Variant classification summary

NM_000314.6:c.253+1G>A

PTEN · NP_000305.3:p.? · NM_000314.6

GRCh37: chr10:89690847 G>A · GRCh38: chr10:87931090 G>A

Gene: PTEN Transcript: NM_000314.6

Final call

Pathogenic

PVS1 very strong PS1 strong PS3 strong PM2 supporting

Variant details

Gene

PTEN

Transcript

NM_000314.6

Protein

NP_000305.3:p.?

gnomAD AF

ClinVar

Pathogenic

OncoKB

Classification rationale

Interpretation summary

Generated evidence synthesis

1

c.253+1G>A is a canonical +1 splice donor variant in PTEN intron 4. Per the PTEN VCEP PVS1 decision tree, canonical GT-AG splice site disruptions at or 5' to p.D375 (c.1121) qualify for PVS1 at very strong strength.

2

Functional characterization in patient-derived lymphoblastoid cells (PMID:28677221) demonstrated that c.253+1G>A causes exon 4 skipping, resulting in a frameshift (p.Ala72Thrfs*5) with predicted NMD. PTEN protein expression was significantly decreased and P-AKT was significantly increased, satisfying PS3 at strong strength per PTEN VCEP specifications.

3

A different nucleotide substitution at the same position, c.253+1G>T, has been established as pathogenic with complete cosegregation with Cowden disease in family N4 (PMID:9259288). SpliceAI predicts a donor loss delta of 0.99 for c.253+1G>A, indicating impact equal to the known pathogenic variant, satisfying PS1 at strong strength.

4

c.253+1G>A is absent from gnomAD v2.1, v4.1, and gnomAD-Canada, satisfying PM2 at supporting strength (allele frequency <0.001% per PTEN VCEP).

5

This variant has been reported in ClinVar as Pathogenic by 7 clinical laboratories (ClinVar Variation ID: 7820) and has been observed in somatic cancers (COSMIC, n=8).

6

Combined classification: PVS1 (very_strong) + PS1 (strong) + PS3 (strong) + PM2 (supporting). Under the ACMG/AMP 2015 combining rules per the PTEN VCEP framework, Rule 1 is satisfied: one very strong criterion (PVS1) plus at least one strong criterion (PS1, PS3) → Pathogenic.

Final determination: Rule1 in the Richards et.al., 2015 - Combining rules v3.2.0 criteria-combination framework is satisfied by the adjudicated criteria, so the variant is classified as Pathogenic.

