Immunogenic Properties of Structurally Modified Human Tissue Plasminogen Activators in Chimpanzees and Mice

Immunogenic properties of second generation human tissue plasminogenactivator (tPA) derivatives were examined in chimpanzee andmouse systems. Five species of modified tPAs (mtPAs) (designated2660, 2663, 2810, 8000, and 9200), recombinant native tPA orbovine serum albumin (BSA) as a positive control were subcutaneouslyinjected nine times at suitable intervals into chimpanzees,genetically the closest species to man. These animals were testedfor antigen(Ag)-specific antibodies to the corresponding proteinsby means of enzyme-linked immunosorbent assay and Western blotanalysis. Neither 9200, one of the five mtPAs tested, nor tPAwas immunogenic, although BSA and the other four mtPAs wereimmunogenic under these conditions. Thus, an antigenic determinant was not exposed by the modification on 9200 and thismodified tPA is expected not to be immunogenic in humans. Inthe mouse studies, mice were immunized with mtPAs. Serum sam-piesfrom these animals were tested for antibodies to the mtPAs whichdid not concomitantly recognize native tPA by immune ad sorptionof the antibodies to tPA. The amount of such antibodies alterthe elimination of native tPA-reactive antibodies was littleor none when the serum samples from 9200 and from the othermtPAs, except 8000, were tested. Taking into consideration theresults of the chimpanzee studies, it can be concluded thatAg-specific antibodies are dominantly produced to unchangedepitopes present in modified proteins in the mouse system, inwhich the native protein is immunogenic. These results suggestthat the chimpanzee model should be useful to predict immunogenicityof second generation recombinant proteins in man, while themouse system adopted by us, which determines the newly generatedepitopes of the modified proteins, is not sufficient.