Synthetic zipper mediated pre-targeting system for near-infrared photoimmunotherapy

Several strategies have gained attraction in cancer diagnosis and therapy, particularly to overcome the limitations associated with direct targeting approaches in radioimmunotherapy and photoimmunotherapy. Here, we have introduced a synthetic zipper-mediated pre-targeting system for near-infrared photoimmunotherapy (NIR-PIT) utilizing the IR700-conjugated Zip1-SNAP protein in combination with three scFv-ZIP2 fusion proteins targeting FOLR1, TROP2, and TF. In this two-step targeting approach, scFv-Zip2 proteins first bind to cancer-specific antigens, followed by hybridization with Zip1-SNAP-IR700 at cell surface via high-affinity zipper interaction. Upon NIR light irradiation, targeted cancer cells underwent selective phototoxicity in a concentration-dependent manner with IC₅₀ range (∼104-1,179 nM). Notably, the treatment induced cell death in ∼86%-94% of cells and triggered ICD, as evidenced by increased (∼4- to 200-fold more) surface exposure of calreticulin, HSP70 and HSP90. These findings demonstrate that the synthetic zipper-mediated pre-targeting platform offers a promising and versatile strategy for enhancing the specificity and immunogenic potential of NIR-PIT in cancer therapy.