siRNA Protection of Islet Grafts in Baboons

点击翻译按钮获取中文摘要

 DESCRIPTION (provided by applicant): Pancreatic islet transplantation (Tx) has a lifesaving potential for Type 1 diabetes patients. However, even with introduction of the immunosuppressive regimen known as Edmonton Protocol, insulin independence was only transient in most recipients due to significant islet loss that starts early after the procedure. Allogeneic immune rejection, one of the major reasons for islet death, requires permanent use of immunosuppressive drugs, which are toxic to islets and cause serious side effects including cancer and infection. Moreover, in addition to its significant short and long-term side effects, there is direct evidence that immunosuppression leads to reemergence of autoreactivity, the condition that Tx is supposed to treat. New strategies to reduce the dose of immunosuppression by providing alternative defense approaches to transplanted islets are urgently needed. RNA interference is a technique that offers great potential for therapeutic gene silencing and could be employed for improving islet graft resistance to damaging factors after transplantation. We have previously demonstrated the utility of siRNA-conjugated iron oxide-based magnetic nanoparticle (MN) probes for siRNA delivery to pancreatic islets prior to transplantation in immunodeficient rodents. siRNA directed towards genes responsible for islet damage silenced these genes and provided islet protection prior to transplantation. By the use of a magnetic reporter, labeled pancreatic islets could be monitored after transplantation by in vivo magnetic resonance imaging (MRI). These preliminary studies in rodents showed significant improvement in graft outcome after the silencing of genes responsible for apoptosis and immune rejection. In this application we propose to extend these studies to the next level and investigate the effectiveness of MN-siRNA in a pre- clinical immunocompetent non-human primate model of islet transplantation. The experimental design will include synthesis of the MN-siRNA probes targeting caspase-3, caspase-8, Fas and beta2-microglobulin genes, which will then be incubated with pancreatic islets. This will lead to mRNA silencing and simultaneous magnetic labeling of islet cells. Allogeneic islets pre-incubated with these MN-siRNA probe(s) will be transplanted into diabetic baboons that will be monitored long-term using magnetic resonance imaging while tapering immunosupression. In vivo MRI will allow us to follow graft outcome and compare it with non- manipulated control grafts. We anticipate that siRNA treatment of the islets will allow for significant improvement in long-term graft outcome and reduction of the effective dose of immunosupressants. Due to the module design of the nanoparticles siRNA molecules to additional targets can be introduced if necessary.