IDO and kynurenines in corneal allograft rejection

Abstract

Transplantation is currently the only form of treatment for many blinding diseases caused by opacity of the cornea. Corneal graft rejection is the single most important reason for corneal graft failure and survival rates have not improved over the decades. Alternative investigative interventions have been studied in experimental models of corneal transplantation for many years. IDO catabolises tryptophan and as a consequence of this and the production of tryptophan catabolites (kynurenines) downregulates T cell responses. Overexpression of IDO by virus-mediated cDNA transfer to donor cornea ex vivo prior to transplantation prolongs graft survival. As the overall objective of this project was to develop IDO or kynurenines as potential therapeutic targets for preventing corneal allograft rejection, the first part of this thesis examined potential methods of upregulation of IDO expression in the cornea, concentrating on the effect of UVB exposure on IDO expression. Upregulation of IDO at the mRNA level but not the protein or functional level was demonstrated in response to UVB exposure. The second part of this thesis examined whether kynurenines could mimic the effect of IDO in prolonging corneal graft survival. It was demonstrated that the effect of IDO overexpression in delaying graft rejection could be replicated by both the systemic and local administration of kynurenines. Furthermore the potential mechanism of action of this observed effect was investigated at the in vitro and in vivo levels with kynurenines demonstrating selective T cell cytotoxicity in vitro as well as local and systemic depletion of lymphocyte counts after both local and systemic administration of kynurenines respectively. Given the potential of using kynurenines as a therapeutic strategy in patients, the third part of this thesis examined the effect of a commercially available tryptophan metabolite- Tranilast- on allograft survival. Significant prolongation in graft survival was observed and the potential mechanisms of action of this drug were analysed. Tranilast inhibited T cell proliferation through cell cycle arrest as a result of upregulation of p21 and p15 (cell cycle inhibitors) and downregulation of Cyclin E. An associated reduction in IL-2 production was also observed allowing us to characterise a novel mechanism for Tranilast induced CD4+ T cell anergy. Further to emerging evidence indicating a role for the aryl-hydrocarbon receptor (AhR) in CD4 T cell differentiation and a tryptophan photo-product as a ligand of the AhR, the fourth part of this thesis examined the potential of kynurenines (3- hydroxykynurenine-3HK) as well as Tranilast as agonists of the AhR. 3HK and Tranilast were found to activate the AhR and the downstream signalling pathway. Collectively, the data represented in this thesis helps to explain the effects of IDO and kynurenines in prolonging corneal allograft survival and helps to shed light on the direct immunoregulatory mechanism of IDO and kynurenines. The identification of a commercially available drug effective in prolonging corneal allograft survival may prove a useful adjunct or alternative to topical corticosteroids that can be transferred from the laboratory to clinical practice.