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Germinality does not necessarily define mAb expression and thermal stability.

Linda Schwaigerlehner1, Patrick Mayrhofer2, Matthias Diem3

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Applied Microbiology and Biotechnology
|July 28, 2019
PubMed
Summary

Somatic mutations in monoclonal antibody (mAb) V(D)J sequences impact expression potential. Reverting mature mAbs to their germline sequences can reveal opportunities for antibody engineering, but results vary by antibody family.

Keywords:
Difficult to expressGerminalizationMammalian expression systemRecombinant antibody productionSecretory pathway

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Area of Science:

  • Biotechnology
  • Molecular Biology
  • Immunology

Background:

  • Recombinant monoclonal antibody (mAb) production relies on cell line expression potential, influenced by antibody structure and amino acid sequence.
  • Somatic mutations within the V(D)J sequence of mature mAbs can alter their intrinsic properties.

Purpose of the Study:

  • To investigate the impact of somatic mutations on the expression potential of four model mAbs.
  • To compare the expression and stability of mature mAbs with their corresponding germline-derived variants.

Main Methods:

  • Development of recombinant Chinese hamster ovary (CHO) cell lines expressing eight mAb variants (four mature mAbs and their germline cognates).
  • Evaluation of critical parameters: productivity, intra- and extracellular product profiles, XBP1 splicing, thermal stability, and in silico hydrophobicity.

Main Results:

  • Significant differences in productivity were observed even between germline-derived mAbs.
  • Back-to-germline mutations did not consistently improve expression or stability, indicating variable effects of engineering.
  • Germinalization strategies revealed both opportunities and limitations in mAb engineering.

Conclusions:

  • Germline reversion (germinalization) can be a strategy to improve mAb properties, but its effectiveness is antibody-specific and depends on the germline family.
  • Each mAb should be individually assessed for optimization potential through germline engineering.