PRPF8 c.3081A>T, p.(Ser1027=)

Reasoning

PM2 is met at supporting strength. The variant is rare in population databases: gnomAD v4.1 overall AF is 0.0000774782 (0.00775%), with maximum subgroup AF 0.000102588 (0.01026%), and gnomAD v2.1 overall AF is 0.0000249093 (0.00249%). These values are below the generic PM2 threshold of <0.1%, and no homozygotes are observed.

Rule

Generic non-VCEP PM2 threshold: population AF <0.1%; absence or very low frequency in population databases supports PM2 at supporting strength.

Evidence considered

• gnomAD v4.1 reports 125/1,613,358 alleles, overall AF 7.74782e-05 (0.00775%), maximum subgroup AF 0.000102588 (0.01026%), and 0 homozygotes.
• gnomAD v2.1 reports 7/281,020 alleles, overall AF 2.49093e-05 (0.00249%), maximum subgroup AF 5.47499e-05 (0.00547%), and 0 homozygotes.
• The variant is absent from gnomAD-Canada v1.0.
• Using the generic non-VCEP population threshold in this case, PM2 requires AF <0.1%; both gnomAD datasets and the maximum observed subgroup frequencies satisfy this rarity threshold.

Sources

Reasoning

NM_006445.3:c.3081A>T (NP_006436.3:p.(Ser1027=)) is a synonymous variant. No PRPF8 VCEP/CSPEC is available (cspec.found = false), so the generic ACMG/AMP in-silico calibration governs: for synonymous variants, SpliceAI is the operative predictor, with max delta < 0.1 supporting a benign splice-impact call. SpliceAI predicts no significant splice impact for this variant (max delta score = 0.005; DS_AG 0.005, DS_AL 0.001, DS_DG 0.0, DS_DL 0.001), well below the 0.1 threshold, consistent with no disruption of splice donor/acceptor sites or creation of a new splice site. BP7 supporting is applied on this basis.

Rule

generic_acmg_pp3_bp4_insilico_calibration

Evidence considered

• SpliceAI predicts no significant splice impact for this variant (max delta score = 0.005; DS_AG 0.005, DS_AL 0.001, DS_DG 0.0, DS_DL 0.001), below the BP4/BP7-supporting threshold of <0.1 in this lab's generic PP3/BP4 in-silico calibration (used here for BP7 since the variant is synonymous).
• Variant is synonymous (p.(Ser1027=)), the variant class this criterion is intended to cover; canonical_splice_consensus flag is false, consistent with SpliceAI's no-impact prediction.

Still missing

• Nucleotide-level conservation (e.g., phyloP/GERP) data for this position was not present in the case evidence bundle, so the classic ACMG BP7 'nucleotide not highly conserved' sub-condition could not be independently verified; the assessment relies on the SpliceAI no-impact prediction as the primary driver per this lab's generic in-silico calibration rule.

Sources

Reasoning

NM_006445.3:c.3081A>T is synonymous (p.(Ser1027=)); no VCEP/CSPEC is available for PRPF8 (cspec.found = false), so the generic ACMG/AMP in-silico calibration rule governs. Per that rule, synonymous/intronic/non-canonical-splice-position variants are assessed with SpliceAI only, never REVEL. SpliceAI predicts no significant splice impact (max delta score = 0.005; DS_AG 0.005, DS_AL 0.001, DS_DG 0.0, DS_DL 0.001), well below the PP3 threshold of >0.2. REVEL/BayesDel are not applicable to this synonymous variant's mechanism and are not used per the same-variant-type rule. No pathogenic computational evidence supports PP3.

Rule

generic_acmg_pp3_bp4_insilico_calibration

Evidence considered

• SpliceAI max delta score = 0.005 (DS_AG 0.005, DS_AL 0.001, DS_DG 0.0, DS_DL 0.001), far below the PP3 threshold of >0.2 defined in this lab's generic PP3/BP4 in-silico calibration (no PRPF8 VCEP CSPEC is available).
• Variant is synonymous (NP_006436.3:p.(Ser1027=)); per the generic calibration rule, only the SpliceAI mechanism is evaluated for non-missense variants, and REVEL/BayesDel are not applicable here.

Sources

Reasoning

PP5 is not met. ClinVar variation 891301 has conflicting single-submitter classifications (uncertain significance and likely benign), criteria provided, with zero expert-panel submissions. The exact-variant expert-panel pathogenic/likely pathogenic prerequisite is absent. Ordinary laboratory submissions, aggregate labels, and cited publications do not trigger PP5.

Rule

PP5 may be applied only when an exact-variant ClinVar expert-panel classification is Pathogenic or Likely pathogenic; non-expert laboratory submissions and aggregate ClinVar classifications are excluded.

