Use ATM PVS1 Decision Tree.

• PVS1: Predicted splice defect 
• PVS1_Strength(RNA): Observed splice defect
• The default RefSeq transcript for nucleotide (c.) annotation is NM_000051.3/ENST00000278616.8. All exons from this transcript can be considered constitutive exons without major alternate splice isoforms that could potentially rescue presumed LoF events (ENIGMA unpublished data).
• Of note, ATM is occasionally annotated with multiple non-coding first exons so exon numbering must be carefully reviewed for variant interpretation using literature sources of data.
• The FAT/PI3K/FATC (collectively the FATKIN) domains are considered critical for ATM protein function (PMID 28508083, 31740029, 31320732). PVS1 alterations that are predicted to escape NMD, but that adversely affect these domains can be granted PVS1 (as opposed to PVS1_Strong as the recommended base-line (PMID 30192042). 
• The HEAT repeat domain is considered important for protein function based on the appearance of many A-T affected individuals harboring a variant resulting in an in-frame, single exon loss in this domain (PMID 10980530, 19535770, 30819809, 15054841, 22927201, 19691550, 10330348, 17124347, 8845835, 16266405, 9463314, 24090759, 22213089). PVS1-eligible alterations that are predicted to escape NMD, but that adversely affect the HEAT repeat domain can be granted PVS1_Strong. They are limited to strong due to a lack of known missense pathogenic alterations in this domain.
• The most 3’/C-Terminal residue considered to be pathogenic is p.R3047 (PMIDs: 8755918, 19691550, 18560558, 10980530, 26628246)
• NOTE: Many diagrams for ATM show the FAT, PI3-K and FATC domains as separated by spacers, however these are not empirically derived and there is evidence of missense pathogenic alterations in the ‘spacer’ regions. This VCEP considers them a contiguous domain (PMID 28508083).
• PVS1 can be applied as per the PVS1 decision tree.
  • PVS1_Variable(RNA) shall be used for observed splice defects, whether from canonical +/-1,2 positions or other spliceogenic regions (including mid-exonic missense/synonymous variants that cause splice defects) with baseline weight as per the below decision tree. Weight can be further modified based on the quality of the RNA study including consideration of concepts such as:
  • Starting material (where patient material is preferable to in vitro minigene)
  • Use of NMD inhibitors where translation does occur such as cell lines⁵⁶
  • Primer design (to make sure it’s comprehensive to capture possible multicassette events)
  • Method of quantification 
    • where e.g. capillary electrophoresis is preferable to estimation by gel band density
    • where SNP analysis is most preferred (where analysis of exonic SNPs and their relative presence in aberrant and WT transcripts is informative)
  • Quantification (where complete effects should have increased weight over incomplete effects)
  • Specific guidance on the use of RNA evidence in variant assessment is not a gene-specific consideration for PALB2 at this time, therefore discretion is left to assessors until further guidance is provided for this general concept from the Sequence Variant Interpretation group.
• In the event that RNA data are available and they reflect a substantial variant-specific impact, do not use both PVS1(RNA) and PP3 or BP4. However, in the event that RNA data are available and they reflect no variant-specific impacts, PP3 or BP4 may be applied in conjunction with BP7(RNA).

• Use as ascribed for missense changes as long as a splice defect is ruled out for both variants;
• Use ATM PS1 Splicing table for splicing variants with similar predictions or observations of splice defect. (PMID: 36865205)

For protein, see detailed notes on ATM-specific assays; For RNA use code PVS1_Strength(RNA) and modulate strength based on assay quality and quantity (curator discretion).

PS4_Moderate: Do not use. Proband counting for genes causing a common disorder need to be calibrated in a population-specific way before use.

• Is not considered a conflicting piece of evidence for variants that otherwise are likely benign/benign 
• Use as PM2_Supporting (not moderate)

See ATM PM3/BP2 table for approach to assign points per proband, and final PM3 code assignment based on the sum of PM3-related points.

Ataxia Telangiectasia (A-T) is a rare, severe, early-onset disease with some exceptions denoted ‘variant’ or ‘atypical’ A-T in which cases phenotypes are more mild with slower progression. Phenotypes associated with A-T are very specific and do not generally require differential diagnosis. Therefore, publications that claim a ‘clinical diagnosis of A-T’ are taken at face value and granted a ‘confident diagnosis. Specific phenotype criteria may qualify for ‘confident or ‘consistent’ diagnosis of A-T based on the below criteria. No additional weight modifications are made for ‘atypical’ cases if they meet ‘confident or ‘consistent’ criteria as although the disease progression is different, the clinical features are the same.

Variant may not exceed general population frequency >0.01%. 

