34131312 full text does not mention ESR1 or p.(E380Q); it is a chromosomal-microarray technical standard and supplies no exact-variant case-control data.

Still missing

• No exact-variant affected-case and control counts or matched-ancestry control frequency were identified.
• No exact-variant odds ratio or 95% confidence interval was available for comparison with the >5.0 and CI-excludes-1.0 requirements.

Sources

Reasoning

Generic ACMG/AMP PP5 concerns a reputable source reporting the variant as pathogenic when the evidence is unavailable for independent evaluation. Under the specified generic rule, only an exact-variant ClinVar expert-panel Pathogenic or Likely pathogenic classification qualifies. ClinVar variation 4539611 is exact-match but has a single-submitter, one-star Likely oncogenic somatic classification, with zero expert-panel submissions; the screened literature candidates PMID:22918138 and PMID:34131312 do not mention the variant. 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 variation 4539611 exactly matches NM_000125.4(ESR1):c.1138G>C (p.Glu380Gln), but the record has one submission, one-star criteria-provided single-submitter review status, somatic Likely oncogenic significance, and zero expert-panel submissions.
• PMID:22918138 and PMID:34131312 were screened because literature_pass.json listed them as PP5 candidates; their full-text extractions found no mention of ESR1 c.1138G>C or p.(E380Q).

Still missing

• No exact-variant ClinVar expert-panel Pathogenic or Likely pathogenic classification was identified.

Sources

Reasoning

BA1 is not met. NM_000125.4:c.1138G>C (p.Glu380Gln) is absent from gnomAD v2.1 and gnomAD v4.1, so no filtering allele frequency or grpmax FAF at or above the generic stand-alone threshold of 0.05 is reported. The available non-cancer subset results also report absence. Because no elevated frequency is observed, the stand-alone benign threshold is not reached.

Rule

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

Evidence considered

• gnomAD v2.1 (exome) reports NM_000125.4:c.1138G>C absent.
• gnomAD v4.1 reports NM_000125.4:c.1138G>C absent.
• The available non-cancer subset results report the variant absent from gnomAD v2.1 non-cancer and gnomAD v3.1 non-cancer.
• The generic BA1 threshold supplied for this case is >=0.05 (ACMG/AMP 2015, PMID:25741868).

Still missing

• The available gnomAD records provide absence results but no allele-count or grpmax FAF fields.

Sources

Reasoning

BS1 is not met on the available population evidence. The variant is absent from gnomAD v2.1 and gnomAD v4.1, and the available non-cancer subset results also report absence; therefore no filtering allele frequency or grpmax FAF demonstrates that the allele is greater than expected for the disorder. The generic 0.01 default is not reached, and no maximum-credible-frequency calculation is needed because no elevated frequency is observed.

Rule

Allele frequency is greater than expected for the disorder.

Evidence considered

• gnomAD v2.1 (exome) reports NM_000125.4:c.1138G>C absent.
• gnomAD v4.1 reports NM_000125.4:c.1138G>C absent.
• The available non-cancer subset results report the variant absent from gnomAD v2.1 non-cancer and gnomAD v3.1 non-cancer.
• The generic BS1 default supplied for this case is >=0.01; the prompt requires consideration of prevalence, penetrance, allelic heterogeneity, and maximum credible allele frequency before applying that default (Whiffin et al. 2017, PMID:28518168).

Still missing

• No disease prevalence, penetrance, allelic heterogeneity, or gene-specific maximum-credible-frequency inputs are available.
• The available gnomAD records provide absence results but no allele-count or grpmax FAF fields.

Sources

Reasoning

The exact ESR1 E380Q substitution was tested in PMID:8114756 with wild-type ER controls and multiple relevant assays. Rather than showing no damaging effect, E380Q produced increased estrogen-independent transcriptional activity, greater estrogen sensitivity, enhanced DNA binding, and reduced hydroxytamoxifen-mediated suppression; its activity without added estrogen was 8-fold higher than wild type. These findings contradict the BS3 requirement for functional studies showing no damaging effect on protein function. The paper does not provide evidence supporting BS3.

