99mTc(I)-Labeled His-Tagged Proteins: Impact in the Development of Novel Imaging Probes and in Drug Discovery.

Technetium-99 m (^99mTc) remains the cornerstone of nuclear medicine for single photon emission computed tomography (SPECT) due to its widespread availability and chemical and physical features. Its multiple oxidation states allow for the design and production of radiopharmaceuticals with versatile properties, namely in terms of pharmacokinetic profile. ^99mTc(V) is the most common oxidation state, but ^99mTc(I) gained traction after the pioneering work of Alberto and colleagues, which resulted in the introduction of the organometallic core fac-[^99mTc(CO)₃(H₂O)₃]⁺. This core is readily available from [^99mTcO₄]^- and displays three labile water molecules that can be easily swapped for ligands with different denticity and/or donor atoms in aqueous environment. This makes it possible to radiolabel small molecules as well as high molecular weight molecules, such as antibodies or other proteins, while assuring biological activity. Direct radiolabelling of those proteins with fac-[^99mTc(CO)₃]⁺ under mild conditions is accomplished through incorporation of a polyhistidine tag (His-tag), a commonly used tag for purification of recombinant proteins. This review aims to address the direct radiolabelling of His-tagged macromolecules with fac-[^99mTc(CO)₃]⁺ for development of molecular imaging agents and the impact of this technology in the discovery and development of imaging and/or therapeutic agents towards clinical application.