Induction of B-cell tolerance by retroviral gene therapy

The primary immunologic barrier to overcome before clinical xenotransplanta tion can be successful is rejection mediated by preformed natural antibodie s in the host, directed toward a single carbohydrate epitope Gal alpha1-3Ga l beta1-4GlcNAc-R (alpha Gal) present on porcine tissue, encoded for by the enzyme glucosyltransferase UDP galactose:beta -D-galactosyl-l,CN-acetyl-D- glucosaminide alpha>(*) over bar * (1-3)galactosyltransferase (EC 2.4.1.151 ) or simply alpha GT. Although we have shown previously that a gene therapy approach could be used to prevent production of natural antibodies specifi c for alpha Gal, the ability to induce and maintain tolerance after rigorou s antigen challenge would be required if similar approaches are to be used clinically. Here, we demonstrate in alpha GT knockout mice (GT(0) mice), wh ich, like humans, contain in their serum antibodies that bind alpha Gal, th at the efficient transduction and expression of a retrovirally transduced a lpha GT gene in bone marrow-derived cells induces stable long-term toleranc e to the alpha Gal epitope. GT(0) mice reconstituted with alpha GT-transduc ed bone marrow cells were unable to produce antibodies that bind alpha Gal after extensive immunization with pig cells. Furthermore, using ELISPOT ass ays, we were unable to detect the presence of B cells that produce alpha Ga l reactive antibodies after immunization, suggesting that such B cells were eliminated from the immunologic repertoire after gene therapy. Interesting ly, after tolerance to alpha Gal is induced by gene therapy, the antiporcin e non-alpha Gal humoral response changes from a predominantly IgM to an IgG response. This suggests that once the natural antibody barrier Is eliminat ed by the induction of tolerance, the antipig response changes to a typical T-cell-dependent response involving isotype switching. Thus, gene therapy approaches may be used to overcome immunologic responses leading to xenogra ft rejection, and similar gene therapy approaches could be used to overcome autoimmunity. (C) 2000 by The American Society of Hematology.