Background

[PubMed]

Optical fluorescence imaging is increasingly used to monitor biological functions of specific targets in small animals (1-3). However, the intrinsic fluorescence of biomolecules poses a problem when fluorophores that absorb visible light (350–700 nm) are used. Near-infrared (NIR) fluorescence (700–1,000 nm) detection avoids the background fluorescence interference of natural biomolecules, providing a high contrast between target and background tissues. NIR fluorophores have wider dynamic range and minimal background as a result of reduced scattering compared with visible fluorescence detection. They also have high sensitivity, resulting from low infrared background, and high extinction coefficients, which provide high quantum yields. The NIR region is also compatible with solid-state optical components, such as diode lasers and silicon detectors. NIR fluorescence imaging is becoming a noninvasive alternative to radionuclide imaging in small animals or with probes in close proximity of the target in humans (4, 5). Among the various optical imaging agents, only indocyanine green (ICG), with NIR fluorescence absorption at 780 nm and emission at 820 nm, is approved by the United States Food and Drug Administration for clinical applications in angiography, blood flow evaluation, and liver function assessment. It is also under evaluation in several clinical trials for other applications, such as optical imaging and mapping of both the lymphatic vessels and lymph nodes in cancer patients for surgical dissection of tumor cells and endoscopic imaging of the pancreas and colon.

Tumor angiogenesis represents a continuous and important process in tumor development in which the tumor attempts to gain an independent blood supply (6). This process is driven by the tumor's chronic overproduction of angiogenic factors, which bind to receptors on nearby vessel endothelial cells. Angiogenesis is essential for the growth of solid tumors and their metastases. Imaging angiogenesis may be useful for monitoring angogeneic treatments of tumors and cardiovascular diseases (7-9). Endostatin was discovered as an endogenous inhibitor of tumor angiogenesis and endothelial-cell growth (10). It is one of the most promising anti-angiogenic cancer drugs. It has been successfully coupled with Cy5.5 NIR dye for optical imaging of endostatin receptor density in tumors in mice (11, 12).

Synthesis

[PubMed]

Cy5.5 monofunctional N-hydroxysuccinimide (NHS) ester was used to conjugate endostatin to form Cy5.5-endostatin, which was purified by gel filtration chromatography (12). Cy5.5-endostatin was adjusted to a final concentration of 1 mg/ml. A molar ratio of Cy5.5 to endostatin was estimated to be 1.0.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

Histoimmunostaining of endostatin and platelet/endothelial cell adhesion molecule 1 (PECAM-1) were performed on tissue sections from tumors (Lewis lung carcinoma, LLC), heart, and muscle of untreated mice bearing the LLC tumors (11). The pattern of endostatin staining of microvasculature revealed endostatin binding to the endothelial cells with no binding to the tumor cells. Staining of PECAM-1, specific for endothelial cells, coincided with endostatin staining on the endothelial cells. There was no detectable endostatin binding in either the skeletal muscle or heart muscle sections. These results suggest that tumor cells induced formation of new blood vessels, which processed receptors for endostatin.

Animal Studies

Human Studies

[PubMed]

No publication is currently available.

References

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