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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-Diethylenetriaminepentaacetic acid-trastuzumab

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

Created: ; Last Update: June 4, 2008.

Chemical name:111In-Diethylenetriaminepentaacetic acid-trastuzumab
Abbreviated name:111In-DTPA-humAb4D5-8
Synonym:111In-Herceptin®, 111In-anti-HER2 MAb, 111In-DTPA-trastuzumab.
Agent Category:Antibody
Target:Human epidermal growth factor receptor 2 (HER2)
Target Category:Antibody to antigen binding
Method of detection:Single-photon emission computed tomography (SPECT), planar imaging
Source of signal:111In
  • Checkbox In vitro
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  • Checkbox Humans
Click on protein, nucleotide (RefSeq), and gene for more information about HER2.



111In-Diethylenetriaminepentaacetic acid-trastuzumab (111In-DTPA-humAb4D5-8), which is formed by the conjugation of 111In with an intact recombinant humanized monoclonal antibody (MAb) against the antigen of human epidermal growth factor receptor 2 (HER2), has been developed for imaging of human breast cancer (1).

The epidermal growth factor receptors (HER/erbB) with tyrosine kinase activity are involved in transmission of signals controlling normal cell development, growth, differentiation, and survival (1-3). This cell-surface receptor family consists of four distinct members, the epidermal growth factor receptors HER1, HER2, HER3, and HER4. These receptors are found in various combinations in different tissues. There are nine ligands that bind directly to HER1, HER3, or HER4. HER2 can be activated as a result of ligand binding to other HER receptors. The ligandless HER2 receptor is encoded by the human gene HER2/c-erbB2 (HER2/neu). It has been found to be overexpressed in a wide variety of human cancers and in 20-30% of primary breast cancers (4). This overexpression is associated with aggressive tumor growth and metastatic activity. In breast cancer, HER2/neu gene amplification can result in 10- to 100-fold higher HER2 protein levels in tumor cells than that in normal breast epithelium. HER2 overexpression is considered a negative prognostic indicator for breast cancer.

Radiolabeled MAbs have been developed for both the diagnosis and treatment of tumors (5-7). Anti-HER2 MAbs have been generated for inhibiting the growth of tumor cells that possess activated HER2/neu receptors (8). Murine 4D5 MAb was raised against human breast and ovarian tumor cell lines overexpressing p185HER2. This MAb was found to recognize an extracellular epitope (amino acids 529 to 627) in the cysteine-rich II domain that resides very close to the transmembrane region (8). Murine 4D5 MAb was fully humanized by Carter et al. (9) to generate the humAb4D5-8 MAb in an effort to improve the safety and tolerance of the antibody for clinical applications. Trastuzumab is of the IgG1 kappa class and binds to the recombinant HER2 extracellular domain with an affinity constant (Kd) of 0.1 nM. Cardiotoxicity is the most serious complication of using trastuzumab in humans (4, 10). One potential application of a radiolabeled anti-HER2 MAb is the pretreatment imaging of breast cancer patients to predict the cardiotoxicity and therapeutic efficacy of unlabeled trastuzumab.

Various anti-HER2 MAbs have been labeled with 131I, 125I, 111In, 86Y, 90Y, 64Cu, 76Br, or Gd for potential imaging and therapeutic applications (11-18). The labeling with radiometals is generally achieved by indirect labeling with use of a bifunctional chelating agent (19, 20). 111In, a photon emitter with a physical half-life (t½) of 2.8 days, is considered one of the radionuclides of choice for antibody imaging studies.



Trastuzumab was synthesized by humanization of murine 4D5 MAb (9). In this procedure, the complementarity-determining regions of the murine 4D5 MAb were transplanted into a human antibody model. A “gene conversion mutagenesis” strategy was used to simultaneously humanize the heavy- and light-chain region genes. A final “humanized” antibody, humAb4D5, containing only the antigen-binding loops from murine 4D5 MAb and human variable region framework residues plus IgG1 constant domains, was constructed. Of 8 humanized variants, humAb4D5-8 was found to bind its antigen 3-fold more tightly than the murine 4D5 MAb.

