PTEN c.388C>T, p.Arg130Ter

Reasoning

NM_000314.8:c.388C>T is a nonsense variant producing NP_000305.3:p.(Arg130Ter)/p.(R130*) in exon 5. The governing PTEN decision tree assigns full PVS1 to nonsense or frameshift variants predicted to undergo NMD when the stop codon is at or 5' to p.D375 (c.1121) and the exon is present in biologically relevant transcript NM_000314.8. The stop at codon 130 is 5' to p.D375, exon 5 is present in NM_000314.8, and no named downgrade rule applies; therefore PVS1 is met at very strong strength.

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

• Variant assessment: NM_000314.8:c.388C>T, NP_000305.3:p.(Arg130Ter)/p.(R130*), nonsense consequence, exon 5.
• PVS1_DecisionTree_PTEN.pdf, page 1: the nonsense/frameshift path with a stop codon at or 5' to p.D375 (c.1121), an exon present in biologically relevant transcript NM_000314.8, and predicted NMD assigns PVS1.
• Mighell et al. supplementary Table S2 in mmc2.xlsx, exact row R130* / Arg130Ter / nonsense for the searched variant notation; this row is consequence corroboration and is not used for a functional criterion in this group.

Sources

Reasoning

The governing PTEN VCEP PS3 rule permits PS3_Moderate for a high-confidence Mighell et al. 2018 phosphatase assay result with cumulative score ≤ -1.11. Table S2 of mmc2.xlsx lists the exact protein-level variant R130* / Arg130Ter, corresponding to NM_000314.8:c.388C>T, with Cum_score -3.488113982 and High_conf True (Pass SE Filter). This is well below the VCEP threshold and directly supports a damaging functional effect; PS3_Moderate is met.

VCEP rule

Well-established _in vitro_ or _in vivo_ functional studies supportive of a damaging effect on the gene or gene product. * Phosphatase activity ≤ -1.11 per Mighell et al. 2018, PMID: 29706350.

Evidence considered

• Mighell et al. 2018 supplementary mmc2.xlsx, Table S2, row R130*: Arg130Ter; Type=nonsense; Cum_score=-3.488113982; Cum_SE=0.394380671; High_conf=True; High_conf_nature=Pass SE Filter.
• The variant was also located in mmc2.xlsx Table S4 as p.Arg130Ter with c.388C>T and annotation stop gained; the governing Table S2 functional row is the R130* entry above.
• The case normalization identifies NM_000314.8:c.388C>T as NP_000305.3:p.(Arg130Ter) / p.(R130*).

Sources

Reasoning

The variant is p.(Arg130Ter), so its affected residue is 130. The PTEN VCEP defines the P-loop/phosphatase core catalytic motif as residues 123-130 of NP_000305.3; residue 130 is within that complete VCEP-defined critical functional region. This satisfies PM1 at Moderate strength through the critical functional domain route.

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

• ClinGen PTEN VCEP Version 3.2 PM1 specification: catalytic motifs include the WPD loop 90-94, P-loop/phosphatase core 123-130, and TI-loop 166-168 (NP_000305.3).
• Case normalization identifies the affected PTEN residue as Arg130 in NP_000305.3:p.(Arg130Ter); residue 130 is included in the VCEP P-loop range 123-130.

Sources

Reasoning

The ClinGen PTEN VCEP applies PM2 at Supporting when the variant is present at allele frequency below 0.00001 in gnomAD, with a subpopulation threshold below 0.00002 if multiple alleles are present in that subpopulation. gnomAD v4.1 reports total AF 2.4787140431544115e-06, below 0.00001. The subpopulation with the highest AF, Ashkenazi Jewish, has one observed allele (1/29600; 3.3783783783783784e-05), so the multiple-allele subpopulation condition is not triggered; the NFE subpopulation has two alleles at 1.6951594721273404e-06, below 0.00002. PM2 Supporting is met. gnomAD v2.1 total AF is 1.1933933742799859e-05 and does not independently meet the VCEP threshold, but the newer gnomAD v4.1 record does.

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 CSPEC PM2 rule: databases present at <0.00001 (0.001%) allele frequency in gnomAD or another large sequenced population; if multiple alleles are present within any subpopulation, that subpopulation's allele frequency must be <0.00002 (0.002%).
• gnomAD v4.1 all-comers record: total AF 2.4787140431544115e-06 (4/1613740 alleles), total homozygotes 0; NFE AF 1.6951594721273404e-06 (2/1179830 alleles); Ashkenazi Jewish AF 3.3783783783783784e-05 (1/29600 alleles).
• gnomAD v2.1 all-comers exome record: total AF 1.1933933742799859e-05 (3/251384 alleles), total homozygotes 0; this exceeds the PM2 total-AF threshold but does not negate the qualifying gnomAD v4.1 result.

