Life supporting pig-to-primate xenotransplantation model using transgenic GalT KO pigs: The Padua experience

Background: Survival of pig organs transplanted into primates is limited due to the early occurrence of acute humoral xenograft rejection (AHXR). Generation of a1,3-galactosyltransferase gene-knockout (GTKO) and/or human complement regulatory protein (hCRP) transgenic pigs avoided occurrence of hyperacute rejection in pig-to-primate xenotransplantation. The aim of this study was to investigate the added value deriving from further genetic modifications, including the addition of coagulation regulating molecules, in a life supporting pig to primate renal xenotransplantation model. Methods: Twelve bilaterally nephrectomized cynomolgus monkeys received a kidney from GTKO pigs. Animals in Group 1 (n = 4) received a kidney from GTKO, CD55 donor pigs. Animals in Group 2 (n = 6) were transplanted with a kidney from GTKO pigs transgenic for human CD39, CD55, CD59 and fucosyltransferase (TF) proteins. Animals in Group 3 (n = 2) received a kidney from GTKO pigs transgenic for human CD55, endothelial protein C receptor (EPCR) and thrombomodulin (TM). All recipients received cyclophosphamide (up to 4 doses perioperatively), cyclosporin A (35 mg/kg twice a day), mycophenolate sodium (40–60 mg/kg daily) and steroids. Complete hematological and biochemical analyses were routinely performed in all xenotransplanted animals. Pre- and post-transplantation sera were tested for binding of IgM and IgG to donor porcine aortic endothelial cells (PAEC) by flow cytometry. Tissue sections from all explanted kidneys were stained with hematoxylin eosin. The xenografts were also examined for the presence of IgM, IgG, C5b-9 and fibrin by immunohystochemistry. In addition, cellular infiltrates were characterized. Results: Median survival of the animals in the study was 8, 16 and 29 days for Group 1, 2 and 3, respectively. Except for one animal euthanized due to abdominal bleeding, all primates were euthanized in the presence of kidney failure. In all cases a humoral immune response against donor PAEC was observed. Indeed, anti PAEC antibodies IgM and IgG increased as early as on day 4 and 7 days, respectively. Antibodies persisted throughout the post-operative period. Higher antibody titers were observed in Group 2 animals. At euthanasia all xenografted kidneys presented grade II and III AHXR with considerable deposition of IgM, IgG, C5b9 and fibrin. Interestingly, failure in animals from Group 3 occurred in the presence of a more pronounced deposition of IgG. Conclusion: Preliminary data suggest that, in our model, the use of Gal deficient donor kidneys multitransgenic pig with endothelial specific human EPCR and TM proteins extend recipient's survival. This in vivo data confirms that control of coagulation derangements is beneficial in solid organ xenotransplantation.