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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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111In-Benzyl-diethylenetriaminepentaacetic acid-ZHER2:342

, PhD
National Center for Biotechnology Information, NLM, NIH
Corresponding author.

Created: ; Last Update: May 30, 2008.

Chemical name:111In-Benzyl-diethylenetriaminepentaacetic acid-ZHER2:342
Abbreviated name:111In-Benzyl-DTPA-ZHER2:342
Agent category:Antibody fragment, Affibody
Target:EGF HER2 receptor
Target category:Receptor
Method of detection:SPECT
Source of signal:111In
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Click on protein, nucleotide (RefSeq), and gene for more information about HER2.



Epidermal growth factor (EGF) is a cytokine that comprises 53 amino acids (6.2 kDa) and is secreted by ectodermic cells, monocytes, kidneys, and duodenal glands (1). EGF stimulates growth of epidermal and epithelial cells. EGF and at least seven other growth factors and their transmembrane receptor kinases play important roles in cell proliferation, survival, adhesion, migration, and differentiation. The EGF receptor (EGFR) family consists of four transmembrane receptors: EGFR (HER1/erbB-1), HER2 (erbB-2/neu), HER3 (erbB-3), and HER4 (erbB-4) (2). HER1, HER3, and HER4 comprise three major functional domains: an extracellular ligand-binding domain, a hydrophobic transmembrane domain, and a cytoplasmic tyrosine kinase domain. No ligand has been clearly identified for HER2; however, HER2 can be activated as a result of ligand binding to other HER receptors with the formation of receptor homodimers and/or heterodimers (3). HER1 and HER2 are overexpressed on many solid tumor cells such as breast, non-small cell lung, head and neck, and colon cancers (4-6). The high levels of HER1 and HER2 expression on cancer cells are associated with a poor prognosis (7-10).

Trastuzumab is a humanized IgG1 monoclonal antibody (mAb) against the extracellular domain of recombinant HER2 with an affinity constant (Kd) of 0.1 nM (11). Trastuzumab is approved for clinical use for anti-cancer therapies in both Europe and North America. 111In-Trastuzumab, Cy5.5-trastuzumab, and 68Ga-trastuzumab-F(ab')2 have been developed for imaging human breast cancer (12-16). However, the pharmacokinetics of the intact radiolabeled mAb, with high liver uptake and slow blood elimination, are generally not ideal for imaging. Smaller antibody fragments, such as Fab or F(ab´)2, have better imaging pharmacokinetics because they are rapidly excreted by the kidneys. A novel class of recombinant affinity ligands (Affibody molecules) for HER2 based on the Z-domain residues (58 amino acids) from one of the IgG-binding domains of staphylococcal protein A was constructed (17). Affibody molecules exhibit high binding affinity (Kd) to HER2 with Kd values <100 pM. Various radiolabeled Affibody molecules have been studied in terms of their ability to image HER2 in tumors [PubMed]. ZHER2:342 was labeled with 111In via isothiocyanate-benzyl-diethylenetriaminepentaacetic acid (DTPA) to form 111In-benzyl-DTPA-ZHER2:342 for single-photon emission computed tomography (SPECT) imaging in nude mice bearing human tumors (18).



Isothiocyanate-benzyl-DTPA and ZHER2:342 (1:1 molar ratio) were incubated in sodium borate buffer (pH 9) at 37°C for 20 h (18). Benzyl-DTPA-ZHER2:342 was isolated from the incubation mixture with a NAP-5 column. High-performance liquid chromatography-mass spectometry analysis revealed that 66% of benzyl-DTPA-ZHER2:342 contained one DTPA moiety, 14% had two DTPA, and 20% was unconjugated. Benzyl-DTPA-ZHER2:342 in 0.2 M ammonium acetate buffer (pH 5.25) was mixed with a predetermined amount of 111InCl3. The mixture was incubated at room temperature for 1 h with a labeling efficiency of >95%. 111In-Benzyl-DTPA-ZHER2:342 was purified with NAP-5 chromatography with a radiochemical purity of >98% with a specific activity of ~8 TBq/mmol (216 Ci/mmol).

In Vitro Studies: Testing in Cells and Tissues


Tolmachev et al. (18) performed binding experiments with In-benzyl-DTPA-ZHER2:342 using a Biacore sensor chip immobilized with extracellular domain of HER2 (HER2ECD). The Kd values of In-benzyl-DTPA-ZHER2:342 and ZHER2:342 were calculated to be 22 pM and 21 pM, respectively. In vitro binding specificity tests showed that binding of 111In-benzyl-DTPA-ZHER2:342 to SKOV-3 cells expressing HER2 was mediated by the receptors because saturation of receptors by preincubation with non-labeled ZHER2:342 significantly decreased binding of 111In-benzyl-DTPA-ZHER2:342. The cell-bound radioactivity remained at 46–52% of the initially bound activity for up to 24 h.

Animal Studies



Tolmachev et al. (18) performed biodistribution studies of 111In-benzyl-DTPA-ZHER2:342 (0.1 MBq (2.7 μCi)) in nude mice (n = 4/group) bearing SKOV-3 xenografts. The initial tracer accumulation in the SKOV-3 tumors was 12% injected dose per gram (ID/g) at 1 h, 12% ID/g at 4 h, 14% ID/g at 12 h, and 9% ID/g at 24 h after injection. The organ with the highest radioactivity at 1 h was the kidney (264% ID/g), followed by the lung (2.3% ID/g) and liver (2.0% ID/g). The radioactivity level in the tumors was higher than in other organs and tissues (except the kidneys) at 4–24 h. The radioactivity in the kidney was 248% ID/g at 24 h. Blood levels were 2.1% ID/g at 1 h, 0.12% ID/g at 4 h, 0.07% ID/g at 12 h, and 0.04% ID/g at 24 h. The elimination half-life of 111In-benzyl-DTPA-ZHER2:342 was 12.6 h. The biodistribution of 111In-benzyl-DTPA-ZHER2:342 was characterized by quick clearance of radioactivity from blood and all normal organs and tissues (except the kidneys). Tumor/blood ratios were 6, 100, 200, and 215 at 1, 4, 12, and 24 h after injection, respectively. Pre-administration of excess ZHER2:342 decreased tumor accumulation by 8.2-fold at 4 h after injection. Some inhibition was observed in the lung, liver, heart, spleen, and intestines. Positron emission tomography analysis was performed in nude mice bearing the SKOV-3 tumors after intravenous injection of 3 MBq (81 μCi) 111In-benzyl-DTPA-ZHER2:342. The tumors were clearly visualized at 4 h along with the kidneys and urinary bladder. The tumor/muscle, tumor/liver, and tumor/kidney ratios were 18, 9, and 0.1, respectively.

Other Non-Primate Mammals


No publication is currently available.

Non-Human Primates


No publication is currently available.

Human Studies


No publication is currently available.


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