Molecular imaging of MDM2 messenger RNA with 99mTc-labeled antisense oligonucleotides in experimental human breast cancer xenografts

Abstract

The mouse double-minute 2 (MDM2) oncogene, amplified or overexpressed in many human cancers, has been suggested to be a novel target for cancer therapy. Visualization of MDM2 expression using radionuclide targeting can provide important diagnostic information in malignant tumors. The overall aim of this study was to evaluate whether liposome-coated (99m)Tc-radiolabeled antisense oligonucleotides (ASONs) targeting MDM2 messenger RNA (mRNA) could be used for imaging of MDM2 expression in vivo.

Methods: ASON and mismatch oligonucleotide (ASONM) targeted to MDM2 mRNA were synthesized and radiolabeled with (99m)Tc using the bifunctional chelator hydrazinonicotinamide (HYNIC). Then the radiolabeled probe was characterized in vitro. Reverse-transcriptase polymerase chain reaction and Western blotting were performed to assay the MDM2 mRNA and protein level after MCF-7 (human breast adenocarcinoma cell line) cells were incubated with liposome-coated (99m)Tc-HYNIC-ASON/ASONM at various concentrations for 24 h. Following established MCF-7-bearing nude mice models, the biodistribution of liposome-coated (99m)Tc-HYNIC-ASON/ASONM was investigated, and in vivo tumor scintigraphic images were acquired for these animal models. All data were analyzed by statistical software.

Results: The labeling efficiencies of (99m)Tc-HYNIC-ASON and (99m)Tc-HYNIC-ASONM were 57.2% ± 2.98% and 56.3% ± 3.01%, respectively; the specific activities were 1,450 ± 60.2 and 1,370 ± 55.4 kBq/μg, respectively; and the radiochemical purity for both was above 95%. The radiolabeled ASON still had the ability to hybridize to the sense oligonucleotide. In comparison with the mismatch probe, the antisense probe had an obvious effect on the levels of MDM2 mRNA and protein. The levels of mRNA and protein were significantly different for different concentration antisense probe groups (P < 0.01). The excretion of the antisense and mismatch probe was mainly through the liver and kidneys. The tumor radioactivity uptake of the antisense probe was significantly higher than that of the mismatch probe (P < 0.01). At 1-10 h after injection of the antisense probe, the tumor could be clearly visualized, whereas the tumors were not imaged at any time after injection of the mismatch probe.

Conclusion: The accumulation of liposome-coated (99m)Tc-labeled ASONs in breast cancer tissue is specific. The antisense imaging with liposome-coated (99m)Tc-HYNIC-ASON may be a promising method for visualization of MDM2 expression in human breast cancer.