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. 2015 Oct 15;5(11):3301-10.
eCollection 2015.

Radiolabeling and evaluation of (64)Cu-DOTA-F56 peptide targeting vascular endothelial growth factor receptor 1 in the molecular imaging of gastric cancer

Hua Zhu  1 , Chuanke Zhao  2 , Fei Liu  1 , Lixin Wang  2 , Junnan Feng  2 , Zheng Zhou  2 , Like Qu  2 , Chengchao Shou  2 , Zhi Yang  1
Affiliations

Radiolabeling and evaluation of (64)Cu-DOTA-F56 peptide targeting vascular endothelial growth factor receptor 1 in the molecular imaging of gastric cancer

Hua Zhu et al. Am J Cancer Res. .

Abstract

Noninvasive imaging of vascular endothelial growth factor receptor 1 (VEGFR1) remains a great challenge in early diagnosis of gastric cancer. Here we reported the synthesis, radiolabeling, and evaluation of a novel (64)Cu-radiolabeled peptide for noninvasive positron emission tomography (PET) imaging of VEGFR1 positive gastric cancer. The binding of modified peptide WHSDMEWWYLLG (termed as F56) to VEGER-1 expressed in gastric cancer cell BCG823 has been confirmed by immune-fluorescence overlap. DOTA-F56 was designed and prepared by solid-phase synthesis and folded in vitro. (64)Cu-DOTA-F56 was synthesized in high radiochemical yield and high specific activity (S.A. up to 255.6 GBq/mmol). It has excellent in vitro stability. Micro-PET imaging of (64)Cu-DOTA-F56 identifies tumor in BCG823 tumor-bearing mice, while that of (18)F-FDG does not. Immunohistochemical analysis of excised BCG823 xenograft showed colocalization between the PET images and the staining of VEGFR1. These results demonstrated that (64)Cu-DOTA-F56 peptide has potential as a noninvasive imaging agent in VEGFR1 positive tumors.

Keywords: F56 peptide; Vascular endothelial growth factor receptor 1; copper-64; gastric cancer; molecular imaging.

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Figures

Figure 1
Figure 1
The Synthetic strategy and radiolabeling of 64Cu-DOTA-F56 peptide.
Figure 2
Figure 2
Fluorescence visualization of FITC-F56 binds to BGC823 cells. BGC823 cells were incubated with 10 μg/ml FITC-F56 peptide for 30 min, fixed, stained with 5 ng/ml DAPI (blue), and observed under a confocal microscopy. Scale bar = 50 μm.
Figure 3
Figure 3
MALDI-TOF mass spectra of DOTA-F56 peptide. The purified DOTA-F56 was characterized by MALDI-TOF-MS and the measured molecular weight was consistent with the expected molecular weight.
Figure 4
Figure 4
HPLC chromatographs of DOTA-F56 and 64Cu-DOTA-F56.
Figure 5
Figure 5
In vitro stability of 64Cu-DOTA-F56. 64Cu-DOTA-F56 was mixed with 0.01 M PBS (pH 7.4), 0.1 M NaOAc (pH 5.5), or 5% HSA, respectively. After incubation at 37°C with shaking in a thermo mixer, the radiochemical purity was measured by Radio-HPLC.
Figure 6
Figure 6
In vitro BCG-823 cell uptake assay. BCG823 cells were seeded in 12-well plate and incubated overnight. After incubation with radiotracer at 37°C for desired time points (10 min, 30 min, 60 min and 120 min), cells were washed and lysed. Radioactivity of the cells was counted using a PerkinElmer 1470 automatic γ-counter. Cell uptake data were expressed as the percentage of the applied radioactivity per total radioactive. ***P < 0.001 student’s t-test.
Figure 7
Figure 7
Micro-PET imaging of a nu/nu mouse bearing BCG823 tumor cells. Images were obtained at 2 h PET imaging of 18F-FDG and 2 h, 24 h after injection of 64Cu-DOTA-F56. Static Micro-PET images are presented in 3D and the green line circles the tumors.
Figure 8
Figure 8
HE and IHC stainings of tissue slides. A. Hematoxylin and eosin staining (HE stain); B. F56 peptide immunohistochemical staining; C. VEGFR1 immunohistochemical staining; Scale bar = 100 μm.
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