Controlled Fab arm exchange

Controlled Fab arm exchange (cFAE) has proven to be a generic and versatile technology for the efficient generation of IgG-like bispecific antibodies (DuoBodies or DBs). This involves the recombination of antigen-binding arms (heavy chain–light chain (H-L) pairs) between individually expressed antibody molecules and is termed Fab-arm exchange, can be modified to make the routine generation of bsAbs feasible. The production of DuoBodies involves (i) separate expression and purification of two parental IgG1s containing two matching point mutations, one in either IgG1, at the CH3:CH3 interface, (ii) mixing of the parental IgGs; (iii) controlled reduction of interchain disulfide bonds in vitro, allowing the recombination of H-L pairs (driven by the matching mutations); and (iv) removal of reducing agent, allowing the reoxidation of interchain disulfide bonds to occur. Fragment-based formats are compatible with well-characterized and cost-effective prokaryotic and lower eukaryotic expression systems, offering the potential advantages of high yields.

Formats of bispecific antibodies (BsAbs)