Treg-expressed CTLA-4 depletes CD80/CD86 by trogocytosis, releasing free PD-L1 on antigen-presenting cells

Abstract

Abstract Foxp3-expressing CD4+CD25+ regulatory T cells (Tregs) are engaged in the maintenance of immunological self-tolerance and homeostasis. They constitutively and highly express the immune checkpoint receptor CTLA-4, whose Treg-specific deficiency causes severe systemic autoimmunity. As a key mechanism of Treg-mediated suppression, CTLA-4 downregulates the expression of CD80/CD86 costimulatory ligands on antigen-presenting cells (APCs). Here we showed that CTLA-4 facilitated the immune synapse formation and conjugation of Tregs and APCs. These conjugates thus provided stable platforms whereby Tregs were able to deplete CD80/CD86 proteins on APCs by uptaking them via CTLA-4-dependent trogocytosis. The depletion occurred even with Tregs expressing a mutant CTLA-4 lacking the cytoplasmic portion required for its endocytosis. The CTLA-4-dependent trogocytosis of CD80/CD86 also accelerated in vitro and in vivo passive transfer of other membrane proteins and lipid molecules from APCs to Tregs without their significant reduction on the APC surface. Furthermore, CD80 downregulation or blockade by the Treg-expressed membrane CTLA-4 or its solubilized form, respectively, disrupted cis-CD80/PD-L1 heterodimers and increased free PD-L1 on dendritic cells. Taken together, Tregs can exert dual suppressive effects on T-cell immune responses by converting highly stimulatory (CD80highfree PD-L1low) DCs to non-stimulatory/inhibitory (CD80lowfree PD-L1high) DCs via CTLA-4-dependent trogocytosis. Immune checkpoint blockade therapies with the combination of anti-CTLA-4 and anti-PD-1/PD-L1 antibodies may therefore synergistically hinder Treg-mediated immune suppression, thereby effectively enhancing tumor immunity.