A Simple Two-Step Approach for Introducing a Protected Diaminedithiol Chelator during Solid-Phase Assembly of Peptides

• Published in: Bioconjugate chemistry 2002-05, Vol.13 (3), p.679-684
• Main Authors: Gariépy, Jean, Rémy, Sandrine, Zhang, Xiuguo, Ballinger, James R, Bolewska-Pedyczak, Eleonora, Rauth, Michael, Bisland, Stuart K
• Format: Article
• Language: English
• Subjects: Animals; Chelating Agents - metabolism; CHO Cells - metabolism; Cricetinae; Diamines - chemistry; Organotechnetium Compounds - chemistry; Peptides - chemical synthesis; Peptides - chemistry; Toluene - analogs & derivatives; Toluene - chemistry
• ISSN: 1043-1802; 1520-4812
• DOI: 10.1021/bc0100814

A simple synthetic strategy is described to incorporate a protected diaminedithiol (N2S2) chelator during Fmoc solid-phase synthesis of short peptides. The resulting constructs could be efficiently labeled with technetium-99m (99mTc). The chelator was assembled at the N-terminus of peptides in a two-step procedure where the deprotected terminal amino group was first reacted with di-Fmoc-diaminopropionic acid (Fmoc-DAP-[Fmoc]-OH). The two protected amino groups were then simultaneously deprotected and subsequently reacted with S-benzoylthiolglycolic acid (TGA) to generate a protected N2S2 chelator. This metal binding site was introduced into di- and tripeptides. Each peptide construct was composed of a C-terminal lysine residue and an N-terminal diaminopropionic moiety modified to create the chelator site. The ε-amino group at the C-terminal lysine was further derivatized with a nitroimidazole group to facilitate cellular retention. The resulting constructs were then cleaved from the resin support, purified, and labeled with [99mTc]pertechnetate. Six constructs were prepared differing by a single amino acid inserted between the diaminopropionic acid and lysine residues. Optimal labeling yields of >70% were achieved around neutral pH and heating at 75 °C for 10 min. Purified 99mTc-labeled constructs were found to accumulate in Chinese hamster ovary (CHO) cells in vitro as a function of charge and hydrophobicity.