LAUSR

Abstract BACKGROUND We and others have identified B7-H3 (CD276) as a promising target for CAR-based immunotherapies for pediatric brain tumors. So far, B7-H3-CAR T cells have only been studied in xenograft models for brain tumors, which do not recapitulate the immunosuppressive tumor microenvironment (TME). To overcome this obstacle, we decided to adapt the immune-competent GL261 murine glioma model which mimics human disease and host immune barriers. METHODS To evaluate the safety and efficacy of antigen-specific CAR T cells, murine B7-H3-CAR T cells were generated using retroviral particles encoding 2nd generation B7-H3-specific CD28.z CAR. Expansion, persistence, and anti-tumor activity were evaluated in vitro and in vivo. Components of the brain TME were then evaluated using flow cytometry and immunostaining. RESULTS B7-H3-CAR T cells only killed B7-H3+ tumor cells, secreted significant levels of IFNγ and IL-2 in an antigen-dependent manner and expanded an average of 33-fold in repeat stimulation assay with B7-H3+ tumor cells in contrast to control CAR T cells. In vivo, intratumoral injection of B7-H3-CAR T cells into orthotopic GL261 glioma induced complete regression in 60% of treated mice. Preliminary studies show numerous infiltration of suppressive tumor-associated macrophages within the tumor and its periphery. CONCLUSIONS In summary, we successfully generated murine B7-H3-CAR T cells and have demonstrated that these cells have potent anti-tumor activity in the immune-competent GL261 glioma model. However, it is likely that the tumor-associated macrophages are mediating immunosuppressive effects on B7-H3-CAR T cells. Therefore, studies focusing on TME/CAR T cell interactions are in progress.