Preparation and biodistribution study of 99mTc-EC-Annexin-SPIO as a tracer of radiation induced apoptosis in mice model

Document Type : Original Article


1 Department of Medical Physics, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran

2 Department of Radioisotope, Nuclear Science and Technology Research Institute, Tehran, Iran

3 Department of Radiology, Sina Hospital, Tehran University of Medical Science, Tehran, Iran


Introduction: Apoptosis is a major consequence of ionizing radiation in proliferative tissues and quantification of the apoptotic cells could be helpful for noninvasive assessment and estimation of the radiation absorbed dose. Annexin V conjugated with super paramagnetic iron oxide nanoparticles (ANX-SPIO) is a biological probe for detection of apoptotic cells using magnetic resonance imaging. This study aimed at assessing the biodistribution alterations of the labeled ANX-SPIO within the mice body shortly after exposure to different doses of ionizing radiation.
Methods: 99mTc-EC-ANX-SPIO was prepared and its in vitro stability was tested. The binding affinity of radiocomplex to apoptotic cells was validated in vitro. Mice irradiated whole body with 2, 4 and 6 Gy (60Co gamma rays) and six hours later, radiocomplex was administrated intravenously and the biodistribution study was conducted 0.5, 1 and 2 hours later.
Results: The radiochemical purity of radiocomplex was 94% ± 3.4% and it showed a good stability in PBS and serum. The radiocomplex maintained its efficacy for in vitro binding to apoptotic cells.radiocomplex accumulated in the bone marrow of all irradiated mice (p <0.05). However, statistical analysis did not show significant correlation between the %ID/g of the femoral bones and the received radiation doses.

Conclusion: Quantification of ANX-SPIO in bone marrow can be used as an indicator for radiation exposure but development and optimization of the assay are necessary for discrimination between different radiation doses.


