owencqueen/cgbench_data huggingface dataset:
Gene: GUCY2D (HGNC:4689)
Evidence Code: PVS1 — Original ACMG Summary
Null variant (nonsense, frameshift, canonical +/−1 or 2 splice sites, initiation codon, single or multi-exon deletion) in a gene where loss of function (LOF) is a known mechanism of disease.
Caveats:
 • Beware of genes where LOF is not a known disease mechanism (e.g. GFAP, MYH7).
 • Use caution interpreting LOF variants at the extreme 3’ end of a gene.
 • Use caution with splice variants that are predicted to lead to exon skipping but leave the remainder of the protein intact.
 • Use caution in the presence of multiple transcripts.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PVS1 — Very Strong
Modification Type: Gene-specific
Null variant in a gene where loss of function is a known mechanism of disease.




Use as defined by ClinGen SVI working group (Abou Tayoun et al., 2018
16
) and as updated by the ClinGen SVI Splicing Subgroup (Walker et al., 2023
17
).


Refer to GUCY2D-specific PVS1 Decision Tree, file attached.


PVS1: Predicted splice defects at +/- 1,2 in exons 2-20


PVS1: Single to multi-exon deletions, with or without predicted NMD. All exons are considered critical to protein function.


PVS1: Applied to variants in the initiation codon. The second in-frame methionine is located at residue 218 and use of this as a start codon would eliminate the leader sequence which targets the protein to the endoplasmic reticulum. There is no known study indicating that this methionine in GUCY2D can be used as start codon. Variants affecting Met1 can lead to a complete absence of the protein product. Additionally, there are multiple variants located upstream of p.Met218 that have been reported as a pathogenic in HGMD and ClinVar, evidence that this entire region of the protein is functionally important.


PVS1: Nonsense or frameshift mutations from p.Thr2 through p.Lys1068


PVS1: Duplications of exons proven in tandem


PVS1(RNA): RNA splicing data with evidence of alternative transcript production at complete levels, relative to normal allele.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PVS1 — Strong
Modification Type: Gene-specific
Null variant in a gene where loss of function is a known mechanism of disease.




Use as defined by ClinGen SVI working group (Abou Tayoun et al., 2018
16
) and as updated by the ClinGen SVI Splicing Subgroup (Walker et al., 2023
17
).


Refer to GUCY2D-specific PVS1 Decision Tree, file attached.


PVS1_Strong: Nonsense or frameshift mutations from p.Pro1069 through p.Ser1103


PVS1_Strong: Duplications of exons presumed in tandem


PVS1(RNA)_Strong: RNA splicing data with evidence of alternative transcript production at near complete levels, relative to normal allele.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS1 — Original ACMG Summary
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.
Caveat: Beware of changes that impact splicing rather than at the amino acid/protein level.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS1 — Strong
Modification Type: Gene-specific
Same amino acid change as a previously established Pathogenic variant regardless of nucleotide change.




Must have one comparison variant that reaches a Pathogenic classification using this rule specification.




Same predicted splicing impact as a previously classified Pathogenic variant. 




Used in conjunction with PP3 for variants located outside the splice donor/acceptor +/-1,2 dinucleotide positions that have a splice AI score ≥0.2 and have a comparable nucleotide variant at the 
same position
 that has been designated 
Pathogenic
.


Used in conjunction with PVS1 for variants located at the splice donor/acceptor +/-1,2 dinucleotide positions and that have a comparable variant within the same splice donor/acceptor +/-1,2 dinucleotide that has been designated Pathogenic.




Specific combinations are found in GUCY2D-specific PVS1 Decision Tree 
part (b)
 (Table 2 from Walker 2023).
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS1 — Moderate
Modification Type: Gene-specific,Strength
Same amino acid change as a previously established Likely Pathogenic variant regardless of nucleotide change.




Must have one comparison variant that reaches a Likely Pathogenic classification using this rule specification.




Same predicted splicing impact as previously classified Likely Pathogenic variant.




