Modulation of MS-like disease by a multi epitope protein is mediated by induction of CD11c+CD11b+Gr1+ myeloid-derived dendritic cells.

Specific neutralization of the pathogenic autoimmune cells is the ultimate goal in therapy of Multiple Sclerosis (MS). However, the pathogenic autoimmunity in MS, can be directed against several major target antigens, and therefore targeting pathogenic T-cells directed against a single target antigen is unlikely to be effective. To overcome this multiplicity and the potential complexity of pathogenic autoreactivities in MS, we have put forward the concept of concomitant multi-antigen/multi-epitope targeting as, a conceivably more effective approach to immunotherapy of MS. We constructed an (Experimental Autoimmune Encephalomeylitis (EAE)/MS-related synthetic human Target Autoantigen Gene (MS-shMultiTAG) designed to encode in tandem only EAE/MS related epitopes of all known encephalitogenic proteins. The MS-related protein product (designated Y-MSPc) was immunofunctional and upon tolerogenic administration, it effectively suppressed and reversed EAE induced by a single encephalitogenic protein. Furthermore, Y-MSPc also fully abrogated the development of "complex EAE" induced by a mixture of five encephalitogenic T-cell lines, each specific for a different encephalitogenic epitope of MBP, MOG, PLP, MOBP and OSP. Strikingly, Y-MSPc was consistently more effective than treatment with the single disease-specific peptide or with the peptide cocktail, both in suppressing the development of "classical" or "complex" EAE and in ameliorating ongoing disease. Overall, the modulation of EAE by Y-MSPc was associated with anergizing the pathogenic autoreactive T-cells, downregulation of Th1/Th17 cytokine secretion and upregulation of TGF-ß secretion. Moreover, we show that both suppression and treatment of ongoing EAE by tolerogenic administration of Y-MSPc is associated also with a remarkable increase in a unique subset of dendritic-cells (DCs), CD11c+CD11b+Gr1+-myeloid derived DCs in both spleen and CNS of treated mice. These DCs, which are with strong immunoregulatory characteristics and are functional in down-modulation of MS-like-disease displayed increased production of IL-4, IL-10 and TGF-ß and low IL-12. Functionally, these myeloid DCs suppress the in-vitro proliferation of myelin-specific T-cells and more importantly, the cells were functional in-vivo, as their adoptive transfer into EAE induced mice resulted in strong suppression of the disease, associated with a remarkable induction of CD4 + FoxP3+ regulatory cells. These results, which highlight the efficacy of "multi-epitope-targeting" agent in induction of functional regulatory CD11c+CD11b+Gr1+myeloid DCs, further indicate the potential role of these DCs in maintaining peripheral tolerance and their involvement in downregulation of MS-like-disease.

Targeting of IL-2 receptor with a caspase fusion protein disrupts autoimmunity in prediabetic and diabetic NOD mice.

AIMS/HYPOTHESIS: Interruption of IL-2 signalling is an attractive therapeutic target in autoimmune disorders. In this study we evaluated the effect of a fusion protein composed of IL-2 and caspase-3 (IL2-cas) on NOD mice, as compared with disease induction by cyclophosphamide. METHODS: IL2-cas was assessed in NOD mice at various ages and in conjunction with cyclophosphamide administration. The effect of IL2-cas on diabetogenic cells was evaluated in adoptive transfer experiments and in cell suspension in vitro. RESULTS: IL2-cas induced apoptosis in T cells expressing the alpha chain of the IL-2 receptor (cluster of differentiation [CD]25) in vitro, with superior survival of T cells expressing CD4 and forkhead box P3 (FOXP3). The fusion protein decreased mixed lymphocyte reactivity, and pretreatment with IL2-cas decreased the efficacy of adoptive transfer of diabetes into NOD severe combined immunodeficiency mice. Administration of one dose of IL2-cas decreased the incidence of diabetes in NOD mice, showing a superior beneficial effect when administered at young age, and effectively blocked induction of hyperglycaemia by cyclophosphamide, reducing the severity of islet inflammation. Administration of IL2-cas caused an acute increase in CD25(-)FOXP3(+) T cells in the lymph nodes, pancreas and thymus in NOD mice, with similar effects in wild-type mice. Administration of IL2-cas after onset of hyperglycaemia resulted in superior survival. CONCLUSIONS/INTERPRETATION: Targeted elimination of cells expressing the IL-2 receptor by this fusion protein disrupts the autoimmune pathogenesis in prediabetic and diabetic NOD mice, despite depletion of CD25(+) regulatory T cells. Furthermore, this particular fusion protein is permissive to the development of FOXP3(+) T cells that might contribute to protracted protection from the progression of insulitis and overt hyperglycaemia.