Potentiating immunotherapy in "immune-cold" solid tumors through orchestrating T cell immunity via tumor-specific genetic engineering.

We engineer a tumor-targeted genetic plasmid vector (PαCD3&LIGHT) to systematically modulate T cell immunity. The tumor-specific telomerase reverse transcriptase (TERT) promoter drives simultaneous expression of tumor necrosis factor superfamily member 14 (LIGHT) and membrane-anchored anti-CD3 single-chain variable fragment (αCD3), which are important immunomodulators with closely clinical relevance. Secreted LIGHT induces high endothelial venule formation and chemokine secretion to recruit circulating lymphocytes, while remodeling extracellular matrix to facilitate immune cell penetration into tumor parenchyma. αCD3 establishes artificial immunological synapses between tumor cells and T lymphocytes. This dual mechanism synergistically establishes tertiary lymphoid structures de novo even within deep tumor regions, harboring stem cell-like CD8+ T cells and driving sustained immunity. Concurrently, αCD3-mediated T cell redirection not only amplifies TCR signaling but also reverses exhausted T cells. The orchestrated T cell immunity significantly potentiates checkpoint inhibitor and chimeric antigen receptor (CAR)-T cell therapies in "immune-cold" tumors without obvious side effects and also remarkably enhances the outcome of human CAR-T cells, demonstrating translational potential in solid tumor immunotherapy.