In Vitro Maturation of a Humanized Shark VNAR Domain to Improve Its Biophysical Properties to Facilitate Clinical Development

John Steven, Mischa R Müller, Miguel F Carvalho, Obinna C Ubah, Marina Kovaleva, Gerard Donohoe, Thomas Baddeley, Dawn Cornock, Kenneth Saunders, Andrew J Porter, Caroline Jane Barelle

Research output: Contribution to journalArticle

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Abstract

Molecular engineering to increase the percentage identity to common human immunoglobulin sequences of non-human therapeutic antibodies and scaffolds has become standard practice. This strategy is often used to reduce undesirable immunogenic responses, accelerating the clinical development of candidate domains. The first humanized shark variable domain (VNAR) was reported by Kovalenko and colleagues and used the anti-human serum albumin (HSA) domain, clone E06, as a model to construct a number of humanized versions including huE06v1.10. This study extends this work by using huE06v1.10 as a template to isolate domains with improved biophysical properties and reduced antigenicity. Random mutagenesis was conducted on huE06v1.10 followed by refinement of clones through an off-rate ranking-based selection on target antigen. Many of these next-generation binders retained high affinity for target, together with good species cross-reactivity. Lead domains were assessed for any tendency to dimerize, tolerance to N- and C-terminal fusions, affinity, stability, and relative antigenicity in human dendritic cell assays. Functionality of candidate clones was verified in vivo through the extension of serum half-life in a typical drug format. From these analyses the domain, BA11, exhibited negligible antigenicity, high stability and high affinity for mouse, rat, and HSA. When these attributes were combined with demonstrable functionality in a rat model of PK, the BA11 clone was established as our clinical candidate.

Original languageEnglish
Article number1361
JournalFrontiers in Immunology
Volume8
DOIs
Publication statusPublished - 23 Oct 2017

Fingerprint

Sharks
Clone Cells
Serum Albumin
Mutagenesis
Dendritic Cells
Half-Life
Immunoglobulins
Antigens
In Vitro Techniques
Antibodies
Serum
Pharmaceutical Preparations
Therapeutics

Keywords

  • Journal Article
  • VNAR
  • soloMER
  • single chain binding domain
  • Shark
  • humanization
  • half-life extension
  • pharmacokinetics

Cite this

Steven, J., Müller, M. R., Carvalho, M. F., Ubah, O. C., Kovaleva, M., Donohoe, G., ... Barelle, C. J. (2017). In Vitro Maturation of a Humanized Shark VNAR Domain to Improve Its Biophysical Properties to Facilitate Clinical Development. Frontiers in Immunology, 8, [1361]. https://doi.org/10.3389/fimmu.2017.01361

In Vitro Maturation of a Humanized Shark VNAR Domain to Improve Its Biophysical Properties to Facilitate Clinical Development. / Steven, John; Müller, Mischa R; Carvalho, Miguel F; Ubah, Obinna C; Kovaleva, Marina; Donohoe, Gerard; Baddeley, Thomas; Cornock, Dawn; Saunders, Kenneth; Porter, Andrew J; Barelle, Caroline Jane.

In: Frontiers in Immunology, Vol. 8, 1361, 23.10.2017.

Research output: Contribution to journalArticle

Steven, John ; Müller, Mischa R ; Carvalho, Miguel F ; Ubah, Obinna C ; Kovaleva, Marina ; Donohoe, Gerard ; Baddeley, Thomas ; Cornock, Dawn ; Saunders, Kenneth ; Porter, Andrew J ; Barelle, Caroline Jane. / In Vitro Maturation of a Humanized Shark VNAR Domain to Improve Its Biophysical Properties to Facilitate Clinical Development. In: Frontiers in Immunology. 2017 ; Vol. 8.
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