Abstract B07: Immunostimulatory gene therapy enhances PD-1 blockade antibody therapy in experimental lung cancer
Authors: Wenthe Jessica , Eriksson Emma , Hellström Ann-Charlotte , Loskog Angelica 
Published: February 2019

Abstract
This study aims to evaluate if immunostimulatory gene therapy is a good enhancer of checkpoint blockade therapy. Lung cancer is a deadly disease responding poorly to conventional therapy. Checkpoint blockade antibodies inhibiting PD-1/PD-L1 signaling have shown significant clinical benefit. Nevertheless, there is still room for improving overall survival rates. Since the success of PD-1 blockade therapy prerequisites the presence of tumor-reactive T cells, we hypothesized that an activating immunotherapy with high potency to stimulate T cell responses would be an ideal candidate to combine with anti-PD-1 antibody treatment. LOAd703 is a replication-restricted adenovirus serotype 5/35. It is armed with two immunostimulatory transgenes, 4-1BB ligand (4-1BBL) and a trimerized, membrane-bound human CD40 ligand (TMZ-CD40L) (PCT/EP2015/057489). Both molecules are key co-stimulatory molecules in the dendritic cell (DC)/T cell synapse. The capacity of LOAd703 was tested using the lung cancer cell lines A549 and H727. Infected tumor cells were killed by oncolysis in a 72hr MTS viability assay with high dose virus (100MOI) which also translated into tumor control in a xenograft mouse model. In tumor cell co-culture with peripheral blood mononuclear cells (PBMCs) (Incucyte), the PBMCs were rapidly tolerized by the tumor microenvironment and did not proliferate, nor kill, the allogeneic tumor. Addition of anti-PD-1 antibody did not increase tumor cell apoptosis as a monotherapy. If OKT3/IL2 was added to the co-culture, apoptosis was confirmed in the cultures peaking at day 5. Combination of OKT3/IL2 with anti-PD-1 antibody significantly enhanced the effect. LOAd703 was added to tumor cell/PBMC co-cultures at a low MOI to delay oncolysis but allow transgene expression. LOAd703 rapidly induced caspase activation in the tumor cells, starting already at day 2 and the activity was continuously rising throughout the experiment. Combining LOAd703 with anti-PD-1 antibody significantly increased apoptosis induction of tumor cells. A control virus lacking immunostimulatory transgenes was less efficient. At endpoint, LOAd703 plus anti-PD-1 was twice as effective as OKT3/IL2 plus anti-PD-1. Immune cells were analyzed by flow cytometry and supernatants taken for cytokine analysis. The T cells were increased in the same groups as tumor cell killing was previously noted. IFNg, TNF, IL2 were all enhanced by LOAd703 and OKT3/IL2. The LOAd703 virus does not infect murine cells since it targets human CD46. However, murine B cells can take up the virus by another route albeit with poor efficacy. Nevertheless, the murine A20 B cell lymphoma was used as a surrogate syngeneic model to test the ability of LOAd703 to potentiate aPD-1 therapy. In line with in vitro data, LOAd703 could enhance in vivo tumor control of anti-PD-1 therapy. IFNg and TNF were enhanced in serum of combination animals. In conclusion, LOAd703 is a potent stimulator of T cell responses and potentiates the effect of aPD-1 therapy. LOAd703 is currently in clinical phase I investigation prior initiating combination studies in suitable indications such as lung cancer.
Citation Format: Jessica Wenthe, Emma Eriksson, Ann-Charlotte Hellström, Angelica Loskog. Immunostimulatory gene therapy enhances PD-1 blockade antibody therapy in experimental lung cancer [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2017 Oct 1-4; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2018;6(9 Suppl):Abstract nr B07.