Sources

Reasoning

The ClinGen PTEN VCEP requires a gnomAD filtering allele frequency above 0.00056 for BA1. gnomAD v4.1 reports filtering allele frequency (grpmax FAF) of 2.8e-07, far below 0.00056; therefore BA1 is not met. The gnomAD v2.1 total allele frequency is also only 1.1933933742799859e-05 and is not a filtering allele frequency above the VCEP cutoff.

VCEP rule

gnomAD Filtering allele frequency >0.00056 (0.056%)

Evidence considered

• ClinGen PTEN VCEP CSPEC BA1 rule: gnomAD Filtering allele frequency >0.00056 (0.056%).
• gnomAD v4.1 all-comers record for NM_000314.8:c.388C>T: total AF 2.4787140431544115e-06; grpmax FAF 2.8000000000000007e-07; homozygotes 0.
• gnomAD v2.1 all-comers exome record: total AF 1.1933933742799859e-05 (3/251384 alleles); homozygotes 0.

Sources

Reasoning

The ClinGen PTEN VCEP requires gnomAD filtering allele frequency from 0.000043 to 0.00056 for BS1 Strong, or allele frequency from 0.0000043 to 0.000043 for BS1 Supporting. The available gnomAD v4.1 filtering allele frequency is 2.8e-07, below even the supporting threshold. gnomAD v2.1 reports total allele frequency 1.1933933742799859e-05, which would fall in the supporting interval if it were a filtering allele frequency, but the case record does not provide a v2.1 filtering allele frequency; that total AF cannot substitute for the VCEP's specified metric.

VCEP rule

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

Evidence considered

• ClinGen PTEN VCEP CSPEC BS1 Strong rule: gnomAD Filtering allele frequency from 0.000043 (0.0043%) up to 0.00056 (0.056%).
• ClinGen PTEN VCEP CSPEC BS1 Supporting rule: allele frequency from 0.0000043 (0.00043%) up to 0.000043 (0.0043%).
• gnomAD v4.1 all-comers record: grpmax FAF 2.8000000000000007e-07, below 0.0000043; total AF 2.4787140431544115e-06; homozygotes 0.
• gnomAD v2.1 all-comers exome record: total AF 1.1933933742799859e-05 (3/251384 alleles), but no filtering allele frequency is reported.

Still missing

• A gnomAD v2.1 filtering allele frequency is not provided; its total allele frequency cannot be substituted for the VCEP-specified filtering allele frequency.

Sources

Reasoning

The ClinGen PTEN VCEP requires observation in the homozygous state in a healthy or PHTS-unaffected individual for BS2. gnomAD v2.1 reports zero homozygotes and gnomAD v4.1 reports zero homozygotes; no qualifying healthy or PHTS-unaffected homozygous individual is available, so BS2 is not met.

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. ... Two homozygous observations with no clinical data provided, or meets criteria for BS2 but BS1 is also applied.

Evidence considered

• ClinGen PTEN VCEP CSPEC BS2 rule: observed in the homozygous state in a healthy or PHTS-unaffected individual; one observation if homozygous status confirmed, two if not confirmed; supporting level if BS1 is also applied.
• gnomAD v2.1 all-comers exome record: homozygotes = 0.
• gnomAD v4.1 all-comers record: homozygotes = 0.

Still missing

• No qualifying homozygous healthy or PHTS-unaffected individual is reported.

Sources

Reasoning

The governing PTEN VCEP BS3 supporting rule requires a Mighell et al. 2018 phosphatase activity score >0. Table S2 of mmc2.xlsx lists the exact R130* / Arg130Ter variant corresponding to NM_000314.8:c.388C>T with High_conf True but Cum_score -3.488113982, which is not >0 and instead indicates damaging functional activity. BS3 is therefore not met.

VCEP rule

_In vitro_ or _in vivo_ functional study or studies showing no damaging effect on protein function. * Phosphatase activity >0 per Mighell et al. 2018, PMID: 29706350.

