Rejection-Resistant Off-the-Shelf T Cells for Adoptive Cell Therapy

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Introduction Autologous therapeutic T cells can cure life-threatening diseases but their manufacturing remains expensive and time-consuming. ‘Off-the-shelf' (OTS) T cell products pre-manufactured from healthy donors are potentially a readily available and less expensive alternative, offering similar therapeutic potency. However, immune rejection by host T- and NK-cells may limit the persistence of OTS cells and compromise their anti-tumor activity. Objectives To enhance the persistence of OTS T cells by engineering resistance to alloimmune rejection. Methods We engineered Alloimmune Defense Receptors (ADRs) that enable OTS T cells to recognize and eliminate alloreactive lymphocytes. We demonstrated ADR-expressing virus-specific (VST) and chimeric antigen receptor (CAR)-expressing T cells resist immune rejection in several in vitro and in vivo models. Results Expression on T cells of ADR specific to 4-1BB and OX40 – costimulatory molecules transiently upregulated on activated T- and NK-cells – enabled specific elimination of activated T cells. Consistent with the expression pattern of ADR targets, 4-1BB ADR T cells preferentially targeted activated CD8+ T cells while OX40 ADR T cells were more selective against CD4+ T cells. Neither ADR produced any discernable activity against resting lymphocytes in peripheral blood. In an in vivo model of allogeneic T-cell rejection, where NSG mice were engrafted with systemic CD19+ leukemia and normal activated PBMC, allogeneic CD19 CAR T cells were rapidly rejected by alloreactive lymphocytes and failed to control systemic leukemia long-term. In contrast, CD19 CAR T cells co-expressing the 4-1BB ADR were fully protected from immune rejection, persisted long term in vivo, and eradicated the tumor (Fig.1). Further, 4-1BB- or OX40- ADR expressing VSTs, which themselves lack alloreactivity when used clinically, were protected from immune rejection by alloreactive T- and NK- cells in an in vitro mixed lymphocyte reaction model, and retained undiminished anti-viral activity. Conclusion These data demonstrate the efficacy of ADRs as first-in-class engineered receptors suppressing immune rejection of OTS VST and CAR-T cell therapies.