Rule

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

Evidence considered

• PMID:8114756 directly evaluated ESR1 E380Q against wild-type ER in estradiol-binding, transcriptional activation, DNA-binding, and hydroxytamoxifen suppression assays.
• The exact variant showed 8-fold higher activity without added estrogen, required two to three times less estradiol for activation, and had enhanced DNA binding relative to wild type, providing evidence of altered function rather than a normal-function result.
• PMID:8114756, Abstract: "Mutation of the one charged amino acid nearest to C381 gave a mutant receptor (E380Q) requiring two to three times less estradiol (E2) than wild type (WT) ER to achieve maximal activity and having activity in the absence of added estrogen that was 8-fold higher than that of WT receptor."

Sources

Reasoning

The variant is a missense substitution, so BP4 must be judged on the generic missense REVEL path only. REVEL is 0.85, which is above the BP4 supporting upper threshold of <=0.29 and is therefore indeterminate for a benign computational effect. The SpliceAI max delta of 0.004 is not used for BP4 because splice prediction is not the governing path for a missense variant; BayesDel is not available for generic ACMG-strength assignment.

Rule

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

Evidence considered

• Local REVEL predictor lookup for NM_000125.4:c.1138G>C (p.Glu380Gln): REVEL = 0.85.
• ClinGen SVI REVEL calibration (Pejaver et al. 2022, PMID:36413997): BP4 supporting threshold <=0.29; the observed score of 0.85 does not meet it and lies outside the benign range.
• Path selection: this is a missense variant, so only REVEL was used for PP3/BP4; the available SpliceAI max delta of 0.004 was not substituted for the indeterminate REVEL result.

Sources

Reasoning

Generic ACMG/AMP BP6 concerns a reputable source reporting the variant as benign when the evidence is unavailable for independent evaluation. Under the specified generic rule, only an exact-variant ClinVar expert-panel Benign or Likely benign classification qualifies. ClinVar variation 4539611 is exact-match but has a single-submitter, one-star Likely oncogenic somatic classification and zero expert-panel submissions, not a benign expert-panel classification; therefore BP6 is not met.

Rule

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

Evidence considered

• ClinVar variation 4539611 exactly matches NM_000125.4(ESR1):c.1138G>C (p.Glu380Gln), but its only submission is one-star, single-submitter, somatic Likely oncogenic, with zero expert-panel submissions.

Still missing

• No exact-variant ClinVar expert-panel Benign or Likely benign classification was identified.

Sources

Reasoning

The variant is NP_000116.2:p.(Glu380Gln), but the available evidence does not establish a previously established pathogenic variant producing the same amino-acid change through a different nucleotide change. The ClinVar record is a single-submitter somatic likely-oncogenic record and is not sufficient evidence for the generic germline PS1 criterion. No validated same-amino-acid pathogenic comparator was identified.

Rule

Same amino acid change as a previously established pathogenic variant regardless of nucleotide change. ... Example: Val->Leu caused by either G>C or G>T in the same codon.

Evidence considered

• ClinVar variation 4539611 is an exact match for NM_000125.4:c.1138G>C (p.Glu380Gln), but its only submission is a single-submitter somatic likely-oncogenic classification, not an established pathogenic germline comparator.

Still missing

• A validated pathogenic ESR1 variant with the same p.Glu380Gln amino-acid change and a different nucleotide change is needed.

Sources

Reasoning

The available evidence identifies ESR1 NM_000125.4:c.1138G>C (p.Glu380Gln) and includes a functional study, but provides no proband phenotype documentation, family-history assessment, or testing results confirming both maternity and paternity. Therefore, the required de novo observation cannot be assessed.

Rule

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

Evidence considered

• PMID:8114756 directly studied p.Glu380Gln in cell-based functional assays but reported no patient observations, family history, parental testing, or de novo data.
• No case-level proband, parental-testing, or family-history evidence establishing confirmed de novo status was present in the reviewed case materials.

