Abstract 5557: EnanDIM, a family of potent enantiomeric TLR9 agonists, modulate the tumor microenvironment and show single-agent antitumor effects in various syngeneic murine models

Abstract

Abstract Introduction: TLR9 agonists, being recognized via non-methylated CG-motifs, target the Toll-like receptor 9 (TLR9). This way, they activate the innate and the adaptive immune system. Two different families of TLR9 agonists are currently used in preclinical and clinical studies: Covalently-closed dumbbell-shaped DNA molecules with natural phosphodiester backbone which are protected from degradation by their conformation (dSLIM®) and single-stranded, oligodeoxynucleotides (CpG-ODN) commonly stabilized by phosphorothioates (PTO). However, chemical PTO-modifications have resulted in off-target effects and an unfavorable risk-to-benefit ratio in clinical trials. EnanDIM® are a new family of TLR9 agonists. EnanDIM® molecules consist of linear single-stranded ODN. Protection against exonucleases is achieved through L-deoxyribose nucleotides instead of the naturally occurring D-deoxyribose nucleotides at their 3'-ends. EnanDIM® lead to pronounced induction of cyto- and chemokines, like IFN-alpha and IP-10, as well as broad activation of immune cells, like monocytes, NK cells and dendritic cells, within PBMC. Therefore, EnanDIM® were evaluated in various syngeneic murine tumor models for their anti-tumor effects. Methods: Mice were subcutaneously inoculated with either CT26 (colon carcinoma), MC38 (colon carcinoma), EMT-6 (breast cancer), B16F10 (melanoma) or A20 (lymphoma) cells for solid tumor growth. EnanDIM® (250 µg per dose, intratumoral) was used to treat established tumors. In addition, the capability of EnanDIM® to modulate the tumor micro-environment (TME) was analyzed via immunohistochemistry in a syngeneic tumor model. Results: Mice in all models showed clear reduction in tumor growth (TGI: 53 to 85%) and, therefore, an increased survival (p ≤0.001, logrank). Notably, in the EMT-6 tumor model 8/10 mice showed complete tumor regression after EnanDIM® injection. Those 8 mice rejected EMT-6 tumor cells in a subsequent re-challenge study, indicating a sustained immune memory against the tumor, which was in contrast to age-matched naïve mice. When analyzing the TME of mice from the CT26 model an increased immune cell infiltration into the tumor by CD3+, especially CD8+ T cells was detected, showing the ability of EnanDIM® to beneficially modulate the TME. Conclusions: The enantiomeric TLR9 agonists EnanDIM® show promising anti-tumor effects in a variety of syngeneic murine tumor models and a beneficial modulation of the TME towards an increase of anti-tumor effector cells. These data provide the basis for a further development of EnanDIM® in cancer. Citation Format: Barbara Volz, Kerstin Kapp, Detlef Oswald, Burghardt Wittig, Manuel Schmidt. EnanDIM, a family of potent enantiomeric TLR9 agonists, modulate the tumor microenvironment and show single-agent antitumor effects in various syngeneic murine models [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 5557.