Bacillus Calmette-Guérin immunotherapy-induced extracellular vesicles promote antitumor immunity in bladder cancer

Intravesical Bacillus Calmette-Guérin (BCG) immunotherapy remains one of the most effective treatments for non-muscle invasive bladder cancer (NMIBC). However, a substantial proportion of patients fail to respond, and the precise mechanisms underlying BCG's therapeutic effects remain incompletely understood. While extracellular vesicles (EVs) can modulate a range of biological processes, their relevance in this context is also poorly understood.

To investigate the interplay between BCG, EVs, and bladder cancer, bladder cancer (BC) cell lines, dendritic cell (DC) and T cell co-culture systems, nanoparticle tracking analysis, molecular profiling, and Western blotting approaches were employed in vitro. Subcutaneous MB49 tumor models in mice were additionally used to assess the impact of exogenously administered BCG-induced EVs on DC activation, T cell proliferation, and antitumor efficacy in vivo.

We identified a novel mechanism of action involving BCG-induced EVs (EVs^BCG). Following BCG internalization by bladder cancer (BC) cells, a robust immune cascade is triggered, accompanied by a surge in EV secretion and altered cargo composition. These EVs^BCG were found to be enriched with key immunomodulatory proteins, including MHC class I and II molecules as well as co-stimulatory molecules CD80 and CD86. Functionally, we found that EVs^BCG were capable of activating DCs, thereby enhancing T cell activity in vitro. Moreover, pre-treatment of mice with EVs^BCG significantly potentiated the therapeutic efficacy of BCG in a syngeneic murine bladder cancer model.

These findings implicate EVs^BCG as critical intermediaries in the anti-tumor immune response and suggest their potential utility as mediators and/or predictive biomarkers for therapeutic efficacy. In summary, we propose a previously unrecognized mechanism of BCG immunotherapy mediated by EVs, establishing them as pivotal components of the immune signaling network driving effective treatment outcomes.