Variant classification summary

NM_006218.3:c.2176G>A

PIK3CA · NP_006209.2:p.(Glu726Lys) · NM_006218.3

GRCh37: chr3:178938934 G>A · GRCh38: chr3:179221146 G>A

Gene: PIK3CA Transcript: NM_006218.3

Final call

Likely Pathogenic

PS2 moderate PS4 strong PM2 supporting PP5 supporting

Variant details

Gene

PIK3CA

Transcript

NM_006218.3

Protein

NP_006209.2:p.(Glu726Lys)

gnomAD AF

ClinVar

Pathogenic

OncoKB

Inconclusive

Classification rationale

Interpretation summary

Generated evidence synthesis

1

NM_006218.3:c.2176G>A (p.Glu726Lys) is a missense variant in PIK3CA exon 14 that is absent from gnomAD population databases (PM2_Supporting).

2

The variant was identified as a confirmed de novo event in one MCAP patient and as a postzygotic mosaic mutation in two additional unrelated MCAP patients, consistent with the known gain-of-function disease mechanism (PS2_Moderate).

3

Four independently reported patients with megalencephaly-capillary malformation syndrome and brain overgrowth phenotypes carry this variant, with strong phenotype specificity for the disorder (PS4_Strong).

4

This variant has been classified as Pathogenic by the ClinGen Brain Malformations Variant Curation Expert Panel after expert panel review (ClinVar Variation ID: 376476).

5

Using the Brain Malformations VCEP Tavtigian point framework (Path Supporting +1, Path Moderate +2, Path Strong +4, Path Very Strong +8), the assigned criteria yield a total of 7 points, which falls within the Likely Pathogenic range (6 to 9 points). The expert panel classification of Pathogenic likely reflects additional criteria such as PM1 or PS3 applied with access to the VCEP functional assay spreadsheet and Table 2A phenotype point values.

Final determination: Brain Malformations Specification Tavtigian point framework v1.1.0 point-based framework yields a total score of 8, which maps to Likely Pathogenic under the specified Tavtigian-style ranges.

Criteria assessment

ACMG/AMP criteria review

Criteria shown when status is available

All criteria require review: For research and educational purposes only.