Still missing

• Clinical documentation showing that the proband has the relevant disease phenotype.
• Documented absence of a relevant family history.
• Variant testing results for the proband and both biological parents.
• Confirmation of both maternity and paternity, with assessment for non-maternity mechanisms where relevant.

Sources

Reasoning

ESR1 has no VCEP domain table in vcep_materials.json. PMID:8114756 places E380 in the estrogen-receptor hormone-binding domain and reports functional effects, but the available evidence does not establish that this domain is a generic ACMG critical and well-established functional domain meeting the PM1 requirement without benign variation. The Cancer Hotspots result cannot be used as a PM1 route here because no VCEP hotspot route governs this gene.

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

• Pakdel et al. studied E380Q in the estrogen-receptor hormone-binding domain and observed altered transcriptional activity, but this does not by itself establish the generic PM1 domain and benign-variation requirements.
• The Cancer Hotspots result reports an E380 hotspot, but the governing generic interpretation has no VCEP-defined hotspot route for ESR1; therefore it is not used to meet PM1.

Still missing

• A documented generic ACMG critical, well-established functional domain designation for ESR1 E380 and evidence of absent benign variation in that domain are needed.

Sources

Reasoning

PM3 cannot be assessed. The available evidence concerns a somatic ESR1 p.(Glu380Gln) functional study and contains no affected-proband observations, recessive ESR1 disease context, pathogenic allele in trans, or parent/offspring testing establishing phase. The paper specifically reports no patient observations or segregation data, so there is no basis to apply PM3.

Rule

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

Evidence considered

• PMID:8114756 directly studied ESR1 p.(Glu380Gln) in engineered CHO-cell assays and reported no patient observations or segregation data.
• The case evidence does not document a recessive ESR1 disease indication, an affected proband carrying this variant with a pathogenic allele, or parent/offspring phase testing.

Still missing

• A defined recessive ESR1 disorder context is not established.
• No affected-proband observation with a second pathogenic variant is documented.
• No parent or offspring testing establishes trans phase with a pathogenic variant.

Sources

Reasoning

The observed variant is p.Glu380Gln, but no different missense change at residue 380 with an established pathogenic determination was identified in the available evidence. The PM5 candidate artifact contains zero same-residue candidates and cannot establish PM5.

Rule

Novel missense change at an amino acid residue where a different missense change determined to be pathogenic has been seen before. ... Example: Arg156His is pathogenic; now you observe Arg156Cys.

Evidence considered

• The PM5 candidate record reports total_candidates_seen=0 and same_residue_candidates=0, so it provides no qualifying pathogenic comparator for residue 380.

Still missing

• A different missense ESR1 variant at residue 380 with an established pathogenic determination is needed.

Sources

Reasoning

The available evidence does not document an assumed de novo occurrence in a patient, and it provides no parental testing or family-history information. The functional study of p.Glu380Gln does not supply case-level evidence for PM6.

Rule

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

Evidence considered

• PMID:8114756 reports functional experiments only and no patient observations, parental testing, or de novo data.
• No case-level evidence supporting an assumed de novo occurrence was present in the reviewed case materials.

Still missing

• A documented affected patient carrying the variant.
• Evidence that the variant arose de novo or was assumed de novo.
• Available parental testing and family-history information.

Sources

Reasoning

No affected family members, segregation pattern, informative meioses, or cosegregation data are reported for this variant. The available variant-specific publication is a functional study and does not provide family data; 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

• PMID:8114756 reports no patient observations or segregation data and therefore does not establish cosegregation.
• No family pedigree, affected-relative testing, or informative meiosis data were present in the reviewed case materials.

Still missing

• Pedigree identifying affected and unaffected relatives.
• Genotype results for informative family members.
• Phenotype consistency and number of informative meioses.

Sources

Reasoning

The available case evidence does not establish both PP2 prerequisites for ESR1: a low rate of benign missense variation and missense variants as a common disease mechanism. Absence of this variant from gnomAD and a somatic gain-of-function annotation do not establish either gene-level prerequisite.

