Molecular imaging and pharmacokinetics of (99m) Tc-hTERT antisense oligonucleotide as a potential tumor imaging probe.

Targeting and visualization of human telomerase reverse transcriptase (hTERT) represents a promising approach for providing diagnostic value. The uptake kinetics and imaging results of (99m) Tc-hTERT antisense oligonucleotides (ASON) in hTERT-expressing cells were examined in vitro and in vivo. The pharmacokinetics and acute toxicity studies of (99m) Tc-hTERT ASON were also performed. The labeling efficiencies of radiolabeled oligonucleotide reached 76 ± 5%, the specific activity was up to 1850 kBq/µg, and the radiochemical purity was above 96%. Radioactivity accumulated to a higher concentration in hTERT-expressing cells with antisense probe than with sense control (p < 0.05). Lipid carrier incorporation significantly increased the transmembrane delivery of radiolabeled probes (p < 0.05). hTERT-expressing xenografts in nude mice were clearly visualized at 6 h postinjection of the antisense probe but not the sense control probe. However, liposome did not increase the radioactivity accumulation of probes in tumors for either antisense or sense probe (p > 0.05). Radioactivity counts per minute versus time profiles for (99m) Tc-hTERT ASON were biphasic, indicative of a three-compartment model. The pharmacokinetics parameters of half-life of distribution (T1/2α ), half-life of elimination (T1/2ß ), total apparent volume of distribution (Vd), and total rate of clearance were 2.04 ± 0.48 min, 24 ± 4.8 min, 109.83 ± 17.20 mL, and 3.19 ± 0.17 mL/min, respectively. The acute toxicity study results showed the safe application of (99m) Tc-hTERT ASON in vivo. This study provides further evidences that (99m) Tc-hTERT ASON should be developed as a safe, potential molecular image-guided diagnostic agent.

Noninvasive visualization of RNA delivery with 99mTc-radiolabeled small-interference RNA in tumor xenografts.

UNLABELLED: Small-interference RNAs (siRNAs) are short, double-strand RNA molecules that target specific messenger RNAs for degradation via the process termed RNA interference. The efficacy of RNA interference depends greatly on effective delivery of siRNA, which calls for noninvasive methods for tracing siRNA in vivo. The purpose of this work was to develop a novel (99m)Tc-radiolabeled method to visualize siRNA targeting of a tumor biomarker of human telomerase reverse transcriptase (hTERT) in HepG2 tumor xenografts. METHODS: After conjugation with S-acetyl N-hydroxysuccinimide-mercaptoacetyltriglycine (NHS-MAG3), antisense RNA with 2'-O-methyl modification was annealed with sense strand to form a duplex and then radiolabeled with (99m)Tc. (99m)Tc-siRNAs were tested for stability in serum by measurement of radiochemical purity and for inhibitory activity by reverse-transcriptase polymerase chain reaction and Western blotting. In vitro cellular uptake was evaluated in HepG2 cells. Biodistribution studies and static imaging were performed in HepG2 tumor-bearing mice. RESULTS: Radiochemical purity remained highly stable in saline and fresh human serum at room temperature and 37 degrees C. Radiolabeled siRNA demonstrated strong inhibitory effects similar to those of unlabeled siRNA on both hTERT messenger RNA and protein in vitro. (99m)Tc-hTERT siRNA showed more uptake than did control siRNA in HepG2 cells after 1 h of incubation. After administration in HepG2 tumor-bearing mice, (99m)Tc-hTERT siRNA had significantly higher accumulation in tumors and a higher tumor-to-blood ratio than did control siRNA (P < 0.05). Scintigraphy of (99m)Tc-hTERT siRNA showed clear tumor images at 0.5, 1, 3, and 6 h after injection. In contrast, (99m)Tc-control siRNA failed to visualize the tumor. Ratios of uptake in tumor to uptake in contralateral region of hTERT-targeted siRNA were significantly higher than those of control siRNA (P < 0.05) at each time point. CONCLUSION: The (99m)Tc radiolabeling method with NHS-MAG3 chelator can be used successfully in siRNA radiolabeling, allowing for the noninvasive visualization of siRNA delivery in vivo.