Transduction of the cytoplasmic domain of CTLA-4 inhibits TcR-specific activation signals and prevents collagen-induced arthritis

Abstract

CTLA-4 (CD152) negatively regulates T cell activation signaling, and the cytoplasmic domain of CTLA-4 (ctCTLA-4) itself has the capacity to inhibit T cell activation in vitro and in vivo. In this study, the inhibitory mechanisms of the cell-permeable recombinant protein Hph-1-ctCTLA-4 on T cell activation and its ability to prevent collageninduced arthritis were analyzed. Hph-1-ctCTLA-4 prevented human and mouse T cell activation and proliferation by inhibition of T cell receptor-proximal signaling and the arrest of the cell cycle. Furthermore, Hph-1-ctCTLA-4 protected human umbilical vein endothelial cells (HUVEC) from the human CTL allo-response. The incidence and severity of collagen-induced arthritis were significantly reduced and the erosion of cartilage and bone was effectively prevented by i.v. injection and transdermal administration of Hph-1-ctCTLA-4. Inflammatory cytokine production (IL-1␤, IL-6, TNF-␣, IL-17A) and collagenspecific antibody levels were significantly reduced, and the numbers of activated T cells and infiltrating granulocytes were substantially decreased. These results demonstrate that systemic or transdermal application of a cell-permeable form of the cytoplasmic domain of CTLA-4 offers an effective therapeutic approach for autoimmune diseases such as rheumatoid arthritis.autoimmune disease ͉ costimulatory molecule R heumatoid arthritis (RA) is a chronic, systemic inflammatory disorder that leads to progressive joint destruction (1). Hallmarks of RA are joint inflammation, proliferation of synovial cells, and attachment and invasion of synovial fibroblasts into adjacent cartilage and bone (2, 3). Cumulative evidence has indicated that CD4 ϩ T cell-mediated autoimmune responses play a critical role in the pathogenesis of RA, in conjunction with activation of B cells and macrophages that infiltrate the synovium (4, 5). Recent biological therapies against IL-1␤, IL-6, or TNF-␣ have demonstrated promising effects against progressive joint destruction (6-8). However, the clinical use of these therapies has been limited because of several issues, including side effects and the increase of therapyinsensitive patients.Recently, several reports demonstrated that regulation of costimulatory signaling in the T cell is an important target for treatment of autoimmune diseases such as arthritis (9, 10). CTLA-4 is an activation-induced surface molecule on T cells and is essential for the negative regulation of T cell activation; its inhibitory effects can be accomplished either by competition with CD28 for their ligands B7-1 and -2 or by transmission of negative signals through its intracellular domain (11,12). The cytoplasmic domain of CTLA-4 (ctCTLA-4), which contains a tyrosine phosphorylation motif, has been found to be 100% conserved among different species, suggesting that this domain is critical for CTLA-4 functions (13,14). In addition, there are several splicing variants of CTLA-4, including full-length CTLA-4, soluble CTLA-4 that lacks the transmembrane domain, ligand-independent CTLA-4 (liC...

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