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Llamanade: An open-source computational pipeline for robust nanobody humanization.

Zhe Sang1, Yufei Xiang2, Ivet Bahar3

  • 1Department of Cell Biology, Pittsburgh, PA, USA; Department Computational and Systems Biology, Pittsburgh, PA, USA; University of Pittsburgh-Carnegie Mellon University Program in Computational Biology, Pittsburgh, PA, USA.

Structure (London, England : 1993)
|December 13, 2021
PubMed
Summary

Camelid-derived nanobodies (Nbs) show therapeutic promise but require humanization. We developed Llamanade, an open-source software, to rationally humanize Nbs, enhancing their potential for clinical trials.

Keywords:
SARS-CoV-2VHH single-domain antibodiesantibody therapeuticshumanizationnanobodiesopen-source software

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Area of Science:

  • Biotechnology and Biologics Development
  • Immunology and Antibody Engineering
  • Computational Biology and Bioinformatics

Background:

  • Nanobodies (Nbs) are single-domain antibody fragments derived from camelids, offering unique therapeutic potential.
  • Despite favorable properties, Nbs often require humanization to mitigate immunogenicity and improve clinical translation.
  • Understanding Nb sequence and structural diversity is crucial for effective humanization strategies.

Purpose of the Study:

  • To systematically analyze Nb sequence and structural properties to identify key differences from human antibodies.
  • To provide insights into conserved residues critical for Nb solubility, stability, and antigen binding.
  • To develop an open-source computational tool, Llamanade, for rational Nb humanization.

Main Methods:

  • Systematic sequence and structural analysis of nanobodies.
  • Big data analysis to identify conserved residues and framework diversities.
  • Development and application of the open-source software 'Llamanade' for Nb humanization, including feature extraction, structural modeling, and optimization.
  • Validation of Llamanade using SARS-CoV-2 neutralizing nanobodies.

Main Results:

  • Identified substantial framework diversities and key differences between Nbs and human IgG antibodies.
  • Discovered conserved residues contributing to enhanced solubility, stability, and antigen binding in Nbs.
  • Developed 'Llamanade', an open-source software enabling rapid, rational humanization of Nbs from sequence input.
  • Successfully humanized a diverse set of potent SARS-CoV-2 neutralizing Nbs using Llamanade.

Conclusions:

  • Rational humanization of nanobodies is feasible and can be significantly aided by computational tools.
  • The developed software, Llamanade, provides a valuable resource for optimizing therapeutic nanobodies.
  • Llamanade facilitates the development of nanobody-based therapeutics for clinical trials by improving their translational potential.