Criterion	Status	Rationale	Evidence used
PVS1	N/A	Not applicable per ClinGen Brain Malformations VCEP: the disease mechanism for PIK3CA-related brain malformations is gain of function, not loss of function.	cspec
PS1	Not met	No alternate nucleotide change producing the same p.Glu726Lys amino acid substitution has been identified as a previously established pathogenic variant. c.2176G>A is the only nucleotide change reported at this residue.
PS2	Met	The variant was identified as de novo in one MCAP patient (162-001P) with confirmed maternity and paternity, satisfying PS2_Moderate per VCEP Criteria 1. Postzygotic mosaicism was demonstrated in two additional unrelated MCAP patients (LR08-261 at 12% in blood and 41% in buccal swab; LR06-333 at 14% in LCL), fulfilling the mosaic nature expected for this disorder.	PMID:22729224 cspec
PS3	Not assessed	No variant-specific functional assay data for NM_006218.3:c.2176G>A (p.Glu726Lys) meeting VCEP BMVCEP quality criteria was identified. PMID:22729224 performed PI3K pathway functional assays on lymphoblastoid lines carrying other PIK3CA mutations (p.Glu453del) but did not test cells harboring E726K. PMID:29549527 identified E726K in Patient 3 primary fibroblasts but reported detailed functional data only for Patient 1 (H1047R).	PMID:22729224 PMID:29549527
PS4	Met	The variant has been observed in four independently reported MCAP patients with megalencephaly and brain malformations, all absent from population controls (PM2 met). Multiple cases with a highly specific phenotype (megalencephaly-capillary malformation with brain overgrowth) support PS4_Strong under the Brain Malformations VCEP point-based framework.	PMID:22729224 PMID:29549527 gnomad_v2 gnomad_v4 cspec
PS5	N/A	PS5 is not a recognized ACMG/AMP criterion. This criterion does not exist in the standard framework.
PM1	Not met	Residue Glu726 lies outside both PIK3CA critical functional domains specified in VCEP Table 4 (kinase domain AA 322-483 and kinase domain AA 797-1068). While the variant resides in a statistically significant Cancer Hotspot, the VCEP restricts PM1_Supporting to Table 4 approved domain membership.	cspec
PM2	Met	The variant is absent from gnomAD v2.1, v4.1, and gnomAD-Canada, meeting the VCEP PM2_Supporting threshold (absent/rare in controls, ≤1 individual).	gnomad_v2 gnomad_v4 gnomad_canada cspec
PM5	Not met	No pathogenic missense variant at residue Glu726, other than the queried E726K, was identified in ClinVar or the reviewed literature. PM5 requires a different pathogenic missense change at the same amino acid residue.
PM6	N/A	Not applicable per ClinGen Brain Malformations VCEP: PM6 is addressed under PS2 and is not used independently.	cspec
PP1	N/A	Not applicable per ClinGen Brain Malformations VCEP: disease-causing variants in these genes are germline mosaic, de novo, or postzygotic mosaic; co-segregation is not expected.	cspec
PP2	Not assessed	The VCEP specifies PP2_Supporting for PIK3CA when the missense z-score exceeds 3.09. PIK3CA is a highly constrained gene with established intolerance to missense variation, but the specific gnomAD missense z-score was not provided in the evidence package and could not be independently verified.	cspec
PP3	N/A	Not applicable per ClinGen Brain Malformations VCEP: traditional in silico pathogenicity prediction algorithms are designed for loss-of-function mechanisms and are not validated for gain-of-function variants.	cspec
PP4	N/A	Not applicable per ClinGen Brain Malformations VCEP: phenotype specificity is accounted for under PS4.	cspec
PP5	Met	Expert panel ClinGen Brain Malformations Variant Curation Expert Panel classified as Pathogenic.	cspec clinvar
BA1	Not met	The variant is absent from gnomAD. The VCEP BA1 threshold of >0.0926% allele frequency is not reached.	gnomad_v2 gnomad_v4 cspec
BS1	Not met	The variant is absent from gnomAD. The VCEP BS1 threshold of >0.0185% allele frequency is not reached.	gnomad_v2 gnomad_v4 cspec
BS2	Not met	The variant is absent from gnomAD with zero homozygotes. No evidence of three or more well-phenotyped heterozygous healthy family members. BS2 criteria not satisfied.	gnomad_v2 gnomad_v4 cspec
BS3	Not met	No well-established in vitro or in vivo functional studies demonstrate a benign effect for this variant. All available functional evidence in the PI3K pathway indicates gain-of-function effects for PIK3CA mutations in this gene region, which is inconsistent with a benign interpretation.
BS4	N/A	Not applicable per ClinGen Brain Malformations VCEP: these are de novo, germline mosaic, or postzygotic mutations; lack of segregation is not expected.	cspec
BP1	N/A	Not applicable per ClinGen Brain Malformations VCEP: loss of function is not the disease mechanism for PIK3CA-related disorders.	cspec
BP2	Not met	No evidence was identified that this variant has been observed in cis or trans with a known pathogenic variant in PIK3CA.
BP4	N/A	Not applicable per ClinGen Brain Malformations VCEP: BP4 is restricted to synonymous, intronic (non-canonical splice), and UTR variants when two of three splicing prediction tools predict no impact. This is a missense variant and does not qualify.	cspec
BP5	Not met	No evidence was identified that any affected individual carrying this variant has an alternate molecular basis for disease.
BP6	N/A	Not applicable per ClinGen Brain Malformations VCEP: BP6 is not for use as recommended by the ClinGen Sequence Variant Interpretation VCEP Review Committee.	cspec
BP7	N/A	Not applicable: BP7 per VCEP is restricted to synonymous, intronic (non-canonical splice), and UTR variants. This is a missense variant.	cspec

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