CNSC-17. UPREGULATION OF CCL4 AFTER TREATMENT WITH A NOVEL MRNA VACCINE SHIFTS THE PHENOTYPE OF CCR5-EXPRESSING MICROGLIA WITHIN TUMOR MICROENVIRONMENT

Abstract INTRODUCTION Intra-tumoral immunosuppression is a major cause of treatment failure for patients with glioblastoma (GBM). Our group has developed a proprietary vaccine that combines mRNA encapsulated in lipid nanoparticles with PEG hydrogel and a chemokine (CXCL9). This hydrogel-chemokine-mRNA (HCM) vaccine is delivered subcutaneously (SQ) and results in significant anti-tumor efficacy in syngeneic murine glioma models. The objective of this study was to evaluate the effects of the vaccine on the tumor microenvironment (TME). METHODS C57BL/6 mice underwent intracranial implantation of KR158-luc tumor cells and underwent SQ injection of the HCM vaccine using total tumor mRNA. Tumors were collected for analysis at different time points. Q-PCR and flow cytometry were done to evaluate the differential gene expression and CCR5 expression (receptor for CCL4) in the TME following HCM vaccination. RESULTS CCL4 was found to be upregulated in tumor-bearing animals treated with HCM vaccine using PCR. In a separate experiment, CCL4 blockade was given prior to HCM vaccination and this resulted in abrogation of the survival benefit of the vaccine. Next, flow cytometry results revealed that microglia in the TME had the highest expression of the CCR5 receptor. Further, QPCR on the isolated microglia from TME revealed significant up-regulation of inflammatory gene CCL5 (25 fold) and down-regulation of anti-inflammatory genes as compared to untreated control and empty hydrogel control. CONCLUSION The HCM vaccine results in significant anti-tumor efficacy in murine syngeneic murine glioma which is dependent on upregulation of CCL4 in the TME. The CCL4 exerts its effect through microglia expressing CCR5 in the TME. HCM vaccination shifted the microglia phenotype from immune-suppression to inflammatory state. Further analysis of the effect of CCR5 knock out on the HCM vaccine survival efficacy are under current investigation.