PTEN c.514_515del, p.Arg172AlafsTer7

Reasoning

NM_000314.8:c.514_515del is a frameshift, NP_000305.3:p.(Arg172AlafsTer7), in exon 6. The variant creates a premature stop/disruption far 5' of the PTEN-specific NMD threshold at p.D375 (c.1121), and exon 6 is present in the biologically relevant NM_000314.8 transcript. The governing PTEN decision-tree path for a nonsense or frameshift predicted to undergo NMD therefore assigns full PVS1, corresponding to the required literal strength "very strong". The searched decision-tree file contains no variant-specific row for c.514_515del, p.Arg172AlafsTer7, or p.R172Afs*7; it provides the governing pre-assigned consequence path rather than a variant table.

VCEP rule

Nonsense or frameshift | Predicted to undergo NMD: stop codon or disruption at or 5' to p.D375 (c.1121) -> exon is present in biologically-relevant transcript NM_000314.8 | PVS1

Evidence considered

• pvs1_variant_assessment: NM_000314.8:c.514_515del; NP_000305.3:p.(Arg172AlafsTer7)/p.(R172Afs*7); frameshift; exon 6; canonical_splice_consensus=false.
• PVS1_DecisionTree_PTEN.pdf, page 1, row "Nonsense or frameshift | Predicted to undergo NMD": the variant is a frameshift at c.514_515del, producing a premature stop/disruption 5' of p.D375 (c.1121) in exon 6 of biologically relevant transcript NM_000314.8; assigned PVS1.
• ClinGen PTEN VCEP Version 3.2 CSPEC PVS1 entry: "Use PTEN PVS1 decision tree."

Still missing

• No variant-specific notation for c.514_515del, p.Arg172AlafsTer7, or p.R172Afs*7 was found in PVS1_DecisionTree_PTEN.pdf.txt; the file is a decision tree, not a variant table.

Sources

Reasoning

The variant is absent from gnomAD v2.1 and gnomAD v4.1 all-comers datasets. The PTEN VCEP permits PM2 at supporting strength when the allele frequency is <0.00001 (0.001%); absence is below that threshold. No VCEP instruction requires a non-cancer or exome-only subset for this criterion, so the default all-comers entries were used.

VCEP rule

Absent in population * Databases present at <0.00001 (0.001%) allele frequency in gnomAD or another large sequenced population. If multiple alleles are present within any subpopulation, allele frequency in that subpopulation must be <0.00002 (0.002%).

Evidence considered

• ClinGen PTEN VCEP Version 3.2, PM2 supporting rule: Absent in population; databases present at <0.00001 (0.001%) allele frequency in gnomAD or another large sequenced population. If multiple alleles are present within any subpopulation, allele frequency in that subpopulation must be <0.00002 (0.002%).
• gnomAD v2.1 (all-comers): variant absent; search status absent; source_data empty, corresponding to observed allele frequency 0.
• gnomAD v4.1 (all-comers): variant absent; search status absent; source_data empty, corresponding to observed allele frequency 0.

Sources

Reasoning

The VCEP defines PM1 to include PTEN catalytic-motif residues 90-94, 123-130, and 166-168 (NP_000305.3). The frameshift begins at residue 172, outside all three specified regions. The VCEP text provides no applicable hotspot route for this PTEN criterion, and the case evidence reports no statistically significant hotspot.

VCEP rule

Located in a mutational hot spot and/or critical and well-established functional domain. Defined to include residues in catalytic motifs: 90-94, 123-130, 166-168 (NP_ 000305.3)

Evidence considered

• PTEN VCEP PM1 definition: catalytic motifs at residues 90-94, 123-130, and 166-168 on NP_000305.3; the variant consequence is p.(Arg172AlafsTer7), beginning at residue 172.
• Cancer Hotspots evidence reports that this variant does not lie in a statistically significant hotspot; the PTEN VCEP PM1 rule does not name a hotspot source or provide a separate hotspot route.

Sources

Reasoning

The variant is absent from the available all-comers gnomAD v2.1 and gnomAD v4.1 records, so its observed filtering allele frequency is 0 and does not exceed the PTEN VCEP BA1 threshold of 0.00056 (0.056%).

