10 Aptamers for Thrombotic Diseases
289
Further studies should be addressed on automatization and continuous monitoring
of the hemostasis for the point-of-care application of the PIEZ system [47].
10.2.2 Anti-thrombin RNA Aptamers
10.2.2.1 R9D-14
Bompianic et al. identified an 80 nucleotide 2
-Fluoropyrimidine modified RNA
aptamer (R9D-14) against prothrombin with high affinity and explored the structure and its anticoagulant mechanism. They also found an optimized truncated 58mer aptamer R9D-14 T as a complementary oligonucleotide antidote. The results
revealed that RNAR9D-14 T recognized and bound both prothrombin and thrombin
pro/exosite I with high affinity and blocked thrombin generation and thrombin exosite
I-mediated activity. Meanwhile, RNAR9D-14 T dramatically prolonged APTT, PT,
and TCT clotting time, and the complementary oligonucleotide antidoted RNAR9D14 T in less than 2 min and durably reversed RNAR9D-14 T anticoagulant activity
for more than 2 h in vitro. In addition, RNAR9D-14 T had a greater anticoagulant
activity compared with the DNA aptamer ARC-183 [48].
10.2.2.2 Toggle 25
Toggle 25 (TOG25) is an RNA aptamer modified with 2
-deoxy-2
-fluoropyrimidine
nucleotides. It targets both human and porcine thrombin. The RNA aptamer was
generated by a new selection strategy termed Toggle SELEX, which “toggled” the
protein target between thrombin of human and porcine during alternating rounds
of selection. The experiments demonstrated that Toggle 25 could inhibit two of the
most important functions of both porcine and human thrombin, including plasma
clot formation and platelet activation, by binding evolutionally conserved regions of
thrombin protein. Further crystal structure analysis revealed that Toggle 25 recognized the exosite II of thrombin through a succession of adenine-arginine stacking
interactions [49, 50].
10.2.3 Anti-thrombin Bimodular Aptamers
Mixed duplex/quadruplex oligonucleotides are attractive biomedical aptamer drugs.
For example, the additive of duplex motif to the G-quadruplex module of TBA
improves the binding affinity, as well as the resistance to physiological biodegradation, suggesting that the bimodular structures is very similar to a single G-quadruplex
but with improved biological activity.
289
Further studies should be addressed on automatization and continuous monitoring
of the hemostasis for the point-of-care application of the PIEZ system [47].
10.2.2 Anti-thrombin RNA Aptamers
10.2.2.1 R9D-14
Bompianic et al. identified an 80 nucleotide 2
-Fluoropyrimidine modified RNA
aptamer (R9D-14) against prothrombin with high affinity and explored the structure and its anticoagulant mechanism. They also found an optimized truncated 58mer aptamer R9D-14 T as a complementary oligonucleotide antidote. The results
revealed that RNAR9D-14 T recognized and bound both prothrombin and thrombin
pro/exosite I with high affinity and blocked thrombin generation and thrombin exosite
I-mediated activity. Meanwhile, RNAR9D-14 T dramatically prolonged APTT, PT,
and TCT clotting time, and the complementary oligonucleotide antidoted RNAR9D14 T in less than 2 min and durably reversed RNAR9D-14 T anticoagulant activity
for more than 2 h in vitro. In addition, RNAR9D-14 T had a greater anticoagulant
activity compared with the DNA aptamer ARC-183 [48].
10.2.2.2 Toggle 25
Toggle 25 (TOG25) is an RNA aptamer modified with 2
-deoxy-2
-fluoropyrimidine
nucleotides. It targets both human and porcine thrombin. The RNA aptamer was
generated by a new selection strategy termed Toggle SELEX, which “toggled” the
protein target between thrombin of human and porcine during alternating rounds
of selection. The experiments demonstrated that Toggle 25 could inhibit two of the
most important functions of both porcine and human thrombin, including plasma
clot formation and platelet activation, by binding evolutionally conserved regions of
thrombin protein. Further crystal structure analysis revealed that Toggle 25 recognized the exosite II of thrombin through a succession of adenine-arginine stacking
interactions [49, 50].
10.2.3 Anti-thrombin Bimodular Aptamers
Mixed duplex/quadruplex oligonucleotides are attractive biomedical aptamer drugs.
For example, the additive of duplex motif to the G-quadruplex module of TBA
improves the binding affinity, as well as the resistance to physiological biodegradation, suggesting that the bimodular structures is very similar to a single G-quadruplex
but with improved biological activity.