| Multiple sclerosis (MS) is considered to be a T cell-mediated autoimmune disease directed against myelinated nerves within the central nervous system. Current therapies available to MS patients have low efficacy and are immunosuppressive. Novel therapies that negatively regulate or delete autoreactive T cells, i.e., induce antigen-specific T cell tolerance, are key for the development of more efficacious and perhaps curative therapies. Our laboratory has developed a vaccine platform comprised of cytokine-antigen fusion proteins to promote T cell tolerance. In this study, GM-CSF and M-CSF cytokines were tested as domains in cytokine-neuroantigen (NAg) fusion proteins to assess targeting of NAg to different antigen presenting cell (APC) subsets. Fusion proteins were designed with a cytokine N-terminal domain and the encephalitogenic peptide 69-88 of guinea pig myelin basic protein (GP69-88) as the C-terminal domain. Studies measuring T cell activation in vitro, prevention of experimental autoimmune encephalomyelitis (EAE), and treatment of EAE showed that GMCSF-NAg was the most potent fusion protein, with the following rank order of activity (GMCSF-NAg > MCSF-NAg > GP69-88). GMCSF-NAg was 1000-fold more potent than GP69-88 in stimulating myelin basic protein (MBP)-specific T cell proliferation. The mechanism by which GMCSF-NAg promoted T cell activation involved cytokine receptor-mediated uptake of NAg by APC, since free GM-CSF inhibited the GMCSF-NAg potentiated response. GMCSF-NAg potently targeted NAg to dendritic cells and macrophages in vitro, but not to B or T cell APC. Covalent linkage between GM-CSF and NAg was required for enhanced potency of GMCSF-NAg in vitro and for the prevention and treatment of EAE in vivo. In conclusion, GMCSF-NAg potently targeted self-antigen to myeloid APC subsets and caused profound antigen-specific tolerance in EAE. In the future, cytokine-NAg fusion proteins may provide a novel tool to develop antigen-specific, tolerogenic vaccines for the treatment of MS. |
