Abstract 297: Immunotherapy with a CD40L/4-1BBL double-armed oncolytic adenovirus drives Th1 immunity and control tumor progression in a pancreas cancer model
Authors: Svensson Emma , Milenova Ioanna , Moreno Rafael , Alemany Ramon , Loskog Angelica 
Published: August 2015

Abstract
We hypothesized that an immunostimulatory oncolytic virus (LOAd703) is a potent inducer of anti-tumor immunity and tested its biological functions in models of pancreas cancer. LOAd703 is a double-armed oncolytic adenovirus that introduces the simultaneous expression of a trimerized membrane-bound CD40L and 4-1BBL locally in the tumor aiming to induce potent Th1-mediated anti-tumor immunity. By utilizing an oncolytic adenovirus serotype 5/35 for gene transfer, virally induced tumor cell oncolysis will provide a broad release of antigens at the site of immune activation. Transgene expression is driven by a separate promoter to allow for efficient expression in both tumor cells and tumor stroma while virus replication is restricted to tumor cells by E1A delta24 deletion. CD40 ligand is a potent stimulator of myeloid cells including dendritic cells (DCs) that in turn induce robust T cell responses. CD40L can also reduce the levels of myeloid suppressor cells and M2 macrophages as well as enhancing T cell infiltration into tumors. Further, CD40-mediated signaling combined with TLR stimuli (such as adenovirus) induces intense Th1 immunity. 4-1BBL is known to provide expansion and survival signaling to pre-activated T- and NK cells.
Pancreatic cancer cell lines and healthy exocrine pancreas cells were transduced with LOAd703 and it efficiently killed tumor cells while normal cells remained intact as evaluated by an MTS assay. CD40L and 4-1BBL were highly expressed by the transduced tumor cells as detected by flow cytometry. Cell lysis by LOAd703 was equally efficient as a similar oncolytic virus without transgenes demonstrating that the double transgene expression did not interfere with viral replication. Repeated (6x) peritumoral injections of LOAd703 in a xenograft Nu/Nu/Panc01 model showed efficient tumor cell growth control and sustained complete responses. The effect could be further enhanced by gemcitabine in both xenograft mice and in a syngeneic immunocompetent C57BL/6/Panc02 model. Transduction of human monocyte-derived immature DCs resulted in a strong CD40L and 4-1BBL expression without lysis of the cells. Instead, the transduced DCs matured as shown by high CD83 and IL12 expression. The addition of 4-1BBL significantly enhanced DC maturation compared to a virus containing only CD40L, or no transgenes, by expressing higher levels of CD70, IL12, TNFa, IFNg and IL21. Further, LOAd703-transduced DCs pulsed with pp65 CMV peptides potently expanded antigen-specific T cells as well as NK cells.
In conclusion, LOAd703 is a novel, double-armed immunostimulatory oncolytic gene therapy that initiates robust Th1 immunity, and eradicates pancreatic cancer in experimental models. A clinical trial using LOAd703 for pancreatic cancer is underway.
Citation Format: Emma Svensson, Ioanna Milenova, Rafael Moreno, Ramon Alemany, Angelica Loskog. Immunotherapy with a CD40L/4-1BBL double-armed oncolytic adenovirus drives Th1 immunity and control tumor progression in a pancreas cancer model. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 297. doi:10.1158/1538-7445.AM2015-297