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In vivo NIRF imaging-guided delivery of a novel NGR–VEGI fusion protein for targeting tumor vasculature

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Abstract

Pathological angiogenesis is crucial in tumor growth, invasion and metastasis. Previous studies demonstrated that the vascular endothelial growth inhibitor (VEGI), a member of the tumor necrosis factor superfamily, can be used as a potent endogenous inhibitor of tumor angiogenesis. Molecular probes containing the asparagine–glycine–arginine (NGR) sequence can specifically bind to CD13 receptor which is overexpressed on neovasculature and several tumor cells. Near-infrared fluorescence (NIRF) optical imaging for targeting tumor vasculature offers a noninvasive method for early detection of tumor angiogenesis and efficient monitoring of response to anti-tumor vasculature therapy. The aim of this study was to develop a new NIRF imaging probe on the basis of an NGR–VEGI protein for the visualization of tumor vasculature. The NGR–VEGI fusion protein was prepared from prokaryotic expression, and its function was characterized in vitro. The NGR–VEGI protein was then labeled with a Cy5.5 fluorophore to afford Cy5.5-NGR–VEGI probe. Using the NIRF imaging technique, we visualized and quantified the specific delivery of Cy5.5-NGR–VEGI protein to subcutaneous HT-1080 fibrosarcoma tumors in mouse xenografts. The Cy5.5-NGR–VEGI probe exhibited rapid HT-1080 tumor targeting, and highest tumor-to-background contrast at 8 h post-injection (pi). Tumor specificity of Cy5.5-NGR–VEGI was confirmed by effective blocking of tumor uptake in the presence of unlabeled NGR–VEGI (20 mg/kg). Ex vivo NIRF imaging further confirmed in vivo imaging findings, demonstrating that Cy5.5-NGR–VEGI displayed an excellent tumor-to-muscle ratio (18.93 ± 2.88) at 8 h pi for the non-blocking group and significantly reduced ratio (4.92 ± 0.75) for the blocking group. In conclusion, Cy5.5-NGR–VEGI provided highly sensitive, target-specific, and longitudinal imaging of HT-1080 tumors. As a novel theranostic protein, Cy5.5-NGR–VEGI has the potential to improve cancer treatment by targeting tumor vasculature.

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Abbreviations

NIRF:

Near-infrared fluorescence

VEGI:

Vascular endothelial growth inhibitor

TNF:

Tumor necrosis factor

NGR:

Asparagine-glycine-arginine

APN:

Aminopeptidase N

TNF-α:

Tumor necrosis factor-alpha

IFN:

Interferon

DOX:

Doxorubicin

NHS:

N-hydroxysuccinimide

DMSO:

Dimethyl sulfoxide

HUVEC:

Human umbilical vein endothelial cell line

DMEM:

Dulbecco’s Modified Eagle’s Medium

FBS:

Fetal bovine serum

PBS:

Phosphate-buffered saline

PFA:

Paraformaldehyde

DAPI:

4′,6-Diamidino-2-phenylindole

pi:

Post-injection

ROIs:

Regions of interest

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Acknowledgments

This work was supported by the USC Department of Radiology, the Major Program of National Natural Science Foundation of China (Grant No. 81230033), the National Basic Research and Development Program of China (Grant No. 2011CB707704), the Major Research Instrumentation Program of National Natural Science Foundation of China (Grant No. 81227901), the General Program of National Natural Science Foundation of China (Grant No. 81371594), and the International Cooperation Program of Xijing Hospital (Grant No. XJZT13G02).

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The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Department of Radiology, Molecular Imaging Center, Keck School of Medicine, University of Southern California, 2250 Alcazar Street, CSC 103, Los Angeles, CA, 90033-9061, USA

    Wenhui Ma, Guoquan Li, Peter S. Conti & Kai Chen

  2. Department of Nuclear Medicine, Xijing Hospital, The Fourth Military Medical University, Xi’an, Shaanxi, 710032, People’s Republic of China

    Wenhui Ma, Guoquan Li, Jing Wang & Weidong Yang

  3. State Key Laboratory of Cancer Biology, Department of Biopharmaceutics, School of Pharmacy, The Fourth Military Medical University, Xi’an, Shaanxi, 710032, People’s Republic of China

    Yingqi Zhang

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  1. Wenhui Ma
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  2. Guoquan Li
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  3. Jing Wang
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  4. Weidong Yang
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  5. Yingqi Zhang
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  6. Peter S. Conti
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  7. Kai Chen
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Correspondence to Jing Wang or Kai Chen.

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W. Ma and G. Li contributed equally to this work.

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Ma, W., Li, G., Wang, J. et al. In vivo NIRF imaging-guided delivery of a novel NGR–VEGI fusion protein for targeting tumor vasculature. Amino Acids 46, 2721–2732 (2014). https://doi.org/10.1007/s00726-014-1828-6

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  • DOI: https://doi.org/10.1007/s00726-014-1828-6

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