In vivo stable bisarylmercury bispidine as a tool for Hg‐197(m) applications

Intro
Reactor‐produced Hg‐197 saw previous medical use in radiolabelled chlormerodrin for SPECT imaging[1] in the 1960s and 70s but was discontinued because of the up-and‐coming Tc‐99m generator system's widespread use, the uncertain in vivo stability of Hg‐197 labelled compounds and the low molar activity of the Hg‐197 itself (<2 GBq/μmol).[2] Cyclotron produced Hg‐197(m) is able to overcome the chemical toxicity issue, due to the much higher molar activity achieved (>500 GBq/μmol),[3] allowing access, at innocuous mercury concentrations, to the theranostically useful decay modes and half‐life of the radiometal's metastable nuclear isomer (γ for SPECT imaging and conversion and auger electrons for tumour therapy. Hg‐197g T1/2 = 64.1 h, Hg‐197m T1/2 = 23.8 h).
Objective
The development of an in vivo stable mercury compound able to conjugate to a cancer‐targeting carrier. Radiopharmaceutical applications obviously require high in vivo stability but the fast metabolism of most mercury compounds in solution is a prevalent issue.[4] However, Hg‐C organometallics show good water‐stability and bypass the issue of Hg‐S bonds suffering from competition by common thiol‐containing biomolecules, e.g., cysteine. Therefore, this project is specifically focussed on the strongest of this kind: the mercury‐phenyl bond.[5] Previous study has shown that the syntheses of monodentate ligands for κ1‐L2Hg species suffer from significant cleavage. [6] For this reason, our research is centred on the synthesis of a bidentate chelator design, due to the entropic advantage, the improved formation kinetics and the stability imparted by steric shielding.
Current Results
Separation from the gold target leaves the produced Hg‐197(m) in an acidic aqueous solution as the chloride salt. Consequently, transmetallation, via boronic acid or stannyl derivatives, was chosen as a feasible route for forming the mercury‐carbon bonds. Following several chelator attempts, all showing low selectivity for the 1:1‐compound, a better fitting structure was found using the bispidine backbone (being known in co‐ordination chemistry for a variety of metals, good bio‐stability and its bridge linking‐functionalisation).[7] After improvement of the reaction conditions, radiolabelling experiments showed good evidence of specific binding with the bispidine derivative 9‐butyl‐1,5‐diphenyl‐3,7‐bis(2‐(trimethylstannyl)benzyl)‐3,7‐diazabicyclo[3.3.1]nonan‐9‐ol “L1(SnMe3)2,” as analysed by thiol‐impregnated radio‐TLC and supported by radio‐HPLC, whilst ESI‐MS of the stable mercury compound provided evidence that the desired 1:1 product had been formed. The in vitro stability tests conducted on the radiolabelled compound showed exciting results, with negligible degradation after 5 days (thereafter the activity was below analytical levels) in solutions with excess glutathione and 2,2′,2″‐nitrilotris (ethane‐1‐thiol). The only noticeable (ca. 5% degradation after 48 h), but expected reaction, was with sodium sulphide. Biodistribution experiments in healthy male Wistar rats showed no build‐up in the kidneys, with excretion occurring through the liver, indicating no de‐metallation.
Summary
The bisarylmercury bispidine (L1Hg) shows good stability in vitro as well as in vivo and is a promising candidate as a biocompatible mercury binding compound. Further
research is continuing into full characterisation, conjugation to peptides and proteins, subsequent in vivo studies and for derivatives based on the structural design.

ACKNOWLEDGEMENTS
Regina Herrlich, Ulrike Gesche, Thomas Wünsche. Hg‐197(m) measurements were carried out at the CANAM infrastructure of the NPI CAS Rez supported through MEYS project no. LM2011019.

REFERENCES
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