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Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2013.

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Molecular Imaging and Contrast Agent Database (MICAD) [Internet].

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99mTc-Tricarbonyl-His6-annexin A5

99mTc(CO)3-His-A5
, PhD
National Center for Biotechnology Information, NLM, NIH
Corresponding author.

Created: ; Last Update: July 19, 2012.

Chemical name:99mTc-Tricarbonyl-His6-annexin A5
Abbreviated name:99mTc(CO)3-His-A5
Synonym:
Agent category:Protein
Target:Phosphatidylserine (PS)
Target category:Phospholipid
Method of detection:Single-photon emission computed tomography (SPECT), gamma planar imaging
Source of signal\contrast:99mTc
Activation:No
Studies:
  • Checkbox In vitro
  • Checkbox Rodents
Click on protein, nucleotide (RefSeq), and gene for more information about annexin V.

Background

[PubMed]

Apoptosis (programmed cell death) plays an important role in the pathophysiology of many diseases, such as cancers, neurodegenerative disorders, vascular disorders, atherosclerosis, and chronic hepatitis, as well as in the biology of normal cells like epithelial cells and immune cells (1). During apoptosis, there is rapid redistribution of phosphatidylserine (PS) from the inner membrane leaflet to the outer membrane leaflet, exposing the anionic head group of PS. PS is also accessible for annexin V (or annexin A5) binding in necrosis because of disruption of the plasma membrane. Annexin V, a 36-kDa protein, binds to PS with high affinity (Kd = 7 nM). Annexin V derivatives have been developed for imaging apoptosis as well as necrosis with different imaging modalities such as single-photon emission computed tomography (SPECT) (2-4), positron emission tomography (5), near-infrared (NIR) fluorescence (6), and magnetic resonance imaging (7). His6-Tagged annexin A5 was radiolabeled with 99mTc using tricarbonyl chemistry (99mTc(CO)3-His-A5) for noninvasive in vivo SPECT imaging of apoptosis in mice (8, 9).

Synthesis

[PubMed]

A solution of [99mTc(H2O)3(CO)3]+ and His6-tagged annexin A5 was incubated for 90 min at 37°C (8). 99mTc(CO)3-His-A5 was isolated with column chromatography with radiochemical purity of >95% and radiochemical yields of 70%–85%. The specific activities were 192–220 MBq/nmol (5.2–6.0 mCi/nmol). 99mTc(CO)3-His-A5 was stable for up to 24 h at 4°C and 37°C in saline and cell culture medium with 10% fetal bovine serum. 99mTc(CO)3-His-A5 was also stable in in human serum for up to 4 h at 37°C.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

Vangestel et al. (8) performed cell-binding assays with 99mTc(CO)3-His-A5 in human Colo205 colon cancer cells in the absence and presence of 1 mM 5-fluorouracil (5-FU) to induce apoptosis at 4 h and 24 h after incubation. The 5-FU-treated cells exhibited 38% and 106% increases in radioactivity (P < 0.05) at 4 h and 24 h, respectively compared with untreated cells at the same time points. The 5-FU-treated cells incubated with annexin A5-FITC showed 24% apoptotic cells at 24 h, whereas only 4% apoptotic cells were shown in the untreated cells (P < 0.05).

Animal Studies

Rodents

[PubMed]

Vangestel et al. (8) performed SPECT imaging studies in nude mice (n = 5) for the first 100 min and then at 4, 8, and 22 h after injection of 37 MBq (1 mCi) 99mTc(CO)3-His-A5. The blood levels were 7% injected dose/gram (ID/g) and 1% ID/g at 10 min and 100 min after injection. The kidneys exhibited the highest radioactivity at all the time points of the studies, with the maximum level (29% ID/g) at 40 min after injection. The initial accumulation in the liver was 13% ID/g at 10 min and declined to 9% at 100 min. Low radioactivity was observed in the thyroid and stomach, suggesting the absence of 99mTc-pertechnetate in the circulation.

In another experiment, imaging studies were performed in nude mice (n = 12) bearing Colo205 tumors to measure spontaneous tumor apoptosis at 3.5 h after injection of 99mTc(CO)3-His-A5. The tumor radioactivity levels ranged from 0.15%–0.35% ID/g. Ex vivo immunohistochemical analysis of the tumor sections showed 1.4 ± 0.4% apoptotic (caspase-3-positive) cells in the tumors and correlated well with 99mTc(CO)3-His-A5 accumulation (R = 0.867, P < 0.01). No blocking and ex vivo biodistribution studies were performed.