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	Met	Canonical +1 splice donor variant (c.253+1G>A) in PTEN intron 4. Per the PTEN VCEP decision tree (NM_000314.8), canonical GT-AG ±1,2 splice site disruptions at or 5' to p.D375 (c.1121) are assigned PVS1. This variant is far 5' of p.D375, and exon 4 skipping with frameshift p.(Ala72Thrfs*5) and predicted NMD was confirmed by RT-PCR in patient-derived lymphoblastoid cells.	PMID:28677221 vcep_pvs1_decisiontree_pten cspec
PS1	Met	A different nucleotide change at the same position, c.253+1G>T, has been established as pathogenic in family N4 with complete cosegregation with Cowden disease. SpliceAI predicts a donor loss delta score of 0.99 for c.253+1G>A, consistent with impact equal to the known pathogenic c.253+1G>T variant, satisfying the PTEN VCEP PS1 rule for same-nucleotide-position splice variants.	PMID:9259288 PMID:28677221 spliceai cspec
PS2	Not assessed	No de novo observations with confirmed maternity and paternity were identified in the reviewed literature for c.253+1G>A.
PS3	Met	PMID:28677221 (Chen et al. 2017) characterized c.253+1G>A in patient-derived lymphoblastoid cells by RT-PCR, demonstrating exon 4 skipping resulting in frameshift p.(Ala72Thrfs*5). PTEN protein expression was significantly decreased (~50% reduction) and downstream P-AKT was significantly increased in the splice-altered group, satisfying the PTEN VCEP PS3 Strong rule for RNA or other assay demonstrating impact on splicing.	PMID:28677221 cspec
PS4	Not assessed	Multiple probands with c.253+1G>A have been reported (PMID:28677221: 1 patient; PMID:20600018: 1 patient with numerous GI polyps; ClinVar: 7 clinical laboratories reporting Pathogenic). However, PTEN VCEP PS4 requires specificity scoring based on detailed phenotype data, which is not available from the reviewed sources. Without computable specificity scores, PS4 strength cannot be assigned.	PMID:28677221 PMID:20600018 clinvar
PS5	Not assessed	No new data for this criterion beyond the assessment skip instruction.
PM1	Not met	c.253+1G>A is an intronic splice site variant in intron 4. The PTEN VCEP defines PM1-applicable catalytic motifs as residues 90-94 (WPD loop), 123-130 (P-loop), and 166-168 (TI-loop) of NP_000305.3. This variant does not lie within any of these catalytic motif residues.	cspec
PM2	Met	c.253+1G>A is absent from gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada v1.0, satisfying the PTEN VCEP PM2_Supporting threshold of allele frequency <0.00001 (0.001%).	gnomad_v2 gnomad_v4 gnomad_canada cspec
PM5	N/A	PM5 applies to missense changes at an amino acid residue where a different pathogenic missense change has been seen. c.253+1G>A is a splice site variant, not a missense change.
PM6	Not assessed	No assumed or confirmed de novo observations for c.253+1G>A were identified in the reviewed literature.
PP1	Not assessed	No co-segregation data are available for c.253+1G>A specifically. While c.253+1G>T at the same position showed complete cosegregation in family N4 (PMID:9259288), this is a different nucleotide change and cannot be directly applied.
PP2	N/A	PP2 applies to missense variants in genes with a low rate of benign missense variation. c.253+1G>A is a canonical splice site variant, not a missense change. PTEN VCEP PP2 specification is explicit for missense variants.	cspec
PP3	N/A	Per ClinGen SVI PVS1 recommendations (PMC6185798), PP3 should not be stacked with PVS1 for the same splice-effect prediction evidence. Additionally, the PTEN VCEP PP3 rule requires concordance of SpliceAI and VarSeak for splicing variants; VarSeak data are not available. SpliceAI max delta is 0.99, but this evidence is already captured by PVS1 for this canonical splice variant.	spliceai pvs1_generic_framework cspec
PP4	N/A	PTEN VCEP specifies PP4 as Not Applicable; phenotype specificity has been incorporated into the rule specifications for PS4.	cspec
PP5	N/A	PTEN VCEP specifies PP5 as Not Applicable for this VCEP, per ClinGen Sequence Variant Interpretation VCEP Review Committee recommendation.	cspec
BA1	Not met	c.253+1G>A is absent from all gnomAD populations. The PTEN VCEP BA1 threshold is filtering allele frequency >0.00056 (0.056%), which is not met.	gnomad_v2 gnomad_v4 cspec
BS1	Not met	c.253+1G>A is absent from all gnomAD populations. The PTEN VCEP BS1 threshold is filtering allele frequency from 0.000043 to 0.00056 (0.0043% to 0.056%), which is not met.	gnomad_v2 gnomad_v4 cspec
BS2	Not met	No observations of c.253+1G>A in the homozygous state in healthy or PHTS-unaffected individuals. The PTEN VCEP BS2 requires homozygous observations for application.	gnomad_v2 gnomad_v4 cspec
BS3	Not met	Functional studies (PMID:28677221) demonstrate a damaging splicing effect for c.253+1G>A (exon 4 skipping, decreased PTEN protein, increased P-AKT). BS3 requires demonstration of no damaging effect on protein function or splicing, which is contradicted by the available evidence.	PMID:28677221 cspec
BS4	Not assessed	No segregation data are available for c.253+1G>A to assess lack of segregation in affected family members. The PTEN VCEP BS4 requires lack of segregation in one or more families.
BP1	N/A	PTEN VCEP specifies BP1 as Not Applicable; this rule is not applicable to PTEN.	cspec
BP2	Not met	No observations of c.253+1G>A in trans with a pathogenic or likely pathogenic PTEN variant, nor in cis with different pathogenic/likely pathogenic PTEN variants. The PTEN VCEP BP2 requires such observations.
BP4	Not met	PTEN VCEP BP4 for splicing variants requires SpliceAI scores of 0-0.2 (evidence of benign) with VarSeak concordance. SpliceAI max delta for c.253+1G>A is 0.99, predicting a strong splicing impact, which contradicts BP4 application. Additionally, functional data (PMID:28677221) confirms splicing disruption.	spliceai PMID:28677221 cspec
BP5	Not met	No evidence was identified of c.253+1G>A occurring in a case with an alternate molecular basis for disease. The PTEN VCEP BP5 requires at least two such cases with a highly penetrant alternate gene/disorder and no phenotypic overlap with PTEN.
BP6	N/A	PTEN VCEP specifies BP6 as Not Applicable for this VCEP, per ClinGen Sequence Variant Interpretation VCEP Review Committee recommendation.	cspec
BP7	N/A	PTEN VCEP BP7 specifies that intronic variants must be positioned at or beyond +7/-21 from the splice junction. c.253+1G>A is at the +1 position, which is the canonical splice donor site, not at or beyond +7/-21. Additionally, BP7 requires splicing prediction algorithms to predict no impact, which is contradicted by SpliceAI delta 0.99.	cspec spliceai

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