Used in conjunction with PP3 for variants located outside the splice donor/acceptor +/-1,2 dinucleotide positions that have a splice AI score ≥0.2 and have a comparable nucleotide variant at the 
same position
 that has been designated 
Likely Pathogenic
.


Used in conjunction with PVS1_(reduced strength) for variants located at the splice donor/acceptor +/-1,2 dinucleotide positions and have a comparable variant 
within the same splice donor/acceptor motif
 (but outside the +/-1,2 dinucleotide) that has been designated 
Pathogenic
.




Specific combinations are found in GUCY2D-specific PVS1 Decision Tree 
part (b)
 (Table 2 from Walker 2023).
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS1 — Supporting
Modification Type: Gene-specific,Strength
Used in conjunction with PP3 for variants located outside the splice donor/acceptor +/-1,2 dinucleotide positions that have a splice AI score ≥0.2 and have a comparable nucleotide variant within the 
same motif
 that has been designated 
Likely Pathogenic
.


Used in conjunction with PVS1 or PVS1_(reduced strength) for variants located at the splice donor/acceptor +/-1,2 dinucleotide positions and have a comparable Likely Pathogenic or Pathogenic variant either within the same splice site donor/acceptor motif, outside the +/-1,2 dinucleotide, or at the +/-1,2 dinucleotide.




 Specific combinations are found in GUCY2D-specific PVS1 Decision Tree 
part (b)
 (Table 2 from Walker 2023).
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS2 — Original ACMG Summary
De novo (both maternity and paternity confirmed) in a patient with the disease and no family history.
Note: Confirmation of paternity only is insufficient. Egg donation, surrogate motherhood, errors in embryo transfer, etc. can contribute to non-maternity.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS2 — Very Strong
Modification Type: Gene-specific
De novo (
both
 maternity and paternity confirmed) in a patient with the disease and no family history.




Use ClinGen SVI's recommendation for assigning weight to the PS2/PM6 codes (See PS2-Table 1 within PS2/PM6 file). 


Total of 4 or more points required for Very Strong level
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS2 — Strong
Modification Type: Gene-specific
De novo (
both
 maternity and paternity confirmed) in a patient with the disease and no family history.




Use ClinGen SVI's recommendation for assigning weight to the PS2/PM6 codes (See PS2-Table 1 within PS2/PM6 file). 


Total of 2.00 - 3.75 points required for Strong level
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS2 — Moderate
Modification Type: Gene-specific
De novo (
both
 maternity and paternity confirmed) in a patient with the disease and no family history.




Use ClinGen SVI's recommendation for assigning weight to the PS2/PM6 codes (See PS2-Table 1 within PS2/PM6 file). 


Total of 1.00 - 1.75 points required for Moderate level
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS2 — Supporting
Modification Type: Gene-specific
De novo (
both
 maternity and paternity confirmed) in a patient with the disease and no family history.




Use ClinGen SVI's recommendation for assigning weight to the PS2/PM6 codes (See Table 1 within PS2/PM6 file). 


Total of 0.50 - 0.75 points required for Supporting level
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS3 — Original ACMG Summary
Well-established in vitro or in vivo functional studies supportive of a damaging effect on the gene or gene product.
Note: 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.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS3 — Strong
Modification Type: Gene-specific,Strength
Well-established in vitro or in vivo functional studies supportive of a damaging effect. 


See attached table for acceptable functional studies. 


Eligible results include 
Gucy2e−/−Gucy2f−/−
 knockout mice subretinally injected with adeno-associated virus-packaged constructs encoding a GUCY2D variant (Boye et al., 2016,
10
) or mice with knock-in of a 
GUCY2D
 variant (not yet available in the published literature). Results should include flat or nearly flat scotopic and photopic electroretinogram responses in comparison to wild-type injected control, absent or near-absent guanylate cyclase activity in comparison to wild-type injected control, and/or absent or near absent GUCY2D expression at the protein level despite approximately wild-type expression at the RNA level.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS3 — Supporting
Modification Type: Gene-specific,Strength
Well-established in vitro or in vivo functional studies supportive of a damaging effect. Not applicable for splicing effects (replaced by PVS1_Strength (RNA))


See attached table for acceptable functional studies.


