Exploration of 68Ga-DOTA-MAL as a Versatile Vehicle for Facile Labeling of a Variety of Thiol-Containing Bioactive Molecules.

Abstract

Efficient and site-specific radiolabeling reactions are essential in molecular probe synthesis. Thus, selecting an effective method for radiolabeling that does not affect bioactivity of the molecule is critical. Varieties of bifunctional chelating agents provide a solution in this matter. As a chemo-specific chelator, maleimido-mono-amide-DOTA (DOTA-Mal) holds significant potential for 68Ga labeling of bioactive molecules; it can react specifically with free sulfhydryl groups under mild conditions. Compared with amino and carboxylic acid groups, free sulfhydryl groups are relatively less common in most biomolecules and can serve as site-specific radiolabeling targets. Labeling of 68Ga usually employs a two-step labeling strategy; first, chelators are conjugated to the biomolecules, which is followed by radiolabeling. However, the bioactivity of biomolecules may be affected by harsh labeling conditions. In this study, three 68Ga-labeled bioactive molecules, namely, 68Ga-DOTA-RGD, 68Ga-DOTA-FA, and 68Ga-DOTA-BSA, were prepared using a novel strategy under mild conditions (pH of 8.0 at room temperature). Using this strategy, DOTA-Mal was labeled by 68Ga before it reacted with the sulfhydryl group-containing biomolecules, which avoided damage to said biomolecules caused by the harsh reaction conditions required in 68Ga-labeling procedures. The biological and chemical properties of these three radiotracers synthesized using this strategy are well manifested. Through a series of experiments, the effectiveness of this strategy is demonstrated, and we believe that this site-specific bioactivity-friendly reaction strategy will facilitate developments and translation applications of varieties of 68Ga-labeled positron emission tomography probes.