Evidence considered

• ClinVar reports the exact matched variant as having one uncertain-significance clinical-laboratory submission and one likely-benign clinical-laboratory submission, with review status criteria provided/single submitter and zero expert-panel submissions.
• The literature candidates flagged for PP5 do not establish PP5: PMID:22234150 is a BEST1 gene card and PMID:20301590 is a nonsyndromic retinitis pigmentosa overview; neither is an exact-variant ClinVar expert-panel assertion for PRPF8 c.3081A>T.

Still missing

• No exact-variant ClinVar expert-panel Pathogenic or Likely pathogenic assertion is available.

Sources

Reasoning

BA1 is not met. The highest observed population frequency is 0.000102588 (0.01026%) in the European (non-Finnish) subgroup of gnomAD v4.1, with overall AF 0.0000774782 (0.00775%); these frequencies are below the generic BA1 threshold of >1%.

Rule

Generic non-VCEP BA1 threshold: population AF >1%.

Evidence considered

• gnomAD v4.1 reports 125/1,613,358 alleles, AF 7.74782e-05, and maximum subgroup AF 0.000102588 in European (non-Finnish) individuals.
• gnomAD v2.1 reports 7/281,020 alleles, AF 2.49093e-05, and maximum subgroup AF 5.47499e-05 in European (non-Finnish) individuals.
• Using the generic non-VCEP population thresholds in this case, BA1 requires AF >1%; the observed frequencies are far below that threshold.

Sources

Reasoning

BS1 is not met. The highest observed population frequency is 0.01026% in gnomAD v4.1, below the generic BS1 threshold of >0.3%.

Rule

Generic non-VCEP BS1 threshold: population AF >0.3%.

Evidence considered

• gnomAD v4.1 reports overall AF 0.00775% and the highest subgroup AF 0.01026% in European (non-Finnish) individuals.
• gnomAD v2.1 reports overall AF 0.00249% and the highest subgroup AF 0.00547% in European (non-Finnish) individuals.
• Using the generic non-VCEP population thresholds in this case, BS1 requires AF >0.3%; the observed frequencies are below that threshold.

Sources

Reasoning

BP6 is not met. Although an exact-variant ClinVar likely-benign submission exists, it is a single clinical-laboratory submission rather than an expert-panel classification. The exact-variant ClinVar expert-panel Benign/Likely benign prerequisite is absent; aggregate labels and ordinary laboratory assertions are excluded.

Rule

BP6 may be applied only when an exact-variant ClinVar expert-panel classification is Benign or Likely benign; non-expert laboratory submissions and aggregate ClinVar classifications are excluded.

Evidence considered

• ClinVar shows the exact variant has a likely-benign single-submitter clinical-laboratory assertion and no expert-panel submissions; the overall record is conflicting rather than an expert-panel Benign/Likely benign classification.

Still missing

• No exact-variant ClinVar expert-panel Benign or Likely benign assertion is available.

Sources

Reasoning

No case-level evidence documents the variant in an affected proband together with confirmed absence in both biological parents. The bundle contains variant normalization and population/ClinVar records but no parental testing, de novo confirmation, maternity/paternity information, or mosaicism assessment.

Rule

Generic ACMG/AMP PS2: de novo occurrence in a patient with the disease and no family history, with maternity/paternity confirmed; stronger use requires confirmed parental absence and appropriate phenotype/relationship evidence.

Evidence considered

• The consolidated case-evidence bundle identifies the variant as NM_006445.3:c.3081A>T in PRPF8, but provides no proband-level or parental segregation/de novo observations.
• The bundle reports no applicable PRPF8 CSPEC/VCEP framework, so generic ACMG/AMP criteria were used only as the fallback framework; generic PS2 evidence requirements are not satisfied by the available records.

Still missing

• Affected proband phenotype and diagnostic context
• Variant detection in the proband
• Tested biological parents and parental genotypes
• Confirmation that the variant is absent in both parents
• Maternity/paternity confirmation and assessment for parental mosaicism

Reasoning

No PRPF8-specific ClinGen VCEP specification was found for this case (cspec.found=false; framework_mode=generic_acmg), so generic ACMG/AMP applies. PS3 requires a well-established functional study (e.g., minigene splicing assay, RT-PCR on patient RNA, or validated in vitro/in vivo functional assay) demonstrating a damaging effect on protein function or splicing for this specific variant. No such wet-lab functional assay data for NM_006445.3:c.3081A>T (p.(Ser1027=)) was found anywhere in the case bundle. The only splicing-relevant datum available is a computational SpliceAI prediction (max delta score = 0.005), which is an in silico prediction, not a validated functional assay, and therefore cannot support PS3. The retrieved literature (PMID:26666451, a CRISPR/Cas9 rat rhodopsin correction study; PMID:22234150, a BEST1 clinical utility gene card) is unrelated to PRPF8 functional characterization of this variant and does not mention this variant.

