Abstract 729: CD40 and 4-1BB pathway activation using oncolytic viruses kills tumor cells by oncolysis and CD40-mediated apoptosis while activating immune cells

Abstract

Abstract LOAd703 is a tumor-selective oncolytic adenovirus (serotype 5/35) encoding two immunostimulatory transgenes: trimerized membrane-bound CD40L and 4-1BBL. We have previously shown the immunostimulatory effect of LOAd703 in pancreatic cancer models and a clinical trial enrolling patients with pancreatic cancer is ongoing. LOAd703 can potentially be used in a variety of cancers and its applicability across various cancer types was investigated herein. The immune function of the transgenes was further elucidated, including the tumor-specific apoptotic effect of TMZ-CD40L. A panel of human tumor cell lines including B-cell lymphoma (Karpas-422, BC-3, Daudi, DG-75, U-698), multiple myeloma (RPMI-8226), colorectal (HT29), ovarian (SKOV3), lung (A549), neuroendocrine (H727), bladder (T24) and pancreas (Panc01) was selected for evaluation of the LOAd703 oncolytic capacity. The killing capability of LOAd viruses was analyzed using CellTiter96AQueous One Solution Cell Proliferation Assay and Incucyte. Further, LOAd703 was investigated in xenograft models (one per indication). Replication was confirmed by virus-specific quantitative PCR in selected lines. Immunomodulatory capacity on myeloid cells including dendritic cells and macrophages was evaluated in in vitro cultures by flow cytometry and cytokine assays. Apoptosis was determined using flow cytometry. LOAd703 infection resulted in robust virus expansion and killing of tumor cell lines across indications while primary cultures obtained from the pancreas, lung, heart and kidney were not affected. Cell lines such as T24 expressing CD40 showed enhanced sensitivity to LOAd703 that contains TMZ-CD40L compared to LOAd(-) lacking transgenes. The enhanced cell death was due to increased apoptosis frequency. In contrast, the CD40 negative cell lines such as SKOV-3 was equally sensitive to LOAd703 and LOAd(-). Likewise, LOAd703 could effectively control established tumors in xenograft models and the effect was more pronounced in CD40+ cell lines. Interestingly, infected B cell lines showed an enhanced immunogenic phenotype (MHC molecules, Fas, ICAM-I, CD80 and CD86) post infection likely due to their origin as antigen-presenting cells. Stimulation of immature DCs with LOAd703 forced differentiation into a mature phenotype with high capacity of antigen-presentation and expansion of both T cells and NK cells while stimulation of macrophages favored a high M1:M2 ratio. In conclusion, the LOAd703 oncolytic virus can kill a broad range of tumor cells both by CD40-mediated apoptosis and replication-induced oncolysis and the effect is most potent in CD40+ tumor cells. CD40+ myeloid cells such as DCs and macrophages are instead activated by LOAd703. Hence, LOAd703 is a potent multi-functional oncolytic virus. Citation Format: Emma Eriksson, Jessica Wenthe, Sedigheh Naseri, Ann-Charlotte Hellström, Angelica Loskog. CD40 and 4-1BB pathway activation using oncolytic viruses kills tumor cells by oncolysis and CD40-mediated apoptosis while activating immune cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 729.