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

• The variant is absent from gnomAD v2.1 and v4.1, which is variant-level population evidence and does not quantify the gene's benign missense rate.
• OncoKB describes this somatic ESR1 p.Glu380Gln variant as likely oncogenic with gain-of-function context, but that does not establish the generic PP2 gene-level prerequisites.

Still missing

• Gene-level evidence for a low rate of benign ESR1 missense variation and a common missense disease mechanism is needed.

Sources

Reasoning

The governing PP4 rule requires a patient's phenotype or family history that is highly specific for a disease with a single genetic etiology. The case evidence provided no patient phenotype, family history, or disease-specific clinical presentation for this ESR1 variant, so PP4 cannot be assessed.

Rule

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

Still missing

• Patient phenotype and family-history details are absent.
• No single-genetic-etiology disease context can be evaluated from the available clinical information.

Reasoning

BS2 cannot be assessed from the available evidence. The gnomAD results report the variant absent, but they do not document an observation in a healthy adult, a homozygote, a heterozygote, or a hemizygote. The available evidence also does not establish a relevant disorder with full penetrance expected at an early age, so the BS2 applicability requirements cannot be confirmed.

Rule

Observed in a healthy adult individual for a recessive (homozygous), dominant (heterozygous), or X-linked (hemizygous) disorder, with full penetrance expected at an early age.

Evidence considered

• gnomAD v2.1 and v4.1 report the variant absent; neither record documents a healthy-adult carrier observation or genotype count.
• The available non-cancer subset results also report absence and do not document a healthy-adult observation.
• No reviewed population evidence establishes full penetrance with early-age manifestation for the relevant disorder.

Still missing

• No individual-level healthy-adult observation or homozygote/heterozygote/hemizygote count is available.
• The relevant disease inheritance, penetrance, and age-of-onset requirements are not established in the available evidence.

Sources

Reasoning

No affected relatives with known negative testing for the variant are documented, and no pedigree or family genotype-phenotype information is available. Consequently, lack of segregation cannot be evaluated and BS4 is not assessed.

Rule

Lack of segregation in affected members of a family.

Evidence considered

• PMID:8114756 reports functional experiments only and no family observations or segregation testing.
• No affected family members with documented absence of the variant were identified in the reviewed case materials.

Still missing

• A pedigree with affected relatives and reliable phenotype assignments.
• Variant testing results in affected relatives.
• Evaluation for phenocopies and for more than one pathogenic variant in the family.

Sources

Reasoning

The available evidence does not establish that ESR1 is a gene for which primarily truncating variants are known to cause disease. The case includes somatic gain-of-function information and no validated gene-level spectrum showing a primarily truncating disease mechanism.

Rule

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

Evidence considered

• OncoKB reports somatic ESR1 p.Glu380Gln gain-of-function context, which does not establish the BP1 requirement concerning the gene's primarily truncating disease mechanism.

Still missing

• A validated ESR1 disease-variant spectrum demonstrating that primarily truncating variants cause disease is needed.

Sources

Reasoning

BP2 cannot be assessed. The available evidence does not document this variant observed in trans with a pathogenic variant in a fully penetrant dominant gene/disorder, nor observed in cis with a pathogenic variant. The available publication is a functional study without patient, segregation, or phase data, and the case record supplies no allelic observations for this variant.

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:8114756 directly studied ESR1 p.(Glu380Gln) functionally but reported no patient observations or segregation data.
• The case evidence contains no documented pathogenic variant in cis or trans with ESR1 p.(Glu380Gln), and no phase information.

Still missing

• No fully penetrant dominant ESR1 disease context and trans observation with a pathogenic variant are documented.
• No cis observation with a pathogenic variant is documented.
• No parent, offspring, tumor-normal, or other phasing evidence is available.

Sources

Reasoning

The governing BP5 rule requires the variant to be found in a case with an alternate molecular basis for disease. The available evidence contains no case-level report linking this exact ESR1 variant to a patient with an independently established alternate molecular cause, so BP5 cannot be assessed.

Rule

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

Still missing

• No exact-variant case report with a documented alternate molecular basis for disease was identified.