Human OX40L-CAR-T(regs) target activated antigen-presenting cells and control T cell alloreactivity.

Regulatory T cells (T(regs)) make major contributions to immune homeostasis. Because T(reg) dysfunction can lead to both allo- and autoimmunity, there is interest in correcting these disorders through T(reg) adoptive transfer. Two of the central challenges in clinically deploying T(reg) cellular therapies are ensuring phenotypic stability and maximizing potency. Here, we describe an approach to address both issues through the creation of OX40 ligand (OX40L)-specific chimeric antigen receptor (CAR)-T(regs) under the control of a synthetic forkhead box P3 (FOXP3) promoter. The creation of these CAR-T(regs) enabled selective T(reg) stimulation by engagement of OX40L, a key activation antigen in alloimmunity, including both graft-versus-host disease and solid organ transplant rejection, and autoimmunity, including rheumatoid arthritis, systemic sclerosis, and systemic lupus erythematosus. We demonstrated that OX40L-CAR-T(regs) were robustly activated in the presence of OX40L-expressing cells, leading to up-regulation of T(reg) suppressive proteins without induction of proinflammatory cytokine production. Compared with control T(regs), OX40L-CAR-T(regs) more potently suppressed alloreactive T cell proliferation in vitro and were directly inhibitory toward activated monocyte-derived dendritic cells (DCs). We identified trogocytosis as one of the central mechanisms by which these CAR-T(regs) effectively decrease extracellular display of OX40L, resulting in decreased DC stimulatory capacity. OX40L-CAR-T(regs) demonstrated an enhanced ability to control xenogeneic graft-versus-host disease compared with control T(regs) without abolishing the graft-versus-leukemia effect. These results suggest that OX40L-CAR-T(regs) may have wide applicability as a potent cellular therapy to control both allo- and autoimmune diseases.