Synthesis of an Immune-Checkpoint Blocker from Engineered Bacteria Elicits a Potent Antitumor Response.

Immune-checkpoint blockade (ICB) reinvigorates T cells from exhaustion and potentiates T-cell responses to tumors. However, most patients do not respond to ICB therapy, and only a limited response can be achieved in a "cold" tumor with few infiltrated lymphocytes. Synthetic biology can be used to engineer bacteria as controllable bioreactors to synthesize biotherapeutics . We engineered attenuated VNP20009 with synthetic gene circuits to produce PD-1 and Tim-3 scFv to block immunosuppressive receptors on exhausted T cells to reinvigorate their antitumor response. Secreted PD-1 and Tim-3 scFv bound PD-1 Tim-3 T cells through their targeting receptors and potentiated the T-cell secretion of IFN-γ. Engineered bacteria colonized the hypoxic core of the tumor and synthesized PD-1 and Tim-3 scFv , reviving CD4 T cells and CD8 T cells to execute an antitumor response. The bacteria also triggered a strong innate immune response, which stimulated the expansion of IFN- CD4 T cells within the tumors to induce direct and indirect antitumor immunity.