IMMU-04. TEMPORAL AND SPATIAL MODULATION OF THE IMMUNE RESPONSE OF THE MURINE GL261 GLIOMA TUMOUR MICROENVIRONMENT

Abstract INTRODUCTION Glioma is a debilitating and early fatal cancer arising in the glial cells of brain. Glioblastoma, the most aggressive form of gliomas, has a 5-year survival rate of 5% with treatment options limited to surgery, radiotherapy or chemotherapy with temozolomide. OBJECTIVE While radiation and immunotherapies are routinely studied in the murine Gl261 glioma model, little is known about its inherent immune response. In this study we quantified the temporal and spatial localisation of immune cell populations and mediators during glioma development. METHODS Mice were inoculated with 1x106 Gl261 cells at AP0.1mm, ML1.0mm, DV2.6mm Bregma. Tumour morphology, local and systemic immune cell populations were assessed at Day-0, 1, 3, 7, 14, and 21 post-inoculation by MRI, immunohistochemistry for Ki67+ proliferation and CD31+ vessel density; local immune infiltrate by multiplex immunofluorescence (VECTRA®;PerkinElmer); systemic immunity in spleen, bone marrow and peripheral blood by 16-parameter flow cytometry (LSRFortessa™;BD Biosciences); and multiplex immunoassay (Bio-Plex®;Bio-Rad) for plasma cytokines/chemokines. RESULTS From Day-3 tumours were distinguishable with high Ki67+ (> 30%) and increased tissue vascularisation (26.38±1.45 vessels/field; p< 0.05). Ki67 remained high until Day-21 with visible necrotic regions, increased vessel density and lumen area within tumour region. Increasing tumour proliferation/malignancy and vascularisation were associated with significant temporal changes in immune cell populations within the tumour (p< 0.05) and systemic compartments (p=0.02- p< 0.0001). Of note, at Day-14 NK, M1, PMN-MDSCs and M-MDSCs in the tumour infiltrate declined, coinciding with a decrease in 16/24 plasma cytokine/chemokines levels. Tumour infiltrating immune cell populations did not correlate with peripheral blood populations. However, a decrease in plasma cytokine/chemokine levels may indicate ‘immune exhaustion’. CONCLUSIONS The data derived provide baseline characteristics to study changes associated with multi-modal treatment strategies and to sequence therapies to maintain immune modulation. This information will contribute to the identification of novel combination therapies.