Reduction of the major porcine xenoantigen Galoc(1,3)Gal by expression of α(1,2)fucosyltransferase

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Abstract
Although removal of the Galα(1,3)Gal antigen from pigs would prevent hyperacute graft rejection, the technique of homologous recombination to knock out the α 1,3 galactosyltransferase gene is not available for pigs, and an alternative strategy is presented. As both α 1,3 galactosyltransferase and α 1,2 fucosyltransferase use the same substrate (N‐acetyl lacto‐samine), competition between the transferases in vitro and in vivo was examined. The data show that there is indeed a hierarchy of these gly‐cosyltransferases competing for the same substrate, and that α 1,2 fuco‐syltransferase takes precedence over α 1,3 galactosyltransferase: a) COS cells simultaneously transfected with cDNA clones encoding α, 2 fuco‐syltransferase and α 1,3 galactosyltransferase show preferential expression of the H substance (synthesised by α 1,2fucosyltransferase) rather than Galα(1,3)Gal (synthesised by α 1,3galactosyltransferase), even though α 1,3galactosyltransferase mRNA and functional enzyme was present, b) In a pig kidney cell line that expressed both the Galα(1,3)Gal and H, the increased expression of H induced by the transfection and stable expression of α 1,2fucosyltransferase resulted in decreased expression of Galα(1,3)Gal. c) Coexpression of α 1,2fucosyltransferase and α 1,3galactosyltransferase in either COS cells or the pig cell line resulted in decreased human antibody binding and complement‐mediated cell lysis, d) Transgenic mice, ubiquitously expressing α 1,2fucosyltransferase show a major decrease in Galα‐(1,3)Gal expression and a decrease in natural human antibody binding. These findings have important implications for xenotransplantation in that α,2fucosyltransferase transgenic pigs could be a source of donors for xenotransplantation to humans.