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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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64Cu-1,4,7-Triazacyclononane-1,4,7-triacetic acid-p-isothiocyanatobenzyl-bevacizumab-IRDye 800CW

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

Created: ; Last Update: March 21, 2013.

Chemical name:64Cu-1,4,7-Triazacyclononane-1,4,7-triacetic acid-p-isothiocyanatobenzyl-bevacizumab-IRDye 800CW
Abbreviated name:64Cu-NOTA-Bev-800CW
Agent category:Antibody
Target:Vascular endothelial growth factor-A (VEGF-A)
Target category:Antigen
Method of detection:Multimodality: positron emission tomography (PET), near-infrared fluorescence (NIR) optical imaging
Source of signal:64Cu, IRDye 800CW
  • Checkbox In vitro
  • Checkbox Rodents
Click on protein, nucleotide (RefSeq), and gene for more information about VEGF-A.



The vascular endothelial growth factor (VEGF) family is composed of five VEGF glycoproteins (VEGF-A, VEGF-B, VEGF-C, VEGF-D, and VEGF-E) and consists of at least six isoforms of various numbers of amino acids (121, 145, 165, 183, 189, and 206 amino acids) produced through alternative splicing (1). VEGF-A is composed of VEGF121, VEGF165, and VEGF189 isoforms, which are secreted by most cell types and are active as homodimers linked by disulfide bonds. VEGF121 does not bind to heparin like other VEGF polypeptides do (2). VEGF is a potent angiogenic factor that induces proliferation, sprouting, migration, and tube formation of endothelial cells. There are three high-affinity tyrosine kinase VEGF receptors (VEGFR-1, Flt-1; VEGFR-2, KDR/Flt-1; and VEGFR-3, Flt-4) on endothelial cells. Several types of non-endothelial cells, such as hematopoietic stem cells, melanoma cells, monocytes, osteoblasts, and pancreatic β cells, also express VEGFRs (1).

VEGF is overexpressed in various tumor cells and tumor-associated endothelial cells (3). Among the at least seven isoforms of VEGF-A, VEGF121 is freely soluble while all VEGF189 is bound to the cell membrane or extracellular matrix (ECM). VEGF165 exhibits an intermediary behavior (i.e. partly diffusible and partly bound). Inhibition of VEGFR function has been shown to inhibit pathological angiogenesis as well as tumor growth and metastasis (4, 5). Radiolabeled VEGF has been developed as a single-photon emission computed tomography tracer for imaging solid tumors and angiogenesis in humans (6-8). However, several studies have shown that cancer treatments (photodynamic therapy, radiotherapy, and chemotherapy) can lead to increased tumor VEGF expression and subsequently to more aggressive disease (9, 10). Therefore, it is important to measure VEGF levels in the tumors to design better anticancer treatment protocols. Bevacizumab (Bev) is a humanized antibody against VEGF-A (11) that binds to all VEGF-A isoforms, and it is approved for clinical use in metastatic colon carcinoma and non-small cell lung cancer (12). VEGFR-2 has been shown to mediate most of the VEGF-A activation in tumor endothelial cells (13, 14). Zhang et al. (15) prepared 64Cu-1,4,7-triazacyclononane-1,4,7-triacetic acid-p-isothiocyanatobenzyl-bevacizumab-IRDye 800CW (64Cu-NOTA-Bev-800CW) for positron emission tomography (PET) and near-infrared fluorescence (NIR) multimodal imaging of VEGF expression in nude mice bearing U87MG human glioblastoma xenografts. U87MG cells express all three isoforms of VEGF-A (VEGF121, VEGF165, and VEGF189).

