¹¹¹In-Diethylenetriaminepentaacetic acid-benzyl-succinamido-Lys-IRDye800-c(Arg-Gly-Asp-D-Phe-Lys)

[¹¹¹In-DTPA-Bz-SA-Lys-IRDye800-c(RGDfK)]

Background

[PubMed]

Optical fluorescence imaging is increasingly being 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 natural background fluorescence interference of biomolecules, providing a high contrast between target and background tissues in small animals. NIR fluorophores have a wider dynamic range and minimal background fluorescence as a result of reduced scattering compared with visible fluorescence detection. NIR fluorophores also have high sensitivity, attributable to low background fluorescence, 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 a non-invasive alternative to radionuclide imaging in small animals (4, 5).

Integrins are a family of heterodimeric glycoproteins on cell surfaces that mediate diverse biological events involving cell–cell and cell–matrix interactions (6). Integrins consist of an α and a β subunit, and they are important for cell adhesion and signal transduction. The α_vβ₃ integrin is the most prominent receptor affecting tumor growth, tumor invasiveness, metastasis, tumor-induced angiogenesis, inflammation, osteoporosis, and rheumatoid arthritis (7-12). Expression of the α_vβ₃ integrin is strong on tumor cells and activated endothelial cells, whereas expression is weak on resting endothelial cells and most normal tissues. Antagonists of α_vβ₃ are being studied as antitumor and antiangiogenic agents and its agonists are being studied as angiogenic agents for coronary angiogenesis (11, 13, 14). A peptide sequence consisting of Arg-Gly-Asp (RGD) has been identified as a recognition motif used by extracellular matrix proteins (vitronectin, fibrinogen, laminin, and collagen) to bind to a variety of integrins, including α_vβ₃. Various radiolabeled antagonists have been introduced for tumor imaging and tumor angiogenesis (15).

Most of the cyclic RGD peptides are composed of five amino acids. Haubner et al. (16) reported that various cyclic RGD peptides exhibit selective inhibition of binding to α_vβ₃ (50% inhibition concentration (IC₅₀), 7–40 nM) but not to α_vβ₅ (IC₅₀, 600–4,000 nM) or α_IIbβ₃ (IC₅₀, 700–5,000 nM) integrins. Various radiolabeled cyclic RGD peptides have been found to have high accumulation in tumors in nude mice (17). Li et al. (18) reported the development of ¹¹¹In-DTPA-Bz-SA-Lys-IRDye800-c(RGDfK), a multimodality imaging agent for imaging α_vβ₃ expression on breast tumors; single-photon emission computed tomography (SPECT) imaging used ¹¹¹In-DTPA-Bz-SA and NIR imaging used IRDye800.

Synthesis

[PubMed]

Li et al. (18) conjugated IRDye800-N-hydroxysuccinimide ester to the free ω-NH₂ group of Lys in DTPA-Bz-SA-Lys-c(RGDfK) by incubation in basic solution (pH 8.5) for 2 h. Unlabeled DTPA-Bz-SA-Lys-IRDye800-c(RGDfK) in 0.1 N sodium acetate was mixed with an aqueous solution of ¹¹¹InCl₃ for 15 min to form ¹¹¹In- DTPA-Bz-SA-Lys-IRDye800-c(RGDfK) with a specific activity of 2.22 MBq/nmol (0.06 mCi/nmol) and a radiochemical purity of >95. No free ¹¹¹In³⁺ ions were found in the preparation. The molar ratio of IRDye800 to c(RGDfK) was ~1. DTPA-Bz-SA-Lys-IRDye800-c(RGDfK) had a maximal excitation wavelength of 764 nm and a maximal emission wavelength of 794 nm; these values are similar to those for IRDye800. DTPA-Bz-SA-Lys-IRDye800-c(RGDfK) (1 µM) had 70% of the signal intensity observed with IRDye800 (1 µM).

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

Li et al. (18) performed cell-adhesion assays to vitronectin using human melanoma cancer cell line M21 (α_vβ₃-positive). Binding of M21 cells to vitronectin was blocked by DTPA-Bz-SA-Lys-IRDye800-c(RGDfK), DTPA-Bz-SA-Lys-IRDye800-c(RGDfK), and c(RGDfK) in a dose-dependent manner with IC₅₀ values of 3.86 ± 0.36 μM, 3.78 ± 0.28 μM, and 3.16 ± 0.09 µM, respectively.

Animal Studies

Human Studies

[PubMed]

No publication is currently available.

NIH Support

R01 EB00174, U54 CA90810

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