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First Pass Optimize...

To automatically scan for and design mutations to resolve severe developability liabilities in a single step, select one or more entries in the Project View, open the Edit menu, and select First Pass Optimize....

An interactive modal allows you to configure severe score cutoffs for AbLang, AbLang2, IgBert, and CVV (defaulting to system severe thresholds) and optionally toggle whether language model recommendations are allowed within CDR regions.

When executed, First Pass Optimize evaluates the target sequences and builds a mutation design set named "First Pass Optimize" in the Engineering workspace using the following rules:

  • Primary Residue Focus & Flanking Evaluation: As a first pass, First Pass Optimize focuses on direct primary residue substitutions (such as \(N \rightarrow Q\) for deamidation or \(D \rightarrow E\) for isomerization). Users should evaluate flanking residues manually in the Engineering workspace when exploring alternative fix pathways or conserving key paratope contacts. Always cross-reference proposed flanking modifications with the Satlawa motif tables in Liabilities to ensure candidate flanking substitutions do not introduce secondary liabilities (e.g. mutating flanking residues in deamidation motifs to Alanine or Serine can generate medium or high deamidation motifs like \(NA\) or \(NS\)).

  • Structural PTM Gating: If a 3D structural model or PDB structure is available for the antibody, relative solvent accessibility (rASA) and sidechain SASA are calculated for all residues. Chemical PTM liabilities (deamidation, isomerization, fragmentation, hydrolysis, oxidation, N-glycosylation) that are buried in the structural core (\(\text{rASA} < 15.0\%\) and \(\text{scASA} < 5.0\text{ \AA}^2\)) are automatically gated out from optimization since they are shielded from solvent-mediated degradation. Structural fold liabilities (such as disrupted CDR3 salt bridges, unusual/missing cysteines, and co-evolutionary/LLM stability outliers) are always retained.

  • Unusual Cysteine Pairing: Mutates non-canonical cysteines to Serine (S). When two or more unusual cysteines are present in the construct, they are paired together (Together, "all or none") so that variant combinations either remove all unusual cysteines together or retain the parental configuration, preventing unpaired free thiols or mismatched disulfides.

  • DMS Library Auto-Pruning: If Deep Mutational Scanning (DMS) / library mutagenesis data is present on the antibody or inherited from its ancestor lineage, proposed mutations are automatically cross-referenced against the library's permissibility matrix. Unsafe mutations (\(\text{safe} = \text{false}\)) are automatically switched to alternative safe substitutions that also resolve the liability (e.g. \(N \rightarrow S/A/T\) for deamidation); if no safe replacement exists at that position, the mutation is pruned from the design set.

  • Disrupted CDR3 Salt Bridge: Recommends Arginine (R) at heavy chain IMGT position 106 (if parent is not R/K) and Aspartate (D) at IMGT position 116 (if parent is not D), pairing both mutations together (Together).

  • Deamidations (N): Mutated to Glutamine (Q) or to a severe model recommendation if available at that position.

  • N-linked Glycosylation (N): Mutated to Glutamine (Q) or to a severe model recommendation (e.g. AbLang recommending S) if available at that position.

  • Isomerizations or Fragmentations (D): Mutated to Glutamic Acid (E).

  • Language Models / CVV: Selects the highest probability or frequency alternative residue based on configured cutoffs (default AbLang \(\ge 5.0\), AbLang2 \(\ge 2.0\), IgBert \(\ge 2.5\), CVV \(\ge 3.0\)). When severe model recommendations are present at a site, the top recommended residue is prioritized.

  • Exclusion Areas & CDR Retention: Preserves the parent residue at structural Honegger exclusion positions, Vernier positions, and CDR positions (setting includeParent: true so the parental residue is kept as an option in the design set).

  • CDR Options: Excludes language model (AbLang/AbLang2/IgBert/CVV) liabilities that fall within CDR regions by default, unless individually enabled in the modal. Region classification uses sequence-specific AntPack region annotations.

  • Target Sequence & Chain Ordering: Reads sequences directly from target antibody entries to evaluate both chains on paired Fvs. Mutated positions are presented with Light chain rows ('L') ordered before Heavy chain rows ('H').