Engineered extracellular vesicles displaying bi-specific T-cell engagers for targeted therapy of B-cell malignancies.

Despite the clinical success of T cell-based immunotherapies such as CAR-T cells and bispecific T cell engagers (BiTEs), therapeutic resistance and immune suppression remain significant barriers in B-cell malignancies. To address these, we developed a novel dual-functional extracellular vesicle (EV) platform, termed BiTE EV@STA, that displays anti-CD3/CD19 BiTE molecules on the EV surface while encapsulating a STING agonist (STA). This strategy enables simultaneous redirection of cytotoxic T cells to tumor cells and stimulation of innate immunity within the tumor microenvironment (TME). BiTE EVs demonstrated favorable pharmacokinetics, enhanced tumor targeting, and robust T cell dependent cytotoxicity and cytokine release. In Nalm6-Luc xenograft models, BiTE EVs significantly inhibited tumor progression and prolonged survival. Further loading of STING agonists into EVs (BiTE EV@STA) activated dendritic cells, and enhanced CD8⁺ T cell infiltration in the TME. Notably, BiTE EV@STA achieved a 4-fold increase in tumor growth inhibition and a marked survival benefit compared to either component alone. This study presents BiTE EV@STA as a promising EV-based immunotherapy that integrates adaptive and innate immune activation to overcome TME-mediated resistance. These findings may have broad implications for enhancing T cell-based therapies in hematologic malignancies and beyond.