| Shortages of human organs for transplantation have made it necessary to examine the possibility of using non-human organs for xenotransplantation—the transplantation of tissues between different species. Pigs are regarded as the most likely species to serve as donors for clinical xenotransplantation. However, rejection of pig tissues, mediated by the host's immune system, remains a major barrier to successful xenotransplantation. The primary immunological barrier to overcome before clinical xenotransplantation can be successful is rejection mediated by preformed natural antibodies in the host directed toward a single carbohydrate epitope Galα1-3Galβ1-4GlcNAc-R (αGal) present on porcine tissue encoded for by the enzyme glucosyltransferase UDP galactose:β-D-galactosyl-1,4-N-acetyl-D-glucosaminide α(1–3)galactosyltransferase (E.C. 2.4.1.151) or simply αGT. While the most stable means of inducing tolerance relies on bone marrow transplantation to achieve a state of mixed hematopoietic cellular chimerism, the use of xenogeneic bone marrow transplantation to induce tolerance is not yet acceptable clinically because of the severity of the host preparative regimen required to allow engraftment; the potential for graft-versus-host disease; and difficulty in establishing bone marrow engraftment across species barriers. Herein is demonstrated in αGT knockout mice (GT0 mice), which like humans contain in their serum antibodies that bind αGal, that efficient transduction and expression of a retrovirally transduced αGT gene in bone marrow derived cells induces stable long-term tolerance to the αGal epitope. GT0 mice reconstituted with αGT transduced bone marrow were unable to produce antibodies that bind αGal after extensive immunization with pig cells. Using ELISPOT assays, B cells that produce αGal reactive antibodies were undetectable following immunization, suggesting that such B cells were eliminated from the immunological repertoire following gene therapy. Furthermore, hyperacute rejection of αGal expressing H-2 matched heart grafts was prevented in GT0 mice that received αGT transduced bone marrow. After tolerance to αGal was induced by gene therapy, the anti-porcine non-αGal humoral response changed from a predominantly IgM to IgG response. This suggests that once the natural antibody barrier is eliminated by the induction of tolerance, the anti-pig response changes to a typical T cell dependent response involving isotype switching. Thus, gene therapy approaches may be used to overcome immunological responses leading to xenograft rejection. |
