title: How do HYNIC-conjugated peptides bind technetium? Insights from LC-MS and stability studies.
abstract: Hydrazinonicotinamide (HYNIC) is an established bifunctional complexing agent for technetium-99m ((99m)Tc) but the structure of the technetium coordination sphere remains uncertain. To gain further insight into this, we have prepared conjugates of HYNIC and hydrazinobenzoic acid (HYBA) with a model peptide, and radiolabelled them with (99m)Tc using three well-established co-ligand systems: EDDA, tricine and tricine-nicotinic acid. The labelled peptides were studied by LC-MS and by subjecting them to serum stability and protein binding assays. For each co-ligand system, HYNIC conjugates formed fewer and more stable labelled species than the corresponding HYBA conjugates. LC-MS analysis showed that all conjugates contained one hydrazine moiety bound to Tc, that binding of Tc to HYNIC-peptide and co-ligand occurs with displacement of 5H(+) indicating a Tc formal oxidation state of +5, and that the Tc has no oxo- or halide ligands. LC-MS also shows that complexes formed with the HYNIC conjugate contain fewer coordinating co-ligand molecules than the HYBA conjugate indicating that HYNIC is able to more effectively satisfy the coordination requirement of technetium, perhaps by binding in chelating mode.
authors.affiliation: Cancer Research UK, Department of Nuclear Medicine, St Bartholomew's Hospital, London, UK EC1A 7BE.
authors.forename: Robert C
authors.initials: RC
authors.lastname: King
authors.forename: M Bashir-Uddin
authors.initials: MB
authors.lastname: Surfraz
authors.forename: Stefano C G
authors.initials: SC
authors.lastname: Biagini
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Blower
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Mather
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D010455:Peptides / Q000737:chemistry*; D013667:Technetium / Q000737:chemistry*
chemical_list: D010455:Peptides; D013667:Technetium