VCEP rule

gnomAD Filtering allele frequency >0.00056 (0.056%)

Evidence considered

• ClinGen PTEN VCEP Version 3.2, BA1 rule: gnomAD Filtering allele frequency >0.00056 (0.056%).
• gnomAD v2.1 (all-comers): variant absent; search status absent; source_data empty.
• gnomAD v4.1 (all-comers): variant absent; search status absent; source_data empty.

Sources

Reasoning

The variant is absent from the available all-comers gnomAD v2.1 and gnomAD v4.1 records, giving no observed allele frequency within the PTEN VCEP BS1 strong range of 0.000043 to 0.00056 or its supporting range of 0.0000043 to 0.000043.

VCEP rule

gnomAD Filtering allele frequency from 0.000043 (0.0043%) up to 0.00056 (0.056%)

Evidence considered

• ClinGen PTEN VCEP Version 3.2, BS1 strong rule: gnomAD Filtering allele frequency from 0.000043 (0.0043%) up to 0.00056 (0.056%).
• ClinGen PTEN VCEP Version 3.2, BS1 supporting rule: Allele frequency from 0.0000043 (0.00043%) up to 0.000043 (0.0043%).
• gnomAD v2.1 (all-comers): variant absent; search status absent; source_data empty.
• gnomAD v4.1 (all-comers): variant absent; search status absent; source_data empty.

Sources

Reasoning

The available evidence does not document a disease-affected proband, confirmed maternity and paternity, absence of family history, or a proven or assumed de novo observation for NM_000314.8:c.514_515del. The PTEN VCEP PS2 rule therefore cannot be evaluated.

VCEP rule

Pathogenic Strong/Very Strong PS2 de novo rule

Evidence considered

• The case evidence contains no proband-level clinical description, family-history assessment, parental genotype results, or de novo observation for NM_000314.8:c.514_515del.
• ClinGen PTEN VCEP Version 3.2 provides PS2 paths for either two proven, four assumed, or one proven plus two assumed de novo observations, or a de novo observation with both maternity and paternity confirmed, but the required case facts are unavailable.

Still missing

• Proband diagnosis and phenotype consistent with PTEN hamartoma tumor syndrome
• Documented absence of family history
• Parental testing confirming or not confirming maternity and paternity
• Number and classification of de novo observations

Sources

Reasoning

The PTEN VCEP permits PS3 for validated functional studies, including the Mighell et al. 2018 massively parallel phosphatase assay. The governing supplementary file mmc2.xlsx is explicitly indexed for PS3 and BS3 and its Table S2 is a missense-variant assay table. Searches of the complete converted mmc2.xlsx text found no exact entry for NM_000314.8:c.514_515del, p.(Arg172AlafsTer7), or p.(R172Afs*7); the file contains nearby R172 missense substitutions and R172* but not this frameshift. Therefore no exact-variant functional result, high-confidence flag, or cumulative score is available to evaluate PS3, and results for nearby substitutions or the different nonsense variant were not substituted.

Evidence considered

• ClinGen PTEN VCEP specification v3.2, PS3: the indexed functional assay path includes Mighell et al. 2018 Table S2 and requires an exact variant match, a TRUE high-confidence flag in column I, and the Table S2 cumulative score in column G.
• Mighell et al. 2018 supplementary data mmc2.xlsx, Table S2: searches for c.514_515del, p.Arg172AlafsTer7, and p.R172Afs*7 found no exact row; nearby rows include R172A and other R172 missense substitutions plus R172* nonsense, none of which is this frameshift.
• The variant is NM_000314.8:c.514_515del, predicted p.(Arg172AlafsTer7)/p.(R172Afs*7), a frameshift; no functional result for this exact variant was identified.

Still missing

• The complete converted VCEP file /Users/gs_agent/LYFESCI_HERMES_v11/vcep_db/PTEN/fulltext/mmc2.xlsx.txt was searched for c.514_515del, p.Arg172AlafsTer7, and p.R172Afs*7; no exact variant entry was found.
• No exact-variant RNA, mini-gene, or other validated functional assay result with the controls and replication required by the PTEN VCEP was available in the consulted case evidence.