For studies reporting guanylate cyclase activity, cutoff is ≤10% of wild-type control.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PS4 — Original ACMG Summary
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. See manuscript for detailed guidance.
Note 2: In instances of very rare variants where case-control studies may not reach statistical significance, the prior observation of the variant in multiple unrelated patients with the same phenotype, and its absence in controls, may be used as moderate level of evidence.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM1 — Original ACMG Summary
Located in a mutational hot spot and/or critical and well-established functional domain (e.g. active site of an enzyme) without benign variation.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM1 — Moderate
Modification Type: Gene-specific
Met by active site residues that bind the GTP substrate or the Mg
2+
 ion; Phe883, Asp885, Thr890, Leu905, Glu925, Ile927, Asp929, Met932, Arg976, Arg995, Cys997, Leu998, Phe999, Gly1000, Val1003, Asn1004, Arg1008, and Glu1010 
13
.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM1 — Supporting
Modification Type: Gene-specific
Met by variants not explicitly listed above, encoding missense substitutions between positions p.873-951. This region forms part of the guanylate cyclase catalytic domain and exhibits intolerance to missense variation 
14
.


Met by variants in the 6 bp CRX binding site located at position 17:8002593-8002598 (hg38) 
18
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM2 — Original ACMG Summary
Absent from controls (or at extremely low frequency if recessive) in Exome Sequencing Project, 1000 Genomes or Exome Aggregation Consortium.
Caveat: Population data for indels may be poorly called by next generation sequencing.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM2 — Supporting
Modification Type: Disease-specific,Strength
Absent/rare from controls in an ethnically-matched cohort population sample.




Used if the gnomAD total allele frequency is ≤ 4.0 x 10
-4
.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM3 — Original ACMG Summary
For recessive disorders, detected in trans with a pathogenic variant
Note: This requires testing of parents (or offspring) to determine phase.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM3 — Very Strong
Modification Type: Disease-specific
Points are calculated using Table 1 from 
https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf
 (or attached file “PM3 Tables”)




≥4 points from Table 1
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM3 — Strong
Modification Type: Disease-specific
Points are calculated using Table 1 from 
https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf
 (or attached file “PM3 Tables”)




 2 to 3.75 points from Table 1
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM3 — Moderate
Modification Type: Disease-specific
Points are calculated using Table 1 from 
https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf
 (or attached file “PM3 Tables”)




 1 to 1.75 points from Table 1
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM3 — Supporting
Modification Type: Disease-specific
Points are calculated using Table 1 from 
https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf
 (or attached file “PM3 Tables”)




 0.5 to 0.75 points from Table 1
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM4 — Original ACMG Summary
Protein length changes due to in-frame deletions/insertions in a non-repeat region or stop-loss variants.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM4 — Moderate
Modification Type: Gene-specific
Protein length changes due to in-frame deletions/insertions in a non-repeat region or stop-loss variants.




 Protein length change of ≥2 amino acids that leads to loss of at least one conserved residue (PhyloP>2.0) or insertion of new amino acids adjacent to at least one conserved residue (PhyloP>2.0).
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM4 — Supporting
Modification Type: Gene-specific,Strength
Protein length changes due to in-frame deletions/insertions in a non-repeat region or stop-loss variants.




 Protein length change of 1 amino acid that leads to loss of at least one conserved residue (PhyloP>2.0) or insertion of new amino acid adjacent to at least one conserved residue (PhyloP>2.0).
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM5 — Original ACMG Summary
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.
Caveat: Beware of changes that impact splicing rather than at the amino acid/protein level.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM5 — Moderate
Modification Type: None
Missense change at an amino acid residue where a different missense change determined to be pathogenic has been seen before.




