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Biochem Biophys Res Commun. 2016 Aug 26;477(3):483-9. doi: 10.1016/j.bbrc.2016.06.068. Epub 2016 Jun 15.

A new fluorescence/PET probe for targeting intracellular human telomerase reverse transcriptase (hTERT) using Tat peptide-conjugated IgM.

Author information

1
Department of Nuclear Medicine, Seoul National University College of Medicine, South Korea; Biomedical Sciences, Seoul National University College of Medicine, South Korea; Cancer Research Institute, Seoul National University College of Medicine, South Korea; Tumor Microenvironment Global Core Research Center, Seoul National University, South Korea.
2
Department of Nuclear Medicine, Seoul National University College of Medicine, South Korea; Cancer Research Institute, Seoul National University College of Medicine, South Korea; Tumor Microenvironment Global Core Research Center, Seoul National University, South Korea; Cancer Imaging Center, Seoul National University Hospital, Seoul, South Korea. Electronic address: hwyoun@snu.ac.kr.
3
Department of Nuclear Medicine, Seoul National University College of Medicine, South Korea; Cancer Research Institute, Seoul National University College of Medicine, South Korea.
4
Department of Nuclear Medicine, Seoul National University College of Medicine, South Korea; Biomedical Sciences, Seoul National University College of Medicine, South Korea; Cancer Research Institute, Seoul National University College of Medicine, South Korea.
5
Department of Nuclear Medicine, Seoul National University College of Medicine, South Korea; Biomedical Sciences, Seoul National University College of Medicine, South Korea; Cancer Research Institute, Seoul National University College of Medicine, South Korea; Tumor Microenvironment Global Core Research Center, Seoul National University, South Korea. Electronic address: jkchung@snu.ac.kr.

Abstract

Despite an increasing need for methods to visualize intracellular proteins in vivo, the majority of antibody-based imaging methods available can only detect membrane proteins. The human telomerase reverse transcriptase (hTERT) is an intracellular target of great interest because of its high expression in several types of cancer. In this study, we developed a new probe for hTERT using the Tat peptide. An hTERT antibody (IgG or IgM) was conjugated with the Tat peptide, a fluorescence dye and (64)Cu. HT29 (hTERT+) and U2OS (hTERT-) were used to visualize the intracellular hTERT. The hTERT was detected by RT-PCR and western blot. Fluorescence signals for hTERT were obtained by confocal microscopy, live cell imaging, and analyzed by Tissue-FAXS. In nude mice, tumors were visualized using the fluorescence imaging devices Maestro™ and PETBOX. In RT-PCR and western blot, the expression of hTERT was detected in HT29 cells, but not in U2OS cells. Fluorescence signals were clearly observed in HT29 cells and in U2OS cells after 1 h of treatment, but signals were only detected in HT29 cells after 24 h. Confocal microscopy showed that 9.65% of U2OS and 78.54% of HT29 cells had positive hTERT signals. 3D animation images showed that the probe could target intranuclear hTERT in the nucleus. In mice models, fluorescence and PET imaging showed that hTERT in HT29 tumors could be efficiently visualized. In summary, we developed a new method to visualize intracellular and intranuclear proteins both in vitro and in vivo.

KEYWORDS:

(64)Cu; IgM antibody; Intracellular and intranuclear protein; Live cell imaging; Tat peptide; hTERT

PMID:
27317485
DOI:
10.1016/j.bbrc.2016.06.068
[Indexed for MEDLINE]

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