10 Aptamers for Thrombotic Diseases
299
10.5 Aptamers Against FX
FX is a vitamin-K-dependent coagulation factor that is synthesized by the liver
and plays a critical role at the beginning of the common pathway in the coagulation cascade. Active FIXa, FVIIIa, calcium, and phospholipids form the complex
of FIXa-VIIIa-Ca
2+ -PF 3 to convert FX to FXa [1]. Active FVIIa, III, calcium, and
phospholipids also form the complex of FVIIa-III-Ca
2+ to activate FX in the extrinsic
coagulation pathway. FXa and FVa form the complex of FXa-Va-Ca
2+ -PF 3 to cleave
prothrombin to thrombin and turn fibrinogen into fibrin [85, 86].
Hemostasis is a highly regulated process associated with the coordination of coagulation factors and platelets when tissue injury or prothrombotic stimuli happen to
generate a fibrin clot and to limit bleeding [87]. Inappropriately activated hemostasis
results in stroke, myocardial infarction, and venous thrombosis. Anticoagulants in
both chronic and acute clinical settings can inhibit the activation of the coagulation
cascade and fibrin clot formation while the inherent bleeding risk of anticoagulants
is difficult to timely reverse in the event of hemorrhage [88].
Potent and rapid-onset anticoagulation is required for some clinical settings, for
example, CPB surgery. Meanwhile, rapid reversal of anticoagulantion is also important for post-surgery resulting from increased bleeding of anticoagulation.UFH is
the standard anticoagulant targeting multiple coagulation enzymes in CPB surgery
with a risk of postoperative bleeding and potential damage in patients with HIT.
Aside from heparin’s side effects, protamine sulfate reversal can result in hypotension, vasodilation, and cardiac dysfunction [89]. Thus, the development of potent
anticoagulant agents with reliable reversal following surgical procedures is needed
[88].
Present FXa antagonists mainly include rivaroxaban, apixaban, and edoxaban
[90]. RNA 11F7t is a truncated RNA aptamer, modified with 2
-flurocytosine, 2
-
flurouracil, and 3
-inverted deoxythymidine, which selectively binds to FXa, block
interactions between the proteinase and cofactor within prothrombinase, and inhibit
prothrombinase catalyzed thrombin formation without interfering the active site of
the enzyme complex of FXa. The high-affinity binding of RNA 11F7t and FXa in a
Ca
2+ -dependent interaction results in competition for enzyme complex assembly.
FVa-dependent inhibition of prothrombinase function revealed that RNA 11F7t can
compete for the FXa-Va interaction, indicating RNA 11F7t significantly interfere with
FXa regions that participate in the proteinase-cofactor interaction. Thus, despite
the complexity of the assembly of prothrombinase, the anticoagulant aptamer can
efficiently inhibit enzyme complex assembly and thrombin formation in a discrete
manner. This study suggested that aptamer therapeutics can be applied to modulate
thrombin formation and other proteinases in the analogous coagulation steps [90].
In a study, two anticoagulant aptamers, the 36-mer FXa aptamer 11F7t and the
58-mer prothrombin/thrombin aptamer R9D-14t, were combined into a single bivalent RNA aptamer molecule. They were as potent as heparin for anticoagulating
blood. The shortest construct 82-mer RNA BA 4 was found to have similar anticoagulant activity as the two parent aptamers at equivalent dosage. They constructed four
299
10.5 Aptamers Against FX
FX is a vitamin-K-dependent coagulation factor that is synthesized by the liver
and plays a critical role at the beginning of the common pathway in the coagulation cascade. Active FIXa, FVIIIa, calcium, and phospholipids form the complex
of FIXa-VIIIa-Ca
2+ -PF 3 to convert FX to FXa [1]. Active FVIIa, III, calcium, and
phospholipids also form the complex of FVIIa-III-Ca
2+ to activate FX in the extrinsic
coagulation pathway. FXa and FVa form the complex of FXa-Va-Ca
2+ -PF 3 to cleave
prothrombin to thrombin and turn fibrinogen into fibrin [85, 86].
Hemostasis is a highly regulated process associated with the coordination of coagulation factors and platelets when tissue injury or prothrombotic stimuli happen to
generate a fibrin clot and to limit bleeding [87]. Inappropriately activated hemostasis
results in stroke, myocardial infarction, and venous thrombosis. Anticoagulants in
both chronic and acute clinical settings can inhibit the activation of the coagulation
cascade and fibrin clot formation while the inherent bleeding risk of anticoagulants
is difficult to timely reverse in the event of hemorrhage [88].
Potent and rapid-onset anticoagulation is required for some clinical settings, for
example, CPB surgery. Meanwhile, rapid reversal of anticoagulantion is also important for post-surgery resulting from increased bleeding of anticoagulation.UFH is
the standard anticoagulant targeting multiple coagulation enzymes in CPB surgery
with a risk of postoperative bleeding and potential damage in patients with HIT.
Aside from heparin’s side effects, protamine sulfate reversal can result in hypotension, vasodilation, and cardiac dysfunction [89]. Thus, the development of potent
anticoagulant agents with reliable reversal following surgical procedures is needed
[88].
Present FXa antagonists mainly include rivaroxaban, apixaban, and edoxaban
[90]. RNA 11F7t is a truncated RNA aptamer, modified with 2
-flurocytosine, 2
-
flurouracil, and 3
-inverted deoxythymidine, which selectively binds to FXa, block
interactions between the proteinase and cofactor within prothrombinase, and inhibit
prothrombinase catalyzed thrombin formation without interfering the active site of
the enzyme complex of FXa. The high-affinity binding of RNA 11F7t and FXa in a
Ca
2+ -dependent interaction results in competition for enzyme complex assembly.
FVa-dependent inhibition of prothrombinase function revealed that RNA 11F7t can
compete for the FXa-Va interaction, indicating RNA 11F7t significantly interfere with
FXa regions that participate in the proteinase-cofactor interaction. Thus, despite
the complexity of the assembly of prothrombinase, the anticoagulant aptamer can
efficiently inhibit enzyme complex assembly and thrombin formation in a discrete
manner. This study suggested that aptamer therapeutics can be applied to modulate
thrombin formation and other proteinases in the analogous coagulation steps [90].
In a study, two anticoagulant aptamers, the 36-mer FXa aptamer 11F7t and the
58-mer prothrombin/thrombin aptamer R9D-14t, were combined into a single bivalent RNA aptamer molecule. They were as potent as heparin for anticoagulating
blood. The shortest construct 82-mer RNA BA 4 was found to have similar anticoagulant activity as the two parent aptamers at equivalent dosage. They constructed four