Purpose: Upregulation of programmed death-ligand 1 (PD-L1) on circulating and tumor-infiltrating myeloid cells is a critical component of GBM-mediated immunosuppression that has been associated with diminished response to vaccine immunotherapy and poor survival. Although GBM-derived soluble factors have been implicated in myeloid PD-L1 expression, the identity of such factors has remained unknown. This study aimed to identify factors responsible for myeloid PD-L1 upregulation as potential targets for immune modulation.Experimental Design: Conditioned media from patientderived GBM explant cell cultures was assessed for cytokine expression and utilized to stimulate na€ ve myeloid cells. Myeloid PD-L1 induction was quantified by flow cytometry. Candidate cytokines correlated with PD-L1 induction were evaluated in tumor sections and plasma for relationships with survival and myeloid PD-L1 expression. The role of identified cytokines on immunosuppression and survival was investigated in vivo utilizing immunocompetent C57BL/6 mice bearing syngeneic GL261 and CT-2A tumors.Results: GBM-derived IL6 was identified as a cytokine that is necessary and sufficient for myeloid PD-L1 induction in GBM through a STAT3-dependent mechanism. Inhibition of IL6 signaling in orthotopic murine glioma models was associated with reduced myeloid PD-L1 expression, diminished tumor growth, and increased survival. The therapeutic benefit of anti-IL6 therapy proved to be CD8 þ T-cell dependent, and the antitumor activity was additive with that provided by programmed death-1 (PD-1)-targeted immunotherapy.Conclusions: Our findings suggest that disruption of IL6 signaling in GBM reduces local and systemic myeloid-driven immunosuppression and enhances immune-mediated antitumor responses against GBM. Ã , P < 0.05; ÃÃ , P < 0.01; ÃÃÃ , P < 0.001; ÃÃÃÃ , P < 0.0001.Lamano et al. Analysis and interpretation of data (e.g., statistical analysis, biostatistics, computational analysis):

Current adjuvant treatment regimens available for the treatment of glioblastoma are widely ineffective and offer a dismal prognosis. Advancements in conventional treatment strategies have only yielded modest improvements in overall survival. Immunotherapy remains a promising adjuvant in the treatment of GBM through eliciting tumor specific immune responses capable of producing sustained antitumor response while minimizing systemic toxicity. Heat Shock Proteins (HSP) function as intracellular chaperones and have been implicated in the activation of both innate and adaptive immune systems. Vaccines formulated from HSP-peptide complexes, derived from autologous tumor, have been applied to the field of immunotherapy for glioblastoma. The results from the phase I and II clinical trials have been promising. Here we review the role of HSP in cellular function and immunity, and its application in the treatment of glioblastoma.

Glioblastoma (GBM) promotes immunosuppression through upregulation of PD-L1 and regulatory T cell (Treg) expansion, but the association of these suppressive factors has not been well elucidated. Here, we investigate a role of PD-L1 in expanding Tregs and the value of targeting the PD-1 receptor to inhibit Treg expansion. Quantitative RNA sequencing data from The Cancer Genome Atlas were evaluated for an association between CD274 and FOXP3 transcript expressions and impact of FOXP3 on clinical outcomes. Peripheral leukocytes from patients with newly diagnosed GBM were profiled for PD-L1 myeloid expressions and Treg abundance. Healthy lymphocytes were assessed for impact of recombinant PD-L1 on expansion of the inducible Treg (iTreg) population. iTreg function was evaluated by the capacity to suppress effector T cell proliferation. Specificity of responses were confirmed by pharmacologic inhibition of the PD-1 receptor. Increased PD-L1 mRNA expression in GBM corresponded to increased FOXP3 mRNA ( = 0.028). FOXP3 elevation had a negative impact on overall survival (HR = 2.0; < 0.001). Peripheral PD-L1 positivity was associated with an increased Treg fraction ( = 0.008). Lymphocyte activation with PD-L1 co-stimulation resulted in greater iTreg expansion compared to activation alone (18.3% vs. 6.5%; < 0.001) and improved preservation of the Treg phenotype. Suppressive capacity on naïve T cell proliferation was sustained. Nivolumab inhibited PD-L1-induced Treg expansion ( < 0.001). These results suggest that PD-L1 may expand and maintain immunosuppressive Tregs, which are associated with decreased survival in glioma patients. Blockade of the PD-L1/PD-1 axis may reduce Treg expansion and further improve T cell function beyond the direct impact on effector cells.