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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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124I-Labeled anti-HER2-specific C6.5 diabody

124I-C6.5db
, PhD
National Center for Biotechnology Information, NLM, NIH, Bethesda, MD
Corresponding author.

Created: ; Last Update: August 4, 2011.

Chemical name:124I-Labeled anti-HER2-specific C6.5 diabody
Abbreviated name:124I-C6.5db
Synonym:
Agent category:Diabody, antibody fragment
Target:Epidermal growth factor receptor (EGFR, HER2)
Target category:Receptor
Method of detection:Positron emission tomography (PET)
Source of signal:124I
Activation:No
Studies:
  • Checkbox In vitro
  • Checkbox Rodents
Click on protein, nucleotide (RefSeq), and gene for more information about HER2.

Background

[PubMed]

Epidermal growth factor (EGF) is a cytokine of 53 amino acids (6.2 kDa) that 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, including 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 as well as 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).

Single-chain variable fragments (scFvs) of antibodies with a molecular mass of 25 kDa are cleared very rapidly from the circulation, but they exhibit poor tumor retention because they have a lower affinity than the parent antibody (11). On the other hand, bivalent antibody fragments possess better tumor-targeting characteristics, including rapid tissue penetration, high target retention, and rapid blood clearance. The diabody (db) fragment (a dimer of scFvs; molecular mass, 52–55 kDa) has been evaluated for targeting in several tumor antigen systems and has demonstrated rapid tumor localization and high-contrast imaging (11, 12). In particular, C6.5db was constructed using the human anti-HER2 C6.5 scFv and radiolabeled with 124/125I for in vivo imaging of HER2 expression in tumors (13). C6.5db binds to an epitope of HER2 distinct from that bound by trastuzumab.

Synthesis

[PubMed]

C6.5db (11.1 nmol) was incubated with 370 MBq (10 mCi) Na124I and Iodogen-coated glass beads for 5 min at room temperature (13). 124I-C6.5db was isolated with size-exclusion column chromatography, with a radiochemical purity of >93.5% and specific activities of 5–15.5 MBq/nmol (0.14–0.42 mCi/nmol). 124I-C6.5db exhibited immunoreactivity of 71%–73%. C6.5db and C6.5 scFv were also radiolabeled with Na125I in the presence of chloramine T with immunoreactivity values of 87.6% and 65.3%, respectively. The specific activities of 125I-C6.5db and 125I-C6.5 scFv were not reported.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

Adams et al. (13) performed binding experiments with C6.5db and C6.5 scFv using a Biacore sensor chip immobilized with extracellular domain of HER2 (4,200 sites/µm2). The dissociation constants (Kd) of C6.5db and C6.5 scFv were calculated to be 0.4 nM and 16 nM, respectively. C6.5db and C6.5 scFv exhibited retention half-life values (t½) of 43 min and 2 min, respectively. In vitro binding of biotinylated C6.5db and C6.5 scFv to SKOV-3 cells expressing HER2 (3,200–4,800 sites/µm2 on the cell surface) was determined with flow cytometry. C6.5db and C6.5 scFv exhibited t½ values of 5 h and 3 min, respectively. Reddy et al. (14) showed that C6.5db bound to an epitope on HER2 distinct from that bound by trastuzumab.

Animal Studies

Rodents

[PubMed]

Adams et al. (13) performed ex vivo biodistribution studies of 125I-C6.5db (0.13 nmol) in severe combined immunodeficient (SCID) mice (n = 5/group) bearing SKOV-3 xenografts at 1, 4, 24, 48, and 72 h after injection. 125I-C6.5db exhibited a rapid equilibration phase (t½α = 0.67 h) and a slower elimination phase from circulation (t½β = 6.42 h). Accumulation values in the tumors were 6.9% injected dose/gram (ID/g), 10.1% ID/g, 6.5% ID/g, 2.4% ID/g, and 1.4% ID/g at 1, 4, 24, 48, and 72 h, respectively. Radioactivity values in the blood were 21.5% ID/ml, 6.7% ID/ml, 0.7% ID/ml, 0.1% ID/ml, and <0.1% ID/ml at these time points, respectively. Accumulation in normal tissues was less than that in the tumor at 24 h and later. As a comparison, 125I-C6.5 scFv was performed at 24 h after injection. Accumulation values were 1.0% ID/g, 0.04% ID/g, and 0.05% ID/ml in the tumor, liver, and blood, respectively. Reddy et al. (14) performed ex vivo biodistribution studies of 125I-C6.5db in SCID mice (n = 5/group) bearing tumors with different HER2 densities: SKOV-3 (1.0 × 106/cell), MDA-MB361/DYT2 (3.7 × 105/cell), and MDA-MB231 (2.3 × 104/cell) with accumulation values of 1.91% ID/g, 1.11% ID/g, and 0.31% ID/g at 24 h after injection, respectively. 125I-C6.5db tumor accumulation correlated with the HER2 tumor density in tumors with high, moderate, and low HER2 expression. No blocking studies were performed.

