PTEN c.276_277insG, p.(His93AlafsTer2)

Reasoning

NM_000314.8:c.276_277insG is a frameshift producing NP_000305.3:p.(His93AlafsTer2) / p.(H93Afs*2), with the predicted truncation beginning at residue 93. The PTEN VCEP decision tree governs this consequence: for a frameshift in the biologically relevant NM_000314.8 transcript, a stop codon or disruption at or 5' to p.D375 (c.1121) that is predicted to undergo NMD receives PVS1. This variant is far 5' of c.1121 and generates a premature termination signal, so the VCEP rule supports full-strength PVS1. No downgrade rule for escape from NMD or a distal non-critical region applies.

Rule

PTEN VCEP PVS1 decision tree: frameshift with predicted NMD at or 5' to p.D375 (c.1121) in NM_000314.8

Evidence considered

• The case normalization identifies NM_000314.8:c.276_277insG as a frameshift yielding NP_000305.3:p.(His93AlafsTer2) / p.(H93Afs*2), beginning at amino-acid residue 93.
• The PTEN VCEP decision tree specifies PVS1 for nonsense or frameshift variants in biologically relevant NM_000314.8 when the stop codon or disruption is at or 5' to p.D375 (c.1121) and NMD is predicted.
• The ClinGen PTEN specification directs users to the PTEN PVS1 decision tree for PVS1 strength assignment; this variant is not a canonical splice-site variant and no VCEP downgrade rule is triggered.

Still missing

• PVS1_DecisionTree_PTEN.pdf was searched for c.276_277insG, p.His93AlafsTer2, and p.H93Afs*2; no variant-specific entry was found, so the governing decision-tree branch was applied rather than a pre-assigned row.

Sources

Reasoning

The PTEN VCEP defines PM1 at moderate strength for a variant in a mutational hotspot and/or critical functional domain, specifically residues 90-94, 123-130, and 166-168 of NP_000305.3. The frameshift begins at His93, within the authoritative flattened VCEP interval 90-94, and the case evidence also records PTEN H93 as a statistically significant hotspot. vcep_materials.json contains a domain_tables key, but it is an empty object, so no gene-specific domain-table entries supersede the VCEP interval. Human review is warranted because the hotspot extraction is flagged as partial and PM1 is being applied to a frameshift rather than a missense variant.

Rule

PTEN VCEP PM1 moderate: located in a mutational hotspot and/or critical, well-established functional domain; defined catalytic-motif residues are 90-94, 123-130, and 166-168 of NP_000305.3.

Evidence considered

• The PTEN VCEP PM1 rule defines critical catalytic-motif residues 90-94, 123-130, and 166-168 on NP_000305.3.
• The variant is NP_000305.3:p.(H93Afs*2), so the frameshift begins at residue His93 within the VCEP interval 90-94.
• The case evidence records PTEN H93 as a statistically significant hotspot, although the hotspot extraction is flagged as partial.
• The vcep_materials.json domain_tables key is present but empty; therefore there are no additional authoritative gene-specific domain-table entries to evaluate.

Still missing

• The hotspot extraction is flagged as partial and should be confirmed against the retained hotspot record or screenshot.
• The empty domain_tables object provides no additional gene-specific entries to check.

Sources

Reasoning

The ClinGen PTEN Expert Panel specifies PM2 at supporting strength when a variant is present at less than 0.00001 (0.001%) allele frequency in gnomAD or another large sequenced population; if multiple alleles occur in a subpopulation, that subpopulation frequency must be less than 0.00002 (0.002%). The variant is absent from gnomAD v2.1 and v4.1, satisfying the specified frequency requirement. The PTEN specification does not require a non-cancer or exome-only subset for PM2, so the all-comers gnomAD results are the applicable default.

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

• The PTEN VCEP specifies PM2 at supporting strength for allele frequency <0.00001 (0.001%) in gnomAD or another large sequenced population, with a <0.00002 (0.002%) subpopulation condition when multiple alleles are present.
• The variant NM_000314.8:c.276_277insG is absent from gnomAD v2.1 and gnomAD v4.1, corresponding to an observed allele frequency of zero in each queried dataset.
• The PTEN VCEP does not specify a non-cancer or exome-only source for PM2, so all-comers gnomAD results are used.

Sources

Reasoning

The ClinGen PTEN Expert Panel specification defines BA1 as a gnomAD filtering allele frequency greater than 0.00056 (0.056%). The variant is absent from gnomAD v2.1 and gnomAD v4.1, so no allele frequency reaches the BA1 threshold. The PTEN specification does not require a non-cancer or exome-only subset for BA1; therefore, the all-comers gnomAD results are the applicable default.

