Abstract 3275: Inhibition of ER-stress factor C/EBP homologous protein (Chop) with LNAplus™ antisense-oligonucleotides to improve immunotherapy of cancer
Authors: Richard Klar, Yu Cao, Eslam Mohamed, Sven Michel, Monika Schell, Lisa Hinterwimmer, Stefanie Raith, Paulo Rodriguez, Frank Jaschinski
Journal: Cancer Research (2019)

Abstract The microenvironment generated by tumor cells and suppressive stromal cells creates unfavorable conditions for effector immune cells in order to escape anti-tumor immunity. Besides suppressive pathways like for example the CD39/CD73 axis, mechanisms including nutrient starvation, hypoxia, exposure to high levels of reactive oxygen species and acidosis contribute to the suppression of tumor-reactive immune cells. We have recently discovered that the ER-stress response, in particular the C/EBP homologous protein (Chop) plays an important role in the suppression of tumor-exposed T cells. Furthermore, it is upregulated in activated T cells in vitro and may hamper the efficacy of adoptive T cell therapies. As Chop is a transcription factor it falls into the category of “difficult to drug” and therefore represents an optimal target for antisense-oligonucleotides. In order to revert Chop-induced suppression of T cell activity, we designed locked nucleic acid (LNA) ASOs with specificity for mouse or human Chop using our Oligofyer™ bioinformatics system. Knockdown on mRNA level was investigated in cancer cell lines and in T cells in vitro and the most potent ASOs were selected for further experiments. Downstream effects of Chop knockdown were investigated by mRNA expression analysis and flow cytometry. We furthermore investigated the effect of ex vivo Chop knockdown on the efficacy of adoptively transferred Pmel-specific T cells (recognized target: gp10025-33 peptide) in a B16 melanoma model. Treatment of cells with selected Chop-specific ASO leads to potent knockdown of Chop in vitro. In accordance with observations made in T cells derived from Chop-knockout mice, we observed an increase in T cell-associated transcription factor (Tbet), Interferon-gamma (IFN-γ) and Granzyme B (GZMB) expression in T cells that have been treated with a Chop ASO compared to control oligo treated cells. Strikingly, this translated into improved tumor control by Pmel-specific T cells. We furthermore observed an increase in IFN-γ producing tumor-infiltrating T cells and increased frequency of IFN-γ producing T cells in gp100 restimulated splenocytes from animals that received Chop ASO-treated Pmel-specific T cells. Of note, the impact of Chop knockdown was comparable to the effects that we observed in T cells derived from Chop knockout animals. Taken together, we herein show that Chop is a highly promising novel target in immunotherapy and can effectively be targeted by LNAplus™ ASOs. As shown in a model of adoptive T cell therapy, Chop ASOs have a high potential to optimize the efficacy of T cell therapies, such as chimeric antigen receptor (CAR)- or T cell receptor (TCR)-transgenic T cells. We are currently investigating the effect of systemic Chop ASO treatment in murine tumor models to expand the spectrum of applications of this innovative therapeutic tool. Citation Format: Richard Klar, Yu Cao, Eslam Mohamed, Sven Michel, Monika Schell, Lisa Hinterwimmer, Stefanie Raith, Paulo Rodriguez, Frank Jaschinski. Inhibition of ER-stress factor C/EBP homologous protein (Chop) with LNAplus™ antisense-oligonucleotides to improve immunotherapy of cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3275.