Positron emission tomography (PET) imaging was performed in SCID mice (n = 7/group) bearing SKOV-3 tumors at 4, 8, 24, and 48 h after injection of 124I-C6.5db (14). The tumors were clearly visualized at 4 h after injection. There was a time-dependent increase in tumor/background signal. The stomach and thyroid were clearly visualized at 24 h and 48 h, respectively. Tumor accumulation (% ID/g) was quantitated with PET and ex vivo necropsy-based analysis: 9.8 ± 0.4 versus 11.3 ± 1.4 at 4 h, 9.4 ± 0.9 versus 10.3 ± 2.3 at 8 h, 5.7 ± 0.7 versus 6.1 ± 2.1 at 24 h, and 3.0 ± 0.1 versus 3.2 ± 0.6 at 48 h. The PET-based quantitation showed a strong correlation with the necropsy-based quantitation (R2 = 0.96). No blocking studies were performed.

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.

NIH Support

CA 65559, CA06927, CA09035

References

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Carpenter G., Cohen S. Epidermal growth factor. J Biol Chem. 1990;265(14):7709–12. [PubMed: 2186024]
2.
Yarden Y. The EGFR family and its ligands in human cancer. signalling mechanisms and therapeutic opportunities. Eur J Cancer. 2001;37 Suppl 4:S3–8. [PubMed: 11597398]
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Rubin I., Yarden Y. The basic biology of HER2. Ann Oncol. 2001;12 Suppl 1:S3–8. [PubMed: 11521719]
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Grunwald V., Hidalgo M. Developing inhibitors of the epidermal growth factor receptor for cancer treatment. J Natl Cancer Inst. 2003;95(12):851–67. [PubMed: 12813169]
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Mendelsohn J. Anti-epidermal growth factor receptor monoclonal antibodies as potential anti-cancer agents. J Steroid Biochem Mol Biol. 1990;37(6):889–92. [PubMed: 2285602]
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Yasui W., Sumiyoshi H., Hata J., Kameda T., Ochiai A., Ito H., Tahara E. Expression of epidermal growth factor receptor in human gastric and colonic carcinomas. Cancer Res. 1988;48(1):137–41. [PubMed: 2446740]
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Ang K.K., Berkey B.A., Tu X., Zhang H.Z., Katz R., Hammond E.H., Fu K.K., Milas L. Impact of epidermal growth factor receptor expression on survival and pattern of relapse in patients with advanced head and neck carcinoma. Cancer Res. 2002;62(24):7350–6. [PubMed: 12499279]
8.
Costa S., Stamm H., Almendral A., Ludwig H., Wyss R., Fabbro D., Ernst A., Takahashi A., Eppenberger U. Predictive value of EGF receptor in breast cancer. Lancet. 1988;2(8622):1258. [PubMed: 2903994]
9.
Ethier S.P. Growth factor synthesis and human breast cancer progression. J Natl Cancer Inst. 1995;87(13):964–73. [PubMed: 7629883]
10.
Yarden Y. Biology of HER2 and its importance in breast cancer. Oncology. 2001;61 Suppl 2:1–13. [PubMed: 11694782]
11.
Wu A.M., Chen W., Raubitschek A., Williams L.E., Neumaier M., Fischer R., Hu S.Z., Odom-Maryon T., Wong J.Y., Shively J.E. Tumor localization of anti-CEA single-chain Fvs: improved targeting by non-covalent dimers. Immunotechnology. 1996;2(1):21–36. [PubMed: 9373325]
12.
Viti F., Tarli L., Giovannoni L., Zardi L., Neri D. Increased binding affinity and valence of recombinant antibody fragments lead to improved targeting of tumoral angiogenesis. Cancer Res. 1999;59(2):347–52. [PubMed: 9927045]
13.
Adams G.P., Schier R., McCall A.M., Crawford R.S., Wolf E.J., Weiner L.M., Marks J.D. Prolonged in vivo tumour retention of a human diabody targeting the extracellular domain of human HER2/neu. Br J Cancer. 1998;77(9):1405–12. [PMC free article: PMC2150193] [PubMed: 9652755]
14.
Reddy S., Shaller C.C., Doss M., Shchaveleva I., Marks J.D., Yu J.Q., Robinson M.K. Evaluation of the anti-HER2 C6.5 diabody as a PET radiotracer to monitor HER2 status and predict response to trastuzumab treatment. Clin Cancer Res. 2011;17(6):1509–20. [PMC free article: PMC3060271] [PubMed: 21177408]

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