PTEN c.254-3C>G, p.?

Reasoning

The PTEN VCEP permits PM2 at supporting strength when a variant is present at <0.00001 (0.001%) allele frequency in gnomAD, with <0.00002 (0.002%) required in any subpopulation if multiple alleles are present. The variant is absent from gnomAD v2.1 and v4.1, corresponding to observed allele frequency 0 and satisfying the threshold. No VCEP instruction requires a non-cancer or exome-only source for PM2, so the default all-comers gnomAD records 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

• gnomAD v2.1 all-comers record for NM_000314.8:c.254-3C>G: absent; observed allele frequency is 0.
• gnomAD v4.1 all-comers record for NM_000314.8:c.254-3C>G: absent; observed allele frequency is 0.
• ClinGen PTEN VCEP Specification v3.2, 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 frequency must be <0.00002 (0.002%).

Sources

Reasoning

The variant is absent from gnomAD v2.1 and gnomAD v4.1, so its observed filtering allele frequency is 0 and does not exceed the PTEN VCEP BA1 threshold of 0.00056 (0.056%). The VCEP does not require a non-cancer or exome-only subset for BA1, so the default all-comers gnomAD records were used.

VCEP rule

gnomAD Filtering allele frequency >0.00056 (0.056%)

Evidence considered

• gnomAD v2.1 all-comers record for NM_000314.8:c.254-3C>G: absent; no allele-frequency observation was reported.
• gnomAD v4.1 all-comers record for NM_000314.8:c.254-3C>G: absent; no allele-frequency observation was reported.
• ClinGen PTEN VCEP Specification v3.2, BA1 rule: gnomAD filtering allele frequency must be >0.00056 (0.056%).

Sources

Reasoning

The variant is absent from gnomAD v2.1 and gnomAD v4.1, so its observed filtering allele frequency is 0 and does not fall within either PTEN VCEP BS1 interval: 0.000043 to 0.00056 for strong or 0.0000043 to 0.000043 for supporting. The VCEP does not require a non-cancer or exome-only subset for BS1, so the default all-comers gnomAD records were used.

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

• gnomAD v2.1 all-comers record for NM_000314.8:c.254-3C>G: absent; observed allele frequency is therefore 0.
• gnomAD v4.1 all-comers record for NM_000314.8:c.254-3C>G: absent; observed allele frequency is therefore 0.
• ClinGen PTEN VCEP Specification v3.2, BS1 strong rule: gnomAD filtering allele frequency from 0.000043 (0.0043%) up to 0.00056 (0.056%).
• ClinGen PTEN VCEP Specification v3.2, BS1 supporting rule: allele frequency from 0.0000043 (0.00043%) up to 0.000043 (0.0043%).

Sources

Reasoning

The PTEN VCEP requires observation of the variant in the homozygous state in a healthy or PHTS-unaffected individual. gnomAD v2.1 and v4.1 report the variant as absent, providing no homozygous observation; therefore BS2 is not met. No BS1 evidence is applied, so the BS2 supporting downgrade is not relevant.

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

• gnomAD v2.1 all-comers record for NM_000314.8:c.254-3C>G: absent, with no homozygous observation.
• gnomAD v4.1 all-comers record for NM_000314.8:c.254-3C>G: absent, with no homozygous observation.
• ClinGen PTEN VCEP Specification v3.2, BS2 rule requires observation in the homozygous state in a healthy or PHTS-unaffected individual; one observation is sufficient when homozygous status is confirmed, otherwise two are required.

Sources

Reasoning

The PTEN VCEP permits PS1 for an intronic splice-site variant when a different variant at the same nucleotide position is a pathogenic splicing variant and in silico splicing impact is equal to or greater than the known variant. The case variant is NM_000314.8:c.254-3C>G, but no pathogenic same-position splice comparator or comparator prediction is available.

Evidence considered

• VariantValidator reports NM_000314.8:c.254-3C>G with predicted protein consequence NP_000305.3:p.? and intronic exon 4i location.
• No same-position pathogenic splice comparator is present in the case evidence.

Still missing

• A previously established pathogenic splicing variant at the same nucleotide position and its comparable in-silico prediction are needed.

