Dipg-36. Drd2 Antagonism Rescues Immune-Tumor Surveillance, Warming the Tumor Immune Microenvironment of Diffuse Midline Glioma Models

Abstract BACKGROUND Less than 10% of patients with diffuse midline glioma (DMG) survive 2-years post-diagnosis, with no approved treatments other than palliative radiotherapy. Immune-checkpoint inhibition has failed in the clinic, likely due to a lack of tumor infiltrating lymphocytes (TILs) and limited/no expression of immune checkpoint proteins or proinflammatory cytokines in the tumor immune microenvironment (TIME). Similarly, glioblastoma patients suffer from a ‘cold’ TIME, linked to the sequestration of T cells in the bone marrow (BM) through β-arrestin-induced internalization of S1PR1. β-arrestin is activated by Dopamine receptors (DRD2); therefore, we hypothesized that systemic DRD2 antagonism using the antagonist ONC201, might reverse lymphopenia, increasing TIL populations. METHODS Flow cytometry analysis was performed on bloods from DMG patients at diagnosis. Circulating immune cells and BM from the immunocompetent PPK model +/-ONC201 and tumor-naïve mice were analyzed by a hematology analyzer and flow cytometry. Immunocompromised SU-DIPG-VI and HSJD-DIPG-007 and immunocompetent XFM murine models were treated +/-ONC201 and tumors analyzed by immunoblotting, immunohistochemistry, and flow cytometry. scRNAseq and scATACseq analysis was performed on tumors from immunocompetent PPK models +/-ONC201. RESULTS DMG patients displayed low levels of total lymphocytes, including CD4+, CD8+ and NK subsets (n=9). Similarly, DMG-bearing mice displayed lower levels of lymphocytes (CD45+, CD4+, CD8+ subsets, n=3) compared to sham-engrafted mice. ONC201 treated mice showed lymphocyte levels comparative to sham-engrafted mice, reversing tumor-induced lymphopenia. A significant increase of T cells expressing S1PR1+ was seen in the BM following ONC201. Tumor immune cell infiltration following ONC201 was confirmed in both immunocompromised and immunocompetent models. Multiomics analysis identified ONC201 to promote pro-inflammatory polarization of microglia/macrophages (MHC II and CD86) and increase expression of immune checkpoints (VISTA and PD-L1). CONCLUSIONS DMG patients suffer from low peripheral lymphocytes, likely contributing to cold TIME. ONC201 reversed tumor-induced lymphopenia, promoting TIL recruitment within the tumor, warming the TIME.