Conditional activation of IL-12 through a Fibronectin-EDB dependent switch gate

Potent immune-activators, such as interleukin-12 (IL-12) have been challenging to develop for the treatment of solid tumors due to high systemic toxicity. To expand the therapeutic window achievable with IL-12, we engineered a novel and reversible antibody format, comprising a switch arm and targeting arm, that permits conditional activation of IL12 only in the presence of the pan-tumor matrix antigen Fibronectin-EDB (FN-EDB). The switch arm is formed by a dual specificity Fab that binds to tethered IL-12 or FN-EDB in a competitive manner. The FN-EDB targeting arm promotes the avidity‑driven unveiling of IL-12 that is tethered to the switch arm. We employed a quantitative systems pharmacology (QSP) model to define binding parameters required for Switch-IL-12 activity and use a phage screening and rational library design process to generate switch binders with the desired binding profiles. We show that our in vitro functional data support FN-EDB dependent Switch-IL-12 activity and incorporate these data into our QSP model to further refine and expand the therapeutic index of Switch-IL-12.