HLA-E single-chain trimer shields gene-edited allogeneic CAR T cells from NK cell attack

Allogeneic chimeric antigen receptor (CAR) T-cell therapies hold promise for certain malignant disorders, as they allow for scalability, on-demand availability, and enhanced product quality. Currently, allogeneic CAR T-cell approaches focus on deleting the αβ T-cell receptor complex and major histocompatibility complex (MHC)-I/II from CAR T cells to prevent graft-versus-host disease and rejection of allogeneic cells by the recipient's immune system. However, genetic ablation of MHC-I leads to natural killer (NK) cell activation in recipients because of the missing-self response. Here, we demonstrate the successful generation of universal allogeneic CAR T (UCART) cells that evade the NK cell's missing-self response and display similar performance to autologous CAR T cells both in vitro and in vivo. An HLA single-chain trimer platform enabled the demonstration that HLA-E presentation of a defined signal peptide sequence, VMAPRTLIL (designated as HLA-ESP-1C), confers resistance to NKG2A+ cell populations with negligible activation of NKG2C+ cell populations. Examination of additional receptor-ligand interactions that may impact NK cell activation confirmed that CD54 and CD58 ablation on UCART cells, in combination with HLA-ESP-1C expression, led to optimal resistance to heterogeneous NKG2A+/C+ NK cell populations. Finally, in a humanized mouse model reconstituted with allogeneic NK cells, we demonstrated that UCART19 cells promote stringent tumor control. Our work suggests an updated approach for allogeneic CAR T-cell therapy in clinical applications.