To sandwich technetium: highly functionalized bis-arene complexes [99mTc(η6-arene)2]+ directly from water and [99mTcO4].

The labeling of (bio)molecules with metallic radionuclides such as 99mTc demands conjugated, multidentate chelators. This is not always necessary since phenyl rings can directly serve as integrated, organometallic ligands. Bis-arene sandwich complexes are generally prepared by the Fischer-Hafner reaction. In extension, we show that [99mTc(η6-C6R6)2]+ type complexes are directly accessible from water and [99mTcO4]-, even with arenes incompatible with Fischer-Hafner conditions. To unambiguously confirm the nature of these unprecedented 99mTc complexes, their rhenium homologous have been prepared by substituting naphthalene ligands in [Re(η6-C10H8)2]+ with the corresponding phenyls. The ease with which highly stable [99mTc(η6-C6R6)2]+ are formed under standard labeling conditions enables a multitude of new potential imaging agents, based on commercial pharmaceuticals or lead structures.

Abstract: The labeling of (bio)molecules with metallic radionuclides such as 99m Tc demands conjugated, multidentate chelators. This is not always necessary since phenyl rings can directly serve as integrated, organometallic ligands. Bis-arene sandwich complexes are generally prepared by the Fischer-Hafner reaction. In extension, we show that [ 99m Tc(η 6 -C6R6)2] + type complexes are directly accessible from water and [ 99m TcO4] -, even with arenes incompatible with Fischer-Hafner conditions. To unambiguously confirm the nature of these unprecedented 99m Tc complexes, their rhenium homologous have been prepared by substituting naphthalene ligands in [Re(η 6 -C10H8)2] + with the corresponding phenyls. The ease with which highly stable [ 99m Tc(η 6 -C6R6)2] + are formed under standard labeling conditions enables a multitude of new potential imaging agents, based on commercial pharmaceuticals or lead structures.
In molecular imaging, targeting vectors or pharmaceutically active compounds are combined with radionuclides for the non-invasive detection of e.g. high proliferation rates, increased receptor densities and other abnormal physiological features. [1][2] The labeling of such vectors, often represented by comparably small molecules, relies typically on the direct formation of covalent bonds such as C-18 F or C-123 I. [3][4] Thereby, the basic lead structure of the molecule stays essentially intact. Many examples for such molecular imaging agents are in clinical application, [ 18 F]-FDG being probably the most prominent example. In contrast, the labeling with metallic radionuclides requests chelators for stabilizing the radionuclide against trans-metalation with ubiquitous and competing ligand sites in e.g. proteins (scheme 1, top). [5] These multidentate ligands are often bulky and their molecular weights may exceed the ones of the lead structures. DOTA for 68 Ga, DTPA or MAG3 for 99m Tc or 111 In and other examples evidence this situation. [6][7] Since the labeled chelator bound to the vector substantially influences pharmacology, rational predictions for successful molecular imaging agents are often affected. Chelators are mandatory for main group metals but may be replaced by comparably small ligands for d-elements. In this respect, cyclopentadiene (HCp) is small and the hypothesis that it can replace phenyls in pharmaceutically active lead structures has been shown in many examples, ferrocifen, an Scheme 1. Classic labelling procedure: i) derivatization of a lead structure with a chelator and ii) subsequent labelling with [ 99m TcO 4 ]-. iii) direct labelling at a phenyl ring without bifunctional chelator with [ 99m TcO 4 ]-in H 2 O leads to "twinning" of the lead structures in sandwich complexes.