Evidence considered

• Mighell et al. 2018 supplementary mmc2.xlsx, Table S2, row R130*: Arg130Ter; Type=nonsense; Cum_score=-3.488113982; High_conf=True; High_conf_nature=Pass SE Filter.
• The negative high-confidence cumulative score does not satisfy the VCEP BS3 supporting requirement of phosphatase activity >0.

Sources

Reasoning

The variant is NP_000305.3:p.(Arg130Ter), but the available evidence does not document a previously established pathogenic PTEN variant producing the same amino-acid/stop change through a different nucleotide change, nor a qualifying pathogenic splice-site comparator at the same nucleotide position. The VCEP PS1 rule therefore cannot be evaluated.

VCEP rule

Same amino acid change as a previously established pathogenic variant regardless of nucleotide change OR different variant at same nucleotide position as a pathogenic splicing variant, where in silico models predict impact equal to or greater than the known pathogenic variant.

Evidence considered

• Case normalization: NM_000314.8:c.388C>T produces NP_000305.3:p.(Arg130Ter) / p.(R130*).
• Available evidence reports the specified R130X variant itself, but does not provide the distinct comparator required by PS1.

Still missing

• A previously established pathogenic PTEN variant with the same amino-acid/stop change but a different nucleotide change, or a qualifying pathogenic splice-site comparator at c.388, is not documented.

Reasoning

PMID:11918710 reports that the affected patient in family 11 carried heterozygous PTEN R130X (equivalent to NM_000314.8:c.388C>T, p.(Arg130Ter)) and that the variant was not detected in either parent. However, the report does not document confirmation of maternity and paternity, which is required by the PTEN VCEP PS2 rule for a Strong call; the higher Very Strong pathways also require multiple proven or assumed de novo observations, and only one relevant observation is documented. The available report also does not explicitly establish no family history for the proband. Therefore PS2 cannot be assigned from the available evidence.

Evidence considered

• Bussaglia et al. 2002, PMID:11918710, Results and Figure 5: family 11's affected patient carried the heterozygous nonsense variant R130X (CGA>TGA at nucleotide 388), and the variant was not detected in either parent; the study used constitutional blood-leukocyte DNA and direct sequencing.
• The parent-negative result supports an assumed de novo observation, but the report does not state that maternity and paternity were confirmed and does not explicitly state that the proband had no family history.

Still missing

• Documented confirmation of both maternity and paternity is unavailable.
• An explicit statement that the affected proband had no family history is unavailable.
• Only one relevant de novo observation is documented; the VCEP Very Strong pathways require additional observations.

Sources

Reasoning

The PTEN VCEP permits PS4 from a sufficiently powered case-control study with odds ratio >2, p<0.05, and 95% confidence-interval lower limit >1.5, or from aggregated phenotype-specificity scores. The reviewed variant-specific literature confirms R130X in affected PTEN/PHTS patients, including a Cowden syndrome family and a pediatric PHTS patient with eosinophilic gastrointestinal disease, but does not provide a valid exact-variant case-control comparison or the VCEP-required Cleveland Clinic or pediatric phenotype scores. The reported odds ratio 272 in Henderson et al. is for eosinophilic gastrointestinal disease enrichment among PHTS patients, not for prevalence of c.388C>T in affected individuals versus controls, so it cannot establish PS4. No PS4 conclusion is made.

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. ... **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. ... 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

• ClinGen PTEN VCEP PS4 rule: the VCEP permits a sufficiently powered case-control study with odds ratio >2, p<0.05, and 95% confidence-interval lower limit >1.5, or phenotype-specificity scoring across independent probands.
• Bussaglia et al. 2002, PMID:11918710, Results and Table III: R130X was identified in one affected Cowden syndrome family; the study tested 50 control patients to exclude common polymorphisms but did not report an exact R130X control count or a valid variant-specific case-control estimate.
• Henderson et al. 2014, PMID:24345843, Abstract and Results: the study reported OR 272 (95% CI 89-831, P<0.0001) for eosinophilic gastrointestinal disease enrichment in PHTS, not for c.388C>T prevalence in affected versus control individuals.
• The reviewed R130X reports provide patient phenotypes but do not report the PTEN VCEP Cleveland Clinic score or pediatric phenotype score needed for the alternate PS4 route.

Still missing

• Exact-variant case and matched-control counts with a valid odds ratio, p value, and 95% confidence interval are unavailable.
• Cleveland Clinic score or pediatric phenotype score for the independent R130X probands is unavailable.

