An engineered 4-1BBL fusion protein with “activity-on-demand”

ABSTRACT Engineered cytokines are gaining importance for cancer therapy but those products are often limited by toxicity, especially at early time points after intravenous administration. 4-1BB is a member of the tumor necrosis factor receptor superfamily, which has been considered as a target for therapeutic strategies with agonistic antibodies or using its cognate cytokine ligand, 4-1BBL. Here we describe the engineering of an antibody fusion protein (termed F8-4-1BBL), which does not exhibit cytokine activity in solution but regains biological activity upon antigen binding. F8-4-1BBL bound specifically to its cognate antigen, the alternatively-spliced EDA domain of fibronectin, and selectively localized to tumors in vivo , as evidenced by quantitative biodistribution experiments. The product promoted a potent anti-tumor activity in various mouse models of cancer, without apparent toxicity at the doses used. F8-4-1BBL represents a prototype for antibody-cytokine fusion proteins, which conditionally display “activity-on-demand” properties at the site of disease upon antigen binding and reduce toxicity to normal tissues.