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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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Avidin-gadolinium

Avidin-Gd
, PhD
National Center for Biotechnology Information, NLM, NIH, Bethesda, MD 20894, vog.hin.mln.ibcn@dacim

Created: ; Last Update: December 26, 2008.

Chemical name:Avidin-gadolinium
Abbreviated name:Avidin-Gd
Synonym:
Agent Category:Protein
Target:Biotinylated anti-HER-2/neu antibody
Target Category:Protein–protein binding
Method of detection:Magnetic resonance imaging (MRI)
Source of Signal/Contrast:Gadolinium
Activation:No
Studies:
  • Checkbox In vitro
  • Checkbox Rodents
No information is available.

Background

[PubMed]

Biotin is a protein found in mammalian cells and is considered to be a vitamin because it has an important function in the cellular metabolism of lipids and carbohydrates. Avidin is a protein that is found primarily in the egg white or in bacteria (known as streptavidin if isolated from bacteria; Streptomyces avidinii is the source of this protein). Although the two proteins have different origins, they are known to have a very strong non-covalent interaction (discovered almost seven decades ago) with a dissociation constant (Kd) of 4 × 10-14 M (1, 2). The avidin–biotin interaction has been described in detail by Wilchek et al. (3). Only harsh conditions (e.g., treatment with formaldehyde at high temperature) are known to break the avidin/streptavidin–biotin bond, and the released proteins cannot be reused for any other work due to deterioration of the samples (4). Because of their strong interaction, the avidin/streptavidin–biotin system has been extensively studied and is widely used after conjugation of either protein to other proteins, nucleic acids, or other matrices for research and diagnostic purposes, including imaging techniques (5-8).

Because of its overexpression, the human epidermal growth factor receptor 2 (HER2/neu) is an important target for the diagnosis, treatment, and prognosis of cancers such as those of the breast, lungs, etc (9, 10). As a result, humanized monoclonal antibodies (MAbs) labeled with different nuclides or imaging and contrast agents targeted toward HER2/neu have been used for the in vitro and in vivo imaging of cancers (11-13). Some humanized MAbs against HER2/neu alone or their conjugated derivatives are used, or are in development, as immunotherapeutic agents for the treatment of this ailment (14, 15). The avidin–biotin system has been used to visualize HER2/neu-positive tumors in transgenic mice by some investigators with magnetic resonance imaging (MRI) (16, 17). To achieve this, the investigators prelabeled the cells expressing HER2/neu, under in vitro and in vivo conditions, with a commercially available biotinylated mouse MAb. The biotinylated MAb was allowed to wash or clear out from the respective test systems, and this was followed by the administration of avidin conjugated to gadolinium (avidin-Gd) to the cells or animals. MRI was subsequently performed, and it was demonstrated that a high positive T1 contrast could be achieved under the in vitro and in vivo conditions used.

Synthesis

[PubMed]

The synthesis of avidin-Gd was performed as described by Artemov et al. using a modified procedure of Hnatowich et al. (13, 18). Briefly, 20-fold excess cyclic anhydride of diethylenetriaminepentaacetic acid (DTPA) was allowed to react with avidin for 24 h at 4°C. Excess low molecular weight reagents from the conjugated avidin (avidin-DTPA) were removed by centrifuge filtration through a device with a 10K cut-off limit. The avidin-DTPA was then reacted with Gd-(1-(2-naphthoyl)-3,3,3-trifluoroacetonate)2 to obtain avidin-GdDTPA (avidin-GdDTPA) as described above. Avidin-GdDTPA was separated from excess low molecular weight compounds as detailed above. On average, 12.5 GdDTPA groups were reported to be attached to each avidin molecule.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

