Targeting of a mutant plasminogen activator to circulating red blood cells for prophylactic fibrinolysis

Sergei Zaitzev, Dirk Spitzer, Juan Carlos Murciano, Bi Sen Ding, Samira Tliba, M. Anna Kowalska, Khalil Bdeir, Alice Kuo, Victoria Stepanova, John P. Atkinson, Mortimer Poncz, Douglas B. Cines, Vladimir R. Muzykantov

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36 Scopus citations

Abstract

Chemical coupling to carrier red blood cells (RBCs) converts tissue type plasminogen activator (tPA) from a problematic therapeutic into a safe agent for thromboprophylaxis. The goal of this study was to develop a more clinically relevant recombinant biotherapeutic by fusing a mutant tPA with a single-chain antibody fragment (scFv) with specificity for glycophorin A (GPA) on mouse RBCs. The fusion construct (anti-GPA scFv/PA) bound specifically to mouse but not human RBCs and activated plasminogen; this led to rapid and stable attachment of up to 30,000 copies of anti-GPA scFv/PA per mouse RBC that were thereby endowed with high fibrinolytic activity. Binding of anti-GPA scFv/PA neither caused RBC aggregation, hemolysis, uptake in capillary-rich lungs or in the reticuloendothelial system nor otherwise altered the circulation of RBCs. Over 40% of labeled anti-GPA scFv/PA injected in mice bound to RBC, which markedly prolonged its intravascular circulation and fibrinolytic activity compared with its non-targeted PA counterpart, anti-GPA scFv/PA, but not its nontargeted PA analog, prevented thrombotic occlusion in FeCl3 models of vascular injury. These results provide proof-of-principle for the development of a recombinant PA variant that binds to circulating RBC and provides thromboprophylaxis by use of a clinically relevant approach.

Original languageEnglish
Pages (from-to)1022-1031
Number of pages10
JournalJournal of Pharmacology and Experimental Therapeutics
Volume332
Issue number3
DOIs
StatePublished - Mar 1 2010

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    Zaitzev, S., Spitzer, D., Murciano, J. C., Ding, B. S., Tliba, S., Kowalska, M. A., Bdeir, K., Kuo, A., Stepanova, V., Atkinson, J. P., Poncz, M., Cines, D. B., & Muzykantov, V. R. (2010). Targeting of a mutant plasminogen activator to circulating red blood cells for prophylactic fibrinolysis. Journal of Pharmacology and Experimental Therapeutics, 332(3), 1022-1031. https://doi.org/10.1124/jpet.109.159194