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

Abstract

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]⁺.