BBB-permeable peptide conjugated cytoplasmic domain of CTLA-4 inhibits Th1 and Th17 responses and pathogenesis of multiple sclerosis

Multiple sclerosis (MS) is one of the most severe autoimmune disease, which cause severe inflammation in central nervous system (CNS). In the MS, CNS-infiltrating effector T cells are regarded that play critical roles. However, current drugs for MS could not targeting infiltrated T cells due to the limitations of drug delivery, which cannot penetrate blood-brain barrier (BBB) and deliver into the CNS. Here, we identified a novel BBB-permeable peptide, dNP2, which effectively delivered proteins into the CNS in vivo . It transduced cargo proteins into the cells with higher efficiency than other cargo delivery peptides. Moreover, it localized in resident neuron, astrocytes and microglia of the mouse brain through blood vessels by penetrating the BBB. Also, we found that dNP2 can deliver its cargo protein into the infiltrated T cells in the CNS of MOG 35-55 immunized MS model mice. When we treated the dNP2-conjugated cytoplasmic domain of cytotoxic T lymphocyte antigen 4 (dNP2-ctCTLA-4), it efficiently downregulated cytokine production in activated T cells, and showed inhibitory impact on experimental autoimmune encephalomyelitis (EAE) in both preventive and therapeutic schemes. This was accompanied with reductions of demyelination and infiltration of T helper 1 (Th1) and T helper 17 (Th17) cells in the CNS. This study suggests that dNP2 is a novel BBB-permeable peptide and that dNP2-ctCTLA-4 could be an effective agent for treating CNS inflammatory diseases such as MS.