Must have one comparison variant that reaches a Pathogenic classification using this rule specification.


SpliceAI scores for both variants should be in the same category (<0.1, between 0.1 and 0.2, >0.2)
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM5 — Supporting
Modification Type: Strength
Missense change at an amino acid residue where a different missense change determined to be pathogenic has been seen before.




Must have one comparison variant that reaches a Likely Pathogenic classification using this rule specification.


SpliceAI scores for both variants should be in the same category (<0.1, between 0.1 and 0.2, >0.2)
Gene: GUCY2D (HGNC:4689)
Evidence Code: PM6 — Original ACMG Summary
Assumed de novo, but without confirmation of paternity and maternity.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP1 — Original ACMG Summary
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.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP1 — Strong
Modification Type: Disease-specific,Strength
Co-segregation with disease in multiple affected family members and evidence that this variant and another GUCY2D variant are 
in trans
.




Requires segregation in one proband plus ≥3 similarly affected relatives
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP1 — Moderate
Modification Type: Disease-specific,Strength
Co-segregation with disease in multiple affected family members and evidence that this variant and another GUCY2D variant are 
in trans
.




Requires segregation in one proband plus 2 similarly affected relatives
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP1 — Supporting
Modification Type: Disease-specific,Strength
Co-segregation with disease in multiple affected family members and evidence that this variant and another GUCY2D variant are 
in trans
.




Requires segregation in one proband plus 1 similarly affected relative
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP2 — Original ACMG Summary
Missense variant in a gene that has a low rate of benign missense variation and where missense variants are a common mechanism of disease.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP3 — Original ACMG Summary
Multiple lines of computational evidence support a deleterious effect on the gene or gene product (conservation, evolutionary, splicing impact, etc.).
Caveat: As many in silico algorithms use the same or very similar input for their predictions, each algorithm should not be counted as an independent criterion. PP3 can be used only once in any evaluation of a variant.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP3 — Moderate
Modification Type: Gene-specific,Strength
For a missense variant use REVEL, requires a score of ≥ 0.774.


Splice variants use PP3 only at Supporting level.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP3 — Supporting
Modification Type: Gene-specific
For a missense variant use REVEL, requires a score of 0.644 - 0.773.


For a variant in an untranslated region use CADD, requires a score of ≥ 20.0


For a predicted splicing variant use SpliceAI with max distance set to 500 bp. Highest delta score from SpliceAI must be ≥ 0.2 to use this code. See GUCY2D-specific PVS1 Decision Tree 
part (a)
 for SpliceAI flowchart.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP4 — Original ACMG Summary
Patient’s phenotype or family history is highly specific for a disease with a single genetic etiology.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP4 — Moderate
Modification Type: Disease-specific,Strength
8 phenotype points or more are required to use this code at the moderate strength for a single proband (see points list below).


Additionally, at least one 
specific
 criterion must be met (see below).


Do not include a proband with a suspected diagnosis of more than one retinal dystrophy.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP4 — Supporting
Modification Type: Disease-specific
4-7.5 phenotype points are required to use this code (see points list below).


Do not include a proband with a suspected diagnosis of more than one retinal dystrophy.
Gene: GUCY2D (HGNC:4689)
Evidence Code: PP5 — Original ACMG Summary
Reputable source recently reports variant as pathogenic, but the evidence is not available to the laboratory to perform an independent evaluation.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BA1 — Original ACMG Summary
Allele frequency is above 5% in Exome Sequencing Project, 1000 Genomes or Exome Aggregation Consortium.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BA1 — Stand Alone
Modification Type: Disease-specific
Use gnomAD Grpmax FAF if available, cutoff of  >0.016




Use large population databases (i.e. gnomAD).
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS1 — Original ACMG Summary
Allele frequency is greater than expected for disorder.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS1 — Strong
Modification Type: Disease-specific
Allele frequency greater than expected for disorder. Use gnomAD Grpmax FAF if available.




