Noninvasive imaging of human telomerase reverse transcriptase (hTERT) messenger RNA with 99mTc-radiolabeled antisense probes in malignant tumors.
The expression of human telomerase reverse transcriptase (hTERT) is present in most malignant cells (>85%) but is undetectable in most normal somatic cells. Visualization of hTERT expression using radionuclide targeting can provide important diagnostic information in malignant tumors. The overall aim of this study was to evaluate whether (99m)Tc-radiolabeled antisense oligonucleotide (ASON) targeting hTERT messenger RNA (mRNA) can be used for imaging of hTERT expression in vivo.


METHODS
One 18-mer antisense or sense uniformly phosphorothioate-modified oligonucleotide targeting hTERT mRNA was radiolabeled with (99m)Tc through the bifunctional chelator N-hydroxysuccinimidyl derivative of S-acetylmercaptoacetyltriglycine (S-acetyl NHS-MAG3). Then the radiolabeled probe was characterized in vitro. Reverse transcription-polymerase chain reaction (RT-PCR) was performed to assay the mRNA level after proliferating cells had been incubated with the antisense and sense probes. (99m)Tc-MAG3-ASON or (99m)Tc-MAG3-SON was injected intravenously in mammary tumor-bearing BALB/c nude mice. Biodistribution and in vivo imaging was performed periodically. All data were analyzed by statistical software.


RESULTS
The labeling efficiencies of (99m)Tc-MAG3-ASON/SON reached 76% +/- 5% (n = 5) within 15-30 min at room temperature, the specific activity was up to 1,850 kBq/mug, and the radiochemical purity was >96% after purification. (99m)Tc-MAG3-ASON showed complete stability at room temperature and in fresh 37 degrees C human serum. In comparison with (99m)Tc-MAG3-SON, (99m)Tc-MAG3-ASON preserved the capacity to bind living hTERT-expressing cells specifically and to inhibit the expression of hTERT mRNA significantly as well as ASON. In nude mice bearing hTERT-expressing MCF-7 xenografts, tumor radioactivity uptake of (99m)Tc-MAG3-ASON after injection was significantly higher than that of (99m)Tc-MAG3-SON after injection (P < 0.05). The hTERT-expressing xenografts were clearly imaged at 4-8 h noninvasively after injection of (99m)Tc-MAG3-ASON, whereas the xenografts were not imaged at any time after injection of (99m)Tc-MAG3-SON.


CONCLUSIONS
This in vivo study provides evidence that (99m)Tc-MAG3-ASON targeting hTERT mRNA can be used as a potential candidate for visualization of hTERT expression in carcinomas.
Liu
Meng
M
Department of Nuclear Medicine, Peking University First Hospital, West District, Beijing, China.
Wang
Rong Fu
RF
Zhang
Chun Li
CL
Yan
Ping
P
Yu
Ming Ming
MM
Di
Li Juan
LJ
Liu
Hong Jie
HJ
Guo
Feng Qin
FQ
D000818:Animals; D045744:Cell Line, Tumor; D004355:Drug Stability; D005260:Female; D006801:Humans; D007553:Isotope Labeling; D051379:Mice; D008807:Mice, Inbred BALB C; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000201:enzymology* / Q000628:therapy; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger / Q000032:analysis*; D011877:Radionuclide Imaging; D019275:Radiopharmaceuticals; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017263:Technetium Tc 99m Mertiatide / Q000493:pharmacokinetics; D019098:Telomerase / Q000037:antagonists & inhibitors* / Q000235:genetics*; D014018:Tissue Distribution; D014183:Transplantation, Heterologous
D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D019275:Radiopharmaceuticals; D017263:Technetium Tc 99m Mertiatide; C509186:TERT protein, human; D019098:Telomerase