EXTH-23. PRECLINICAL ANALYSIS OF COMBINATORIAL GLIOBLASTOMA THERAPY WITH THE PRODRUG-MEDIATED GENE THERAPY VECTOR AdV-tk AND IMMUNE CHECKPOINT INHIBITION

Abstract

While the brain has traditionally been considered to be an immune-privileged site, evidence supporting the use of immunotherapeutics has been rapidly accumulating. Given that virus-based cancer therapies can be immunostimulatory and immune-checkpoint inhibitors block tumor-induced T-cell exhaustion, the combination of these two approaches offers a potentially synergistic interaction. One of the molecular underpinnings of T-cell exhaustion is the expression of Programmed Death-1 (PD1) on T-cells that recognizes its ligand PD-L1. AdV-TK is an immunostimulatory virus-based approach, known as Gene-Mediated Cytotoxic Immunotherapy (GMCI), that involves the intra-tumoral delivery of a non-replicating adenoviral vector carrying the Herpes virus thymidine kinase gene(TK) followed by administration of an anti-herpetic prodrug(ganciclovir-GCV) and recently showed encouraging results in a Phase II trial in glioblastoma(Wheeler et al.,2016). To provide a rationale for this therapeutic combination we investigated PD-L1 expression during GMCI therapy in human and mouse glioma cells in vitro and found that there was a consistent increase in cell surface PD-L1 levels. Interestingly, this was not associated with an increase of mRNA or protein. We also show that GMCI induces a type-I interferon response, and that the release of IFNβ is at least partially responsible for autocrine/paracrine PD-L1 up-regulation. In vivo studies using an intracranial GL261 model showed high levels of long term survivors in the GMCI/PD1 combination(11/14), compared with GMCI(6/16), anti-PD1(5/12) and controls(0/11). In addition, tumor infiltrating lymphocytes after GMCI showed an increase in CD8+, CD8+/GranzymeB+, and CD8+/IFNγ+/TNFα+cells suggestive of cytotoxic T-cell activation. However, there was also a significant increase in CD4+, CD4+/FoxP3+, and IL-10 indicating a significant infiltration by Tregs, releasing immunosuppressive cytokines. Additionally, there was a significant increase in PD1+/TIM3+ T-cells, indicative of an immunosuppressive microenvironment. Overall, our data show that GMCI/anti-PD1 combinatorial therapy is effective in a syngeneic tumor model, and strongly support clinical trials of GMCI/checkpoint inhibitor combinations in glioblastoma patients.