Main Subjects

Blankenberg FG, Katsikis PD, Tait JF, Davis RE, Naumovski L, Ohtsuki K, Kopiwoda S, Abrams MJ, Strauss HW. Imaging of apoptosis (programmed cell death) with 99mTc annexin V. J Nucl Med. 1999 Jan;40(1):184-91.
Story MD, Voehringer DW, Malone CG, Hobbs ML, Meyn RE. Radiation-induced apoptosis in sensitive and resistant cells isolated from a mouse lymphoma. Int J Radiat Biol. 1994 Dec;66(6):659-68.
Lin KJ, Wu CC, Pan YH, Chen FH, Fu SY, Chiang CS, Hong JH, Lo JM. In vivo imaging of radiation-induced tissue apoptosis by (99m)Tc(I)-his (6)-annexin A5. Ann Nucl Med. 2012 Apr;26(3):272-80.
Strauss HW, Blankenberg F, Vanderheyden JL, Tait J. Translational imaging: Imaging of apoptosis.  In: München FH. Handbook of experimental pharmacology. Heidelberg: Springer-Verlag Berlin Heidelberg; 2008. p. 259-75.
Blankenberg FG. In vivo detection of apoptosis. J Nucl Med. 2008 Jun;49 Suppl 2:81S-95S.
Verhoven B, Schlegel RA, Williamson P. Mechanisms of phosphatidylserine exposure, a phagocyte recognition signal, on apoptotic T lymphocytes. J Exp Med. 1995 Nov 1;182(5):1597-601.
Blankenberg FG, Katsikis PD, Tait JF, Davis RE, Naumovski L, Ohtsuki K, Kopiwoda S, Abrams MJ, Darkes M, Robbins RC, Maecker HT, Strauss HW. In vivo detection and imaging of phosphatidylserine expression during programmed cell death. Proc Natl Acad Sci U S A. 1998 May 26;95(11):6349-54.
Zhang R, Lu W, Wen X, Huang M, Zhou M, Liang D, Li C. Annexin A5-conjugated polymeric micelles for dual SPECT and optical detection of apoptosis. J Nucl Med. 2011 Jun;52(6):958-64.
Yang SK, Attipoe S, Klausner AP, Tian R, Pan D, Rich TA, Turner TT, Steers WD, Lysiak JJ. In vivo detection of apoptotic cells in the testis using fluorescence labeled annexin V in a mouse model of testicular torsion. J Urol. 2006 Aug;176(2):830-5.
Vermes I, Haanen C, Steffens-Nakken H, Reutelingsperger C. A novel assay for apoptosis. Flow cytometric detection of phosphatidylserine expression on early apoptotic cells using fluorescein labelled Annexin V. J Immunol Methods. 1995 Jul 17;184(1):39-51.
Dash R, Chung J, Chan T, Yamada M, Barral J, Nishimura D, Yang PC, Simpson PC. A molecular MRI probe to detect treatment of cardiac apoptosis in vivo. Magn Reson Med. 2011 Oct;66(4):1152-62.
Sosnovik DE, Nahrendorf M, Weissleder R. Molecular magnetic resonance imaging in cardiovascular medicine. Circulation. 2007 Apr 17;115(15):2076-86.
Khoda Me, Utsunomiya K, Ha-Kawa S, Kanno S, Kono Y, Sawada S. An investigation of the early detection of radiation induced apoptosis by 99mTc-Annexin V and 201thallium-chloride in a lung cancer cell line. J Radiat Res. 2012;53(3):361-7.
Kurihara H, Yang DJ, Cristofanilli M, Erwin WD, Yu DF, Kohanim S, Mendez R, Kim EE. Imaging and dosimetry of 99mTc EC annexin V: preliminary clinical study targeting apoptosis in breast tumors. Appl Radiat Isot. 2008 Sep;66(9):1175-82.
Cauchon N, Langlois R, Rousseau JA, Tessier G, Cadorette J, Lecomte R, Hunting DJ, Pavan RA, Zeisler SK, van Lier JE. PET imaging of apoptosis with (64)Cu-labeled streptavidin following pretargeting of phosphatidylserine with biotinylated annexin-V. Eur J Nucl Med Mol Imaging. 2007 Feb;34(2):247-58.
Murakami Y, Takamatsu H, Taki J, Tatsumi M, Noda A, Ichise R, Tait JF, Nishimura S. 18F-labelled annexin V: a PET tracer for apoptosis imaging. Eur J Nucl Med Mol Imaging. 2004 Apr;31(4):469-74.
Lee CM, Jeong HJ, Kim EM, Kim DW, Lim ST, Kim HT, Park IK, Jeong YY, Kim JW, Sohn MH. Superparamagnetic iron oxide nanoparticles as a dual imaging probe for targeting hepatocytes in vivo. Magn Reson Med. 2009 Dec;62(6):1440-6.
Andersen LB, Frayne R. Applications of molecular imaging with MR. In: Hallgr´ımsson CWSaB, editor. Advanced imaging in biology and medicine: Springer-Verlag Berlin Heidelberg; 2009. p. 363-393.
Kettunen M, Brindle K. Apoptosis detection using magnetic resonance imaging and spectroscopy. Prog Nucl Mag Reson Spectrosc. 2005;47:175-85.
Zhao M, Beauregard DA, Loizou L, Davletov B, Brindle KM. Non-invasive detection of apoptosis using magnetic resonance imaging and a targeted contrast agent. Nat Med. 2001 Nov;7(11):1241-4.
Ahrens ET, Feili-Hariri M, Xu H, Genove G, Morel PA. Receptor-mediated endocytosis of iron-oxide particles provides efficient labeling of dendritic cells for in vivo MR imaging. Magn Reson Med. 2003 Jun;49(6):1006-13.
Wilkins RC, Kutzner BC, Truong M, Sanchez-Dardon J, McLean JR. Analysis of radiation-induced apoptosis in human lymphocytes: flow cytometry using Annexin V and propidium iodide versus the neutral comet assay. Cytometry. 2002 May 1;48(1):14-9.
Pellmar TC, Rockwell S; Radiological/Nuclear Threat Countermeasures Working Group. Priority list of research areas for radiological nuclear threat countermeasures. Radiat Res. 2005 Jan;163(1):115-23.
Thorek DL, Chen AK, Czupryna J, Tsourkas A. Superparamagnetic iron oxide nanoparticle probes for molecular imaging. Ann Biomed Eng. 2006 Jan;34(1):23-38.
Wolters SL, Corsten MF, Reutelingsperger CP, Narula J, Hofstra L. Cardiovascular molecular imaging of apoptosis. Eur J Nucl Med Mol Imaging. 2007 Jun;34 Suppl 1:S86-98.
Paris F, Fuks Z, Kang A, Capodieci P, Juan G, Ehleiter D, Haimovitz-Friedman A, Cordon-Cardo C, Kolesnick R. Endothelial apoptosis as the primary lesion initiating intestinal radiation damage in mice. Science. 2001 Jul 13;293(5528):293-7.
Liu WC, Wang SC, Tsai ML, Chen MC, Wang YC, Hong JH, McBride WH, Chiang CS. Protection against radiation-induced bone marrow and intestinal injuries by Cordyceps sinensis, a Chinese herbal medicine. Radiat Res. 2006 Dec;166(6):900-7.
Smith BR, Heverhagen J, Knopp M, Schmalbrock P, Shapiro J, Shiomi M, Moldovan NI, Ferrari M, Lee SC. Localization to atherosclerotic plaque and biodistribution of biochemically derivatized superparamagnetic iron oxide nanoparticles (SPIONs) contrast particles for magnetic resonance imaging (MRI). Biomed Microdevices. 2007 Oct;9(5):719-27.
Meyn RE, Milas L, Ang KK. The role of apoptosis in radiation oncology. Int J Radiat Biol. 2009 Feb;85(2):107-15.