Arene complexes of rhenium and even more for technetium are scarcely i nvestigated, despite their discovery some 60 years ago. [21][22] Being inspired by the opportunity of labeling molecules directly via phenyl groups and without chelators, we recently reported about the syntheses and properties of a variety of [M(η 6arene)2] + ( M= 99 Tc, 99m Tc, Re) complexes. [23][24][25] The syntheses followed classical Fischer-Hafner conditions, i.e. with AlCl3 and Zn° or Al° as reductants. The extreme stabilities towards base, acid, air and water encouraged us to aim at a direct labeling of functionalized phenyls beyond the simple alkyl-bearing ones (scheme 1, bottom). Oxygen-or nitrogen functionalities are incompatible with Fischer-Hafner conditions. Furthermore, for binding e.g. pharmaceuticals to 99m Tc, the reactions have to be performed in water, even more incompatible with Fischer-Hafner conditions. We report here about a new method for preparing [ 99m Tc(η 6arene)2] + s andwiches with N-and O-functionalized arenes directly from water and [ 99m TcO4]in moderate to very good yields and high radiochemical purities.
The reaction of [Re(η 6 -naphthalene)2] + with the neat arenes, e.g. paracetamol or p-cresol, at high temperatures lead to the desired products [Re(η 6 -C6R6)2] + , albeit in variable yields. Separation by preparative HPLC and subsequent crystallization gave the pure products, which could now be compared by analytical HPLC with the compounds formed with 99m Tc. Figure 1 shows the X-ray  Arenes are thus directly labeled, even if they are functionalized with potentially coordinating groups such as amino-or hydroxyl functions, this is a core message from this report. This mode of reactivity is unprecedented and has not been considered at all in labeling procedures so far. The formation of [M(η6-arene)2]+ complexes may well account for unidentified side products formed during standard labeling reactions of molecules comprising phenyl groups.
The reaction mechanisms consist of an in total 6ereduction from Tc VII to Tc I , in the transfer of eight protons for the formation of four H2O molecules together with the coordination of the two arenes.
Many competing ligands such as halides, water and electrolyte anions are present in solution. Still, the arene as an atypical ligand in water coordinates to the once reduced 99m Tc center. Multiple elementary steps must be involved in this highly complex reaction scheme. We can only speculate about the mechanism but three stereochemical features should still be noted; bis-or multi-functionalized phenyls with ortho-, meta-or multisubstituents will yield two stereoisomers as observed with 2,4dimethylaniline (scheme 4, upper line for 99m Tc complex, Scheme S7 for Re-complex). The HPLC γ-trace of the 99m Tc complex and the UV/vis trace of the Re homologue show one peak but the NMR spectrum clearly evidenced two compounds (Figures S14 and S33). In case of chiral side chains, three stereoisomers are expected due to the prochirality of the 99m Tc center. A reaction with two different arenes of about the same reactivities towards bis-arene complex formation and solubilities, p-cresol and paracetamol, gave the three complexes [ 99m Tc(η 6 -p-cresol)2] + , [ 99m Tc(η 6 -paracetamol)2] + and the mixed arene complex [ 99m Tc(η 6 -p-cresol)(η 6 -paracetamol)] + (scheme 4 lower line and figures S2-S6). As shown in figure 3, these complexes are clearly discernible on the HPLC analysis. We note that the traces remain unchanged when the solution was kept in air over night, corroborating the stabilities of bis-arene complexes of 99m Tc. Whereas these expected and actually found stereochemical or stochastic features serve as indirect proofs for the authenticities of [ 99m Tc(η 6 -arene)2] + complexes, only a comparison with a fully characterized 99 Tc or Re complex will confirm unambiguously their identities . Scheme 4. Stereochemical and stochastic features for multifunctionalized arenes and unsymmetrical products when labeling two different arenes the same time with 99m Tc.
In conclusion, sandwich complexes of the bis-arene-type can be prepared with 99m Tc directly from [ 99m TcO4]and in water in a convenient procedure. Potentially, technetium can be sandwiched with many organic molecules, comprising phenyls without the need for conjugating additional chelators. Exact conditions need to be optimized from phenyl to phenyl but the given reaction schemes can be applied to a wide variety of phenyls. These general routes allow access to a multitude of new molecular imaging agents and are applicable to the labeling of biomolecules. We emphasize that unidentified side products in other standard labeling procedures may well account for sandwich-type 99m Tc complexes.