Sources

Reasoning

The available case evidence does not document a proband with PTEN hamartoma tumor syndrome, confirmed maternity and paternity, absence of family history, or any de novo observation for NM_000314.8:c.254-3C>G. Therefore, the PTEN VCEP PS2 rule cannot be evaluated.

VCEP rule

De novo (both maternity and paternity confirmed) observation in a patient with the disease and no family history.

Evidence considered

• No proband-level, parental-testing, family-history, or de novo evidence for NM_000314.8:c.254-3C>G is present in the reviewed case materials.

Still missing

• A proband-level disease phenotype and family-history assessment are not documented.
• Maternity and paternity confirmation are not documented.
• No de novo observation is documented.

Sources

Reasoning

The PTEN VCEP permits PS3 for well-established functional studies, including an RNA, mini-gene, or other assay showing splicing impact, and permits PS3_Moderate from Mighell et al. 2018 Table S2 only when the variant is listed as high confidence and has phosphatase activity <= -1.11. The variant is intronic, NM_000314.8:c.254-3C>G (NP_000305.3:p.?), and the governing mmc2 full text was searched for c.254-3C>G, c.254-3, p.?, and 254-3 with no matching entry. No qualifying RNA, mini-gene, or other variant-specific functional assay result is present in the available case evidence, so PS3 cannot be assigned.

VCEP rule

Well-established _in vitro_ or _in vivo_ functional studies supportive of a damaging effect on the gene or gene product. * RNA, mini-gene, or other assay shows impact on splicing ... 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

• ClinGen PTEN Expert Panel Specifications v3.2: PS3 allows RNA, mini-gene, or other assays demonstrating splicing impact; PS3_Moderate requires a high-confidence Mighell et al. 2018 Table S2 entry with cumulative phosphatase score <= -1.11.
• Mighell et al. 2018 supplementary data mmc2.xlsx, Table S2: searches for c.254-3C>G, c.254-3, p.?, and 254-3 returned no matching row; the table is keyed by protein-level missense variants.
• Case normalization identifies NM_000314.8:c.254-3C>G as intronic with predicted protein consequence NP_000305.3:p.?.

Still missing

• No variant-specific RNA, mini-gene, or other splicing assay result is available.
• No matching entry for c.254-3C>G, c.254-3, p.?, or 254-3 was found in the governing mmc2.xlsx converted full text; Table S2 is a protein-level missense assay table.

Sources

Reasoning

The governing PTEN VCEP rule permits strong PS4 when the prevalence of the variant in affected individuals is significantly increased compared with controls, and also provides phenotype-specific proband specificity-score paths. The available evidence reports no affected-case/control counts, matched-ancestry control frequency, odds ratio, confidence interval, p-value, or phenotype-specific proband score, so neither path can be evaluated. The literature packet reports total_pmids=0 and contains no packet.ai_classified_papers entries to verify for PS4.

VCEP rule

Probands with specificity score ≥16 (see text). ... Probands with specificity score 4-15.5 (see text) OR The prevalence of the variant in affected individuals is significantly increased compared with the prevalence in controls. ... Probands with specificity score of 2-3.5 (see text). ... Phenotype specific for disease with single genetic etiology. Proband(s) with specificity score of 1-1.5 (see text).

Evidence considered

• ClinGen PTEN VCEP CSPEC v3.2 PS4 rule: very strong requires probands with specificity score ≥16; strong requires probands with specificity score 4-15.5 or significantly increased prevalence in affected individuals compared with controls; moderate requires a score of 2-3.5; supporting requires a phenotype-specific single-genetic-etiology presentation or score of 1-1.5.
• gnomAD v2.1 and v4.1 both report NM_000314.8:c.254-3C>G as absent, but no control denominator or matched-ancestry frequency is provided, so an odds ratio and confidence interval cannot be calculated.
• literature_pass.json reports total_pmids=0, with no packet.ai_classified_papers entries available for PS4 verification.

Still missing

• Affected-case and unaffected/control counts or matched-ancestry case and control frequencies.
• A calculable case-control significance assessment, including sufficient counts for an odds ratio and 95% confidence interval.
• Independent proband specificity scores and their aggregate score, if using the phenotype-specific path.
• Variant-specific literature with affected and control observations, if available.