For in vitro studies, NT-5 cells expressing HER-2/neu were derived from spontaneous mammary tumors of female HER-2/neu transgenic mice (13). In the same study, mouse carcinoma EMT-6 cells that do not express HER-2/neu were used as controls. Expression of HER-2/neu by the two cell lines was confirmed with fluorescence-activated cell-sorting analysis using appropriate reagents (13). The cells were exposed to biotinylated anti-HER-2/neu MAb in phosphate-buffered saline (PBS) containing bovine serum albumin (BSA) for 30 min at room temperature. The cells were then washed with PBS several times and incubated with avidin-GdDTPA for 5 min at room temperature. Control cells were exposed to a non-specific MAb or to avidin-GdDTPA. The cells were washed again as detailed above and fixed in 2% paraformaldehyde in PBS in a nuclear magnetic resonance tube. Subsequently, T1-weighted images (repetition time, TR = 1 s; spin echo time, TE = 8 ms) were obtained with an MRI probe. Only the NT-5 cells that were exposed to the biotinylated anti-HER-2/neu MAb showed a positive T1 contrast with avidin-GdDTPA, and the EMT-6 cells had a low background signal.

Animal Studies

Rodents

[PubMed]

In vivo MRI studies were performed with severe combined immunodeficient mice bearing NT-5 and EMT-6 cell tumors (13). All experimental animals (n = 5/time point) were treated with the biotinylated anti-HER-2/neu antibodies, and the control animals bearing NT-5 tumors (n = 3) were treated with BSA in saline. The test animals were injected with the avidin-GdDTPA conjugate 12 h after the biotinylated MAb treatment. Control animals were given avidin-GdDTPA via intravenous injection. Parameters for the T1-weighted images were as follow: TR = 300 ms, TE = 15 ms, flip angle = 90° with a field view of 32 mm; 16 slice images were acquired over 15 min with a slice thickness of 2 mm. MRI images were obtained at 1, 8, 24, and 48 h after contrast treatment. The NT-5 tumors were reported to retain the contrast for 8–24 h after treatment, but the contrast decreased in the EMT-6 cell tumors 1 h after treatment. Also, the NT-5 tumors that were not pre-treated with the biotinylated MAb were reported to show no retention of the contrast agent. A statistically significant (P < 0.05) increase in signal was observed in the tumors treated with NT-5 (~1.7 relative units for the cells treated with NT-5 versus ~1 relative unit for the NT-5 control and EMT-6 cells) under these experimental conditions.

Other Non-Primate Mammals

[PubMed]

No references are currently available.

Non-Human Primates

[PubMed]

No references are currently available.

Human Studies

[PubMed]

No references are currently available.

Supplemental Information

[Disclaimers]

