ng2-car-macrophages Demonstrate Robust Anti-tumor Effects in Glioblastoma Models

Introduction: The immunosuppressive tumor microenvironment of glioblastoma, primarily comprised of tumor-associated macrophages, poses a significant challenge when designing glioblastoma therapeutics. Chimeric antigen receptor (CAR) technology is a type of cellular immunotherapy that involves engineering the patient's immune cells to target and kill cancer cells, overcoming the immunosuppressive microenvironment. While successful in treating several liquid tumors, the success of CAR-T cells has been limited in the treatment of glioblastoma. CAR-Macrophages (CAR-M) represent a promising alternative that takes advantage of the glioblastoma microenvironment rich in macrophages. Our team has developed a CAR-M that specifically targets neuron-glial antigen 2 (NG2), a cell surface proteoglycan expressed on glioblastoma, to explore the anti-tumor efficacy of CAR-Ms in glioblastoma.

Methods: Using primary human macrophages, we have designed NG2-CAR-Ms to recognize and phagocytose glioblastoma cells. The CAR contains an extracellular single-chain variable fragment (scFv) that binds NG2 and an FcR gamma intracellular domain with an EGFP tag to trigger phagocytosis and allow for downstream analysis. We utilized glioblastoma spheroid models to perform spheroid adhesion, invasion, phagocytosis, cell death, and tumor spheroid growth assays in vitro. For the in vivo portions of this study, we utilized PDX mouse models for tumor growth and survival studies.

Results: We have demonstrated that NG2-CAR-Ms specifically and effectively target and infiltrate glioblastoma spheroids in vitro. Additionally, infiltration by NG2-CAR-Ms is associated with increased levels of phagocytosis of glioblastoma cells by NG2-CAR-Ms and inhibited tumor spheroid growth. Our in vivo studies demonstrated decreased tumor volumes in mice injected with NG2-CAR-Ms compared to controls. Together, these results suggest that NG2-CAR-Ms impede the growth of glioblastoma cells in vitro and in vivo, directly through phagocytosis and indirectly by inducing tumor cell death.

Conclusion : CAR-Macrophages are a new technology currently underexplored in glioblastoma. Our study suggests NG2-CAR-Ms have robust anti-tumor effects in in vitro and in vivo models of glioblastoma. Further work is underway exploring the interplay between NG2-CAR-Ms and anti-tumor immune education in glioblastoma.