2 Aptamer-Based Probes for Molecular Imaging
35
as well as several tumors are proved to overexpress TN-C [20]. This 2
fluoropyrimidine RNA oligomer was generated by a cross-over SELEX referring the purified recombinant TN-C protein and TN-C-positive U251 glioblastoma cells. Then,
the aptamer was further synthesized with chemically modified nucleotides and a
5
amine functional group. Chemically modified nucleotides offered better resistance against degradation by nucleases. The amine group acted as a conjugation site
for the chelators 2-mercaptoacetylglycylglycylglycine (MAG2), MAG2-PEG3400
and diethylenetriaminepentaacetic acid (DTPA). In order to confirm the nucleases
resistance of aptamer TTA1 in vivo, the previously described anti-elastase aptamer
[18] without chemically modified was used as control aptamer. The biodistribution
of aptamer TTA1radiolabeled with
99m Tc was visualized by SPECT imaging after
intravenous injection in U251 glioblastoma xenografted mice (Fig. 2.2b). Aptamer
99m Tc -MAG2-TTA1 showed extremely rapid blood clearance and deep tumor penetration. The half-life of this aptamer was <2 min, and a maximum uptake of 6%
injected dose/g was achieved in the tumor at 10 min after injection. In contrast to
the anti-TN-C antibody BC-8 which required 40 h to reach a tumor-to-blood ratio
of 5, the tumor-to-blood ratio of
99m Tc -MAG2-TTA1 reached 50 just at 3 h after
injection. Such rapid clearance can be explained by both renal and hepatic elimination pathways, and nuclease degradation. In fact, when the aptamer was completely
degraded in the blood at 3 h after injection, the radioactivity presented in the tumor
was consistent mostly to nondegraded aptamer. This study also suggested the significant contribution of radiometal chelators on the aptamer’s biodistribution. Although
three different chelators had less of an effect on tumor targeting, there was a markedly
difference on the clearance of aptamers.
Mostly aptamers using SPECT for molecular cancer imaging were selected against
proteins that are overexpressed at the surface of cancer cells. MUC1 is a large glycoprotein that provided a protective barrier for the outer surface of epithelial cells and
its overexpression has been associated with poor clinical prognosis, such as human
breast cancer and other cancers [21]. Two aptamers against the MUC1 protein core
(AptA) or the glycosylated MUC1 protein (AptB) were selected [22]. Da Pieve
et al. compared the biodistribution of radiolabeled AptA and AptB with
99m Tc
conjugated chelator MAG2b in MCF7 xenografted mice [23]. The tumor uptake
of
99m Tc -MAG2-AptB was slightly higher at 5 h after injection. But the aptamer
was completely cleared from the system at 3 h after injection, which prevents the
aptamer from adequately penetrating the tumor tissue. In another study, aptamers
targeting MUC1 were radiolabeled with
99m Tc and loaded on silica nanoparticles
(SBA-15) [24]. In the presence of stannous chloride, radiolabeling with
99m Tc was
highly efficient (the average radiolabeling yield was 99.4%) and stable up until 8 h
after radiolabeling. Nucleolin, one of the major proteins of the nucleolus, could be
translocated to the external side of the cell membrane. Overexpressed nucleolin on
the cell membrane served as a receptor for several molecules implicated in tumor
development [25]. AS1411was a 26-base guanine-rich oligonucleotide aptamer with
a high affinity to nucleolin and the first aptamer to reach phase I and II clinical
trials due to its unexpected ability to inhibit cancer cell growth [26]. Noaparast et al.
labeled a modified AS1411 with
99m Tc and evaluated it as a potential tumor-targeting
35
as well as several tumors are proved to overexpress TN-C [20]. This 2
fluoropyrimidine RNA oligomer was generated by a cross-over SELEX referring the purified recombinant TN-C protein and TN-C-positive U251 glioblastoma cells. Then,
the aptamer was further synthesized with chemically modified nucleotides and a
5
amine functional group. Chemically modified nucleotides offered better resistance against degradation by nucleases. The amine group acted as a conjugation site
for the chelators 2-mercaptoacetylglycylglycylglycine (MAG2), MAG2-PEG3400
and diethylenetriaminepentaacetic acid (DTPA). In order to confirm the nucleases
resistance of aptamer TTA1 in vivo, the previously described anti-elastase aptamer
[18] without chemically modified was used as control aptamer. The biodistribution
of aptamer TTA1radiolabeled with
99m Tc was visualized by SPECT imaging after
intravenous injection in U251 glioblastoma xenografted mice (Fig. 2.2b). Aptamer
99m Tc -MAG2-TTA1 showed extremely rapid blood clearance and deep tumor penetration. The half-life of this aptamer was <2 min, and a maximum uptake of 6%
injected dose/g was achieved in the tumor at 10 min after injection. In contrast to
the anti-TN-C antibody BC-8 which required 40 h to reach a tumor-to-blood ratio
of 5, the tumor-to-blood ratio of
99m Tc -MAG2-TTA1 reached 50 just at 3 h after
injection. Such rapid clearance can be explained by both renal and hepatic elimination pathways, and nuclease degradation. In fact, when the aptamer was completely
degraded in the blood at 3 h after injection, the radioactivity presented in the tumor
was consistent mostly to nondegraded aptamer. This study also suggested the significant contribution of radiometal chelators on the aptamer’s biodistribution. Although
three different chelators had less of an effect on tumor targeting, there was a markedly
difference on the clearance of aptamers.
Mostly aptamers using SPECT for molecular cancer imaging were selected against
proteins that are overexpressed at the surface of cancer cells. MUC1 is a large glycoprotein that provided a protective barrier for the outer surface of epithelial cells and
its overexpression has been associated with poor clinical prognosis, such as human
breast cancer and other cancers [21]. Two aptamers against the MUC1 protein core
(AptA) or the glycosylated MUC1 protein (AptB) were selected [22]. Da Pieve
et al. compared the biodistribution of radiolabeled AptA and AptB with
99m Tc
conjugated chelator MAG2b in MCF7 xenografted mice [23]. The tumor uptake
of
99m Tc -MAG2-AptB was slightly higher at 5 h after injection. But the aptamer
was completely cleared from the system at 3 h after injection, which prevents the
aptamer from adequately penetrating the tumor tissue. In another study, aptamers
targeting MUC1 were radiolabeled with
99m Tc and loaded on silica nanoparticles
(SBA-15) [24]. In the presence of stannous chloride, radiolabeling with
99m Tc was
highly efficient (the average radiolabeling yield was 99.4%) and stable up until 8 h
after radiolabeling. Nucleolin, one of the major proteins of the nucleolus, could be
translocated to the external side of the cell membrane. Overexpressed nucleolin on
the cell membrane served as a receptor for several molecules implicated in tumor
development [25]. AS1411was a 26-base guanine-rich oligonucleotide aptamer with
a high affinity to nucleolin and the first aptamer to reach phase I and II clinical
trials due to its unexpected ability to inhibit cancer cell growth [26]. Noaparast et al.
labeled a modified AS1411 with
99m Tc and evaluated it as a potential tumor-targeting