A Universally Applicable 68Ga-Labeling Technique for Proteins

Although protein-based PET imaging agents are projected to become important tracer molecules in the future, the labeling of complex biomolecules with PET radionuclides is inexpedient and, most of the time, challenging. Methods: Here we present a straightforward labeling chemistry to attach the versatile radionuclide Ga-68 to proteins. Introducing the Ga-68 chelating agent NODA-GA-T (2,2'-(7-(1-carboxy-4-(2-mercaptoethylamino)-4-oxobutyl)- 1,4,7-triazonane-1,4-diyl) diacetic acid) by reaction with proteins chemically processed with sulfo-SMCC (4-(N-maleimidomethyl) cyclohexane-1-carboxylic acid 3-sulfo-N-hydroxysuccinimide ester sodium salt) results in labeling precursors, enabling a simple and rapid kit-labeling procedure that requires no workup of the radiolabeled proteins. Various Ga-68-proteins were labeled using this method, and the radiochemical yields and specific activities of the labeled proteins were determined. To show that the radiotracers are applicable for in vivo studies, proof-of-concept small-animal PET images were acquired in healthy rats using Ga-68 rat serum albumin for blood-pool imaging and Ga-68-annexin V for apoptosis imaging in mice with a left ventricular myocardial infarction. Results: The proteins could be modified, yielding 1.2-1.7 Ga-68-labeling sites per protein molecule. All investigated proteins could be labeled in high radiochemical yields of 95% or more in less than 10 min in 1 step, using acetate-buffered medium (pH 3.5-4.0) at room temperature without any further purification. The labeled proteins displayed specific activities of 20-45 GBq/mmol (540-1,200 Ci/mmol). In the proof-of-concept in vivo studies, Ga-68 rat serum albumin and Ga-68-annexin V were successfully used for in vivo imaging. Both radiotracers showed a favorable biodistribution in the animal models, thus demonstrating the usefulness of the developed approach for the kit Ga-68 labeling of proteins. Conclusion: The preprocessing of proteins proceeds in high chemical yields and with high protein recovery rates after purification. These precursors can be stored for several months at -20 degrees C without degradation, and Ga-68 labeling can be performed in a 1-step kit-labeling reaction in high radiochemical yields. Two of the derivatized model proteins were successfully used in proof-of-concept in vivo imaging studies to prove the applicability of this kit Ga-68-labeling technique.