Still missing

• No RNA-based (RT-PCR/minigene) or protein functional assay data specific to NM_006445.3:c.3081A>T was located in this case bundle.
• SpliceAI is a computational predictor and cannot be used to satisfy PS3, which requires empirical functional assay evidence.

Reasoning

No qualifying case-control enrichment or reliable affected-case series for the exact PRPF8 variant NM_006445.3:c.3081A>T was identified. The literature candidates in the bundle concern different genes/variants and cannot establish PS4.

Rule

Generic ACMG/AMP PS4: prevalence of the variant in affected individuals is significantly increased compared with controls; no qualifying exact-variant enrichment evidence is available here.

Evidence considered

• The supplied case summary identifies the variant as NM_006445.3:c.3081A>T in PRPF8, synonymous p.(Ser1027=), with no case-level phenotype or case-control dataset.
• The reviewed candidate publications were not evidence for this criterion: PMID:22234150 concerns BEST1-related dystrophies, and PMID:26666451 concerns a RHO S334ter rat model; neither reports the exact PRPF8 variant.

Still missing

• An exact-variant case series or case-control study with affected individuals and appropriate controls is not available in the supplied evidence.

Reasoning

No applicable PRPF8 VCEP or gene-specific framework was retrieved, so the generic ACMG/AMP framework applies. The case-evidence bundle provides no affected-proband observations, second pathogenic/likely pathogenic allele, phase information, cis/trans determination, parental testing, or inheritance data for NM_006445.3:c.3081A>T (p.Ser1027=). Therefore PM3 cannot be assessed.

Rule

Generic ACMG/AMP PM3: evidence that the variant is observed in trans with a pathogenic variant in a recessive disorder; phase and affected-proband context must be established.

Evidence considered

• The consolidated case-evidence bundle identifies PRPF8 NM_006445.3:c.3081A>T (p.Ser1027=) but contains no proband-level allelic observations or phase/inheritance data relevant to PM3.
• The bundle states that no official CSPEC/VCEP or gene-specific framework was found; generic ACMG/AMP fallback is therefore applicable, but its PM3 evidence requirements are not met by the available case data.

Still missing

• Affected-proband evidence involving this variant
• A pathogenic or likely pathogenic variant in the same gene in the relevant proband(s)
• Validated phase information demonstrating the variants are in trans
• Parental or segregation data and inheritance mode

Reasoning

No case report or clinical record in the available bundle documents a presumed de novo occurrence of NM_006445.3:c.3081A>T without parental testing. The literature packet does not provide a relevant exact-variant de novo observation.

Rule

Generic ACMG/AMP PM6: presumed de novo occurrence in a patient with the disease, without confirmation of maternity/paternity and/or without parental molecular testing; the evidence must still document the affected case and a credible de novo assessment.

Evidence considered

• The consolidated case-evidence bundle contains no clinical case narrative, pedigree, parental genotype result, or presumed-de-novo statement for this variant.
• The listed literature records are not documented as reports of this exact PRPF8 variant in a de novo case, so no publication was used to support PM6.

Still missing

• A specific affected proband carrying the variant
• Clinical evidence supporting the relevant phenotype
• Evidence that the variant was presumed de novo
• Parental testing or an explicit reason testing was unavailable

Reasoning

No affected or unaffected relatives, pedigrees, variant genotypes, phenotype status, or informative meioses are reported for this variant. Therefore, no cosegregation evidence can be counted.

Rule

Generic ACMG/AMP PP1: cosegregation with disease in multiple affected family members in a gene definitively known to cause the disease; strength depends on the number and informativeness of observed meioses and phenotype consistency.

Evidence considered

• The consolidated case-evidence bundle provides no family segregation data for NM_006445.3:c.3081A>T.
• Population frequency and ClinVar submission records do not establish cosegregation and were not treated as PP1 evidence.

Still missing

• Family pedigree and relationship structure
• Genotypes of informative relatives
• Affected/unaffected status and phenotype consistency of relatives
• Number of informative meioses
• Independent segregation observations

Reasoning

The bundle does not provide a phenotype description for an individual carrying the exact variant, nor a highly specific PRPF8 disease phenotype that can be matched to this variant. PP4 therefore cannot be assessed.

Rule

Generic ACMG/AMP PP4: the patient's phenotype or family history is highly specific for a disease with a single-gene etiology; no patient phenotype is supplied.

Evidence considered

• The supplied evidence identifies the variant and its synonymous consequence but contains no patient phenotype, diagnostic findings, or phenotype specificity assessment for a carrier of NM_006445.3:c.3081A>T.

Still missing

• Individual-level clinical phenotype and a sufficiently specific phenotype-to-gene correlation for this exact variant are absent.

