Toward Bifunctional Chelators for Thallium-201 for Use in Nuclear Medicine

Auger electron therapy exploits the cytotoxicity of low-energy electrons emitted during radioactive decay that travel very short distances (typically <1 microm). ²⁰¹Tl, with a half-life of 73 h, emits ∼37 Auger and other secondary electrons per decay and can be tracked in vivo as its gamma emissions enable SPECT imaging. Despite the useful nuclear properties of ²⁰¹Tl, satisfactory bifunctional chelators to incorporate it into bioconjugates for molecular targeting have not been developed. H₄pypa, H₅decapa, H₄neunpa-NH₂, and H₄noneunpa are multidentate N- and O-donor chelators that have previously been shown to have high affinity for ¹¹¹In, ¹⁷⁷Lu, and ⁸⁹Zr. Herein, we report the synthesis and serum stability of [^nat/201Tl]Tl³⁺ complexes with H₄pypa, H₅decapa, H₄neunpa-NH₂, and H₄noneunpa. All ligands quickly and efficiently formed complexes with [²⁰¹Tl]Tl³⁺ that gave simple single-peak radiochromatograms and showed greatly improved serum stability compared to DOTA and DTPA. [^natTl]Tl-pypa was further characterized using nuclear magnetic resonance spectroscopy (NMR), mass spectroscopy (MS), and X-ray crystallography, showing evidence of the proton-dependent presence of a nine-coordinate complex and an eight-coordinate complex with a pendant carboxylic acid group. A prostate-specific membrane antigen (PSMA)-targeting bioconjugate of H₄pypa was synthesized and radiolabeled. The uptake of [²⁰¹Tl]Tl-pypa-PSMA in DU145 PSMA-positive and PSMA-negative prostate cancer cells was evaluated in vitro and showed evidence of bioreductive release of ²⁰¹Tl and cellular uptake characteristic of unchelated [²⁰¹Tl]TlCl. SPECT/CT imaging was used to probe the in vivo biodistribution and stability of [²⁰¹Tl]Tl-pypa-PSMA. In healthy animals, [²⁰¹Tl]Tl-pypa-PSMA did not show the myocardial uptake that is characteristic of unchelated ²⁰¹Tl. In mice bearing DU145 PSMA-positive and PSMA-negative prostate cancer xenografts, the uptake of [²⁰¹Tl]Tl-pypa-PSMA in DU145 PSMA-positive tumors was higher than that in DU145 PSMA-negative tumors but insufficient for useful tumor targeting. We conclude that H₄pypa and related ligands represent an advance compared to conventional radiometal chelators such as DOTA and DTPA for Tl³⁺ chelation but do not resist dissociation for long periods in the biological environment due to vulnerability to reduction of Tl³⁺ and subsequent release of Tl⁺. However, this is the first report describing the incorporation of [²⁰¹Tl]Tl³⁺ into a chelator-peptide bioconjugate and represents a significant advance in the field of ²⁰¹Tl-based radiopharmaceuticals. The design of the next generation of chelators must include features to mitigate this susceptibility to bioreduction, which does not arise for other trivalent heavy radiometals.