N^ε functionalization of metal and organic protected L-histidine for a highly efficient, direct labeling of biomolecules with [Tc(OH₂)₃(CO)₃]⁺

Two different pathways for the introduction of an acetyl group at N^ε in a N^α, N^δ, and -COO protected histidine to afford N^ε-(CH₂COOH)-histidine derivative 7b are presented. The purpose of this study is the coupling of 7b to amino groups in bioactive molecules such as peptides. After full deprotection of such a bioconjugate, histidine provides three coordination sites which efficiently coordinate to [^99mTc(OH₂)₃-(CO)₃]⁺ or [Re(OH₂)₃(CO)₃]⁺ in a facial geometry. This allows the development of novel radiopharmaceuticals. Selective derivatization at the N^ε position has conveniently been achieved by concomitant protection of N^α and N^δ with a carbonyl group forming a six-membered urea. Cyclic urea ring opening with Fm-OH, coupling of phenylalanine as a model to 7b through its primary amine and removing of all protecting groups in one step gave a histidine derivative of phenylalanine which could be labeled at 10^-5M with ^99mTc in very high yield and even in about 50% yield at 10^-6M. The X-ray structure of a complex with [Re-(CO)₃]⁺ in which anilin is coupled to 7b confirms the facial arrangement of histidine. A second pathway applies directly the [Re(CO)₃]⁺ moiety as a protecting group. This is one of the rare examples in which a metal fragment is used as a protecting group for organic functionalities. The coordination to histidine protects the N^α, N^δ and COO group in one single step, subsequent alkylation with BrCH₂COOH(R) at N^ε, coupling to phenylalanine and oxidative deprotection of [Re(CO)₃]⁺ to [ReO₄]^- gave the corresponding bioconjugate in which histidine is coupled to phenylalanine through an acetylamide at N^ε. Both methods offer convenient pathways to introduce histidine in a biomolecule under retention of its three coordination sites. The procedures are adaptable to any biomolecule with pendant amines and allow the development of novel radiopharmaceuticals or inversed peptides.