Dual-targeted contrast agent for US assessment of tumor angiogenesis in vivo

Radiology. 2008 Sep;248(3):936-44. doi: 10.1148/radiol.2483072231.


Purpose: To develop and validate a dual-targeted ultrasonographic (US) imaging agent with microbubbles (MBs) that attaches to both vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2) and alpha(v)beta(3) integrin and to compare the US imaging signal obtained from dual-targeted MBs (MB(D)) with that from single-targeted MBs (MB(S)) in a murine model of tumor angiogenesis.

Materials and methods: Animal protocols were approved by the institutional Administrative Panel on Laboratory Animal Care. Single- and dual-targeted US imaging agents were prepared by attaching anti-VEGFR2, anti-alpha(v)beta(3) integrin, or both antibodies to the shell of perfluorocarbon-filled MBs. Binding specificities of targeted MBs compared with isotype-matched immunoglobulin G-labeled control MBs (MB(C)) and nontargeted nonlabeled MBs (MB(N)) were tested with VEGFR2-positive and alpha(v)beta(3) integrin-positive cells (mouse SVR cells) and control cells (mouse 4T1 cells). In vivo imaging signals of contrast material-enhanced US by using anti-VEGFR2-targeted MBs (MB(V)), anti-alpha(v)beta(3) integrin-targeted MBs (MB(I)), MB(D), and MB(C) were quantified in 49 mice bearing SK-OV-3 tumors (human ovarian cancer). Tumor tissue was stained for VEGFR2, alpha(v)beta(3) integrin, and CD31.

Results: Attachment of MB(D) to SVR cells (mean, 0.74 MBs per cell +/- 0.05 [standard deviation]) was significantly higher than attachment to 4T1 cells (mean, 0.04 +/- 0.03), and attachment to SVR cells was higher for MB(D) than for MB(V) (mean, 0.58 +/- 0.09), MB(I) (mean, 0.42 +/- 0.21), MB(C) (mean, 0.11 +/- 0.13), and MB(N) (mean, 0.01 +/- 0.01) (P < .05). Imaging signal in the murine tumor angiogenesis model was significantly higher (P < .001) for MB(D) (mean, 16.7 +/- 7.2) than for MB(V) (mean, 11.3 +/- 5.7), MB(I) (mean, 7.8 +/- 5.3), MB(C) (mean, 2.8 +/- 0.9), and MB(N) (mean, 1.1 +/- 0.4). Immunofluorescence confirmed expression of VEGFR2 and alpha(v)beta(3) integrin on tumor vasculature.

Conclusion: Dual-targeted contrast-enhanced US directed at both VEGFR2 and alpha(v)beta(3) integrin improves in vivo visualization of tumor angiogenesis in a human ovarian cancer xenograft tumor model in mice.

Supplemental material: http://radiology.rsnajnls.org/cgi/content/full/248/3/936/DC1.

Publication types

  • Comparative Study
  • Evaluation Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Validation Study

MeSH terms

  • Biomarkers, Tumor / metabolism
  • Cell Line, Tumor
  • Contrast Media / metabolism*
  • Drug Delivery Systems / methods
  • Female
  • Humans
  • Image Enhancement
  • Integrin alphaVbeta3 / metabolism*
  • Microbubbles
  • Neovascularization, Pathologic / diagnostic imaging*
  • Neovascularization, Pathologic / metabolism*
  • Ovarian Neoplasms / blood supply
  • Ovarian Neoplasms / diagnostic imaging*
  • Ovarian Neoplasms / metabolism*
  • Reproducibility of Results
  • Sensitivity and Specificity
  • Ultrasonography
  • Vascular Endothelial Growth Factor Receptor-2 / metabolism*


  • Biomarkers, Tumor
  • Contrast Media
  • Integrin alphaVbeta3
  • Vascular Endothelial Growth Factor Receptor-2