If the variant under assessment has co-occurred with at least 2 different P/LP variants, one co-occurrence must be weighed as phase unknown while the remaining can be assumed in trans

Multiple unrelated cases are additive. 

• For example, one individual with a ‘confident A-T phenotype’ is homozygous for a variant scores 2.0 points. Another individual who has a ‘consistent A-T phenotype’ and has the same variant and another phase-unknown truncating ATM variant scores 1.0 points. The total points towards PM3 are 3.0 points leading to PM3 used as its baseline moderate strength.

CONFIDENT PHENOTYPE (must include Laboratory result)

• Presence of ≥2 Laboratory results 1-4 (see notes) -OR- 
• Presence of Clinical feature 1a or 1b AND presence of Laboratory result 1 or 2 -OR-
• Presence of Clinical feature 2 or 3 AND Laboratory result 1 or 2

CONSISTENT PHENOTYPE (does not require laboratory result)

• Presence of two or more Clinical features of ataxia (1a-1e) -OR-
• Presence of one Clinical feature 1a or 1b AND either Clinical feature 2 or 3

Clinical features (Neurological and MRI findings):

1. Progressive cerebellar ataxia, manifesting as:
   • a: Progressive truncal/limb ataxia 
   • b: Cerebellar degeneration (atrophy of the frontal and posterior vermis and both hemispheres by MRI).
   • c: Oculomotor apraxia (inability to follow an object across visual fields) or abnormal ocular saccades (rapid refixation from one object to another).
   • d: Choreoathetosis or dystonia (involuntary movements; twisting and repetitive movements, abnormal postures).
   • e: Peripheral axonal neuropathy OR Anterior horn cell neuronopathy
2. Oculocutaneous telangiectasia of the conjunctivae, ears, or face.
3. Immunodeficiency (often frequent infections) and/or leukemia/lymphoma.

Laboratory Results:

1. ATM protein levels ≤ 15% of controls in patient fibroblast or lymphoblastoid cell lines. If ATM protein levels are slightly greater than 15%, the ATM kinase activity must be shown to be "negative or low or residual" (see notes).
2. Elevated serum alpha-fetoprotein (AFP) levels >65ug/L in a patient ≥ 2 years old.
3. Increased sensitivity to ionizing radiation in patient fibroblast or lymphoblastoid cell lines.
4. Presence of a 7;14 chromosomal translocation in patient peripheral blood cells (≥ 5% of cells).

Notes:

1. ATM protein levels ≤15% of control levels show >95% sensitivity and >98% specificity for diagnosing ataxia-telangiectasia (A-T). Protein levels >15% may arise due to a missense variant, a leaky splicing variant, a variant resulting in a kinase-dead protein (where protein levels may not be affected), or a diagnosis other than A-T.
2. When assigning case report criteria based solely on laboratory results (i.e., presence of TWO or more of laboratory results 1-4), there is a greater likelihood that the most specific laboratory results #1 and #2 will be available, and that there will be some clinical indication that the individual(s) has A-T.

Do not use for in-frame insertions or deletions less than a single exon; Use for stop-loss variants, only.

• Based on location of the most C-terminal known pathogenic variant, p.Arg3047*.
• Use as PM5_Supporting (not moderate)
• Do not use for start-loss variants
• Do not use for missense changes: Multiple amino acid substitutions at the same residue can be pathogenic or benign and bioinformatic tools cannot yet confidently distinguish them

• AR Condition: Affected relatives must have both variants identified in proband.
• AD Condition – Do not use: Co-segregation analysis in lower-penetrance genes can lead to false positive results (PMID 32773770)

• NOTE: Splice analysis needs to be considered for all variant types (including missense, frameshift, nonsense, etc. as any variant has the potential to impact splicing which may preclude any expected protein effects) 
• NOTE: PP3 for splice predictions may not be applied in addition to PVS1 or PVS1_Variable(RNA) codes.
• Use caution in applying the wrong type of computational evidence (protein vs. RNA) towards the cumulative body of evidence for the opposite mechanism.
• The VCEP uses SpliceAI as a sole predictor due to its ability to accurately predict loss of native splice sites and creation of cryptic sites (Jaganathan et al., 2019). This VCEP recommends SpliceAI thresholds set forth by the SVI in applying PP3 and BP4 to non-canonical splice variants: Apply PP3 for SpliceAI scores ≥0.2 and apply BP4 for SpliceAI scores ≤0.1 (Walker et al., 2023).
• In the event that RNA data are available and they reflect a substantial variant-specific impact, do not use both PVS1(RNA) and PP3 or BP4. However, in the event that RNA data are available and they reflect no variant-specific impacts, PP3 or BP4 may be applied in conjunction with BP7(RNA).