COMPARATIVE ANTIGENICITY OF RECOMBINANT WILD-TYPE STAPHYLOKINASE (SAKSTAR) AND A SELECTED MUTANT (SAKSTAR.M38) IN A BABOON THROMBOLYSIS MODEL

Staphylokinase, a bacterial plasminogen activator, is a potent, highly fibrin-specific but antigenic thrombolytic agent in humans. In an eff ort to attenuate the antigenicity of wild-type staphylokinase (SakSTAR variant), 2 of its 3 immunodominant epitopes were altered by substitu ting clusters of 2 or 3 charged amino acids with alanine, yielding the mutant SakSTAR.M38 (K35A, E38A, K74A, E75A, R77A), which was less ant igenic in inbred New Zealand White rabbits. In the present study, grou ps of 6 baboons (Papio hamadryas) were randomized to SakSTAR (group 1) or SakSTAR.M38 (group 2). The thrombolytic potencies of 50 mu g/kg co mpound at baseline, assessed in an extracorporeal thrombosis model, we re similar: 77 +/- 2.9% (mean +/- SEM) clot lysis in group 1 and 83 +/ - 3.6% in group 2. Groups 1 and 2 were immunized subcutaneously at 2, 3, and 5 weeks with 500 mu g SakSTAR or SakSTAR.M38, respectively. Fro m 6 weeks, group 1 developed significantly more antibody-related neutr alizing activity than group 2 (maximal titer at 8 weeks of 100 +/- 23 mu g SakSTAR and of 22 +/- 7.1 mu g SakSTAR.M38 neutralized per millil iter of plasma, respectively). Neutralizing activities subsequently de creased gradually to 10-20% of peak values at 18 weeks. At 6 weeks, bo th groups were resistant to thrombolysis with 50 mu g/kg of either com pound. Rechallenge at 18 weeks with 250 mu g/kg of the immunizing comp ound showed a significantly better recovery of the thrombolytic potenc y of SakSTAR.M38 (68 +/- 4.5% clot lysis) than of SakSTAR (39 +/- 5.3% clot lysis). Neither agent degraded fibrinogen or depleted alpha(2)-a ntiplasmin. Therefore, SakSTAK.M38 is comparably active and fibrin-spe cific but less antigenic than wild-type SakSTAR. These findings in out bred primates confirm and extend earlier observations in inbred rabbit s and provide a basis for the further development of staphylokinase va riants with reduced antigenicity in humans.