Abstract 5598: Engineered hypoimmune allogeneic CAR T cells as potential off-the-shelf CAR T cell immunotherapies

Off-the-shelf CAR T cells may offer advantages over autologous strategies, including ease of manufacturing, improved quality control with avoidance of malignant contamination and T cell dysfunction, and the ability to generate a final product from healthy T cells. However, host-versus-graft immune response against histoincompatible T cells prevents the expansion and persistence of allogeneic CAR T cells and mitigates the efficacy of this approach. A major challenge is that, while HLA deletion can result in adaptive immune evasion, innate reactivity is enhanced. While T cells express CD47, we demonstrate here that CD47 expression above endogenous levels is important for immune evasion. We describe here the engineering of human immune evasive CAR T cells building on our previously described hypoimmune technology (Nat Biotechnol 2019;37(3):252-258 and Proc Natl Acad Sci U S A 2021;118(28):e2022091118). The goal is to achieve improved rates of durable complete remissions by improving allogeneic CD19CAR persistence, since it has been shown that autologous CAR T cells have greater durability over years than allogeneic CAR T cells. Human T cells from healthy donors were obtained by leukapheresis. To generate hypoimmune CD19CAR T cells, gene editing was used to eliminate HLA-I/II and TCR expression and lentiviral transduction was used to express CD47 and CD19CAR containing a 4-1BB costimulatory domain to generate hypoimmune CD19CAR T cells. Control CD19CAR T cells were unmanipulated, i.e., unedited, except for lentiviral transduction used to express CD19CAR. Hypoimmune CD19CAR T cells persist in allogeneic humanized mice and lack T cell activation measured using bioluminescence imaging and ELISPOT analysis, respectively. In contrast, transplantation of control CD19CAR T cells generated from the same human donor resulted in rejection (ELISPOT mean 59 and 558 spot frequencies for hypoimmune CD19CAR T cells and control CD19CAR T cells, respectively; p