owencqueen/cgbench_data huggingface dataset:
Gene: GATM (HGNC:4175)
Evidence Code: PVS1 — Very Strong
Modification Type: None
Nonsense-mediated decay predicted.


CCDS VCEP notes:
Loss of function (LOF) of GATM is a known mechanism of disease for arginine:glycine amidinotransferase deficiency (AGAT-D). There are examples of various LOF variants, including nonsense and frameshift, in GATM in individuals with AGAT-D (
https://databases.lovd.nl/shared/variants/GATM/unique
). The specifications below are based on the PVS1 decision tree (see Appendix 1) (based on Figure 1, Abou Tayoun et al, 2018, PMID 30192042).


GATM specifications:
Nonsense and frameshift variants
* All nonsense and frameshift variants will meet PVS1 unless a premature termination codon is predicted to be missed by nonsense-mediated decay (NMD) because it is located in the last exon (exon 9) or the last 50 bases of the penultimate exon (exon 8, 3’ of c.1109). In that case, please refer to the PVS1 flowchart for guidance on PVS1 weight (Appendix 1).


Splice site variants (+1, +2, -1, -2)
* All canonical splice site pairs in GATM are GT-AG.
* For any canonical splice site variant (+1, +2, -1, -2), the exon immediately adjacent to the variant is predicted to be skipped i.e. upstream exon skipped for canonical donor splice site variants and downstream exon skipped for canonical acceptor splice site variants. However, to apply PVS1 at the very strong level, splice site variants must have no detectable nearby (+/- 20 nucleotides) strong consensus splice sequence that may reconstitute in-frame splicing, as predicted by SpliceAI. Otherwise, the PVS1 strength should be reduced accordingly.
* For the predicted in frame/out of frame consequences of exon skipping and considerations for strength of PVS1, see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss).
* If this criterion is applied, PP3 (in silico splice site prediction tools) should not be used.
* Non-canonical splice variants, such as +3 or -3, may also meet PVS1 - refer to Walker et al, PMID: 37352859.


Deletions (single or multi exon)
* If a single or multi-exon deletion results in an out of frame consequence, use PVS1 at the very strong level unless not predicted to undergo NMD. If not predicted to undergo NMD, please refer to Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss).
* If the consequence is in frame, the deletion must encompass one or more exons for PVS1 to apply. For weight of PVS1, see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss) .
* If the in frame deletion is smaller than one exon, PVS1 does not apply; consider using PM4.


Duplications
* To assess the impact of duplications, see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss).
Gene: GATM (HGNC:4175)
Evidence Code: PVS1 — Strong
Modification Type: Strength
In frame loss of >10% of the protein.
CCDS VCEP notes:
Loss of function (LOF) of GATM is a known mechanism of disease for arginine:glycine amidinotransferase deficiency (AGAT-D). There are examples of various LOF variants, including nonsense and frameshift, in GATM in individuals with AGAT-D (
https://databases.lovd.nl/shared/variants/GATM/unique
). The specifications below are based on the PVS1 decision tree (see Appendix 1) (Figure 1, Abou Tayoun et al, 2018, PMID 30192042).


GATM specifications:
Nonsense and frameshift variants
* All nonsense and frameshift variants will meet PVS1 unless a premature termination codon is predicted to be missed by nonsense-mediated decay (NMD) because it is located in the last exon (exon 9) or the last 50 bases of the penultimate exon (exon 8, 3’ of c.1109). In that case, please refer to the PVS1 flowchart for guidance on PVS1 weight (Appendix 1).


Splice site variants (+1, +2, -1, -2)
* All canonical splice site pairs in GATM are GT-AG.
* For any canonical splice site variant (+1, +2, -1, -2), the exon immediately adjacent to the variant is predicted to be skipped i.e. upstream exon skipped for canonical donor splice site variants and downstream exon skipped for canonical acceptor splice site variants. However, it is recommended to use SpliceAI to look for nearby (+/- 20 nucleotides) strong consensus splice sequence that may reconstitute in-frame splicing. 
* For considerations for strength at which PVS1 may be applied see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss) .
* If this criterion is applied, PP3 (in silico splice site prediction tools) should not be used.
* Non-canonical splice variants, such as +3 or -3, may also meet PVS1 - refer to Walker et al, PMID: 37352859.


Deletions (single or multi exon)
* If a single or multi-exon deletion results in an out of frame consequence, use PVS1 at the very strong level unless not predicted to undergo NMD. If not predicted to undergo NMD, please refer to Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss).
* If the consequence is in frame, the deletion must encompass one or more exons for PVS1 to apply. For weight of PVS1, see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss) .
* If the in frame deletion is smaller than one exon, PVS1 does not apply; consider using PM4.


