MDB-97. INVESTIGATING THE IMMUNOMODULATORY ROLE OF V-DOMAIN IG SUPPRESSOR OF T-CELL ACTIVATION IN AN AGGRESSIVE MURINE MODEL OF PEDIATRIC MEDULLOBLASTOMA

Abstract

Abstract BACKGROUND Efforts to treat relapsed medulloblastoma patients with immune checkpoint inhibitors aim to enhance T-cell recognition and elimination of tumor cells. However, significant barriers to treatment efficacy include low numbers of infiltrating effector T-cells and paucity of PD-L1 expression in the tumor microenvironment. Thus, identifying alternative immune checkpoints on the predominant immune cells that infiltrate tumors represents an attractive therapeutic alternative. METHODS Medulloblastoma tumor cells (mCB MYC DNp53) are orthotopically implanted into the right cerebellum of C57/BL6J mice. Once moribund, tumors are harvested and analyzed by flow cytometry. For in vitro studies, tumor cells are co-cultured at increasing concentrations with naïve splenic CD4+ T-cells, CD3/CD28 beads, and rIL-2 for 5 days and then analyzed by flow cytometry. RESULTS Flow cytometric analysis of tumors reveals an abundance of CD45highCD11bhigh tumor-associated macrophages (TAMs), CD45intCD11bint tumor-associated microglia, and CD4+FoxP3+Treg cells. Across all three immune populations, we observe high expression of the inhibitory immune checkpoint, V-domain Ig Suppressor of T-cell Activation (VISTA), with the highest MFI observed on Mø-like TAMs (n=9 tumors; p<0.05). Compared to sham controls, infiltration of Mø-like TAMs is specific to the tumor and express higher levels of MHCII than Mg-like cells (n=14 tumors, 6 sham; p <0.05). Additionally, we find that medulloblastoma tumor cells (mCB MYC DNp53) express VISTA’s binding partner, VSIG3. Tumor cells co-cultured with CD4+ T-cells can inhibit T-cell proliferation in vitro. Preliminary results suggest that addition of 2ug/mL of anti-VISTA blocking antibody (13F3) can rescue proliferation. CONCLUSIONS Tumors are highly infiltrated by VISTA+ T-regs and TAMs. These TAMs upregulate MHCII, suggesting that they are presenting antigen in the context of an inhibitory molecule. Furthermore, our in vitro results warrant further investigation into the role of the VISTA/VSIG3 axis in inhibiting T-cells and supports blocking VISTA as a potential strategy to promote T-cell activation and an anti-tumor response.