Abstract Therapeutic approaches that target the immune system in cancer are gaining traction, with agents that disrupt the PD-L1/PD-1 axis, i.e. immune checkpoint inhibitors, showing success in a growing list of malignancies. Despite preclinical studies suggesting activity of PD-1 blockade in malignant gliomas, initial clinical trial results in glioblastoma (GBM) patients have demonstrated that the majority of patients do not respond to PD-1 blockade monotherapy. These results suggest other immunosuppressive pathways operative in malignant gliomas may impart resistance to immune checkpoint inhibitors, highlighting a need for combinatorial approaches to overcome glioma-induced immunosuppression. Our studies establish that tumor infiltrative myeloid cells constitute a targetable axis within resistant gliomas. By utilizing genetic tools and clinically available inhibitors of myeloid cell trafficking and function, we aim to advance a novel combinatorial strategy which may hold relevance for effective immune checkpoint blockade in human GBM tumors. Immune suppressive myeloid-derived cells within the tumor microenvironment are a major contributor to the inability of the immune system to mount an effective anti-tumor response. As such, they constitute a promising cell type to target in order to enhance anti-tumor immune-based therapies. This project will address an important question: does inhibiting the tumor promoting activities of glioma-associated myeloid derived cells, provide a viable strategy for enhancing anti-GBM immune-based therapies? The migration and function of myeloid cells are controlled by chemokines/chemokine receptors, with the CCL2/CCR2 system being a major pathway utilized by these cells to access tissues. CCR2+ myeloid cells are present within human GBM tumors, and pre-clinical glioma models, where they exhibit immunosuppressive characteristics. We have determined that the glioma presence of these cells is dependent on CCR2. We provide compelling results that CCR2-deficiency promotes efficacy of immune checkpoint inhibitors in α-PD-1 insensitive gliomas, as well as enhanced activity in anti-PD- 1 sensitive gliomas. Moreover, treatment of glioma-bearing mice with novel CCR2 antagonists also similarly overcomes resistance of glioma to α-PD-1 inhibitory antibodies. We hypothesize that pharmacologic antagonism of CCR2 will augment the efficacy of immune targeted anti-glioma therapies by inhibiting immune suppressive myeloid-derived cells. The hypothesis will be addressed by the Aims 1) Determine the role of CCR2-expressing immune suppressive myeloid cells in PD-1 resistant glioma, 2) Determine impact of CCR2 antagonism on the adaptive immune response in PD-1 resistant glioma, and 3) Determine efficacy of CCR2 antagonists in human GBM pre-clinical models. These first ever studies will provide clear pre-clinical proof of principle for using CCR2 antagonists as an adjunctive therapeutic modality for immune checkpoint inhibitor- resistant ...