The induction of tolerance using the brand new gene therapy in transplantation setting and clarification of the mechanism.

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1. To develop innovative strategies for transplant immunosuppression, we tested the effect of the blockade of costimulatory pathways by using adenovirus vector coding CTLA4Ig(AdCTLA4Ig) or CD40Ig(AdCD40Ig)genes in ACI to LEW rat organ transplantations.A) Heart transplantation : A single treatment with AdCTLA4Ig or AdCD40Ig prolonged graft survival. Although the combined gene therapy allowed long term graft survival, this strategy was not enough to induce tolerance and to avoid chronic rejection.B) Liver transplantation : A single treatment with AdCTLA4Ig or AdCD40Ig prolonged graft survival, and could induce donor-specific tolerance.C) Small bowel transplantation : Although the combined gene therapy prolonged graft survival, this strategy was not enough to induce tolerance and to avoid chronic rejection.D) Xeno heart transplantation(Hamster to Rat) : DXR and cellular rejection were controlled successfully and all xenografts were accepted for over 100 days when AdCTLA4Ig and AdCD40Ig we … More re administered under FK779 induction therapy. However, chronic rejection was inevitable in the long-term surviving xenografts.2. Several strategies have been made in order to avoid the toxicity of adenoviral vector and to establish an efficient and safety gene transfer.A) Ex-vivo transfer : In rat liver transplantation, the graft with 1x10^9pfu AdCTLA4Ig in 4℃ UW solution for more than 8hrs could get efficient transfection rate and tolerance.B) Modified Clamp Technique : Using the short term CT transduction method (under the condition of 4℃ cold preservation for 10 minutes) long term graft survival are achieved.C) Cre/loxP : The administration of adenovirus vector containing Cre recombinase gene(AdexCACre) at the desired time induced Cre-mediated recombination within a gene derived from Adex1CALoxCTLA4IgGLox vector, and the cDNA of CTLA4IgG was excised from the transduced gene and terminated the expression of CTLA4IgG in vitro and in vivo.3. Immunosuppressive mechanism of blockade of CD40-CD 154A) CD40Ig has bi-directional effects, i.e., one direction towards CD4 + T cells that enhances IL-2 production by cross-linking CD40L, and the other to APCs, which results in generation of CD4 + CD25 + T cells. These events, in concert, lead to generation of functional CD4 + CD25 + T cells that regulate CTL activity in an antigen-specific manner. Less