Overview #
The PS1 and PM5 criteria assess missense variants based on known pathogenic changes affecting the same codon in a gene.
Both criteria leverage prior knowledge from ClinVar and other curated variant databases to infer the potential significance of new missense substitutions.
These rules are particularly powerful because they connect novel observations to well-established pathogenic mechanisms at the protein level.
1. What Are PS1 and PM5? #
| Criterion | Definition | Evidence Direction |
| PS1 – Pathogenic Strong 1 | “Same amino acid change as a previously established pathogenic variant, regardless of nucleotide change.” | Strong pathogenic evidence — applies when a variant produces the same amino acid substitution as a known pathogenic variant. |
| PM5 – Pathogenic Moderate 5 | “Novel missense change at an amino acid residue where a different missense change determined to be pathogenic has been seen before.” | Moderate pathogenic evidence — applies when a different amino acid change occurs at a codon already shown to harbor pathogenic variants. |
Both criteria are limited to missense variants, as they rely on amino acid substitutions and codon-level protein impact.
2. Conceptual Examples #
PS1 Example – Identical Amino Acid Change #
| Variant | cDNA Change | Protein Change | ClinVar Classification | Applied Criterion |
| Variant A | c.500G>A | p.Arg167His | Pathogenic | Reference |
| Variant B | c.501C>T | p.Arg167His | Novel | ✅ PS1 |
Variant B causes the same amino acid change (Arg→His) as the known pathogenic Variant A, but via a different nucleotide substitution.
PM5 Example – Different Amino Acid Change, Same Codon #
| Variant | cDNA Change | Protein Change | ClinVar Classification | Applied Criterion |
| Variant A | c.500G>A | p.Arg167His | Pathogenic | Reference |
| Variant B | c.500G>C | p.Arg167Pro | Novel | ✅ PM5 |
Both variants affect codon 167. Since another amino acid substitution at this codon (Arg→His) is already known to be pathogenic, PM5 applies to the novel change (Arg→Pro).
3. SeqSMART’s Automated Assessment #
SeqSMART automates the evaluation of PS1 and PM5 through a multi-step process integrating codon-level mapping, ClinVar cross-referencing, and rigorous filtering.
Step 1: Variant Type Verification #
SeqSMART first identifies whether the input variant is a missense substitution.
Only missense variants are eligible for PS1 or PM5; other variant types (e.g., nonsense, frameshift, splice) are excluded.
Step 2: Codon and Protein-Level Mapping #
For each missense variant:
- The affected codon is identified.
- The resulting amino acid change is determined.
- SeqSMART then generates a complete list of possible single-nucleotide variants (SNVs) that could:
- Produce the same amino acid change (→ PS1 candidates).
- Produce a different amino acid change at the same codon (→ PM5 candidates).
- Produce the same amino acid change (→ PS1 candidates).
Step 3: ClinVar Cross-Referencing #
SeqSMART then cross-references ClinVar for all known variants affecting the same codon.
For each codon:
- The system searches for ClinVar entries with classifications of Pathogenic or Likely Pathogenic (P/LP).
- It identifies whether those known variants produce the same or different amino acid substitutions.
Decision logic:
| Scenario | Codon Match | Amino Acid Change | ClinVar Status | Criterion |
| Same codon | Same amino acid change | Pathogenic / LP | ✅ PS1 | |
| Same codon | Different amino acid change | Pathogenic / LP | ✅ PM5 | |
| No match | – | – | ❌ None |
Step 4: Conflict Filtering and Quality Control #
To maintain high reliability:
- ClinVar entries with conflicting interpretations are excluded from automated PS1/PM5 application.
- Variants must have a clear pathogenic or likely pathogenic classification to qualify.
- The algorithm automatically avoids using entries annotated as Uncertain Significance or Conflicting.
4. Strength Adjustment Logic #
SeqSMART follows ACMG/ClinGen recommendations for adjusting evidence strength:
| Criterion | Condition for Upgrade | Strength Level |
| PS1 | Multiple independent reports of identical amino acid change with consistent pathogenic evidence | PS1_Strong |
| PM5 | Multiple different pathogenic amino acid changes reported at the same codon | PM5_Strong |
| PM5 | Only one previously pathogenic change at the same codon | PM5 (Moderate) |
Strength upgrades occur automatically when multiple pathogenic substitutions are validated within the same codon.
5. Manual Review and Expert Flexibility #
All PS1/PM5 findings in SeqSMART are:
- Fully traceable to the corresponding ClinVar records (with variant IDs and evidence links).
- Editable by users — experts may downgrade, remove, or strengthen these criteria based on internal data or functional studies.
- Displayed transparently within the variant detail page under the “Amino Acid Evidence” section.
Users can also incorporate laboratory-confirmed family data or experimental evidence to refine or override the automated classification.
6. Important Notes for Users #
- PS1 and PM5 apply only to missense variants.
- Variants in genes with noncanonical translation models or alternative transcripts may require manual validation.
- When ClinVar evidence is inconsistent, SeqSMART flags the variant for manual review rather than automatic classification.
- All codon-level mappings are recalculated whenever ClinVar updates are imported to ensure results remain current.
PS1 and PM5 leverage existing clinical and experimental data to infer the significance of new missense variants.
SeqSMART automates their evaluation by integrating codon-level mapping, ClinVar-based pathogenic references, and dynamic evidence filtering, ensuring reliable and reproducible interpretation.
SeqSMART Codon-Level Evidence Principle:
Different variants, same story — codon context reveals the truth.