Metallic radionuclides conjugated to biological vectors via an appropriate chelator are employed in nuclear medicine for the diagnosis (imaging) and radiotherapy of diseases. For the application of radiolabeled antibodies using positron emission tomography (immunoPET), zirconium-89 has gained increasing interest over the last decades as its physical properties (t_1/2 = 78.4 h, 22.6% β⁺ decay) match well with the slow pharmacokinetics of antibodies (t_biol. = days to weeks) allowing for late time point imaging. The most commonly used chelator for ⁸⁹Zr in this context is desferrioxamine (DFO). However, it has been shown in preclinical studies that the hexadentate DFO ligand does not provide ⁸⁹Zr-complexes of sufficient stability in vivo and unspecific uptake of the osteophilic radiometal in bones is observed. For clinical applications, this might be of concern not only because of an unnecessary dose to the patient but also an increased background signal. As a consequence, next generation chelators based on hydroxamate scaffolds for more stable coordination of ⁸⁹Zr have been developed by different research groups. In this review, we describe the progress in this research field until end of 2020, including promising examples of new candidates of chelators currently in advanced stages for clinical translation that outrun the performance of the current gold standard DFO.

DFO* is an octadentate chelator able to form highly stable chelates with Zirconium‐89 (⁸⁹Zr) for nuclear medicinal applications in Positron Emission Tomography (PET).^[1,2] The synthesis of DFO* and its scale‐up remains challenging by reported synthetic protocols. For this reason, we set out to develop a de novo synthesis of a hydroxamate‐containing building block suitable for the coupling to the commercially available DFO (desferrioxamine B, mesylate salt) yielding, after deprotection, the desired chelator DFO* in a more efficient procedure. Highlights of the new synthesis of DFO* reported herein are less synthetic steps and the isolation of the desired product DFO* by using solid phase extraction (SPE), thus avoiding tedious HPLC purification. DFO* is obtained in excellent purity (92‐98 %) and an overall yield of approximately 29 %. In addition, the isolated trifluoroacetic acid (TFA)‐salt of DFO* displays an improved solubility in organic solvents (DMSO, DMF, methanol), which will facilitate its use for the preparation of structurally diverse derivatives suitable for bioconjugation chemistry and the development of ⁸⁹Zr‐labeled radiotracers.

Radiolabeled peptides play a key role in nuclear medicine to selectively deliver radionuclides to malignancies for diagnosis (imaging) and therapy. Yet, their efficiency is often compromised by low metabolic stability. The use of 1,4‐disubstituted 1,2,3‐triazoles (1,4‐Tzs) as stable amide bond bioisosteres can increase the half‐life of peptides in vivo while maintaining their biological properties. Previously, the amide‐to‐triazole substitution strategy was used for the stabilization of the pansomatostatin radioligand [¹¹¹In]In‐AT2S, resulting in the mono‐triazolo‐peptidomimetic [¹¹¹In]In‐XG1, a radiotracer with moderately enhanced stability in vivo and retained ability to bind multiple somatostatin receptor (SSTR) subtypes. However, inclusion of additional 1,4‐Tz led to a loss of affinity towards SST₂R, the receptor overexpressed by most SSTR‐positive cancers. To enhance further the stability of [¹¹¹In]In‐XG1, alternative modifications at the enzymatically labile position Thr¹⁰‐Phe¹¹ were employed. Three novel 1,4,7,10‐tetraazacyclododecane‐1,4,7,10‐tetraacetic acid (DOTA)‐peptide conjugates were synthesized with a 1,4‐Tz (Asn⁵‐Ψ[Tz]‐Phe⁶) and either a β‐amino acid (β‐Phe¹¹), reduced amide bond (Thr¹⁰‐Ψ[NH]‐Phe¹¹), or N‐methylated amino acid (N‐Me‐Phe¹¹). Two of the new peptidomimetics were more stable in blood plasma in vitro than [¹¹¹In]In‐XG1. Yet none of them retained high affinity towards SST₂R. We demonstrate for the first time the combination of the amide‐to‐triazole substitution strategy with alternative stabilization methods to improve the metabolic stability of tumor‐targeting peptides.

