Enzymatic removal of alphaGal antigen in pig kidneys by ex vivo and in vivo administration of endo-beta-galactosidase C.

Abstract

Xenotransplantation using the pig as a donor species is considered to be a promising solution to the serious shortage of organ donors. Both hyperacute and acute vascular rejection (AVR) are believed to be associated with xenoreactive antibody binding to alphaGal epitopes on the pig vascular endothelial cells. Thus, suppression of this antigen-antibody reaction would appear essential for successful long-term xenograft survival. The purpose of this study was to examine the efficacy of ex vivo and in vivo administration of recombinant endo-beta-galactosidase C (EndoGalC which, in previous in vitro studies, has been proven to digest alphaGal antigens completely) on alphaGal epitopes expressed in pig kidneys. Excised pig kidneys were perfused with University of Wisconsin solution containing EndoGalC and preserved for 4 h. After cold storage, the pig kidney was transplanted into another pig. Ex vivo perfusion and cold storage with EndoGalC reduced alphaGal epitope expression on vascular endothelial cells to an undetectable level. However, alphaGal antigens began to be expressed again as early as 1 day after transplantation. The digestion of alphaGal epitopes by EndoGalC did not cause any damage to the kidney graft. EndoGalC was intravenously administered to two pigs (15 kg), without causing any serious adverse effect. Twelve hours later, >98% of alphaGal antigens on pig red blood cells (RBCs) had been digested. Immunohistochemical study revealed almost complete elimination of alphaGal expression on vascular endothelial cells of the kidney graft 4 and 8 h after in vivo administration, but reappearance within 24 h. EndoGalC was administered to a baboon after an interval of 2 months. The second administration did not result in any serious toxicity or reduction in efficacy. These results suggest that ex vivo and in vivo administration of EndoGalC is simple and useful in removing alphaGal epitopes from pig organs. As the effect of EndoGalC is temporary, multiple in vivo administrations of EndoGalC would be required to inhibit the reappearance of alphaGal epitopes. Alternatively, transgenic techniques of introducing the gene for EndoGalC into the donor organ might permanently prevent alphaGal expression.