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NOTA-p-Bn-SCN-Bev (NOTA-Bev) was prepared by conjugation of p-SCN-Bn-NOTA to Bev in 25:1 molar ratio at pH 9 (15). NOTA-Bev was purified with column chromatography. NOTA-Bev and IRDye 800CW-N-hydroxysuccinimide ester were incubated at 2:1 molar ratio at pH 8.5 for 2 h at room temperature. NOTA-Bev-800CW was purified with column chromatography. There were ~5 NOTA and 0.8 IRDye 800CW moieties per antibody conjugate. NOTA-Bev-800CW (0.162 nmol) was radiolabeled using 37 MBq (1 mCi) 64CuCl2 in sodium acetate buffer (pH 5.0) for 30 min at 37°C. 64Cu-NOTA-Bev-800CW was purified with a PD-10 column. Total preparation time of the tracer was 60 ± 10 min (n = 10). The radiochemical yield was >80%. The radiochemical purity was >98%. The specific activity of 64Cu-NOTA-Bev-800CW was reported to be 170 MBq/nmol (4.6 mCi/nmol).

In Vitro Studies: Testing in Cells and Tissues


64Cu-NOTA-Bev-800CW had immunoreactivity similar to the unconjugated mAb as determined by binding to U87MG tumor cells with flow cytometry analysis (15).

Animal Studies



Zhang et al. (15) studied the whole-body distribution of 5–10 MBq (0.14–27 mCi) 64Cu-NOTA-Bev-800CW (0.3 nmol IRDye 800CW) with PET and NIR fluorescence imaging in nude mice (n = 4/group) bearing human U87MG xenografts at 4, 24, 48, and 72 h. The U87MG tumors were clearly visualized at 24–72 h after injection. The tumor accumulation of radioactivity levels were 4.6 ± 0.7% injected dose per gram (ID/g) at 4 h, 16.3 ± 1.6% ID/g at 24 h, 18.1 ± 1.4% ID/g at 48 h, and 20.7 ± 3.7% ID/g at 72 h. The liver accumulation of radioactivity levels were 14.4 ± 3.7% ID/g at 4 h, 17.2 ± 2.0% ID/g at 24 h, 13.8 ± 2.9% ID/g at 48 h, and 12.8 ± 2.7% ID/g at 72 h. The radioactivity level in the blood was 20.5% ID/g at 4 h and decreased to 13.0% ID/g at 72 h. Pretreatment (6 h) with 13 nmol Bev inhibited accumulation of radioactivity in the U87MG tumors to <7% ID/g at 24–72 h (P < 0.01), whereas little inhibition was observed in the liver and blood. NIR fluorescence imaging was performed immediately after the PET scans, and the tumors were clearly visualized at 24–72 h. Region-of-interest analysis showed tumor NIR fluorescence radiant intensity values of 12.0 ± 3.9 × 106, 19.2 ± 3.7 × 106, 15.2 ± 1.9 × 106, and 13.6 ± 3.9 × 106 at 4, 24, 48, and 72 h, respectively. Pretreatment with Bev resulted in reduced tumor signal intensity values of 11.6 ± 1.0 × 106, 8.93 ± 1.0 × 106, and 7.7 ± 1.0 × 106 at 24, 48, and 72 h (P < 0.05), respectively. Ex vivo PET and NIR fluorescence imaging studies were performed at 72 h. There was a linear correlation between ex vivo tumor NIR fluorescence data and in vivo PET tumor radioactivity level (% ID/g) (R2 = 0.93, P < 0.001). 64Cu-NOTA-Bev-800CW bound to VEGF that is bound to the VEGFR on the tumor cells, tumor endothelial cells and tumor ECM.

Other Non-Primate Mammals


No publication is currently available.

Non-Human Primates


No publication is currently available.

Human Studies


No publication is currently available.

