Allogeneic Chimeric Antigen Receptor-Invariant Natural Killer T Cells Exert Both Direct and Indirect Antitumor Effects through Host CD8 T Cell Cross-Priming

Abstract

Introduction Chimeric antigen receptor (CAR) T cells have shown impressive results in B-cell malignancies but, unfortunately, their use is still limited by the logistical and financial burdens related to the need of generating autologous cell products. The development of universal allogeneic CAR T cells to be used off-the-shelf has faced major limitations, namely the risk of Graft-versus-Host-Disease (GvHD) induction and the rejection of the cells by the host immune system. Invariant Natural Killer-T (iNKT) cells are innate lymphocytes deprived of GvHD induction potential and displaying antitumor effects, both directly and indirectly, through enhancement of CD8 T cell responses. Preclinical studies in xenogeneic mouse models demonstrated the feasibility of using iNKT cells as a platform for CAR-based therapies, and two clinical trials are currently ongoing. Objective To assess the immunoadjuvant effect of allogeneic CD19-specific iNKT CAR on the host immune system. Methods We transduced murine iNKT cells from FVB/N (H-2Kq) mice with a CD19-specific CD28/CD3ζ CAR and assessed their antitumor effect in vitro and in vivo. Results CD19-iNKT CAR had a direct cytotoxic effect in vitro against the CD19+ A20 lymphoma cells and significantly improved survival of mice after administration to major histocompatibility complex (MHC)–mismatched, immunodeficient BALB/c (H-2Kd) Rag2−/− gamma-chain−/− mice receiving A20 cells (2 × 10e4; Figure 1A) without inducing any signs of GvHD. To test the efficacy of iNKT CAR cells in the presence of host immune cells, we employed BALB/c mice receiving sublethal irradiation (4.4 Gy), resulting in only a partial and transient lymphopenia. The antitumor effect of allogeneic iNKT CAR cells was greatly enhanced (Figure 1B) suggesting the participation of host cells in the antitumor effect. Interestingly, the iNKT CAR effect was partially abrogated when we employed as recipients BALB/c BATF3−/− mice, in which CD8 T cell cross-priming is impaired as a result of the absence of BATF3-dependent CD103+ CD8a+ dendritic cells known to play a role in iNKT cell interactions with other immune effector cells (Figure 1C). Co-administration of allogeneic FVB/N iNKT CAR with autologous BALB/c CD8 T cells at the time of transfer of T-cell-depleted autologous bone marrow cells and A20 lymphoma cells into lethally irradiated BALB/c recipients resulted in a synergistic effect (Figure 1D). Due to their immuno-adjuvant effect on host CD8 T cells, low numbers of allogeneic iNKT CAR outperformed allogeneic conventional CAR T (Tcon CAR) when administered to sublethally irradiated BALB/c mice receiving A20 cells (Figure 1E-F). Conclusion Collectively, these results demonstrate the potent immunoadjuvant effect exerted by allogeneic iNKT CAR cells toward the host immune system suggesting that iNKT CAR cells are an attractive off the shelf product for adoptive immunotherapy.