Split-Protein Therapeutic Platforms: Identifying Binder Pairs

Therapeutic proteins, including enzymes, interferons, interleukins, and growth factors, are emerging as important modalities to treat many diseases that elude management by small molecule drugs. One challenge of protein treatment is the propensity for off-target or systemic activity. A promising approach to overcome such toxicity is to create conditionally active constructs by splitting the therapeutic protein into two, or more, inactive fragments and by fusing these fragments to binders (e.g., antibodies) that target distinct epitopes on a cell surface. When these antibodies bind to their respective targets, the protein fragments are brought into proximity and then reconstitute into the active form of the therapeutic protein. In this chapter, we describe approaches to determine antibody pairs that enable the reconstitution of the active protein. General computational and empirical methods are provided to facilitate the identification of pairs starting only from protein sequence data.