Extensive sequence and structural evolution of Arginase 2 inhibitory antibodies enabled by an unbiased approach to affinity maturation

Proc Natl Acad Sci U S A. 2020 Jul 21;117(29):16949-16960. doi: 10.1073/pnas.1919565117. Epub 2020 Jul 2.

Abstract

Affinity maturation is a powerful technique in antibody engineering for the in vitro evolution of antigen binding interactions. Key to the success of this process is the expansion of sequence and combinatorial diversity to increase the structural repertoire from which superior binding variants may be selected. However, conventional strategies are often restrictive and only focus on small regions of the antibody at a time. In this study, we used a method that combined antibody chain shuffling and a staggered-extension process to produce unbiased libraries, which recombined beneficial mutations from all six complementarity-determining regions (CDRs) in the affinity maturation of an inhibitory antibody to Arginase 2 (ARG2). We made use of the vast display capacity of ribosome display to accommodate the sequence space required for the diverse library builds. Further diversity was introduced through pool maturation to optimize seven leads of interest simultaneously. This resulted in antibodies with substantial improvements in binding properties and inhibition potency. The extensive sequence changes resulting from this approach were translated into striking structural changes for parent and affinity-matured antibodies bound to ARG2, with a large reorientation of the binding paratope facilitating increases in contact surface and shape complementarity to the antigen. The considerable gains in therapeutic properties seen from extensive sequence and structural evolution of the parent ARG2 inhibitory antibody clearly illustrate the advantages of the unbiased approach developed, which was key to the identification of high-affinity antibodies with the desired inhibitory potency and specificity.

Keywords: Arginase 2; affinity maturation; antibody engineering; inhibitory antibodies; ribosome display.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies / chemistry*
  • Antibodies / genetics
  • Antibodies / immunology
  • Antibody Affinity*
  • Arginase / immunology*
  • Binding Sites, Antibody
  • Complementarity Determining Regions / chemistry*
  • Complementarity Determining Regions / immunology
  • Humans

Substances

  • Antibodies
  • Complementarity Determining Regions
  • ARG2 protein, human
  • Arginase