Medicine (St Vincent's) - Theses
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ItemTargeting CD8+ T cells to protect beta cells in type 1 diabetes( 2016)Type 1 diabetes results from destruction of pancreatic beta cells by autoreactive T cells. CD8+ T cells play central role in beta cell destruction. The T cell receptor on CD8+ T cells engages with peptide-MHC class I molecules present on beta cells, and deliver cytotoxic molecules though the immunological synapse. Inhibiting the interaction between CD8+ T cells and beta cells, or blocking cytotoxic pathways could prevent beta cell destruction and hence type 1 diabetes. In this thesis I have used novel small molecule inhibitors to block recognition and killing of beta cells by CD8+ T cells. To achieve this goal in an antigen specific manner for future immunotherapy, I have also investigated the antigens recognized by islet-infiltrating CD8+ T cells from type 1 diabetic donors. In chapter 2, I investigated role of perforin as the major killing mechanism used by CD8+ T cells to kill beta cells. I confirmed that perforin is essential to facilitate beta cell destruction in vivo. In addition, perforin-deficient beta cell antigen-specific CD8+ T cells from NOD8.3 mice were activated more in response to antigen, indicating that perforin may regulate the activation of cytotoxic T lymphocytes. There are currently no therapies available that directly target cytotoxic CD8+ T cells. In chapter 3, I have tested the use of novel small molecule perforin inhibitors for prevention of beta cell death in autoimmune diabetes. Perforin inhibitors protected beta cells from CD8+ T cell killing in vitro and blocked antigen specific CD8+ T cell mediated killing of target cells in vivo. These studies pave the way for testing perforin inhibitors in mouse models of diabetes. Blocking the interaction between CD8+ T cells and beta cells holds promise for prevention of beta cell death, In chapter 4, I showed that small molecule JAK1/JAK2 inhibitors successfully blocked the interaction between beta cells and CD8+ T cells and protected beta cells from CD8+ T cell mediated killing in vitro. When used in mice JAK1/JAK2 inhibitors reduced migration of T cells to islets and prevented cytokine mediated MHC class I upregulation on beta cells, even at later stages of autoimmune diabetes in mice. These inhibitors significantly protected mice from development of autoimmune diabetes. In chapter 5, human islet-infiltrating CD8+ T cell clones from organ donors who died with type 1 diabetes were used to discover beta cell antigens. COS-7 cells co-transfected with donor specific HLA class I alleles and plasmids encoding beta cell antigens were used as antigen presenting cells. While this method worked well to identify the antigen specificity of a CD8+ T cell clone for which the antigen was already known, none of the 24 islet-infiltrating clones tested recognized any of the beta cell antigen and donor specific HLA class I encoding plasmids. This thesis shows that the use of small molecule inhibitors may be effective in protecting protect beta cells from CD8+ T cells in type 1 diabetes. Identifying beta cell antigens recognized by CD8+ T cells will help to develop therapies where these inhibitors can be used in combination with antigen-specific therapy.