Abstract 2310: Functional demonstration of CD19 chimeric antigen receptor (CAR) engineered Epstein-Barr virus (EBV) specific T cells: An off-the-shelf, allogeneic CAR T-cell immunotherapy platform
Authors: Shen Rhine R. , Pham Christina D. , Wu Michelle Min , Munson Daniel J. , Aftab Blake T. 
Published: August 2019

Chimeric antigen receptor (CAR) T cells have transformed the treatment of advanced B cell malignancies, but broad application has been limited due to technical and operational challenges in the autologous approach. Virus-specific T cells maintain native T cell receptors and have an established clinical profile and potential utility as an off-the-shelf, allogeneic CAR T-cell immunotherapy platform.

Tabelecleucel (tab-cel®) is an investigational off-the-shelf, allogeneic T-cell immunotherapy that utilizes endogenous T cell receptors targeting EBV antigens associated with select lymphomas and solid tumors. Tab-cel® functions through endogenous TCRs restricted to EBV antigens and has been shown to be generally well tolerated with low incidence of GvHD and cytokine release syndrome, as well as efficacy in EBV+ post-transplant lymphoproliferative disorders (PTLD). Introduction of CAR transgenes into EBV-specific T cells provides an appealing approach for developing off-the-shelf, allogeneic CAR T immunotherapies.

To evaluate feasibility for this platform, we engineered EBV-specific T cells to express second-generation CD19 CARs, utilizing CD28 or 4-1BB co-stimulatory domains. Resulting allo-EBV.CD19.CAR T cells exhibit high expression of both the CD19 CAR and EBV TCR. Additionally, allo-EBV.CD19.CAR T cells demonstrate an enriched central memory phenotype with higher frequency expression of CD62L, CCR7, and CD45RO. Allo-EBV.CD19.CAR T cells also show increased frequency of activation markers CD25 and 4-1BB, and produce activation-associated levels of IFN-γ comparable to EBV-specific T cells. Allo-EBV.CD19.CAR T cells exert potent and specific cytotoxicity against CD19-positive Nalm6 and Raji cells but have limited activity against CD19-negative K562 cells. Both CD19.CAR T and allo-EBV.CD19.CAR T-mediated cytotoxicity displayed rapid and comparable kinetics. Stimulation of allo-EBV.CD19.CAR T cells with either EBV- or CD19-positive targets also induced robust proliferation over several days of antigen challenge. Like unmodified EBV-specific T cells, allo-EBV.CD19.CAR T cells retain the ability to kill B-lymphoblastoid cell lines (BLCL) but spare autologous and allogeneic PHA-blast targets lacking CD19 and EBV antigen expression. This specificity in cytotoxicity is further supported by inflammatory cytokine profiles of effector-target cultures.

From these results, allo-EBV.CD19.CAR T cells demonstrate efficient and effective lysis of antigen-positive target cells, while maintaining low allo-cytotoxic potential. These findings establish feasibility for engineering EBV-specific T cells by leveraging next-generation CAR technologies, and support further development as an off-the-shelf, allogeneic CAR-T immunotherapy platform to generate IND clinical candidates.

Citation Format: Rhine R. Shen, Christina D. Pham, Michelle Min Wu, Daniel J. Munson, Blake T. Aftab. Functional demonstration of CD19 chimeric antigen receptor (CAR) engineered Epstein-Barr virus (EBV) specific T cells: An off-the-shelf, allogeneic CAR T-cell immunotherapy platform [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 2310.