Vangestel et al. (9) performed SPECT imaging studies in nude mice (n = 3/group) bearing Colo205 tumors to measure tumor apoptosis after treatments with buffer, 5-FU (250 mg/kg), irinotecan (100 mg/kg), oxaliplatin (30 mg/kg), anti-VEGF bevacizumab (5 mg/kg), and anti-EGFR panitumumab (6 mg/kg) at 4, 8, 12, 24, and 48 h after injection of 99mTc(CO)3-His-A5. Bevacizumab-treated mice showed no increase in accumulation of 99mTc(CO)3-His-A5, and a peak of 99mTc(CO)3-His-A5 uptake in panitumumab-treated mice was observed at 24 h after injection, as confirmed with caspase-3 immunostaining. For irinotecan-, oxaliplatin-, and bevacizumab-treated tumors, a significant correlation was observed between the 99mTc(CO)3-His-A5 accumulation and caspase-3 immunostaining (r = 0.8, P < 0.05; r = 0.9, P < 0.001; r = 0.9, P < 0.001, respectively). For 5-FU- and panitumumab-treated mice, the correlation coefficients were not significant (r = 0.7, P = 0.18; r = 0.7, P = 0.19, respectively).

Other Non-Primate Mammals

[PubMed]

No publication is currently available.

Non-Human Primates

[PubMed]

No publication is currently available.

Human Studies

[PubMed]

No publication is currently available.

References

1.
Thompson C.B. Apoptosis in the pathogenesis and treatment of disease. Science. 1995;267(5203):1456–62. [PubMed: 7878464]
2.
Blankenberg F.G. Recent advances in the imaging of programmed cell death. Curr Pharm Des. 2004;10(13):1457–67. [PubMed: 15134569]
3.
Keen H.G., Dekker B.A., Disley L., Hastings D., Lyons S., Reader A.J., Ottewell P., Watson A., Zweit J. Imaging apoptosis in vivo using 124I-annexin V and PET. Nucl Med Biol. 2005;32(4):395–402. [PubMed: 15878509]
4.
Lahorte C., Slegers G., Philippe J., Van de Wiele C., Dierckx R.A. Synthesis and in vitro evaluation of 123I-labelled human recombinant annexin V. Biomol Eng. 2001;17(2):51–3. [PubMed: 11163751]
5.
Murakami Y., Takamatsu H., Taki J., Tatsumi M., Noda A., Ichise R., Tait J.F., Nishimura S. 18F-labelled annexin V: a PET tracer for apoptosis imaging. Eur J Nucl Med Mol Imaging. 2004;31(4):469–74. [PubMed: 14666384]
6.
Schellenberger E.A., Sosnovik D., Weissleder R., Josephson L. Magneto/optical annexin V, a multimodal protein. Bioconjug Chem. 2004;15(5):1062–7. [PubMed: 15366960]
7.
van Tilborg G.A., Mulder W.J., Deckers N., Storm G., Reutelingsperger C.P., Strijkers G.J., Nicolay K. Annexin A5-functionalized bimodal lipid-based contrast agents for the detection of apoptosis. Bioconjug Chem. 2006;17(3):741–9. [PubMed: 16704213]
8.
Vangestel C., Peeters M., Oltenfreiter R., D'Asseler Y., Staelens S., Van Steenkiste M., Philippe J., Kusters D., Reutelingsperger C., Van Damme N., Van de Wiele C. In vitro and in vivo evaluation of [99mTc]-labeled tricarbonyl His-annexin A5 as an imaging agent for the detection of phosphatidylserine-expressing cells. Nucl Med Biol. 2010;37(8):965–75. [PubMed: 21055628]
9.
Vangestel C., Van de Wiele C., Mees G., Mertens K., Staelens S., Reutelingsperger C., Pauwels P., Van Damme N., Peeters M. Single-Photon Emission Computed Tomographic Imaging of the Early Time Course of Therapy-Induced Cell Death Using Technetium 99m Tricarbonyl His-annexin A5 in a Colorectal Cancer Xenograft Model. Mol Imaging. 2012;11(2):135–47. [PubMed: 22469241]

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