Extending the lifetime of anticoagulant oligodeoxynucleotide aptamers in blood

We have investigated 123I and 125I DNA aptamer analogs of anticoagulant DNA aptamers to thrombin exosite 1 and exosite 2 for thrombus imaging potential. Two severe problems are rapid clearance from circulating blood and blood nuclease. With aptamers (unlike antisense) the nucleotide analogs used in polymerase chain reaction-selection cycles also must be used in the radiotracer. We investigated 3ʹ-biotin-streptavidin (SA) bioconjugates of the aptamers to alleviate these problems. Blood nuclease assays and biodistribution analysis were used in the mouse and rabbit. We found that 3ʹ-biotin protected the aptamers significantly from blood nuclease in vitro, but it did not slow in vivo clearance. In contrast, the 3ʹ-biotin-SA bioconjugates were resistant to blood nuclease in vitro and were also longer-lived (10¯20 times) in vivo. Bioconjugate aptamers retained affinity for thrombin. Two solutions emerge: 1) In noncirculating blood (within a thrombus) 3ʹ-biotin extends aptamer lifetime, whereas 2) in circulating blood (the transport medium), where more aggressive clearance is encountered, 3ʹSA extends aptamer lifetime.