Sources

Reasoning

The PTEN VCEP limits PM1 to residues in the WPD loop 90-94, P-loop/phosphatase core 123-130, or TI-loop 166-168. This intronic variant has no assigned protein residue (NP_000305.3:p.?), so its location relative to every specified catalytic motif cannot be evaluated.

Evidence considered

• The VCEP defines the approved PTEN PM1 regions as WPD loop residues 90-94, P-loop/phosphatase core residues 123-130, and TI-loop residues 166-168.
• VariantValidator reports the intronic variant's predicted protein consequence as NP_000305.3:p.?; vcep_materials.json has no PTEN domain_tables entry.

Still missing

• A validated protein consequence or residue assignment is needed to compare the variant with all VCEP-defined catalytic motifs.

Sources

Reasoning

The available case evidence does not document an assumed de novo occurrence in a proband with disease and no family history, nor any confirmed or unconfirmed parental testing. Therefore, the PTEN VCEP PM6 rule cannot be evaluated.

VCEP rule

Assumed _de novo,_ but without confirmation of paternity and maternity, in proband with the disease and no family history.

Evidence considered

• No affected proband, family-history assessment, assumed de novo observation, or maternity/paternity testing status for NM_000314.8:c.254-3C>G is present in the reviewed case materials.

Still missing

• An affected proband and absence of family history are not documented.
• No assumed de novo observation is documented.
• Confirmation status for maternity and paternity is not documented.

Sources

Reasoning

The available case evidence contains no affected family members carrying NM_000314.8:c.254-3C>G, no segregation analysis, and no observed meioses. The PTEN VCEP PP1 thresholds of 3-4, 5-6, or at least 7 meioses therefore cannot be evaluated.

VCEP rule

Co-segregation with disease in multiple affected family members, with ≥7 meioses observed across at least two families. ... Co-segregation with disease in multiple affected family members, with 5 or 6 meioses observed. ... Co-segregation with disease in multiple affected family members, with 3 or 4 meioses observed.

Evidence considered

• No affected-family carrier results, segregation observations, or informative meioses for NM_000314.8:c.254-3C>G are present in the reviewed case materials.

Still missing

• Affected family members with variant testing results are not documented.
• The number of informative meioses is not documented.
• Segregation across one or more families is not documented.

Sources

Reasoning

The VCEP applies PP3 to splicing variants when there is concordance of SpliceAI and VarSeak. This intronic variant is within the splicing-variant scope, and SpliceAI is available with a maximum delta score of 0.962, but no VarSeak result is present; therefore the required concordance cannot be evaluated and PP3 is not_assessed.

VCEP rule

Multiple lines of computational evidence support a deleterious effect on the gene or gene product. ... * Splicing variants: Concordance of SpliceAl and VarSeak

Evidence considered

• ClinGen PTEN VCEP PP3 rule: for splicing variants, supporting evidence requires concordance of SpliceAI and VarSeak.
• SpliceAI Lookup for NM_000314.8:c.254-3C>G reports DS_AG 0.903, DS_AL 0.962, DS_DG 0.0, DS_DL 0.002, and max delta score 0.962.
• No VarSeak prediction is available in the case evidence.

Still missing

• VarSeak prediction for NM_000314.8:c.254-3C>G is unavailable.

Sources

Reasoning

The PTEN VCEP requires an RNA, mini-gene, or other splicing assay demonstrating no splicing impact for BS3 in intronic or synonymous variants. It also permits BS3_Supporting from Mighell et al. 2018 Table S2 only for a high-confidence variant row with phosphatase activity > 0. The variant is intronic, NM_000314.8:c.254-3C>G (NP_000305.3:p.?), and searches of the governing mmc2 full text for c.254-3C>G, c.254-3, p.?, and 254-3 found no matching entry. No qualifying benign functional assay result is available, so BS3 cannot be assigned.

