Adnectins: engineered target-binding protein therapeutics

Abstract

Adnectins™ are a new family of therapeutic proteins based on the 10th fibronectin type III domain, and designed to bind with high affinity and specificity to therapeutically relevant targets. Adnectins share with antibody variable domains a beta-sheet sandwich fold with diversified loops, but differ from antibodies in primary sequence and have a simpler, single-domain structure without disulfide bonds. As a consequence, Adnectins bind targets with affinity and specificity as high as those of antibodies, but are easier to manipulate genetically and compatible with bacterial expression systems. Adnectins that bind macromolecular targets with nanomolar and picomolar affinity have been selected using in vitro evolution methods, including mRNA display, phage display and yeast display. CT-322, a PEGylated, anti-angiogenic Adnectin that binds vascular endothelial growth factor (VEGF) receptor 2 and blocks its interaction with VEGF A, C and D, is being evaluated in Phase II clinical trials for efficacy in several oncology indications.

Fig. 1.

Engineered antibodies and ¹⁰Fn3-based target-binding proteins in context. (A) Comparison of three-dimensional structures of a full-length monoclonal, IgG, antibody (PDB ID: 1ITGY; Harris et al., 1998), single-chain antibody (scFv) and domain antibody (V_H; shown in teal). Complementarity-determining regions of the heavy-chain variable domains are shown in blue (CDR-H1), green (CDR-H2) and red (CDR-H3). (B) Comparison of three-dimensional structures of a fragment of human fibronectin (type III domains 7−10) (PDB ID: 1FNF; Leahy et al., 1996) and of a single 10th fibronectin type III domain (¹⁰Fn3; shown in orange). The three loops analogous to CDRs typically diversified in ¹⁰Fn3-based libraries are shown in blue (BC), green (DE) and red (FG). (C) Detailed comparison of three-dimensional structures of V_H and of ¹⁰Fn3, showing diversified loops [colored as in (a) and (b)] and the disulfide bond in the V_H domain (black).