Duplications
* To assess the impact of duplications, see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss).
Gene: GATM (HGNC:4175)
Evidence Code: PVS1 — Moderate
Modification Type: Strength
Single exon or larger deletion resulting in loss of <10% of the protein, and initiator codon variants.


CCDS VCEP notes:
Loss of function (LOF) of GATM is a known mechanism of disease for arginine:glycine amidinotransferase deficiency (AGAT-D). There are examples of various LOF variants, including nonsense and frameshift, in GATM in individuals with AGAT-D (
https://databases.lovd.nl/shared/variants/GATM/unique
). The specifications below are based on the PVS1 decision tree (Figure 1, Abou Tayoun et al, 2018, PMID 30192042).  


GATM specifications:


Nonsense and frameshift variants
* All nonsense and frameshift variants will meet PVS1 unless a premature termination codon is predicted to be missed by nonsense-mediated decay (NMD) because it is located in the last exon (exon 9) or the last 50 bases of the penultimate exon (exon 8, 3’ of c.1109). In that case, please refer to the PVS1 flowchart for guidance on PVS1 weight (Appendix 1).


Splice site variants (+1, +2, -1, -2)
* All canonical splice site pairs in GATM are GT-AG.
* For any canonical splice site variant (+1, +2, -1, -2), the exon immediately adjacent to the variant is predicted to be skipped i.e. upstream exon skipped for canonical donor splice site variants and downstream exon skipped for canonical acceptor splice site variants. However, it is recommended to look for nearby (+/- 20 nucleotides) strong consensus splice sequence that may reconstitute in-frame splicing. 
* For considerations for strength at which PVS1 may be applied see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss).
* If this criterion is applied, PP3 (in silico splice site prediction tools) should not be used.
* Non-canonical splice variants, such as +3 or -3, may also meet PVS1 (refer to Walker et al, PMID: 37352859).


Initiator codon variants
* All initiator codon variants will meet PVS1_Moderate. The next in-frame methionine is at amino acid position 130 (based on NP_001473). 


Deletions (single or multi exon)
* If a single or multi-exon deletion results in an out of frame consequence, use PVS1 at the very strong level unless not predicted to undergo NMD. If not predicted to undergo NMD, please refer to Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss).
* If the consequence is in frame, the deletion must encompass one or more exons for PVS1 to apply. For weight of PVS1, see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss) .
* If the in frame deletion is smaller than one exon, PVS1 does not apply; consider using PM4.


Duplications
* To assess the impact of duplications, see Appendix 1 (PVS1 flowchart) and Appendix 2 (predicted impact of exon loss).
Gene: GATM (HGNC:4175)
Evidence Code: PS1 — Strong
Modification Type: General recommendation
This criterion is applicable as is for any variant resulting in the same amino acid change as a previously established pathogenic variant regardless of nucleotide change.


If the variant is in the last 3 nucleotides of an exon, further analysis using splicing site prediction algorithms (see PP3) and data from the literature (if available) is required to investigate the impact on splicing.


PS1 may also be applied for splicing variants under specific circumstances (see Table 3 in PMID: 37352859).
Gene: GATM (HGNC:4175)
Evidence Code: PS1 — Moderate
Modification Type: General recommendation,Strength
This criterion is applicable as for any variant resulting in the same amino acid change as a previously established likely pathogenic variant regardless of nucleotide change.


If the variant is in the last 3 nucleotides of an exon, further analysis using splicing site prediction algorithms (see PP3) and data from the literature (if available) is required to investigate the impact on splicing.


PS1_Moderate may also be applied for splicing variants under specific circumstances (see Table 3 in PMID: 37352859).
Gene: GATM (HGNC:4175)
Evidence Code: PS1 — Supporting
Modification Type: General recommendation,Strength
PS1_Supporting may be applied for splicing variants under specific circumstances (see Table 3 in PMID: 37352859).
Gene: GATM (HGNC:4175)
Evidence Code: PS3 — Supporting
Modification Type: Disease-specific,Strength
AGAT activity data from an in vitro assay in which GATM variants were overexpressed in HeLa cells has been published (DesRoches et al, 2016; PMID 27233232). Any variant with AGAT activity at or below 15% of normal in this paper meets PS3_Supporting (see Appendix 3 for further details on AGAT functional assays).
Gene: GATM (HGNC:4175)
Evidence Code: PM2 — Supporting
Modification Type: Disease-specific,Strength
Allele frequency <0.000055 (<0.0055%) in all populations in gnomAD.


CCDS VCEP notes: It is acceptable for a GATM variant to be present in controls, if heterozygous, because AGAT-D is a recessive disorder. Homozygotes should not be seen in a population database, such as gnomAD, because the penetrance of this condition in individuals with biallelic pathogenic variants is expected to be 100%. 