Sources

Reasoning

PMID:11918710 reports one affected family-11 patient with R130X, equivalent to NM_000314.8:c.388C>T, and states that the mutation was not detected in either parent. This is compatible with an assumed de novo observation, but the PTEN VCEP PM6 rule requires an affected proband with no family history, and the report does not explicitly document no family history. It also does not provide the additional observations required for the Very Strong or Strong PM6 pathways. PM6 is therefore not assessed.

Evidence considered

• Bussaglia et al. 2002, PMID:11918710, Results and Figure 5: family 11's affected patient carried R130X and the mutation was not detected in either parent.
• The report documents one affected patient and parent-negative testing, but does not explicitly state that there was no family history.

Still missing

• An explicit statement that the affected proband had no family history is unavailable.
• Additional proven or assumed de novo observations required for stronger PM6 levels are unavailable.

Sources

Reasoning

The available variant-specific evidence documents one de novo family-11 patient with R130X and does not document multiple affected relatives carrying this same variant across three or more meioses. The statement in PMID:11918710 that mutations segregated with disease in affected members of eight kindreds refers to the different mutations identified in those kindreds, not to repeated segregation of R130X. The PTEN VCEP PP1 rule therefore cannot be evaluated for this variant.

Evidence considered

• Bussaglia et al. 2002, PMID:11918710, Table III and Results: R130X was identified in family 11, while the paper's broader segregation statement concerns PTEN mutations across eight kindreds and does not establish segregation of R130X across multiple affected relatives or three meioses.
• The separate variant-specific family report indexed in the case materials was not available as a source-registry record with an auditable full-text extract, so it was not used to assign PP1.

Still missing

• Variant-specific testing results for multiple affected relatives are unavailable.
• A documented count of at least three meioses for this variant is unavailable.

Sources

Reasoning

No affected individual documented to lack NM_000314.8:c.388C>T is reported. PMID:11918710 states that unaffected individuals did not carry germline mutations, but this is not lack of segregation in affected members and does not establish BS4. The PTEN VCEP BS4 rule therefore cannot be evaluated from the available evidence.

Evidence considered

• Bussaglia et al. 2002, PMID:11918710, Results: the study states that none of the unaffected individuals presented germline mutations, but it does not report an affected individual who lacked the familial R130X variant.
• The available evidence therefore does not demonstrate lack of segregation in affected members of one or more families.

Still missing

• A documented affected family member who tested negative for c.388C>T is unavailable.
• Evidence from one or more families showing lack of segregation is unavailable.

Sources

Reasoning

The PTEN VCEP requires either observation of the tested variant in trans with a pathogenic or likely pathogenic PTEN variant, or at least three observations in cis and/or phase unknown with different pathogenic or likely pathogenic PTEN variants. The available exact-variant report in PMID:11918710 describes heterozygous germline R130X and a de novo observation in family 11, but does not document a second PTEN variant in trans or three qualifying cis/phase-unknown observations. The BP2 rule therefore cannot be evaluated from the available evidence.

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

• ClinGen PTEN Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines for PTEN Version 3.2, CSPEC BP2 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."
• PTEN mutations in eight Spanish families and one Brazilian family with Cowden syndrome, PMID:11918710: the paper reports heterozygous R130X and states that, in family 11, the affected patient had a de novo mutation not detected in either parent; the extracted report does not document a second PTEN variant or qualifying phase observation.

Still missing

• Phasing and paired-variant data are unavailable for the reported c.388C>T observations, including whether another pathogenic or likely pathogenic PTEN variant is present in trans, cis, or unknown phase.

Sources

Reasoning

The PTEN VCEP BP5 rule requires the variant to be found in a case with an alternate molecular basis for disease, with at least two such cases, a highly penetrant alternate gene/disorder, and personal or family history showing no overlap between that disorder and PTEN. The available evidence reports PTEN-associated phenotypes and does not document two independent cases with a pathogenic alternate molecular basis meeting all of these requirements. BP5 cannot be evaluated to met or not_met from the available evidence.

VCEP rule

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

• ClinGen PTEN VCEP BP5 instruction requires at least two cases and additionally requires a highly penetrant other gene/disorder plus a personal/family history demonstrating no overlap with PTEN.
• The reviewed case literature documents PTEN/PHTS patients but no qualifying alternate-gene diagnosis meeting the complete BP5 requirements.

Still missing

• Two qualifying cases with an alternate pathogenic molecular basis are not documented.
• Evidence that the alternate disorder is highly penetrant and clinically non-overlapping with PTEN is not documented.

Sources