NIH Support

Supported by NIH grant P20 CA86346

References

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Green N.M. Avidin and streptavidin. Methods Enzymol. 1990;184:51–67. [PubMed: 2388586]
2.
Holmberg A. , Blomstergren A. , Nord O. , Lukacs M. , Lundeberg J. , Uhlen M. The biotin-streptavidin interaction can be reversibly broken using water at elevated temperatures. Electrophoresis. 2005;26(3):501–10. [PubMed: 15690449]
3.
Wilchek M. , Bayer E.A. , Livnah O. Essentials of biorecognition: the (strept)avidin-biotin system as a model for protein-protein and protein-ligand interaction. Immunol Lett. 2006;103(1):27–32. [PubMed: 16325268]
4.
Tong X. , Smith L.M. Solid-phase method for the purification of DNA sequencing reactions. Anal Chem. 1992;64(22):2672–7. [PubMed: 1294003]
5.
Hama Y. , Urano Y. , Koyama Y. , Choyke P.L. , Kobayashi H. Activatable fluorescent molecular imaging of peritoneal metastases following pretargeting with a biotinylated monoclonal antibody. Cancer Res. 2007;67(8):3809–17. [PubMed: 17440095]
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7.
Xu H. , Regino C.A. , Koyama Y. , Hama Y. , Gunn A.J. , Bernardo M. , Kobayashi H. , Choyke P.L. , Brechbiel M.W. Preparation and preliminary evaluation of a biotin-targeted, lectin-targeted dendrimer-based probe for dual-modality magnetic resonance and fluorescence imaging. Bioconjug Chem. 2007;18(5):1474–82. [PubMed: 17711320]
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Kanehira K. , Hu J. , Pier T. , Sebree L. , Huang W. High endogenous avidin binding activity: an inexpensive and readily available marker for the differential diagnosis of kidney neoplasms. Int J Clin Exp Pathol. 2008;1(5):435–9. [PMC free article: PMC2480577] [PubMed: 18787626]
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Stipsanelli E. , Valsamaki P. Monoclonal antibodies: old and new trends in breast cancer imaging and therapeutic approach. Hell J Nucl Med. 2005;8(2):103–8. [PubMed: 16142251]
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Barrett T. , Koyama Y. , Hama Y. , Ravizzini G. , Shin I.S. , Jang B.S. , Paik C.H. , Urano Y. , Choyke P.L. , Kobayashi H. In vivo diagnosis of epidermal growth factor receptor expression using molecular imaging with a cocktail of optically labeled monoclonal antibodies. Clin Cancer Res. 2007;13(22 Pt 1):6639–48. [PubMed: 17982120]
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Gee M.S. , Upadhyay R. , Bergquist H. , Alencar H. , Reynolds F. , Maricevich M. , Weissleder R. , Josephson L. , Mahmood U. Human breast cancer tumor models: molecular imaging of drug susceptibility and dosing during HER2/neu-targeted therapy. Radiology. 2008;248(3):925–35. [PMC free article: PMC2798096] [PubMed: 18647846]
12.
Sampath L. , Kwon S. , Ke S. , Wang W. , Schiff R. , Mawad M.E. , Sevick-Muraca E.M. Dual-labeled trastuzumab-based imaging agent for the detection of human epidermal growth factor receptor 2 overexpression in breast cancer. J Nucl Med. 2007;48(9):1501–10. [PubMed: 17785729]
13.
Artemov D. , Mori N. , Okollie B. , Bhujwalla Z.M. MR molecular imaging of the Her-2/neu receptor in breast cancer cells using targeted iron oxide nanoparticles. Magn Reson Med. 2003;49(3):403–8. [PubMed: 12594741]
14.
Lewis Phillips G.D. , Li G. , Dugger D.L. , Crocker L.M. , Parsons K.L. , Mai E. , Blattler W.A. , Lambert J.M. , Chari R.V. , Lutz R.J. , Wong W.L. , Jacobson F.S. , Koeppen H. , Schwall R.H. , Kenkare-Mitra S.R. , Spencer S.D. , Sliwkowski M.X. Targeting HER2-positive breast cancer with trastuzumab-DM1, an antibody-cytotoxic drug conjugate. Cancer Res. 2008;68(22):9280–90. [PubMed: 19010901]
15.
Nahta R. , Esteva F.J. Trastuzumab: triumphs and tribulations. Oncogene. 2007;26(25):3637–43. [PubMed: 17530017]
16.
Artemov D. , Mori N. , Ravi R. , Bhujwalla Z.M. Magnetic resonance molecular imaging of the HER-2/neu receptor. Cancer Res. 2003;63(11):2723–7. [PubMed: 12782573]
17.
Reilly R.T. , Gottlieb M.B. , Ercolini A.M. , Machiels J.P. , Kane C.E. , Okoye F.I. , Muller W.J. , Dixon K.H. , Jaffee E.M. HER-2/neu is a tumor rejection target in tolerized HER-2/neu transgenic mice. Cancer Res. 2000;60(13):3569–76. [PubMed: 10910070]
18.
Hnatowich D.J. , Virzi F. , Rusckowski M. Investigations of avidin and biotin for imaging applications. J Nucl Med. 1987;28(8):1294–302. [PubMed: 3612292]

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