Rule

gnomAD Filtering allele frequency >0.00056 (0.056%)

Evidence considered

• The PTEN VCEP specifies BA1 at gnomAD filtering allele frequency >0.00056 (0.056%).
• The variant NM_000314.8:c.276_277insG is absent from gnomAD v2.1 and gnomAD v4.1.
• The PTEN VCEP does not specify a non-cancer or exome-only source for BA1, so all-comers gnomAD results are used.

Sources

Reasoning

The ClinGen PTEN Expert Panel specification defines BS1 at gnomAD filtering allele frequency from 0.000043 through 0.00056 (0.0043% through 0.056%), with BS1_Supporting from 0.0000043 through 0.000043. The variant is absent from gnomAD v2.1 and gnomAD v4.1, so neither BS1 interval is met. The PTEN specification does not require a non-cancer or exome-only subset for BS1; therefore, the all-comers gnomAD results are the applicable default.

Rule

gnomAD Filtering allele frequency from 0.000043 (0.0043%) up to 0.00056 (0.056%); BS1_Supporting from 0.0000043 (0.00043%) up to 0.000043 (0.0043%).

Evidence considered

• The PTEN VCEP specifies BS1 at filtering allele frequency 0.000043-0.00056 (0.0043%-0.056%) and BS1_Supporting at 0.0000043-0.000043.
• The variant NM_000314.8:c.276_277insG is absent from gnomAD v2.1 and gnomAD v4.1.
• The PTEN VCEP does not specify a non-cancer or exome-only source for BS1, so all-comers gnomAD results are used.

Sources

Reasoning

The ClinGen PTEN Expert Panel requires observation of the variant in the homozygous state in a healthy or PHTS-unaffected individual for BS2; one observation is sufficient when homozygosity is confirmed, and two are required when it is not confirmed. The variant is absent from gnomAD v2.1 and v4.1, with no homozygous observation or healthy-individual evidence, so BS2 is not met.

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

• The PTEN VCEP requires a homozygous observation in a healthy or PHTS-unaffected individual for BS2.
• The variant is absent from gnomAD v2.1 and gnomAD v4.1; therefore, no population homozygous observation is available.
• No separate healthy-individual homozygous observation is provided in the reviewed case evidence.

Sources

Reasoning

No proband-level parental testing result is documented in the available case evidence, so a confirmed de novo observation cannot be established. The PTEN VCEP specifies PS2 at strong strength when both maternity and paternity are confirmed in a patient with disease and no family history; stronger PS2 levels require multiple proven or assumed de novo observations.

Rule

De novo (both maternity and paternity confirmed) observation in a patient with the disease and no family history; very strong requires two proven OR four assumed OR one proven + two assumed de novo observations.

Evidence considered

• The available case materials contain no documented maternity confirmation, paternity confirmation, parental variant results, or explicit confirmed de novo observation.
• PTEN VCEP version 3.2: PS2 strong requires de novo occurrence with both maternity and paternity confirmed in a patient with disease and no family history; PS2 very strong requires two proven, four assumed, or one proven plus two assumed de novo observations.

Still missing

• Documented variant testing results for both biological parents are required.
• The proband's disease phenotype and family history must be documented to apply the PTEN PS2 rule.

Sources

Reasoning

PS3 cannot be assigned from the governing PTEN functional specification. The variant is a frameshift, NP_000305.3:p.(His93AlafsTer2), whereas the governing Mighell et al. assay resource is a missense saturation assay requiring an exact variant entry in Table S2 with High_conf=TRUE and a Cum_score threshold of <= -1.11 for PS3_Moderate. Searches of mmc2.xlsx.txt for c.276_277insG, p.His93AlafsTer2, p.H93Afs*2, and the corresponding H93 entries found no entry for this frameshift; the H93 entries present are different missense substitutions and cannot be substituted for the case variant. The two fetched functional-theme papers were also checked through their pre-computed extracts and neither mentions this variant or provides variant-specific functional evidence.

Rule

PTEN VCEP PS3: exact Mighell et al. 2018 Table S2 missense entry with High_conf=TRUE; PS3_Moderate when Cum_score <= -1.11.

Evidence considered

• The PTEN VCEP Version 3.2 specification permits PS3_Moderate for Mighell et al. 2018 Table S2 only when the exact missense variant is listed with High_conf=TRUE and Cum_score <= -1.11.
• The variant normalizes to the frameshift NP_000305.3:p.(His93AlafsTer2) / p.(H93Afs*2), not a missense substitution represented by the Mighell saturation assay.
• The governing mmc2.xlsx full-text conversion was searched for c.276_277insG, p.His93AlafsTer2, p.H93Afs*2, and H93 variants; only different H93 missense substitutions were present, so none is evidence for this frameshift.
• PMID:11237521 describes PTEN phosphatase biology but its extraction states that the specified variant is not mentioned and no variant-specific assay is reported.
• PMID:17218262 describes functional effects of other PTEN disruptions but its extraction states that the specified variant is not mentioned.

