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Streptokinase-polyethylene glycol conjugates with increased stability and reduced side effects

Abstract

Covalent SK-PEG2 and SK-PEG5 conjugates with various degrees of modification of the protein amino groups were obtained by variation of the duration of streptokinase (SK) incubation with activated polyethylene glycol (M 2 and 5 kDa, PEG2 and PEG5); their properties were studied in comparison with the properties of unmodified SK in vitro. SK-PEG2 and SK-PEG5 conjugates with the highest stability in plasma retaining 80% of initial fibrinolytic activity were formed at modification degrees of 54 and 52%, respectively. Interaction of the conjugates with equimolar plasminogen resulted in the formation of plasmin (Pm) activator complexes Pm·SK-PEG2 and Pm·SK-PEG5 with the maximum amidase activity being the same as that of Pm complex with native SK. Catalytic efficiency of plasminogen activation (k Pg/K Pg) was found to be slightly higher (2.84 min−1 μM−1) in case of Pm·SK-PEG2 complex and slightly lower, in case of the Pm·SK-PEG5 complex (1.17 min−1 μM−1), if compared to that of the unmodified complex Pm·SK (2.1 min−1 μM−1). Investigation of lysis kinetics of human plasma clot and depletion of plasminogen and fibrinogen plasma levels under the effect of equal doses of SK in free and conjugated forms demonstrated that SK-PEG2 and SK-PEG5 conjugates possess high thrombolytic activity (89 and 72% to the activity of free SK, respectively) and cause 3.5–4-fold lower side effects than free SK. The SK-PEG2 and SK-PEG5 conjugates with increased stability in plasma and reduced side effects may be used in therapy of thrombotic disorders.

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Abbreviations

pNA:

p-nitroaniline

TNBS:

trinitrobenzenesulfonic acid

PA:

plasminogen activators

Pg:

plasminogen

Pm:

plasmin

PEG:

polyethylene glycol

SK:

streptokinase

SK-PEG:

covalent conjugate of streptokinase and polyethylene glycol

Fg:

fibrinogen

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Correspondence to R. B. Aisina.

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Original Russian Text © R.B. Aisina, L.I. Mukhametova, D.V. Tyupa, K.B. Gershkovich, D.A. Gulin, S.D. Varfolomeev, 2014, published in Bioorganicheskaya Khimiya, 2014, Vol. 40, No. 5, pp. 560–570.

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Aisina, R.B., Mukhametova, L.I., Tyupa, D.V. et al. Streptokinase-polyethylene glycol conjugates with increased stability and reduced side effects. Russ J Bioorg Chem 40, 516–525 (2014). https://doi.org/10.1134/S1068162014050021

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  • DOI: https://doi.org/10.1134/S1068162014050021

Keywords

  • streptokinase
  • plasminogen
  • polyethylene glycol conjugate
  • activity
  • stability
  • polyethylene glycol
  • fibrinogen