Oncolytic adenovirus-based therapeutics to target or reprogram glioma-associated macrophages

BackgroundGlioblastoma (GBM) is a highly aggressive and heterogeneous tumor, whose immunosuppressive tumor microenvironment (TME) is a major determinant of resistance to CAR T cell therapy. To address this, targeting glioma-associated macrophages (GAMs) is of significant interest as GAMs comprise a substantial part of the GBM TME and are involved in generating an immunosuppressive phenotype detrimental to T cell function. Here, we investigate an oncolytic adenovirus ICO15K-based therapy to target GAMs, first, by investigating ICO15K-induced reprograming of GAMs and, second, by arming ICO15K with GAM-specific cytotoxic fusion proteins (FPs).MethodsICO15K was previously engineered to specifically infect and replicate in cancer cells. We used glioma patient-derived cell lines to assess its susceptibility to ICO15K in vitro. To characterize ICO15K-induced M1 to M2 transition, monocytes-derived macrophages were obtained and evaluated by qRT-PCR. In an effort to target GAMs, we analyzed gene expression data of sorted immune cell populations found in the TME of tissue specimens from non-tumor brain tissue, IDH-mutated, and wild type gliomas to identify GAM potential surface markers. Versions of GB-CSF1 FPs were constructed using Granzyme B linked to CSF1 ligand, and evaluated in vitro by flow cytometry.ResultsWe show that ICO15K is able to infect, replicate and exert a significant cytotoxicity effect in a panel of glioma patient-derived cell lines. In addition, M2 macrophages populations showed ICO15K-induced transition from a pro-tumoral M2 towards an M1-like pro-inflammatory phenotype. In parallel, GB-CSF1 FPs were successfully designed and expressed, showing specific cytotoxic capacity of CSF1R-positive acute myeloid cell lines in vitro. Finally, transcriptomic analysis resulted in the identification of FCGR2B as a new potential macrophage target for the design of new anti-GAM FPs.ConclusionsAltogether, here we present preliminary data of ICO15K-mediated TME modulation strategy, identification of potential specific targets for GAMs, and the development of anti-GAM FPs. Further preclinical development of these strategies could help to better understand its therapeutic impact for the improvement of GBM patient outcomes.Legal entity responsible for the studyBrain Tumor and Immune Cell Engineering Laboratory.FundingISREC – Swiss Institute for Experimental Cancer Research.DisclosureAll authors have declared no conflicts of interest. BackgroundGlioblastoma (GBM) is a highly aggressive and heterogeneous tumor, whose immunosuppressive tumor microenvironment (TME) is a major determinant of resistance to CAR T cell therapy. To address this, targeting glioma-associated macrophages (GAMs) is of significant interest as GAMs comprise a substantial part of the GBM TME and are involved in generating an immunosuppressive phenotype detrimental to T cell function. Here, we investigate an oncolytic adenovirus ICO15K-based therapy to target GAMs, first, by investigating ICO15K-induced reprograming of GAMs and, second, by arming ICO15K with GAM-specific cytotoxic fusion proteins (FPs). Glioblastoma (GBM) is a highly aggressive and heterogeneous tumor, whose immunosuppressive tumor microenvironment (TME) is a major determinant of resistance to CAR T cell therapy. To address this, targeting glioma-associated macrophages (GAMs) is of significant interest as GAMs comprise a substantial part of the GBM TME and are involved in generating an immunosuppressive phenotype detrimental to T cell function. Here, we investigate an oncolytic adenovirus ICO15K-based therapy to target GAMs, first, by investigating ICO15K-induced reprograming of GAMs and, second, by arming ICO15K with GAM-specific cytotoxic fusion proteins (FPs). MethodsICO15K was previously engineered to specifically infect and replicate in cancer cells. We used glioma patient-derived cell lines to assess its susceptibility to ICO15K in vitro. To characterize ICO15K-induced M1 to M2 transition, monocytes-derived macrophages were obtained and evaluated by qRT-PCR. In an effort to target GAMs, we analyzed gene expression data of sorted immune cell populations found in the TME of tissue specimens from non-tumor brain tissue, IDH-mutated, and wild type gliomas to identify GAM potential surface markers. Versions of GB-CSF1 FPs were constructed using Granzyme B linked to CSF1 ligand, and evaluated in vitro by flow cytometry. ICO15K was previously engineered to specifically infect and replicate in cancer cells. We used glioma patient-derived cell lines to assess its susceptibility to ICO15K in vitro. To characterize ICO15K-induced M1 to M2 transition, monocytes-derived macrophages were obtained and evaluated by qRT-PCR. In an effort to target GAMs, we analyzed gene expression data of sorted immune cell populations found in the TME of tissue specimens from non-tumor brain tissue, IDH-mutated, and wild type gliomas to identify GAM potential surface markers. Versions of GB-CSF1 FPs were constructed using Granzyme B linked to CSF1 ligand, and evaluated in vitro by flow cytometry. ResultsWe show that ICO15K is able to infect, replicate and exert a significant cytotoxicity effect in a panel of glioma patient-derived cell lines. In addition, M2 macrophages populations showed ICO15K-induced transition from a pro-tumoral M2 towards an M1-like pro-inflammatory phenotype. In parallel, GB-CSF1 FPs were successfully designed and expressed, showing specific cytotoxic capacity of CSF1R-positive acute myeloid cell lines in vitro. Finally, transcriptomic analysis resulted in the identification of FCGR2B as a new potential macrophage target for the design of new anti-GAM FPs. We show that ICO15K is able to infect, replicate and exert a significant cytotoxicity effect in a panel of glioma patient-derived cell lines. In addition, M2 macrophages populations showed ICO15K-induced transition from a pro-tumoral M2 towards an M1-like pro-inflammatory phenotype. In parallel, GB-CSF1 FPs were successfully designed and expressed, showing specific cytotoxic capacity of CSF1R-positive acute myeloid cell lines in vitro. Finally, transcriptomic analysis resulted in the identification of FCGR2B as a new potential macrophage target for the design of new anti-GAM FPs. ConclusionsAltogether, here we present preliminary data of ICO15K-mediated TME modulation strategy, identification of potential specific targets for GAMs, and the development of anti-GAM FPs. Further preclinical development of these strategies could help to better understand its therapeutic impact for the improvement of GBM patient outcomes. Altogether, here we present preliminary data of ICO15K-mediated TME modulation strategy, identification of potential specific targets for GAMs, and the development of anti-GAM FPs. Further preclinical development of these strategies could help to better understand its therapeutic impact for the improvement of GBM patient outcomes.