# Labeling Peptides with Radioiodine: An Overview of Traditional and Emerging Techniques
## 4 Indirect Radioiodination Methods: The Labelling of the Prosthetic Groups
### Iodonium Salts  
Iodonium salts are highly reactive electrophilic reagents widely employed in radioiodination reactions due to their excellent leaving group ability and compatibility with mild conditions. Their use enables efficient labeling of aromatic compounds with radioiodine isotopes, facilitating the synthesis of radiotracers for imaging and therapeutic applications [2]. [PERSON]'s study provides several new alternatives to conventional and sub-optimal approaches currently in use for radioiodination and astatination of biomolecules (Figure 13) [56]. The study proposes the use of \({}^{125}\)I- and \({}^{211}\)At-labeled azide and tetrazine prosthetic groups for bioorthogonal conjugation, designed and tested in a comparative study of five bioorthogonal systems. Radiolabeling, in particular, was performed from precursors of \({}^{125}\)I airlidonium salts. All five biocconjugation reactions conducted on a clickable model peptide produced quantitative yields in times ranging from less than a minute to several hours, depending on the system used. Finally, one of the bioorthogonal systems was used for tagging an IgG.  
Figure 12: Standard example of radioiodination through diazonium salts.  
A comparative study of the radiolabogenation of arylhalogen salts with \({}^{125}\)I and \({}^{211}\)At was performed by [PERSON] and colleagues [57]. Initial experiments were performed on a model compound, which showed the higher reactivity of astatide compared with iodide. From the kinetic studies, a significant difference in the activation energy (Ea = 23.5 and 17.1 kcal mol\({}^{-1}\) with \({}^{125}\)I and \({}^{211}\)At, respectively) was inferred along with computational studies. The good to excellent regioselectivity of the halogenation and the high yields obtained with variously substituted arylodonium salts indicate that this class of compounds is a promising alternative to the stannan chemistry currently used for radiolabogen labeling of tracers in nuclear medicine. The same group describes the development of an alternative approach to arylstannan chemistry for radiolabeling antibodies with radioiodine or astatine, based on precursors of arylodium salts [58]. Bifunctional aryl salts were designed and tested for the synthesis of \({}^{125}\)I- and \({}^{211}\)At labeled prosthetic groups for bioconjugation (Figure 14). The nature of the electron-rich aryl group was varied and its impact on the regioselectivity of radiolabogenation was evaluated. Under optimized conditions, both radioiodination and astatination were performed very efficiently under mild conditions (radiochemical yields > 85%). The ionic nature of the precursors was exploited to develop an efficient purification approach: the HPLC step, usually required in conventional approaches to optimize removal of toxic organotin precursors and side products, was replaced by filtration through a silica cartridge with significantly reduced loss of radiolabeled product. The radioiodinated and astatinated prosthetic groups were then efficiently conjugated to an anti-CD138 monoclonal antibody (conjugation yield of 75-80%), exploiting the \(N\)-hydroxysuccinidyl ester group.  
Figure 13: \({}^{125}\)I-labeled azide and tetrazine prosthetic groups for bioorthogonal conjugation.