Development of 99mTc-Labeled Human Serum Albumin with Prolonged Circulation by Chelate-then-Click Approach: A Potential Blood Pool Imaging Agent

Technetium-99m-labeled human serum albumin (99mTc-HSA) has been utilized as a blood pool imaging agent in the clinic for several decades. However, 99mTc-HSA has a short circulation time, which is a critical shortcoming for a blood pool imaging agent. Herein, we developed a novel 99mTc-labeled HSA with a long circulation time using click chemistry and a chelator, 2,2′-dipicolylamine (DPA), (99mTc-DPA-HSA). Specifically, we examined the feasibility of copper-free strain-promoted alkyne-azide cycloaddition (SPAAC) for the incorporation of HSA to the [99mTc (CO)3(H2O)3]+ system by adopting a chelate-then-click approach. In this strategy, a potent chelate system, azide-functionalized DPA, was first complexed with [99mTc (CO)3(H2O)3]+, followed by the SPAAC click reaction with azadibenzocyclooctyne-functionalized HSA (ADIBO–HSA) under biocompatible conditions. Radiolabeling efficiency of azide-functionalized DPA (99mTc-DPA) was >98%. Click conjugation efficiency of 99mTc-DPA with ADIBO–HSA was between 76 and 99% depending on the number of ADIBO moieties attached to HSA. In whole-body in vivo single photon emission computed tomography images, the blood pool uptakes of 99mTc-DPA-HSA were significantly enhanced compared to those of 99mTc-HSA at 10 min, 2, and 6 h after the injection (P < 0.001, 0.025, and 0.003, respectively). Furthermore, the blood activities of 99mTc-DPA-HSA were 8 times higher at 30 min and 10 times higher at 3 h after the injection compared to those of conventional 99mTc-HSA in ex vivo biodistribution experiment. The results exhibit the potential of 99mTc-DPA-HSA as a blood pool imaging agent and further illustrate the promise of the pre-labeling SPAAC approach for conjugation of heat-sensitive biological targeting vectors with [99mTc (CO)3(H2O)3]+.