Still missing

• No exact c.276_277insG entry was found in mmc2.xlsx.txt.
• No exact p.His93AlafsTer2 entry was found in mmc2.xlsx.txt.
• No exact p.H93Afs*2 entry was found in mmc2.xlsx.txt.
• No variant-specific experimental functional study for this frameshift was identified in the reviewed case sources.

Sources

Reasoning

The PTEN VCEP permits PS4 from a sufficiently powered case-control study with odds ratio >2 for a PHTS component phenotype, p<0.05, and a 95% confidence-interval lower limit >1.5, or from accumulated phenotype-specific proband scores: 1 point for each adult Cleveland Clinic score ≥30, 0.5 for scores 25-29, 1 point for each pediatric score ≥5, or 0.5 for pediatric score 4. The framework assigns PS4 very strong at a specificity score ≥16, strong at 4-15.5, moderate at 2-3.5, and supporting at 1-1.5; it also states PS4 cannot be combined with PP4 and cannot be applied if BS1 applies. This case contains no case-control enrichment statistics, proband phenotype specificity scores, or aggregate score, so PS4 cannot be assigned.

Rule

PTEN VCEP PS4: case-control odds ratio >2 with p<0.05 and 95% confidence-interval lower limit >1.5, or phenotype specificity score supporting 1-1.5, moderate 2-3.5, strong 4-15.5, or very strong ≥16; PS4 may not be combined with PP4 or used if BS1 applies.

Evidence considered

• The PTEN Expert Panel specification states that a sufficiently powered case-control study may support PS4 when odds ratio is >2, p<0.05, and the 95% confidence-interval lower limit is >1.5.
• The PTEN Expert Panel specification assigns PS4 strength from summed phenotype-specific proband scores: supporting 1-1.5, moderate 2-3.5, strong 4-15.5, and very strong ≥16.
• No exact-variant case-control statistics or phenotype-specific proband scoring data were provided for NM_000314.8:c.276_277insG.

Still missing

• Obtain case-control odds ratio, p value, and 95% confidence interval for this exact variant or independent proband phenotype scores using the PTEN VCEP rubric.

Sources

Reasoning

No assumed or presumed de novo observation in an affected proband is documented in the available case evidence. The PTEN VCEP permits PM6 moderate for an assumed de novo observation without maternity and paternity confirmation, strong for two such probands or one highly specific phenotype, and very strong for the specified multiple-observation combinations.

Rule

Assumed de novo without confirmation of paternity and maternity in a proband with disease and no family history; strong requires two presumed de novo probands or one highly specific phenotype, and very strong requires two proven OR four assumed OR one proven + two assumed observations.

Evidence considered

• The available case materials contain no documented presumed de novo occurrence, parental testing status, or family-history statement for this variant.
• PTEN VCEP version 3.2: PM6 moderate applies to an assumed de novo observation without confirmed maternity and paternity; PM6 strong requires two presumed de novo probands or one proband with a highly specific phenotype; PM6 very strong requires two proven, four assumed, or one proven plus two assumed observations.

Still missing

• A documented affected proband with an assumed de novo variant is required.
• Parental testing status and family history are required to determine the applicable PM6 strength.
• Additional independent presumed de novo probands are required for PM6 strong or very strong.

Sources

Reasoning

No affected relatives, informative meioses, or family-level cosegregation data are documented in the available case evidence. Under the PTEN VCEP, PP1 requires 3 or 4 meioses for supporting strength, 5 or 6 for moderate, and at least 7 meioses across at least two families for strong strength.

Rule

Cosegregation with disease in multiple affected family members: supporting for 3 or 4 meioses, moderate for 5 or 6 meioses, and strong for at least 7 meioses across at least two families.

Evidence considered

• The available case materials contain no pedigree, affected-relative testing results, segregation counts, or informative meioses for this variant.
• PTEN VCEP version 3.2: PP1 supporting requires 3 or 4 meioses, moderate requires 5 or 6 meioses, and strong requires at least 7 meioses across at least two families with cosegregation in multiple affected family members.

Still missing

• Pedigree and clinical status of relatives must be documented.
• Variant and phenotype results for informative relatives are required.
• The number of informative meioses and number of families must be documented.