GATM specifications:




All subpopulations in gnomAD must have a maximum allele frequency less than 0.000055 (based on the prevalence of the most common suspected pathogenic variants, c.484+1G>T and p.Arg169Ter) (see Appendix 4). Use the current version recommended by SVI; version number will be stated in classification summary.


Note – PM2 will NOT be used at moderate strength; PM2 will only be applied as a Supporting criterion.


If homozygotes are observed, the variant will meet BS2 (assuming 100% penetrance for an individual with 2 pathogenic variants in trans).
Gene: GATM (HGNC:4175)
Evidence Code: PP3 — Moderate
Modification Type: General recommendation,Strength
Missense variant with a REVEL score >0.773 (based on guidance from Pejaver et al, 2022, PMID: 36413997).
Gene: GATM (HGNC:4175)
Evidence Code: PP3 — Supporting
Modification Type: General recommendation
Missense variant with a REVEL score 0.644-0.773 (based on guidance from Pejaver et al, 2022, PMID: 36413997).


In frame deletion or insertion predicted deleterious by PROVEAN and MutationTaster. Results must be consistent to count.


For non-canonical splice site variants (e.g., +3, -3), predict the impact on splicing by using SpliceAI, 
https://spliceailookup.broadinstitute.org/
 , and apply PP3 for a score equal to or >0.2 (as indicated in PMID: 37352859, Table 1 and Figure 4). Assess the possibility of activation of cryptic splice sites.
Gene: GATM (HGNC:4175)
Evidence Code: BA1 — Stand Alone
Modification Type: Disease-specific
GrpMax >0.0005 (0.05%) in gnomAD




Any variant with a GrpMax (lower bound 95%ile) >0.0005 in gnomAD. Use the current version recommended by SVI; version number will be stated in classification summary. 


Threshold based on (max allelic contribution = 100% and max genetic contribution = 100% based on estimated prevalence of 1 in 3,450,000 (PMID 27233232), and penetrance of 100%) (PMID 30311383) (see Appendix 3)
Gene: GATM (HGNC:4175)
Evidence Code: BS1 — Strong
Modification Type: Disease-specific
GrpMax >0.0001 (0.01%) in gnomAD




Any variant with a GrpMax (lower bound 95%ile) >0.0001 in gnomAD. Use the current version recommended by SVI; version number will be stated in classification summary.


 Threshold based on max allelic contribution = 25% and max genetic contribution = 100% based on estimated prevalence of 1 in 3,450,000, (PMID 27233232), and penetrance of 100%) (PMID 30311383) (see Appendix 3).
Gene: GATM (HGNC:4175)
Evidence Code: BS3 — Supporting
Modification Type: Disease-specific,Strength
In vitro assays in which a variant is expressed in AGAT-deficient cultured cells (e.g. AGAT-deficient fibroblasts) or in-fusion High-Fidelity cloning of GATM transcript and site directed mutagenesis to generate missense variant overexpressed in HeLa cells and measurement of AGAT activity in cells for wild-type and missense variant. Any variant with enzyme activity at or above 30% of normal in DesRoches et al, 2016, PMID 27233232, meets BS3_Supporting.
Gene: GATM (HGNC:4175)
Evidence Code: BP2 — Supporting
Modification Type: None
Observed in cis with a pathogenic variant (to take AR inheritance for AGAT deficiency into account).
Gene: GATM (HGNC:4175)
Evidence Code: BP4 — Supporting
Modification Type: General recommendation,None
REVEL score <0.29 for missense variants (based on guidance from Pejaver et al, 2022, PMID: 36413997).


In frame deletion or insertion predicted benign by MutPredIndel and MutationTaster.


No predicted impact on splicing by SpliceAI, 
https://spliceailookup.broadinstitute.org/
 , based on a score <0.1 (as indicated in PMID: 37352859, Table 1 and Figure 4).
Gene: GATM (HGNC:4175)
Evidence Code: BP7 — Strong
Modification Type: General recommendation,Strength
Experimental evidence, such as RT-PCR, shows no impact on splicing. Follow the decision tree outlined in Figure 5, Walker et al, 2023, PMID: 37352859. Note that splicing may appear normal in compound heterozygous patients with one allele that is degraded by nonsense-mediated decay as the results of a frameshift and premature termination codon due to splicing defect. Therefore. caution must be used in asessing the data prior to applying this code.
Gene: GATM (HGNC:4175)
Evidence Code: BP7 — Supporting
Modification Type: None
A synonymous (silent) variant OR an intronic variant at or beyond positions +7 and -21, for which SpliceAI, 
https://spliceailookup.broadinstitute.org/
, predicts no impact on splicing (score <0.1) (see Walker et al, 2023, PMID: 37352859).