DNA aptamers for human thrombin with high anticoagulant activity demonstrate target- and species-specificity

G-quadruplex-based DNA aptamers for human thrombin represent promising pharmaceuticals having high anticoagulant activity, rapid clearance from the bloodstream, and availability of DNA antidote. High specificity of aptamers minimizes side-effects but at the same time leads to difficulties when choosing animal model for preclinical trials. Well described minimal G-quadruplex 15-TBA, elongated with hinge and duplex regions 31-TBA, and developed recently modular aptamer RA-36 were investigated thoroughly and compared in this paper. To estimate the species-specificity of aptamers the inhibition constants and types were determined for human fibrinogen hydrolysis with human, bovine, porcine, rabbit, rat, and mouse thrombins using turbidimetric assay. Coagulation tests on human fibrinogen, factor II deficient human plasma and animal plasmas were conducted to verify turbidimetric data and reveal either the concentration effects or the influence of other plasma components. And finally, a set of tests on blood clotting cascade was performed to investigate target-specificity of RA-36 aptamer.