Sources

Reasoning

The PTEN VCEP permits PS4 from a case-control study with an odds ratio >2 for a PHTS component phenotype, p<0.05, and a 95% confidence-interval lower limit >1.5, or from aggregated phenotype-specificity scores. The available literature contains no verified counts for this exact variant in affected versus unaffected/control subjects, so no odds ratio or 95% confidence interval can be computed. It also contains no Cleveland Clinic or pediatric specificity scores for independent probands carrying NM_000314.8:c.514_515del. The VCEP additionally states that PS4 cannot be applied if BS1 applies; BS1 does not apply because the variant is absent from gnomAD v2.1, v4.1, and gnomAD-Canada, but the PS4 evidence requirements themselves remain unmet with the available data.

VCEP rule

**PTEN EP Commentary:** This criterion is unlikely to be used in this manner for a condition as rare as PHTS. However, if sufficiently powered, a case-control study finding an odds ratio >2 for a PHTS component phenotype with p<0.05 and 95% confidence interval with lower limit >1.5, this criteria may be applied. However, this criterion may _not_ be applied in combination with PP4.  * Use 2: Patient’s phenotype or family history is highly specific for a disease with a single genetic etiology. **PTEN EP Specifications:** This criterion may not be applied if BS1 applies.  Phenotype specificity scores are added across independent probands and calculated as follows: Adults:  * 1 point per proband with Cleveland Clinic (CC) score ≥ 30 (Tan 2011) * 0.5 points per proband with CC score of 25-29. Children: * 1 point per proband with pediatric phenotype score ≥ 5 (please see supplementary information in manuscript for scoring rubric). * 0.5 points per proband with pediatric phenotype score of 4, but autism/developmental delay/intellectual disability may not contribute to the score. PS4\_Very Strong: Probands with specificity score ≥16 (see text). PS4: Probands with specificity score of 4-15.5. PS4\_Moderate: Probands with specificity score of 2-3.5. PS4\_Supporting: Proband(s) with specificity score of 1-1.5.

Evidence considered

• The PTEN VCEP instruction states: a sufficiently powered case-control study may apply PS4 when the odds ratio is >2 for a PHTS component phenotype, p<0.05, and the 95% confidence-interval lower limit is >1.5.
• The VCEP specifies phenotype-specificity scoring as 1 point per adult proband with Cleveland Clinic score >=30, 0.5 points for scores 25-29, 1 point per child with pediatric phenotype score >=5, and 0.5 points for score 4 under the stated autism/developmental-delay restriction; no such score is reported for this exact variant.
• PMID:21194675 abstract: the study conducted a multicenter prospective study of 3042 probands to develop a clinical scoring system for PTEN mutation testing, but the available case material does not provide exact-variant carrier counts or an exact-variant specificity score.
• PMID:9467011 full-text extraction: the exact c.514_515del/p.(R172Afs*7) variant is not mentioned; the study reports broader PTEN mutation and family cohorts without variant-specific evidence for this deletion.
• PMID:23788249 full-text extraction: the paper does not mention PTEN or this variant.
• PMID:25190698 full-text extraction: the paper discusses germline PTEN mutation syndrome risk but does not mention this exact variant or provide variant-specific case-control data.
• The variant is absent from gnomAD v2.1, gnomAD v4.1, and gnomAD-Canada; therefore the VCEP's BS1 exclusion condition is not triggered, but absence alone is not the PS4 case-control or specificity evidence required here.

Still missing

• Exact-variant affected-case and unaffected/control counts, or a matched-ancestry control frequency sufficient to calculate an odds ratio and 95% confidence interval.
• Cleveland Clinic scores or pediatric phenotype scores for independent probands carrying NM_000314.8:c.514_515del.

Sources

Reasoning

The available evidence does not document an assumed de novo occurrence in a disease-affected proband, the absence of family history, or the number of proven or assumed de novo observations. The PTEN VCEP PM6 rule therefore cannot be evaluated.

