Antiangiogenic cancer therapy: monitoring with molecular US and a clinically translatable contrast agent (BR55).
OBJECTIVE
To develop and test human kinase insert domain receptor (KDR)-targeted microbubbles (MBs) (MB(KDR)) for imaging KDR at the molecular level and for monitoring antiangiogenic therapy in a human colon cancer xenograft tumor model in mice.


METHODS
Animal studies were approved by the Institutional Administrative Panel on Laboratory Animal Care. A heterodimeric peptide that binds to human KDR with low nanomolar affinity (K(D) = 0.5 nmol/L) was coupled onto the surface of perfluorobutane-containing lipid-shelled MBs (MB(KDR)). Binding specificity of MB(KDR) to human KDR and cross-reactivity with murine vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2) were tested in cell culture under flow shear stress conditions (at 100 sec(-1)). In vivo binding specificity of MB(KDR) to VEGFR2 was tested in human LS174T colon cancer xenografts in mice with a 40-MHz ultrasonographic (US) transducer. Targeted contrast material-enhanced US imaging signal by using MB(KDR) was longitudinally measured during 6 days in tumors with (n = 6) and without (n = 6) antiangiogenic treatment (anti-VEGF antibody). Ex vivo VEGFR2 staining and microvessel density analysis were performed. Significant differences were evaluated (t, Mann-Whitney, or Wilcoxon test).


RESULTS
Cell culture experiments showed four times greater binding specificity of MB(KDR) to human KDR and cross-reactivity to murine VEGFR2 (P < or = .01). In vivo imaging signal was more than three times higher (P = .01) with MB(KDR) compared with control MBs and decreased significantly (approximately fourfold lower, P = .03) following in vivo receptor blocking with anti-VEGFR2 antibody. One day after initiation of antiangiogenic therapy, imaging signal was significantly decreased (approximately 46% lower, P = .02) in treated versus untreated tumors; it remained significantly lower (range, 46%-84% decreased; P = .038) during the following 5 days. Microvessel density was significantly reduced (P = .04) in treated (mean, 7.3 microvessels per square millimeter +/- 4.7 [standard deviation]) versus untreated tumors (mean, 22.0 microvessels per square millimeter +/- 9.4); VEGFR2 expression was significantly decreased (>50% lower, P = .03) in treated tumors.


CONCLUSIONS
Human MB(KDR) allow in vivo imaging and longitudinal monitoring of VEGFR2 expression in human colon cancer xenografts.
Pysz
Marybeth A
MA
Department of Radiology, Molecular Imaging Program at Stanford, Stanford University School of Medicine, 300 Pasteur Dr, Room H1307, Stanford, CA 94305, USA.
Foygel
Kira
K
Rosenberg
Jarrett
J
Gambhir
Sanjiv S
SS
Schneider
Michel
M
Willmann
Jürgen K
JK
D020533:Angiogenesis Inhibitors / Q000008:administration & dosage*; D000818:Animals; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000000981:diagnostic imaging* / Q000188:drug therapy*; D003287:Contrast Media; D016503:Drug Delivery Systems / Q000379:methods; D006801:Humans; D051379:Mice; D045423:Microbubbles; D015336:Molecular Probe Techniques; D010455:Peptides; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014463:Ultrasonography / Q000379:methods*
D020533:Angiogenesis Inhibitors; D003287:Contrast Media; D010455:Peptides