VCEP rule

Well-established _in vitro_ or _in vivo_ functional studies shows no damaging effect on protein function. To be applied to intronic or synonymous variants, RNA, mini-gene or other splicing assay demonstrating no splicing impact. ... _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

• ClinGen PTEN Expert Panel Specifications v3.2: BS3 requires an RNA, mini-gene, or other assay demonstrating no splicing impact for intronic or synonymous variants; BS3_Supporting requires a high-confidence Mighell et al. 2018 Table S2 result with phosphatase activity > 0.
• Mighell et al. 2018 supplementary data mmc2.xlsx, Table S2: searches for c.254-3C>G, c.254-3, p.?, and 254-3 returned no matching row; the table is keyed by protein-level missense variants.
• Case normalization identifies NM_000314.8:c.254-3C>G as intronic with predicted protein consequence NP_000305.3:p.?.

Still missing

• No variant-specific RNA, mini-gene, or other assay demonstrating no splicing impact is available.
• No matching entry for c.254-3C>G, c.254-3, p.?, or 254-3 was found in the governing mmc2.xlsx converted full text; therefore no phosphatase score can be evaluated.

Sources

Reasoning

The available case evidence does not document affected family members who lack NM_000314.8:c.254-3C>G, or any family-level non-segregation observation. Consequently, neither the one-family supporting nor the two-or-more-family strong BS4 rule can be evaluated.

VCEP rule

Lack of segregation in affected members of two or more families. ... Lack of segregation in affected members of one family.

Evidence considered

• No affected-family non-segregation or variant-negative affected relative is documented for NM_000314.8:c.254-3C>G in the reviewed case materials.

Still missing

• Affected family members tested for the variant are not documented.
• No non-segregation observation in one family is documented.
• No non-segregation observations across two or more families are documented.

Sources

Reasoning

The PTEN VCEP BP2 rule requires either observation 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 case evidence contains no affected-proband observation or documented cis, trans, or phase-unknown observation meeting either path, so 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

• ClinGen PTEN Expert Panel Specifications to the ACMG/AMP Variant Interpretation Guidelines for PTEN Version 3.2, BP2 rule: the criterion requires observation 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 case evidence contains no documented affected-proband, cis, trans, or phase-unknown observation involving a pathogenic or likely pathogenic PTEN variant.

Still missing

• Documented phase and paired pathogenic or likely pathogenic PTEN variant observations are unavailable.

Sources

Reasoning

The VCEP applies BP4 to splicing variants when there is concordance of SpliceAI and VarSeak. This intronic variant is within the splicing-variant scope, and SpliceAI is available with a maximum delta score of 0.962, but no VarSeak result is present; therefore the required concordance cannot be evaluated and BP4 is not_assessed.

VCEP rule

Multiple lines of computational evidence suggest no impact on gene or gene product. ... * Splicing variants: Concordance of SpliceAl and VarSeak

Evidence considered

• ClinGen PTEN VCEP BP4 rule: for splicing variants, supporting evidence requires concordance of SpliceAI and VarSeak.
• SpliceAI Lookup for NM_000314.8:c.254-3C>G reports DS_AG 0.903, DS_AL 0.962, DS_DG 0.0, DS_DL 0.002, and max delta score 0.962.
• No VarSeak prediction is available in the case evidence.

Still missing

• VarSeak prediction for NM_000314.8:c.254-3C>G is unavailable.

Sources

Reasoning

The governing PTEN VCEP BP5 rule requires a variant found in a case with an alternate molecular basis, a highly penetrant other gene/disorder, and personal or family history demonstrating no overlap between that disorder and PTEN. The available case evidence documents no alternate molecular diagnosis, highly penetrant alternate disorder, or qualifying personal/family-history comparison, so BP5 cannot be evaluated.

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 CSPEC v3.2 BP5 rule requires an alternate molecular basis, a highly penetrant other gene/disorder, and no overlap between the other disorder and PTEN in the patient's personal or family history.
• The available case evidence contains no documented alternate molecular basis, penetrance evidence, or relevant personal/family-history comparison.

Still missing

• A case with the variant and a documented alternate molecular basis for disease.
• Evidence that the alternate gene/disorder is highly penetrant.
• Personal and family histories demonstrating no overlap between the alternate disorder and PTEN-related disease.

Sources