VCEP rule

Pathogenic Moderate/Strong/Very Strong PM6 de novo rule

Evidence considered

• The case evidence contains no proband-level clinical description, family-history assessment, parental genotype results, or de novo observation for NM_000314.8:c.514_515del.
• ClinGen PTEN VCEP Version 3.2 defines PM6 using assumed or presumed de novo observations in patients with disease and no family history, with strength determined by the qualifying observation count; those facts are unavailable.

Still missing

• Disease status and phenotype of the proband or probands
• Documented absence of family history
• Maternity and paternity confirmation status
• Number of proven and assumed de novo observations

Sources

Reasoning

No affected family members carrying NM_000314.8:c.514_515del and no informative meioses are documented. PMID:9467011 was reviewed, but the exact variant is not mentioned and its general segregation findings cannot establish segregation for this case. The PTEN VCEP PP1 rule cannot therefore be applied.

VCEP rule

Pathogenic Supporting/Moderate/Strong PP1 co-segregation rule

Evidence considered

• No affected or unaffected relatives, variant-positive relatives, or informative meioses are reported for NM_000314.8:c.514_515del.
• Marsh et al. 1998, Results, PTEN mutation analysis in Cowden disease and Bannayan-Zonana syndrome, page 509: the study analyzed 37 Cowden disease families and seven Bannayan-Zonana families and assessed segregation when additional family members were available, but the exact c.514_515del variant is not mentioned and no case-specific meiosis count is provided.
• ClinGen PTEN VCEP Version 3.2 assigns PP1 Supporting for 3 or 4 observed meioses, Moderate for 5 or 6, and Strong for at least 7 observed across at least two families; none are documented here.

Still missing

• Affected relatives tested for NM_000314.8:c.514_515del
• Unaffected relatives tested for the variant where relevant
• Informative meioses count
• Family-level phenotype and segregation records

Sources

Reasoning

The available gnomAD results establish absence of the variant but provide no homozygous observation and no confirmation that any individual was healthy or PHTS-unaffected. Therefore the PTEN VCEP BS2 requirement cannot be evaluated.

VCEP rule

Observed in the homozygous state in a healthy or PHTS-unaffected individual. One observation if homozygous status confirmed, two if not confirmed. To be applied at supporting evidence level if BS1 is also applied.

Evidence considered

• ClinGen PTEN VCEP Version 3.2, BS2 rule: Observed in the homozygous state in a healthy or PHTS-unaffected individual. One observation if homozygous status confirmed, two if not confirmed. To be applied at supporting evidence level if BS1 is also applied.
• gnomAD v2.1 (all-comers): variant absent; no homozygote count or clinical-status observation provided.
• gnomAD v4.1 (all-comers): variant absent; no homozygote count or clinical-status observation provided.

Still missing

• A confirmed homozygous observation in a healthy or PHTS-unaffected individual, or two independent unconfirmed homozygous observations, is not available.

Sources

Reasoning

The PTEN VCEP permits BS3 for functional studies showing no damaging effect, including the Mighell et al. 2018 phosphatase assay path. The governing supplementary file mmc2.xlsx is explicitly indexed for BS3 and its Table S2 is a missense-variant assay table. Searches of the complete converted mmc2.xlsx text found no exact entry for NM_000314.8:c.514_515del, p.(Arg172AlafsTer7), or p.(R172Afs*7); the file contains nearby R172 missense substitutions and R172* but not this frameshift. Consequently, there is no exact-variant benign functional result, high-confidence flag, or cumulative score supporting BS3, and results for nearby substitutions or the different nonsense variant were not substituted.

Evidence considered

• ClinGen PTEN VCEP specification v3.2, BS3: the indexed functional assay path includes Mighell et al. 2018 Table S2 and requires an exact variant match, a TRUE high-confidence flag in column I, and a cumulative score greater than 0 for BS3 supporting.
• Mighell et al. 2018 supplementary data mmc2.xlsx, Table S2: searches for c.514_515del, p.Arg172AlafsTer7, and p.R172Afs*7 found no exact row; nearby rows include R172A and other R172 missense substitutions plus R172* nonsense, none of which is this frameshift.
• The variant is NM_000314.8:c.514_515del, predicted p.(Arg172AlafsTer7)/p.(R172Afs*7), a frameshift; no functional result demonstrating preserved function for this exact variant was identified.

