Background.

Anti-Galα1-3Gal antibodies cause hyperacute rejection (HAR) in pig-to-primate xenotransplantation. Long-term graft survival has not been achieved despite abrogation of HAR using transgenic pigs. IgG and IgM anti-Galα1-3Gal also play a role in the events following abrogation of HAR. Characterizing these antibodies and developing a system for their removal is therefore crucial to future success in xenotransplantation.

Methods and Results.

We have developed a neoglycoprotein enzyme-linked immunosorbent assay to probe the precise antigenic requirements for the binding of anti-Galα1-3Gal and have analyzed 77 normal sera. Sixty-six percent of individuals have IgG that recognizes the Galα1-3Gal di-, tri-, and pentasaccharides (D, T, and P, respectively), termed DTP phenotype. The frequency of other phenotypes was--P, 13%;-TP, 12%; DP, 8%; and DT-, 1%. The IgG subclasses found were IgG 2 (95%), IgG 3 (34%), IgG 1 (31%), and IgG 4 (17%). IgM in 91% of individuals recognized all three antigens. Further antibody heterogeneity was demonstrated when immunoadsorbents derived from Galα1-3Galβ1-4GlcNAcβ1-3Galβ1-4Glc (PENTA) were tested. Galα1-3Galβ1-4Glc (TRI 6) or PENTA agarose were effective for IgG removal in all individuals. For IgM removal, two deoxy derivatives were completely successful in 73% of individuals. Combining the Galα1-3Gal (DI) and TRI 6 agarose produced an adsorbent that completely removed anti-Galα1-3Gal IgG and IgM in all individuals tested.

Conclusions.

Although the polymorphism in the anti-Galα1-3Gal repertoire, which we have demonstrated, represents a major obstacle to the development of an effective immunoadsorbent, the combination of DI and TRI 6 agarose appears sufficient for pig-to-human xenotransplantation.