Endostatin is a potent inhibitor of angiogenesis currently in phase I clinical trials. Imaging technologies that use near-infrared
fluorescent probes are well suited to the laboratory setting. The goal of this study was to determine whether
endostatin labeled with a near-infrared probe (
Cy5.5) could be detected in an animal and whether it would selectively localize to a
tumor.
Endostatin was conjugated to
Cy5.5 monofunctional
dye and injected into mice bearing
Lewis lung carcinoma tumors (350 mm2). Mice were imaged at various time points while under sedation using a lightproof box affixed to a fluorescent microscope mounted with a filter in the near-infrared bandwidth consistent with
Cy5.5 fluorescence. After i.p. injection, endostatin-Cy5.5 was absorbed producing a near-infrared fluorescent image within the
tumors at 18 h reaching a maximum at 42 h after injection. No signal was emitted from mice injected with unlabeled
endostatin or
Cy5.5 dye alone or those that received no injection. Further results show that a dose response exists with injection of endostatin-Cy5.5. Mimicking the clinical route of administration, an i.v. injection had a peak signal emission at 3 h but also persisted to 72 h. Finally, to determine the intratumoral binding site for
endostatin, we performed immunofluorescence on
tumor specimens and demonstrated that
endostatin binds to
tumor vasculature and colocalizes with
platelet/endothelial cell adhesion molecule 1 expression. This study demonstrates that
endostatin covalently bound to
Cy5.5 will migrate from a distant i.p. injection site to a
tumor. These data indicate that endostatin-Cy5.5 is appropriate for selectively imaging
tumors in uninjured experimental animals.