432. Molecular Chimerism Prevents Experimental Autoimmune Encephalomyelitis (EAE)

Abstract

Molecular chimerism in the hematopoietic system is usually associated with immune tolerance to the transgene products. Experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis (MS), is a CD4+T cell mediated inflammatory and demyelinating disease that can be induced in susceptible animals upon immunization with certain proteins or peptides present in the myelin sheet, which results in an ascending paralysis. We hypothesized that stably expressing the encephalitogenic peptide containing 16 aminoacids of the myelin oligodendrocyte glycoprotein (MOG40-55) in the murine marrow cells will induce specific immune tolerance to the antigen, which will prevent or reduce susceptibility to the disease. To this end, we constructed a retroviral vector containing the coding sequence of the invariant chain (CD74), in which the CLIP region was replaced by a sequence encoding the MOG40-55 peptide, in order to target its expression to the MHC class II compartment. EGFP was also included in the vector as a reporter gene. Groups of mice (n = 4 and 8, respectively, in two separate experiments) were conditioned with non-myeloablative doses of busulfan (20 mg/Kg) on days -3 and -2, and transplanted with 0.8-0.9 |[times]| 106 unfractionated bone marrow cells transduced with either the vector encoding the antigenic peptide or a control vector containing only EGFP. Three weeks after transplantation the animals were immunized with the MOG40-55 peptide for EAE induction and clinically scored for 30 days. Animals transplanted with marrow cells transduced with the vector encoding the encephalitogenic peptide showed significant protection from EAE (11 of 13 mice did not show any clinical sign of EAE), whereas most animals of the control groups (a total of 30 of 34 mice) developed the disease. Analysis of hematopoietic chimerism and gene transfer rates (EGFP) in the peripheral blood of the animals 21 days after marrow transplantation showed an average level of engraftment of 16.25% (range 0.7 - 66.9%), and a mean transduction rate in vivo of 36,1% (range 7.7 - 75.9%). These results indicate that creating molecular chimerism in the murine hematopoietic system using non-myeloablative conditioning is a powerful tool to induce immune tolerance to the transgene product and that this strategy can be applied to tolerize to antigens involved in autoimmune diseases.