Gallium-68 complexes with tripodal NS₃ ligands: in vitro and in vivo stability studies

Introduction:

We report on formation and structures of ⁶⁸Ga '4+1' complexes, [Ga(NS₃)R], with the tripodal ligand tris(2-mercaptoethyl) amine (NS₃) and water or secondary amines as co-ligands (R). To improve the bio-behavior of such compounds new hydrophilic NS₃ chelators have been introduced. Challenge experiments with transferrin have been performed to study the influence of the co-ligand on the in vitro stability of the gallium complexes. Their in vivo stability was evaluated in the rat using HSA microspheres.

Experimental:

⁶⁸Ga was eluted from a ⁶⁸Ge/⁶⁸Ga generator (Obninsk, Russia) as GaCl₃ in hydrochloric solution (0.1 M) and added to the aqueous solution of the oxalate salt of the NS₃ ligand in presence of the corresponding amine. Ga complexes were formed in aqueous solution at ambient temperature. Challenge experiments were carried out at 37°C with the plasma protein transferrin (5 mg/ml), the exchange was controlled by thin layer chromatography on silicagel (Merck) and methanol as eluent.

Results and Discussion:

The labelling procedure with ⁶⁸Ga generator eluate runs in high yield under weakly alkaline conditions within some minutes. It is not necessary to heat the reaction mixture so that sensitive molecules can also be labelled. Challenge experiments with apotransferrin were carried out for several hours. Amine-coordinated complexes as well as non-amine compounds showed high in vitro stability within 240 minutes. The in vivo stability of the complexes was evaluated in the rat using HSA microspheres. ⁶⁸Ga-labelled HSA microspheres were completely accumulated in the lung. In small animal PET studies no significant loss of ⁶⁸Ga activity was observed within three hours.

Conclusion:

The '4+1' mixed-ligand approach enables the ⁶⁸Ga-labelling of biomolecules under physiological conditions. The lipophilicity of the complexes can be controlled by introducing pharmacological modifiers (e.g. PEG, glucosamine, crown ether) into the tetradentate ligand and/or derivatization of the monodentate ligand. Moreover, the described approach allows the functionalization of a biomolecule with the tripodal as well as with the monodentate ligand.