Abstract 3221: Oncolytic virotherapy combined with T-cell co-signaling antibodies for intracranial gliomas
Authors: Belcaid Zineb , Berrevoets Cor , Kloezeman Jenneke , van der Kaaij Marielle , Mathios Dimitrios , Lim Michael , Dirven Clemens , Debets Reno , Lamfers Martine 
Published: July 2016

Abstract
Introduction: The tumor-selective human adenovirus Delta24-RGD (DNX-2401) is currently under investigation in phase II clinical trials for recurrent glioblastoma. In recent preclinical studies, our group demonstrated that the immune system, and CD8+ T cells in particular, plays an important role in the establishment of a protective anti-glioma immune memory (Kleijn, PLoS ONE, 2014). In order to improve anti-tumor efficacy of Delta24-RGD therapy with antibody immunotherapy, we are currently investigating the role of T cell co-signaling molecules in our model and their contribution toward an anti-glioma T cell response.
Methods: Immune-competent C57bl/6 mice were intracranially injected with GL261 glioma tumor cells and treated on day 5 with an intra-tumoral injection of Delta24-RGD virus. Single cell suspensions of brain tumors and spleens were stained for flow cytometry with antibodies against CD3, CD4, CD8 as well as ten selected T cell co-signaling molecules: ICOS, 4-1BB, CD28, OX-40, CD40L, CTLA-4, PD-1, LAG-3, TIM-3, BTLA. Tumor cell lysis and IFNy production was assessed in ex vivo cultures of brain tumor cells and immune cells. In addition, IFNy production was assessed from co-cultures with splenocytes and GL261 glioma tumor cells treated with either a PD-1 blocking antibody or an agonist ICOS antibody.
Results: In brain tumors, we observed an increase in numbers of CD8+, but not CD4+ T cells, on day 9 and 14 after virus-treatment. Expression analysis of the co-signaling molecules in infiltrating brain T cells revealed higher expression levels of the co-signaling molecules ICOS, CD28 and PD-1. In the ex vivo tumor cell and immune cell cultures derived from Delta24-RGD treated mice, there is a significant positive correlation between the observed tumor cell lysis and IFNy production ex vivo (Spearman r = 0,9524; p &lt; 0.01). Interestingly, the amount of tumor cell lysis inversely correlated with a low density of CD3+ PD-1+ T cells in vivo (Spearman r = -0,7619; p &lt; 0.05). Co-cultures of splenocytes from virus-treated mice and GL261 glioma tumor cells showed a significant increase in IFNy production when treated with a PD-1 blocking antibody (p &lt; 0.01), but not when treated with an ICOS agonistic antibody.
Conclusion: The inverse relationship between the presence of a low density of PD-1+ TILs in vivo and a higher T cell functionality ex vivo provides rationale for inhibiting the PD-1 receptor with antibody immunotherapy in combination with Delta24-RGD therapy in order to improve therapeutic efficacy. In vivo studies combining these therapies are underway.
Citation Format: Zineb Belcaid, Cor Berrevoets, Jenneke Kloezeman, Marielle van der Kaaij, Dimitrios Mathios, Michael Lim, Clemens Dirven, Reno Debets, Martine Lamfers. Oncolytic virotherapy combined with T-cell co-signaling antibodies for intracranial gliomas. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3221.