Variant: NM_000152.4(GAA):c.-32-3C>A

CA16041878

371622 (ClinVar)

Gene: GAA

Condition: glycogen storage disease II

Inheritance Mode: Autosomal recessive inheritance

UUID: d7050ce4-1aa4-4158-9721-09823c6c79f9

HGVS expressions

NM_000152.4:c.-32-3C>A
NM_000152.4(GAA):c.-32-3C>A
NC_000017.11:g.80104552C>A
CM000679.2:g.80104552C>A
NC_000017.10:g.78078351C>A
CM000679.1:g.78078351C>A
NC_000017.9:g.75692946C>A
NG_009822.1:g.7997C>A
ENST00000302262.8:c.-32-3C>A
ENST00000302262.7:c.-32-3C>A
ENST00000390015.7:c.-32-3C>A
ENST00000570803.5:c.-32-3C>A
ENST00000577106.5:c.-32-3C>A
NM_000152.3:c.-32-3C>A
NM_001079803.1:c.-32-3C>A
NM_001079804.1:c.-32-3C>A
NM_001079803.2:c.-32-3C>A
NM_001079804.2:c.-32-3C>A
NM_000152.5:c.-32-3C>A
NM_001079803.3:c.-32-3C>A
NM_001079804.3:c.-32-3C>A
NM_000152.5(GAA):c.-32-3C>A

Pathogenic

• Met criteria codes: PM3 PP4_Moderate PM2_Supporting PS1_Supporting PVS1_Strong

• Not Met criteria codes: PP3

Expert Panel

Lysosomal Diseases VCEP

Criteria Specification Information

Criteria Specification: ClinGen Lysosomal Storage Disorders Variant Curation Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines Version 2

Evidence submitted by expert panel

Lysosomal Diseases VCEP

The NM_000152.5:c.-32-3C>A variant is located in the splice acceptor region of intron 1 and has been shown to impact splicing. In fibroblasts from an individual with Pompe disease who was compound heterozygous for the variant and a missense variant in GAA, RT-PCR revealed loss of exon 2 (p.M1_T182del, which includes the 27 residues of signal sequence, the 42 residues of the propeptide and the first 113 amino acids of the mature protein (from 70 to 182) (PMID: 18429042). A later study, in which the variant was expressed in a minigene, showed that the variant results in about 20% of transcripts with normal splicing of exon 2 (of note, this is a slightly lower level of normal splicing than observed for a well known pathogenic variant in the same splice region, c.-32-13T>G). Abnormal splicing events included complete loss of exon 2 (r.-32_546del) and activation of a cryptic splice site (r.‐32_486del) (PMID: 31301153). Due to loss of a critical region (the signal sequence) in aberrantly spliced transcripts, but with presence of some normal transcript (albeit at a lower level than a known pathogenic variant in the same splice region), PVS1 was applied at the strong level (PMID: 37352859) (PVS1_Strong, PS1_Supporting applied based on recommendations in PMID: 37352859). This variant has been reported in at least 6 probands with late onset Pompe disease including two individuals with documented GAA activity <30% normal in fibroblasts (PMID: 19046416, 21550241) (PP4_Moderate). Two of these probands have been reported to have this variant in compound heterozygosity, phase unknown, with another variant in GAA that has been classified as pathogenic by the ClinGen LD VCEP including c.1655T>C (p.Leu552Pro) (PMID: 18429042) and c.482_483del (PMID: 21550241). In addition, three affected siblings have been reported to be homozygous for the variant (PMID: 19588081, 20464284, 25681614) (PM3). Another three patients are compound heterozygous for the variant and a missense variant (either c.1905C>A (p.Asn635Lys), c.1447G>A (p.Gly483Arg), or c.2173C>T (p.Arg725Trp)) (PMID: 19588081). The allelic data from these patients will be used in the classification of the second variant and is not included here to avoid circular logic. The highest population minor allele frequency in gnomAD v4.0. is 0.000007314 (8/1093866 alleles) in the European non-Finnish populationn, which is lower than the ClinGen Lysosomal Diseases VCEP’s threshold for PM2_Supporting (<0.001), meeting this criterion (PM2_Supporting). There is a ClinVar entry for this variant (Variation ID: 371662). In summary, this variant meets the criteria to be classified as pathogenic for Pompe disease. GAA-specific ACMG/AMP criteria applied, as specified by the ClinGen LSD VCEP (Specifications Version 2.0): PVS1_Strong, PM3, PS1_Supporting, PP4_Moderate, PM2_Supporting. (Classification approved by the ClinGen Lysosomal Diseases Variant Curation Expert Panel on November 21, 2023).