Still missing

• The complete converted VCEP file /Users/gs_agent/LYFESCI_HERMES_v11/vcep_db/PTEN/fulltext/mmc2.xlsx.txt was searched for c.514_515del, p.Arg172AlafsTer7, and p.R172Afs*7; no exact variant entry was found.
• No exact-variant functional assay demonstrating no damaging effect on PTEN protein function or splicing was available in the consulted case evidence.

Sources

Reasoning

The available evidence does not identify any affected family member who lacks NM_000314.8:c.514_515del, and it provides no family-level non-segregation analysis. The PTEN VCEP BS4 rule cannot therefore be evaluated.

VCEP rule

Benign Supporting/Strong BS4 lack-of-segregation rule

Evidence considered

• The case evidence contains no affected relatives with documented negative testing for NM_000314.8:c.514_515del and no non-segregation analysis across one or more families.
• ClinGen PTEN VCEP Version 3.2 specifies BS4 Supporting for lack of segregation in affected members of one family and BS4 Strong for lack of segregation in affected members of two or more families; neither situation is documented.

Still missing

• Affected family members tested for the exact variant
• Negative genotype results in affected relatives
• Number of families showing lack of segregation

Sources

Reasoning

The PTEN VCEP permits BP2 at supporting strength when the variant is observed in trans with a pathogenic or likely pathogenic PTEN variant, or when there are at least three observations in cis and/or phase unknown with different pathogenic or likely pathogenic PTEN variants. The available case evidence documents the target variant as NM_000314.8:c.514_515del (NP_000305.3:p.(Arg172AlafsTer7)) but provides no qualifying observation of a second pathogenic or likely pathogenic PTEN variant, no phase relationship, and no three-observation cis or unknown-phase series. Therefore BP2 cannot be evaluated.

VCEP rule

Observed in trans with a pathogenic or likely pathogenic PTEN variant OR at least three observations in cis and/or phase unknown with different pathogenic/likely pathogenic PTEN variants.

Evidence considered

• The case evidence identifies NM_000314.8:c.514_515del / NP_000305.3:p.(Arg172AlafsTer7), but contains no documented pathogenic or likely pathogenic PTEN comparator variant in trans, cis, or unknown phase.
• No affected-proband observation, parental/offspring phase result, or qualifying three-observation cis/unknown-phase series is provided for BP2.

Still missing

• Documented co-occurrence with a different pathogenic or likely pathogenic PTEN variant is needed, including confirmed trans phase or at least three cis/unknown-phase observations.

Sources

Reasoning

The PTEN VCEP requires a variant to be found in a case with an alternate molecular basis, with at least two such cases, a highly penetrant alternate gene/disorder, and personal or family history demonstrating no overlap between that disorder and PTEN. The available case evidence identifies no qualifying alternate gene/disorder, two qualifying cases, or non-overlapping personal/family histories, so BP5 cannot be evaluated.

VCEP rule

**PTEN EP Specifications:** At least two such cases are required for criteria to apply.  In addition, the other gene/disorder must be considered highly penetrant AND the patient’s personal/family history must demonstrate no overlap between the other gene and _PTEN_. Variant found in a case with an alternate molecular basis for disease. Other gene/disorder must be considered highly penetrant AND patient’s personal/family history must demonstrate no overlap between other gene and PTEN.

Evidence considered

• The VCEP instruction requires at least two such cases and additionally requires that the other gene/disorder be highly penetrant and that the patient's personal/family history demonstrate no overlap between the other gene and PTEN.
• No qualifying alternate molecular basis or corresponding clinical histories are present in the available case evidence.

Still missing

• At least two cases carrying this variant with a documented alternate molecular diagnosis.
• Evidence that the alternate gene/disorder is highly penetrant and that each patient's personal or family history does not overlap PTEN-associated disease.

Sources