Novel Bifunctional Cyclic Chelator for (89)Zr Labeling-Radiolabeling and Targeting Properties of RGD Conjugates

Abstract

Within the last years (89)Zr has attracted considerable attention as long-lived radionuclide for positron emission tomography (PET) applications. So far desferrioxamine B (DFO) has been mainly used as bifunctional chelating system. Fusarinine C (FSC), having complexing properties comparable to DFO, was expected to be an alternative with potentially higher stability due to its cyclic structure. In this study, as proof of principle, various FSC-RGD conjugates targeting αvß3 integrins were synthesized using different conjugation strategies and labeled with (89)Zr. In vitro stability, biodistribution, and microPET/CT imaging were evaluated using [(89)Zr]FSC-RGD conjugates or [(89)Zr]triacetylfusarinine C (TAFC). Quantitative (89)Zr labeling was achieved within 90 min at room temperature. The distribution coefficients of the different radioligands indicate hydrophilic character. Compared to [(89)Zr]DFO, [(89)Zr]FSC derivatives showed excellent in vitro stability and resistance against transchelation in phosphate buffered saline (PBS), ethylenediaminetetraacetic acid solution (EDTA), and human serum for up to 7 days. Cell binding studies and biodistribution as well as microPET/CT imaging experiments showed efficient receptor-specific targeting of [(89)Zr]FSC-RGD conjugates. No bone uptake was observed analyzing PET images indicating high in vivo stability. These findings indicate that FSC is a highly promising chelator for the development of (89)Zr-based PET imaging agents.

Keywords: 89Zr; RGD peptide; fusarinine C; positron emission tomography (PET); triacetylfusarinine C.

Figure 1

Chemical structures of DFO and FSC derivatives.

Figure 2

Time course of ⁸⁹Zr complexation of FSC(succ-RGD)₃ (58 μM, pH 7, at RT).

Figure 3

Internalization of [⁸⁹Zr]FSC(RGDfE)₃, [⁸⁹Zr]FSC(succ-RGD)₃, and [⁸⁹Zr]FSC(Mal-RGD)₃ in α_vβ₃ integrin positive M21 tumor cells.

Figure 4

Biodistribution of [⁸⁹Zr]/[⁶⁸Ga]FSC(succ-RGD)₃ at 1, 2, and 4 h p.i. in BALB/C nude mice bearing the α_vβ₃ integrin positive M21 cell tumor on the right flank and the α_vβ₃ integrin negative control tumor M21-L on the left flank, and [⁸⁹Zr]TAFC at 6 h in normal BALB/c mice. Significant differences in uptake are marked with an asterisk (P = 0.05). Each data point represents an average of biodistribution data in four animals (n = 4).

Figure 5

Three-dimensional volume projections of fused microPET/CT images of M21/M21-L tumor xenograft bearing nude mouse ([⁸⁹Zr]FSC(succ-RGD)₃, 5 μg, 5 MBq) at 1, 4, and 24 h p.i. Red arrow, α_vβ₃ integrin positive M21 tumor; blue arrow, α_vβ₃ integrin negative M21-L tumor.