EXTH-71. CHEMOTACTIC RECOMBINANT ADENO-ASSOCIATED VIRUS (RAAV) VIROTHERAPY IMPROVES LYMPHOCYTE RECRUITMENT IN GLIOBLASTOMA (GBM)

Abstract

Abstract BACKGROUND Although immunotherapy holds promise for producing durable outcomes in GBM, a major obstacle is the limited ability of cytotoxic T lymphocytes (CTLs) to home to the tumor. CTLs are recruited via long-range signaling mediated by the diffusion of chemokines present in inflammatory environments. During glioma formation, tumors manufacture immune-suppressive chemokines and cytokines that co-opt resident cells, resulting in the preferential recruitment of immune suppressor cells from the periphery. We explored a novel strategy to selectively reprogram the tumor microenvironment (TME) using focal delivery of rAAV encoding a powerful call-and-receive chemokine for CTLs: CXCL9. HYPOTHESIS: rAAV transduction of the GBM TME to express CXCL9 (rAAV6-CXCL9) will enhance CTL tumor infiltration and improve responsiveness to immunotherapy. METHODS Proteomic arrays were used to identify CTL chemokines absent in human glioma. 3D immunohistochemistry and flow cytometry was used to evaluate geospatial rAAV transgene expression and lymphocyte trafficking in vitro and in vivo. Survival studies of rAAV6-CXCL9 alone and in combination with anti-PD-1 checkpoint blockade was performed in preclinical models of GBM. Tumor immunogenicity following treatment was evaluated by scRNAseq and proteomic analysis. RESULTS We identify rAAV6 as a reliable capsid for transducing murine tumor-reactive astrocytes. Transgene expression following a single intra-tumor injection is focal, stable, and results in high levels of CXCL9 production. CXCL9 is effective at promoting tumor T-cell chemotaxis in vitro and in vivo. rAAV6-CXCL9 treatment sensitizes GBM to PD-1 blockade, improving survival in two distinct syngeneic preclinical models- an effect that is largely dependent on CD8 T-cells. CONCLUSIONS We combine the excellent safety profile of rAAV with focal delivery, enhancing exposure within the tumor compartment and mitigating systemic toxicities seen with conventional therapies. rAAV therapy was successful at stimulating the intended biological response to our encoded transgene (CXCL9): increased lymphocyte recruitment. This strategy has far-reaching potential across other immunotherapy platforms.