Allo-defensive, multiplex base-edited, anti-CD38 CAR T cells for 'off-the-shelf' immunotherapy

Chimeric antigen receptor (CAR) T cell therapies are being widely investigated in both autologous and allogeneic settings, with gene editing providing new strategies to address barriers to mismatched cell therapies. Currently 'universal' donor derived T cell therapies require intensive lymphodepletion and are still prone to hostmediated rejection. CD38, a transmembrane glycoprotein involved in cell activation and bioenergetics, is a promising immunotherapy target for haematological malignancies. Disruption of CD38 expression using base editing prevented fratricide between T cells expressing anti-CD38 CAR (CAR38). Additional base editing enabled generation of 'universal' donor CAR38-T cells, devoid of endogenous TCRαβ and Human Leukocyte Antigen (HLA) molecules after disruption of T Cell Receptor Beta Constant (TRBC), Beta-2 microglobulin (B2M), and Regulatory Factor X5 (RFX5). Removal of cell surface HLA expression enabled evasion of anti-HLA antibodies in sera from sensitised donors and reduced allo-stimulation in mixed lymphocyte cultures (MLCs), while TCRαβ disruption prevented allo-reactivity. In MLCs, CAR38 expression enabled potent 'allo-defense' activity against CD38+ alloreactive cells. Multiplex-base-edited CAR38-T cells exhibited antigen-specific antileukemic activity against human B, T, and myeloid malignancies and inhibited disease progression in humanised murine xenograft models. CAR38-T cells offer a potent 'off-the-shelf' strategy against CD38+ haematological malignancies and longlived plasma cells which can be associated with autoantibody production.