Sources

Reasoning

BS3 cannot be assigned because no variant-specific benign functional result was identified. The PTEN VCEP specification describes BS3_Supporting for phosphatase activity >0 in Mighell et al. 2018, but that resource is a missense assay and contains no exact entry for c.276_277insG, p.His93AlafsTer2, or p.H93Afs*2. Different H93 missense rows cannot establish benign function for this frameshift, and the reviewed papers do not report this variant.

Rule

PTEN VCEP BS3_Supporting: phosphatase activity >0 per Mighell et al. 2018, PMID: 29706350, for the exact tested variant.

Evidence considered

• The PTEN VCEP Version 3.2 specification permits BS3_Supporting for Mighell et al. 2018 phosphatase activity >0, but only an exact assay result for the variant can support BS3.
• The governing mmc2.xlsx full-text conversion was searched under all supplied variant notations; the frameshift was absent, while H93 missense substitutions were distinct variants and were not used as proxies.
• PMID:11237521 and PMID:17218262 were reviewed through their pre-computed paper extracts; neither reports the specified frameshift or a benign functional result for it.

Still missing

• No exact c.276_277insG entry was found in mmc2.xlsx.txt.
• No exact p.His93AlafsTer2 entry was found in mmc2.xlsx.txt.
• No exact p.H93Afs*2 entry was found in mmc2.xlsx.txt.
• No variant-specific assay showing preserved function or a benign effect was identified in the reviewed case sources.

Sources

Reasoning

No affected family members carrying the variant and no demonstrated lack of segregation are documented in the available case evidence. The PTEN VCEP specifies BS4 supporting for lack of segregation in one family and BS4 strong for lack of segregation in affected members of two or more families.

Rule

Lack of segregation in affected members of one family supports BS4 at supporting strength; lack of segregation in affected members of two or more families supports BS4 at strong strength.

Evidence considered

• The available case materials contain no documented affected-relative genotypes, phenotype assessments, or family-level non-segregation observations.
• PTEN VCEP version 3.2: BS4 supporting is lack of segregation in affected members of one family; BS4 strong is lack of segregation in affected members of two or more families, with caution for phenocopies and multiple pathogenic variants.

Still missing

• Documented affected relatives who carry or lack the variant are required.
• The family count and clinical assessment of affected relatives must be documented.
• Potential phenocopies or alternative genetic diagnoses should be considered when evaluating apparent non-segregation.

Sources

Reasoning

The ClinGen PTEN Expert Panel Version 3.2 specification permits BP2 at supporting strength when the variant is observed in trans with a pathogenic or likely pathogenic PTEN variant, or in at least three observations in cis and/or phase unknown with different pathogenic or likely pathogenic PTEN variants. The available case evidence contains no affected-proband observation, paired PTEN variant, parental testing, offspring testing, or documented cis/trans/unknown-phase observation for NM_000314.8:c.276_277insG; therefore BP2 cannot be assessed.

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 PTEN Expert Panel BP2 rule requires either one trans observation with a pathogenic or likely pathogenic PTEN variant or at least three cis and/or phase-unknown observations with different pathogenic or likely pathogenic PTEN variants.
• The case materials document no paired PTEN variant, affected-proband allelic observation, parental or offspring phase testing, or cis/trans observation for NM_000314.8:c.276_277insG.
• The available literature extractions concern functional or gene-level findings and do not report this variant or allelic phase observations.

Still missing

• A documented affected-proband or family observation with the paired PTEN variant and phase determination is needed; alternatively, at least three cis or phase-unknown observations with different pathogenic or likely pathogenic PTEN variants are required.

Sources

Reasoning

The PTEN VCEP requires at least two cases for BP5, an alternate molecular basis involving a highly penetrant other gene or disorder, and personal or family history demonstrating no phenotype overlap between that disorder and PTEN. The available case evidence does not document two qualifying cases, an alternate molecular diagnosis, penetrance, or the required non-overlapping history; BP5 therefore cannot be assigned.

Rule

PTEN VCEP BP5: variant found in a case with an alternate molecular basis; at least two cases are required, the other disorder must be highly penetrant, and personal/family history must demonstrate no overlap with PTEN.

Evidence considered

• The PTEN Expert Panel specification requires at least two cases for BP5.
• The PTEN Expert Panel specification additionally requires the other gene or disorder to be highly penetrant and the patient's personal or family history to show no overlap between that disorder and PTEN.
• No qualifying alternate molecular basis or required non-overlapping clinical history is documented for this variant.

Still missing

• Obtain at least two exact-variant cases documenting a highly penetrant alternate molecular diagnosis and personal or family history with no overlap with PTEN hamartoma tumor syndrome.

Sources