Lub-de Hooge et al. (1) described radiolabeling of the intact humAb4D5-8 MAb with 111In by conjugation using a modified cyclic diethylenetriamine pentaacetic acid (DTPA) anhydride method (1, 20). The initial DTPA conjugation was performed at a 1:1 molar ratio with 10 mg of humAb4D5-8 MAb in 1.4% sodium bicarbonate (pH 8, 200 µl) (1). Unbound DTPA was removed by ultrafiltration (twice for 30 min at 2,684g). The purified immunoconjugate, DTPA-humAb4D5-8 MAb, was either stored at −20 °C or used immediately for radiolabeling. Radiolabeled indium chloride (111InCl3; pH 1.5-1.9) solution was first buffered with an equal volume of 0.1 M sodium acetate to achieve a final pH of 5.5. The buffered 111InCl3 was then added to the DTPA-humAb4D5-8 MAb, and the reaction mixture was incubated at room temperature for 5 min. The labeling yield was 92.3 ± 2.3% (n = 4), and the radiochemical purity was 97.0 ± 1.5%. The specific activity of 111In-DTPA-trastuzumab was 18.5 MBq (0.5 mCi)/mg (2.8 GBq (0.075 Ci)/μmol based on a molecular weight of 150,000 for the intact MAb).

In Vitro Studies: Testing in Cells and Tissues


Lu-de Hooge et al. (1) determined the radioimmunoreactivity of 111In-DTPA-trastuzumab in cell-binding assays using HER2-overexpressing SK-OV-3 (human ovarian cancer) and SK-BR-3 (human breast cancer) cells. Both assays showed that the immunoreactive fraction was 0.87 ± 0.06 (n = 3). The nonspecific binding was less than 3%. An internalization assay with the SK-OV-3 cells showed that internalization was 25% after 1 h and 45% after 4 h.

Animal Studies



Lu-de Hooge et al. (1) studied the biodistribution of 111In-DTPA-trastuzumab in mice bearing HER2-positive SK-OV-3 or HER2-negative GLC4 tumors. Each mouse with a 0.5-0.8 cm diameter s.c. tumor received an i.v. dose of 25 μg (0.2 ml) of MAb protein (450 ± 25 kBq (12.2 ± 0.68 μCi)). The tumor radioactivity uptakes (percentage injected dose (% ID)/g; n = 3-7) in mice with HER2-positive tumors were 9.77 ± 1.14, 13.71 ± 0.57, 16.30 ± 0.64, 15.29 ± 1.57, 8.31 ± 1.66, and 9.80 ± 3.22 at 5, 24, 48, 72, 96, and 168 h after injection, respectively. The tumor was best demonstrated in gamma imaging at 72 h after injection. The tumor radioactivity uptakes (% ID/g; n = 3-6) in the mice with HER2-negative tumors ranged from 3.17 ± 1.19 at 72 h to 5.70 ± 1.11 at 96 h. In the mice with HER2-positive tumors, the blood radioactivities (% ID/g) were 12.75 ± 1.50, 8.76 ± 0.46, 7.15 ± 0.69, 2.33 ± 0.46, 1.36 ± 0.51, and 1.63 ± 0.83 at 5, 24, 48, 72, 96, and 168 h after injection, respectively. At 5 h, the radioactivity uptakes (% ID/g) in well-perfused organs were 13.97 ± 2.52, 8.40 ± 1.67, 6.46 ± 0.47, 3.87 ± 1.00, and 3.83 ± 0.15 for the liver, spleen, kidney, lung, and heart, respectively. At 24 h, these values were 7.89 ± 1.80, 2.68 ± 0.48, 5.42 ± 0.58, 4.09 ± 0.69, and 1.90 ± 0.05% ID/g, respectively. No radioactivity was found in the brain, and the pancreas, stomach, bowel, bone, and skeletal muscle all demonstrated low uptake. A similar pattern of distribution was seen in mice bearing HER2-negative tumors. Metastases in the abdominal cavities of two mice with HER2-positive tumors also showed 15.4 and 11.2% ID/g at 24 and 96 h, respectively.

Other Non-Primate Mammals


No publication is currently available.

Non-Human Primates


No publication is currently available.

Human Studies


Behr et al. (10) performed a preliminary single-photon emission computed tomography (SPECT) imaging study with 111In-DTPA-trastuzumab in 20 patients with metastatic breast cancer that expressed HER2. The radioactive tracer dose was coadministered with a loading dose of 4 mg of unlabeled trastuzumab. Subsequently, all patients were treated with unlabeled trastuzumab either as monotherapy (n = 4) or in combination with other chemotherapeutic agents. They reported that 7 of the 20 patients showed evidence of myocardial uptake. Six of these 7 patients reported cardiotoxicity after trastuzumab treatment, and the 7th patient suffered episodes of cardiac arrhythmia during trastuzumab administration. In contrast, the other 13 patients without myocardial uptake of 111In-DTPA-trastuzumab had reported no adverse cardiac effects. All 11 patients who showed intense tumor uptake of 111In-DTPA-trastuzumab had objective responses to trastuzumab therapy. The authors suggested that pretreatment imaging with 111In-DTPA-trastuzumab could predict the cardiotoxicity and therapeutic efficacy of unlabeled trastuzumab.


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