NIH Support

5 T32 CA009206-32


Ferrara N. Vascular endothelial growth factor: basic science and clinical progress. Endocr Rev. 2004;25(4):581–611. [PubMed: 15294883]
Cohen T., Gitay-Goren H., Sharon R., Shibuya M., Halaban R., Levi B.Z., Neufeld G. VEGF121, a vascular endothelial growth factor (VEGF) isoform lacking heparin binding ability, requires cell-surface heparan sulfates for efficient binding to the VEGF receptors of human melanoma cells. J Biol Chem. 1995;270(19):11322–6. [PubMed: 7744769]
Soria J.C., Fayette J., Armand J.P. Molecular targeting: targeting angiogenesis in solid tumors. Ann Oncol. 2004;15 Suppl 4:iv223–7. [PubMed: 15477311]
Ferrara N. Vascular endothelial growth factor as a target for anticancer therapy. Oncologist. 2004;9 Suppl 1:2–10. [PubMed: 15178810]
Hicklin D.J., Ellis L.M. Role of the vascular endothelial growth factor pathway in tumor growth and angiogenesis. J Clin Oncol. 2005;23(5):1011–27. [PubMed: 15585754]
Li S., Peck-Radosavljevic M., Koller E., Koller F., Kaserer K., Kreil A., Kapiotis S., Hamwi A., Weich H.A., Valent P., Angelberger P., Dudczak R., Virgolini I. Characterization of (123)I-vascular endothelial growth factor-binding sites expressed on human tumour cells: possible implication for tumour scintigraphy. Int J Cancer. 2001;91(6):789–96. [PubMed: 11275981]
Li S., Peck-Radosavljevic M., Kienast O., Preitfellner J., Hamilton G., Kurtaran A., Pirich C., Angelberger P., Dudczak R. Imaging gastrointestinal tumours using vascular endothelial growth factor-165 (VEGF165) receptor scintigraphy. Ann Oncol. 2003;14(8):1274–7. [PubMed: 12881392]
Li S., Peck-Radosavljevic M., Kienast O., Preitfellner J., Havlik E., Schima W., Traub-Weidinger T., Graf S., Beheshti M., Schmid M., Angelberger P., Dudczak R. Iodine-123-vascular endothelial growth factor-165 (123I-VEGF165). Biodistribution, safety and radiation dosimetry in patients with pancreatic carcinoma. Q J Nucl Med Mol Imaging. 2004;48(3):198–206. [PubMed: 15499293]
Gorski D.H., Beckett M.A., Jaskowiak N.T., Calvin D.P., Mauceri H.J., Salloum R.M., Seetharam S., Koons A., Hari D.M., Kufe D.W., Weichselbaum R.R. Blockage of the vascular endothelial growth factor stress response increases the antitumor effects of ionizing radiation. Cancer Res. 1999;59(14):3374–8. [PubMed: 10416597]
Solban N., Selbo P.K., Sinha A.K., Chang S.K., Hasan T. Mechanistic investigation and implications of photodynamic therapy induction of vascular endothelial growth factor in prostate cancer. Cancer Res. 2006;66(11):5633–40. [PubMed: 16740700]
Wang Y., Fei D., Vanderlaan M., Song A. Biological activity of bevacizumab, a humanized anti-VEGF antibody in vitro. Angiogenesis. 2004;7(4):335–45. [PubMed: 15886877]
Ferrara N., Hillan K.J., Gerber H.P., Novotny W. Discovery and development of bevacizumab, an anti-VEGF antibody for treating cancer. Nat Rev Drug Discov. 2004;3(5):391–400. [PubMed: 15136787]
Folkman J. Role of angiogenesis in tumor growth and metastasis. Semin Oncol. 2002;29(6) Suppl 16:15–8. [PubMed: 12516034]
Ferrara N., Gerber H.P., LeCouter J. The biology of VEGF and its receptors. Nat Med. 2003;9(6):669–76. [PubMed: 12778165]
Zhang Y., Hong H., Engle J.W., Yang Y., Barnhart T.E., Cai W. Positron Emission Tomography and Near-Infrared Fluorescence Imaging of Vascular Endothelial Growth Factor with Dual-Labeled Bevacizumab. Am J Nucl Med Mol Imaging. 2012;2(1):1–13. [PMC free article: PMC3249831] [PubMed: 22229128]


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