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Journal of Nuclear Medicine

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Meeting ReportPoster Presentations - Physicians/Scientists/Pharmacists

PET imaging of angiogenesis with scVEGF-DOTA-PEG-Cu-64

Francis Blankenberg, Zoia Levashova, Marina Backer and Joseph Backer
Journal of Nuclear Medicine May 2006, 47 (suppl 1) 503P;
Francis Blankenberg
1Radiology, Stanford University, Stanford, California
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Zoia Levashova
1Radiology, Stanford University, Stanford, California
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Marina Backer
2Research, SibTech, Inc., Newington, Connecticut
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Joseph Backer
2Research, SibTech, Inc., Newington, Connecticut
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Abstract

1841

Objectives: We wished to test a novel single-chain(sc) VEGF expressed with 15-aa cysteine-containing tag (C-tag) that was site-specifically labeled with PEG-DOTA conjugate and radiolabled with Cu-64 (scVEGF/Cu) for PET imaging of angiogenesis.

Methods: scVEGF consisting of two 3-112aa fragments of VEGF121 genetically fused head-to-tail was expressed with an N-terminal C-tag for site-specific modification. NHS-PEG-maleimide dissolved in DMSO was mixed with 2-(4-aminobenzyl)-DOTA (molar ratio of 5:1) in carbonate buffer pH 8.0, and incubated for 1 h. Unreacted NHS was quenched with 0.1 M Tris-HCl pH 8.0 (30 min at RT), and then mixed with scVEGF to a final protein to DOTA molar ratio of 1:10 and incubated for 1 h. A portion of purified scVEGF-PEG-DOTA was incubated with 90-fold excess of biotin-NHS in order to inactivate scVEGF. Purified functional and inactive scVEGF-PEG-DOTA were incubated with 400-500 µCi of Cu-64 in 0.1 M NaAc buffer pH 5.5 for 1 h at 55°C followed by quenching with 1 mM EDTA and purification by gel-filtration, labeling 40% of the total added protein. Balb/c mice were implanted with 1000 4T1luc mouse mammary adenocarcinoma cells injected into the left mammary fat pad and underwent imaging 14 days later.

Results: scVEGF-PEG-DOTA retained VEGF functional activity in tissue culture assays, while biotinylated scVEGF-PEG-DOTA was inactive, providing an authentic negative control for imaging experiments. One hour after injection of 1-2 µCi of tracers into tumor-bearing mice tumor/muscle uptake of radioactivity was 2.9 ± 0.6 %ID/g / 0.54 ± 0.15 %ID/g (n=10) for scVEGF/Cu and 0.81 ± 0.13 %ID/g / 0.34 ± 0.26%ID/g (n=9) for biotin-inactivated probe (Bt-VEGF/Cu). Transverse and coronal PET tomographic slices, respectively, are shown in Figure 1 19 hours after injection of 100 to 200uCi of VEGF/Cu or Bt-VEGF/Cu. White arrows mark the location of tumor in the mouse injected with Bt-VEGF/Cu. Tumor uptake of scVEGF/Cu was 2 to 3 times greater than for Bt-VEGF/Cu.

Conclusions: C-tagged scVEGF can be site-specifically modified with PEG-DOTA without affecting VEGF activity and radiolabeled with Cu-64 for PET imaging of tumor angiogenesis.

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Journal of Nuclear Medicine
Vol. 47, Issue suppl 1
May 1, 2006
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PET imaging of angiogenesis with scVEGF-DOTA-PEG-Cu-64
Francis Blankenberg, Zoia Levashova, Marina Backer, Joseph Backer
Journal of Nuclear Medicine May 2006, 47 (suppl 1) 503P;

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PET imaging of angiogenesis with scVEGF-DOTA-PEG-Cu-64
Francis Blankenberg, Zoia Levashova, Marina Backer, Joseph Backer
Journal of Nuclear Medicine May 2006, 47 (suppl 1) 503P;
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