Allele frequency of 0.0016 - 0.016
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS2 — Original ACMG Summary
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.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS2 — Strong
Modification Type: Disease-specific,Strength
Variant is present in ≥ 3 homozygotes without any features of the phenotype. This rule applies to individuals found in the literature who have been well-phenotyped and are unaffected by age 40. Alternatively, this strength can be applied if the variant is present in ≥ 6 homozygotes in gnomAD v.4.1.0 or later.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS2 — Supporting
Modification Type: Disease-specific,Strength
Variant is present in ≥ 3 homozygotes in gnomAD v.4.1.0 or later.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS3 — Original ACMG Summary
Well-established in vitro or in vivo functional studies show no damaging effect on protein function or splicing.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS3 — Supporting
Modification Type: Gene-specific,Strength
Not applicable for splicing effects (replaced by BP7_Strong (RNA)).


For studies reporting guanylate cyclase activity, cutoff is >50% of wild-type control. Studies included; Peshenko et al., 2020
1
, Feng et al., 2020, Jacobson et al., 2021
2
, Jacobson et al., 2020
3
, Tucker et al., 2004
4
, Jacobson et al., 2013
5
, and Peshenko et al., 2015
7


See attached table of Approved PS3 Functional Assays for a list of acceptable functional studies including those that:




Report failure of GUCY2D to localize to the cell surface : Zulliger et al., 2015
6
.


Report failure of GUCY2D to bind RD3: Zulliger et al., 2015
6
. 


Report failure of GCAP1, GCAP2, or RD3 to co-localize with GUCY2D at the cell surface: Peshenko et al., 2020
1
, Jacobson et al., 2020
3
, and Peshenko et al., 2015
7
.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS4 — Original ACMG Summary
Lack of segregation in affected members of a family.
Caveat: The presence of phenocopies for common phenotypes (i.e. cancer, epilepsy) can mimic lack of segregation among affected individuals. Also, families may have more than one pathogenic variant contributing to an autosomal dominant disorder, further confounding an apparent lack of segregation.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BS4 — Strong
Modification Type: Clarification
Lack of segregation in affected members of a family.




One or both variants are absent in a similarly affected family member.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BP1 — Original ACMG Summary
Missense variant in a gene for which primarily truncating variants are known to cause disease.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BP2 — Original ACMG Summary
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.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BP2 — Supporting
Modification Type: Disease-specific
Observed in cis with a Pathogenic variant.




Use code if the variant of interest is 
in cis
 with a Pathogenic or Likely Pathogenic variant. The other variant must meet a Likely Pathogenic or Pathogenic classification using these rule specifications.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BP3 — Original ACMG Summary
In frame-deletions/insertions in a repetitive region without a known function.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BP4 — Original ACMG Summary
Multiple lines of computational evidence suggest no impact on gene or gene product (conservation, evolutionary, splicing impact, etc)
Caveat: As many in silico algorithms use the same or very similar input for their predictions, each algorithm cannot be counted as an independent criterion. BP4 can be used only once in any evaluation of a variant.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BP4 — Moderate
Modification Type: Gene-specific,Strength
For a missense variant use REVEL, requires a score of ≤0.183. In addition, highest SpliceAI delta score should also be below cutoff of 0.1.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BP4 — Supporting
Modification Type: Gene-specific
For a missense variant use REVEL, requires a score between 0.183 - 0.290. In addition, highest SpliceAI delta score should also be below cutoff of 0.1.


For a silent / intronic variant outside the designated splice region (conservatively at or beyond positions +7/-21) and synonymous (silent) exonic variants located outside of the first and the last 3 bases of the exon, BP4 can be met if the highest of the four delta scores within SpliceAI is below the cutoff of ≤0.1. See GUCY2D-specific PVS1 Decision Tree 
part (a)
 for SpliceAI flowchart.




Please note that BP7 can be met as well.
Gene: GUCY2D (HGNC:4689)
Evidence Code: BP5 — Original ACMG Summary
Variant found in a case with an alternate molecular basis for disease.