US imaging of tumor angiogenesis with microbubbles targeted to vascular endothelial growth factor receptor type 2 in mice.
OBJECTIVE
To prospectively evaluate contrast material-enhanced ultrasonography (US) with microbubbles targeted to vascular endothelial growth factor receptor type 2 (VEGFR2) for imaging tumor angiogenesis in two murine tumor models.


METHODS
Animal protocols were approved by the Institutional Administrative Panel on Laboratory Animal Care. A US contrast agent, consisting of encapsulated gaseous microbubbles, was developed specifically to bind to VEGFR2 (by using anti-VEGFR2 antibodies and biotin-streptavidin interaction) which is up-regulated on endothelial cells of tumor blood vessels. VEGFR2-targeted microbubbles (MB(V)), control microbubbles (MB(C)), and nonlabeled microbubbles (MB(N)) were tested for binding specificity on cells expressing VEGFR2 (mouse angiosarcoma SVR cells) and control cells (mouse skeletal myoblast C2C12 cells). Expression of mouse VEGFR2 in culture cells was tested with immunocytochemical and Western blot analysis. Contrast-enhanced US imaging with MB(V) and MB(C) was performed in 28 tumor-bearing nude mice (mouse angiosarcoma, n = 18; rat malignant glioma, n = 10). Differences were calculated by using analysis of variance.


RESULTS
In cell culture, adherence of MB(V) on SVR cells (2.1 microbubbles per SVR cell) was significantly higher than adherence of control microbubbles (0.01-0.10 microbubble per SVR cell; P < .001) and significantly more MB(V) attached to SVR cells than to C2C12 cells (0.15 microbubble per C2C12 cell; P < .001). In vivo, contrast-enhanced US imaging showed significantly higher average video intensity when using MB(V) compared with MB(C) for angiosarcoma and malignant glioma tumors (P < .001). Results of immunohistochemical analysis confirmed VEGFR2 expression on vascular endothelial cells of both tumor types.


CONCLUSIONS
US imaging with contrast microbubbles targeted to VEGFR2 allows noninvasive visualization of VEGFR2 expression in tumor vessels in mice.
Willmann
Jürgen K
JK
Department of Radiology and Bio-X Program, Stanford University School of Medicine, James H. Clark Center, 318 Campus Dr, East Wing, 1st Floor, Stanford, CA 94305-5427, USA.
Paulmurugan
Ramasamy
R
Chen
Kai
K
Gheysens
Olivier
O
Rodriguez-Porcel
Martin
M
Lutz
Amelie M
AM
Chen
Ian Y
IY
Chen
Xiaoyuan
X
Gambhir
Sanjiv S
SS
D000818:Animals; D003287:Contrast Media; D016503:Drug Delivery Systems / Q000379:methods; D005260:Female; D006394:Hemangiosarcoma / Q000098:blood supply / Q000000981:diagnostic imaging* / Q000378:metabolism*; D007089:Image Enhancement / Q000379:methods; D051379:Mice; D008819:Mice, Nude; D045423:Microbubbles; D009389:Neovascularization, Pathologic / Q000000981:diagnostic imaging* / Q000378:metabolism*; D011379:Prognosis; D014463:Ultrasonography / Q000379:methods*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism*
D003287:Contrast Media; D040301:Vascular Endothelial Growth Factor Receptor-2