Met criteria codes

• PM3: Two patients with Pompe disease have been reported to have this variant in compound heterozygosity, phase unknown, with another variant in GAA that has been classified as pathogenic by the ClinGen LD VCEP; the second variant is either c.1655T>C (p.Leu552Pro) (PMID: 18429042) (0.5 points), or c.482_483del (PMID: 21550241) (0.5 points). In addition, three affected siblings have been reported to be homozygous for the variant (PMID: 19588081, 20464284, 25681614) (0.5 points). Another three patients are compound heterozygous for the variant and a missense variant (either c.1905C>A (p.Asn635Lys), c.1447G>A (p.Gly483Arg), or c.2173C>T (p.Arg725Trp)) (PMID: 19588081). The allelic data from these patients will be used in the classification of the second variant and is not included here to avoid circular logic. Total 1.5 points (PM3).
• PP4_Moderate: This variant has been reported in at least 6 probands with later-onset symptoms of Pompe disease including two individuals with documented GAA activity <30% normal in fibroblasts (PMID: 19046416, 21550241) (PP4_Moderate).
• PM2_Supporting: The highest population minor allele frequency in gnomAD v4.0. is 0.000007314 (8/1093866 alleles) in the European non-Finnish populationn, which is lower than the ClinGen Lysosomal Diseases VCEP’s threshold for PM2_Supporting (<0.001), meeting this criterion (PM2_Supporting).
• PS1_Supporting: Other variants in the same splice region have been reported, including c.-32-13T>G, which has a similar impact on splicing and is the most common variant identified in patients with late-onset Pompe disease (PS1_Supporting applied based on recommendations in PMID: 37352859; pathogenic variant at a different nucleotide within the same splice region; reduced PS1 shown in Table 3 by one strength level).
• PVS1_Strong: The NM_000152.5:c.-32-3C>A variant is located in the splice acceptor region of intron 1 and has been shown, experimentally, to impact splicing. In fibroblasts from an individual with Pompe disease who was compound heterozygous for the variant and a missense variant in GAA, RT-PCT showed loss of exon 2 (pM1_T182, which includes the 27 residues of signal sequence, the 42 residues of the propeptide and the first 113 amino acids of the mature protein (from 70 to 182) (PMID: 18429042). A later study, in which the variant was expressed in a minigene, showed that the variant results in about 20% of transcripts with normal splicing of exon 2 (of note, this is a slightly lower level of normal splicing than observed for the well known pathogenic variant, c.-32-13T>G), with abnormal splicing event including complete loss of exon 2 (r.-32_546del) and activation of a cryptic splice site (r.‐32_486del) (PMID: 31301153). Due to loss of a critical region (the signal sequence) in aberrantly splice transcripts, but with presence of some normal transcript (albeit at a lower level than a known pathogenic variant in the same splice region), PVS1 was applied at the strong level (PMID: 37352859) (PVS1_Strong).

Not Met criteria codes

• PP3: The computational splicing predictor SpliceAI gives a score of 0.19 for acceptor loss, therefore does not provide a clear prediction on the impact of this variant on splicing. Because PVS1 will be applied, this data will not be included (based on the recommendations in PMID: 37352859). Note SpliceAI score for c.-32-13T>G (pathogenic variant) is 0.06 for acceptor loss.

Approved on: 2023-11-21

Published on: 2023-12-07