Reasoning

BS2 cannot be assessed from the available evidence. gnomAD v2.1 and v4.1 report zero homozygotes, but there is no documented observation of the variant in healthy adults with an established age-appropriate phenotype, and no disease-specific penetrance, inheritance, or prevalence framework is available for PRPF8 in this case.

Rule

Generic BS2 requires observation of the variant in healthy adults for a condition with expected age-appropriate penetrance; absence of homozygotes alone is insufficient.

Evidence considered

• gnomAD v4.1 reports 0 homozygotes for the variant across 1,613,358 alleles.
• gnomAD v2.1 reports 0 homozygotes for the variant across 281,020 alleles.
• Zero observed homozygotes is not itself evidence that the variant occurs in healthy adults; the case bundle does not provide the clinical phenotype, age, penetrance, or inheritance information required for BS2.

Still missing

• No phenotype-confirmed healthy-adult carriers or age-appropriate unaffected individuals are documented.
• No applicable PRPF8 disease penetrance, inheritance, or prevalence specification is available to determine whether population carriers would support BS2.

Sources

Reasoning

No PRPF8-specific ClinGen VCEP specification was found for this case (cspec.found=false; framework_mode=generic_acmg), so generic ACMG/AMP applies. BS3 requires a well-established functional study showing no damaging effect on protein function or splicing for this specific variant. No wet-lab functional assay (e.g., RNA splicing assay, protein activity assay) addressing NM_006445.3:c.3081A>T (p.(Ser1027=)) was found in the case bundle. The available SpliceAI prediction (max delta score = 0.005, suggesting no significant computational splice impact) is an in silico prediction only and does not constitute a validated functional study; it cannot be used to satisfy BS3. The retrieved literature (PMID:26666451, PMID:22234150) does not report functional characterization of this variant.

Still missing

• No RNA-based or protein functional assay data specific to NM_006445.3:c.3081A>T was located in this case bundle.
• SpliceAI in silico prediction alone is insufficient for BS3, which requires empirical functional assay evidence.

Reasoning

No informative affected relatives known to carry the variant, no unaffected relatives tested for the variant, and no family phenotype/genotype data are available. Absence of segregation therefore cannot be evaluated.

Rule

Generic ACMG/AMP BS4: lack of segregation in affected members of a family; requires informative affected relatives and reliable genotype/phenotype data, with consideration of reduced penetrance, phenocopies, and misclassification of phenotype.

Evidence considered

• The consolidated case-evidence bundle contains no family-based non-segregation observations for NM_006445.3:c.3081A>T.
• The presence of the variant in population databases and conflicting single-submitter ClinVar classifications does not demonstrate lack of cosegregation with disease.

Still missing

• A pedigree with informative relatives
• Reliable phenotype classification for relatives
• Variant testing results in affected and unaffected relatives
• Assessment of whether affected relatives lack the variant

Reasoning

No applicable PRPF8 VCEP or gene-specific framework was retrieved, so the generic ACMG/AMP framework applies. The case-evidence bundle provides no affected or unaffected individual-level observations, no second variant with documented phase, and no evidence that NM_006445.3:c.3081A>T (p.Ser1027=) is in trans with a pathogenic variant or in cis with a pathogenic variant in a relevant disease context. Therefore BP2 cannot be assessed.

Rule

Generic ACMG/AMP BP2: the variant is observed in trans with a pathogenic variant for a fully penetrant dominant disorder or in cis with a pathogenic variant for any disorder; the qualifying phase and disease context must be documented.

Evidence considered

• The consolidated case-evidence bundle contains no phase-resolved allelic data for NM_006445.3:c.3081A>T (p.Ser1027=) and no qualifying cis/trans observation for BP2.
• The available literature packet concerns unrelated genes or general background and does not document this PRPF8 variant in an allelic observation relevant to BP2; no publication was used for this criterion.

Still missing

• Individual-level observations documenting the relevant second variant
• Phase information establishing cis or trans configuration
• Pathogenicity classification of the other allele(s)
• Phenotype and inheritance context needed to interpret the allelic observation

Reasoning

No individual-level molecular finding or established alternate molecular diagnosis is supplied to show that the patient's phenotype is better explained by another pathogenic variant. Population frequency and the conflicting ClinVar submissions do not establish BP5.

Rule

Generic ACMG/AMP BP5: a pathogenic variant is found in an alternative gene that explains the patient's phenotype; no such case-level evidence is supplied.

Evidence considered

• The case bundle contains no affected-person phenotype paired with an alternate pathogenic molecular diagnosis that would explain the phenotype better than PRPF8 c.3081A>T.

Still missing

• A documented alternate pathogenic variant or molecular diagnosis in an affected individual is not available.