The role of extracellular vesicles and PD-L1 in glioblastoma-mediated immunosuppressive monocyte induction.

BACKGROUND: Immunosuppression in glioblastoma (GBM) is an obstacle to effective immunotherapy. GBM-derived immunosuppressive monocytes are central to this. Programmed death ligand-1 (PD-L1) is an immune checkpoint molecule, expressed by GBM cells and GBM extracellular vesicles (EVs). We sought to determine the role for EV-associated PD-L1 in the formation of Immunosuppressive monocytes. METHODS: Monocytes collected from healthy donors were conditioned with GBM-derived EVs to induce the formation of immunosuppressive monocytes, which were quantified via flow cytometry. Donor-matched T cells were subsequently co-cultured with EV-conditioned monocytes in order to assess effects on T cell proliferation. PD-L1 constitutitive overexpression or shRNA-mediated knockdown was used to determined the role of altered PD-L1 expression. RESULTS: GBM EVs interact with both T cells and monocytes but do not directly inhibit T cell activation. However, GBM EVs induce immunosuppressive monocytes including myeloid-derived suppressor cells (MDSCs) and non-classical monocytes (NCMs). MDSCs and NCMs inhibit T cell proliferation in vitro and are found within GBM in situ. EV PD-L1 expression induces NCMs but not MDSCs, and does not affect EV-conditioned monocytes' T cell inhibition. CONCLUSION: These findings indicate GBM EV-mediated immunosuppression occurs through induction of immunosuppressive monocytes rather than direct T cell inhibition and that, while PD-L1 expression is important for the induction of specific immunosuppressive monocyte populations, immunosuppressive signaling mechanisms through EVs are complex and not limited to PD-L1.