Background: Actinium-225 (²²⁵Ac) has gained interest in nuclear medicine for use in targeted alpha therapy (TAT) for the treatment of cancer. However, the number of suitable chelators for the stable complexation of ²²⁵Ac³⁺ is limited. The promising physical properties of ²²⁵Ac result in an increased demand for the radioisotope that is not matched by its current supply. To expand the possibilities for the development of ²²⁵Ac-based TAT therapeutics, a new hydroxamate-based chelator, DFO*¹², is described. We report the DFT-guided design of dodecadentate DFO*¹² and an efficient and convenient automated solid-phase synthesis for its preparation. To address the limited availability of ²²⁵Ac, a small-scale ²²⁹Th/²²⁵Ac generator was constructed in-house to provide [²²⁵Ac]AcCl₃ for research. Methods: DFT calculations were performed in ORCA 5.0.1 using the BP86 functional with empirical dispersion correction D3 and Becke–Johnson damping (D3BJ). The monomer synthesis over three steps enabled the solid-phase synthesis of DFO*¹². The small-scale ²²⁹Th/²²⁵Ac generator was realized by extracting ²²⁹Th from aged ²³³U material. Radiolabeling of DFO*¹² with ²²⁵Ac was performed in 1 M TRIS pH 8.5 or 1.5 M NaOAc pH 4.5 for 30 min at 37 °C. Results: DFT calculations directed the design of a dodecadentate chelator. The automated synthesis of the chelator DFO*¹² and the development of a small-scale ²²⁹Th/²²⁵Ac generator allowed for the radiolabeling of DFO*¹² with ²²⁵Ac quantitatively at 37 °C within 30 min. The complex [²²⁵Ac]Ac-DFO*¹² indicated good stability in different media for 20 h. Conclusions: The novel hydroxamate-based dodecadentate chelator DFO*¹², together with the developed ²²⁹Th/²²⁵Ac generator, provide new opportunities for ²²⁵Ac research for future radiopharmaceutical development and applications in TAT.

The use of metabolically stabilized, radiolabeled somatostatin (SST) analogs ([⁶⁸Ga]Ga/[¹⁷⁷Lu]Lu-DOTA-TATE/TOC/NOC) is well established in nuclear medicine. Despite the pivotal role of these radioligands in the diagnosis and therapy of neuroendocrine tumors (NETs), their inability to interact with all five somatostatin receptors (SST_1–5R) limits their clinical potential. [¹¹¹In]In-AT2S is a radiolabeled DOTA-conjugate derived from the parent peptide SST-14 that exhibits high binding affinity to all SSTR subtypes, but its poor metabolic stability represents a serious disadvantage for clinical use. In order to address this issue, we have replaced strategic trans-amide bonds of [¹¹¹In]In-AT2S with metabolically stable 1,4-disubstituted 1,2,3-triazole bioisosteres. From the five cyclic triazolo-peptidomimetics investigated, only [¹¹¹In]In-XG1 combined a preserved nanomolar affinity for the SST_1,2,3,5R subtypes in vitro and an improved stability in vivo (up to 17% of intact peptide 5 min postinjection (pi) versus 6% for [¹¹¹In]In-AT2S). The involvement of neprilysin (NEP) in the metabolism of [¹¹¹In]In-XG1 was confirmed by coadministration of Entresto^®, a registered antihypertensive drug, in vivo releasing the selective and potent NEP-inhibitor sacubitrilat. A pilot SPECT/CT imaging study conducted in mice bearing hSST₂R-positive xenografts failed to visualize the xenografts due to the pronounced kidney uptake (>200% injected activity (IA)/g at 4 h pi), likely the result of the formation of cationic metabolites. To corroborate the imaging data, the tumors and the kidneys were excised and analyzed with a γ-counter. Even if receptor-specific tumor uptake for [¹¹¹In]In-XG1 could be confirmed (1.61% IA/g), further optimization is required to improve its pharmacokinetic properties for radiotracer development.