A multifunctional T-cell engager containing CD80 enhances prostate cancer treatment.

BACKGROUND: T-cell engagers (TCEs), the most extensively studied class of bispecific antibodies, redirect effector T cells to tumor cells to induce immune synapse formation, T-cell activation, and subsequent tumor cell killing. However, their therapeutic efficacy in solid tumors is limited by immunosuppressive mechanisms within the tumor microenvironment (TME). To address this challenge, we engineered an enhanced PSMA/CD3 bispecific antibody by incorporating the extracellular domain of CD80, which provides a co-stimulatory signal to counteract T-cell inhibition in prostate cancer. This trifunctional antibody is designated as TriTE-N13. METHODS: We engineered the fully humanized fusion antibody TriTE-N13 using gene editing and eukaryotic expression systems. In vitro, we evaluated its ability to activate and proliferate T cells, as well as its cytotoxicity against PSMA(+) tumor cells in both 2D co-culture and 3D spheroid models. Additionally, we assessed the in vivo antitumor efficacy and safety of TriTE-N13 using human immune-reconstituted mouse models bearing prostate cancer xenografts. RESULTS: In vitro, TriTE-N13 demonstrated robust activation of human T cells and high-affinity binding to both human PSMA and CD3 antigens. Furthermore, TriTE-N13 effectively mediated T-cell-dependent cytotoxicity against PSMA-positive prostate cancer cells. In vivo studies revealed that TriTE-N13 achieved significantly greater tumor volume reduction compared to conventional bispecific TCEs, particularly in established large tumors. CONCLUSIONS: Our findings indicate that incorporating CD80 as a co-stimulatory signal substantially enhances the antitumor efficacy of TCEs against solid tumors. These results position TriTE-N13 as a promising immunotherapeutic candidate for advanced prostate cancer treatment.