Glioblastoma (GBM) is the most common and aggressive brain malignancy in adults and is currently incurable with conventional therapies. The use of chimeric antigen receptor (CAR) modified T cells has been successful in clinical treatment of blood cancers, except solid tumors such as GBM. This study generated two third-generation CARs targeting different epitopes of ephrin type-A receptor 2 (EphA2) and examined their anti-GBM efficacy in vitro and in tumor-bearing mice. We observed that these two types of T cells expressing CAR (CAR-T) targeting EphA2 could be activated and expanded by EphA2 positive tumor cells in vitro. The survival of tumor-bearing mice after EphA2 CAR-T cell treatment was significantly improved. T cells transduced with one of the two EphA2 CARs exhibited better anti-tumor activity, which is related to the upregulation of CXCR-1/2 and appropriate interferon-γ (IFN-γ) production. CAR-T cells expressed excessively high level of IFN-γ exhibited poor anti-tumor activity resulting from inducing the upregulation of PD-L1 in GBM cells. The combination of CAR-T cells with poor anti-tumor activity and PD1 blockade improved the efficacy in tumor-bearing mice. In conclusion, both types of EphA2 CAR-T cells eliminated 20%-50% of GBM in xenograft mouse models. The appropriate combination of IFN-γ and CXCR-1/2 levels is a key factor for evaluating the antitumor efficiency of CAR-T cells.

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Background:Chimeric antigen receptor (CAR)-T cell therapy is a powerful adoptive immunotherapy against both B cell malignancies and some types of solid tumors. Interleukin (IL) -15 is an important immune stimulator that may offer ideal long-term persistent CAR-T cells. However, higher base line or peak serum IL-15 levels also relate to severe toxicity, such as cytokine release syndrome (CRS), graft-versus-host disease (GVHD) and neurotoxicity.Methods:We successfully constructed CD19 specific armored CAR-T cells overexpressing IL-I5 and IL-15 receptor alpha (IL-15Ra). In vitro cell differentiation and viability were monitored by flow cytometry, and an in vivo xenograft mouse models was used to evaluate the anti-tumor efficiency and toxicity of CAR-T cells. Results:CAR-T cells overexpressing just IL-15 demonstrated enhanced viability, retarded exhaustion in vitro and superior tumor inhibitory effects in vivo. However, these tumor-cured mice had lower survival rate, because of, to some extent, toxicity. Nevertheless, CAR-IL-15 T cells transduced with IL-15Ra had reduced CD132 expression and released less cytokines (IFNγ, IL-2 and IL-15) in vitro, and improved mouse survival compared to CAR-IL-15 T cells.Conclusion:These results indicated that the importance of IL-15 in enhancing T cells persistence and IL-15Ra in reducing the adverse events of IL-15, with superior tumor retardation during CAR-T therapy, which paves the way for the rapid exploitation of IL-15 in adoptive cell therapy.