Phosphonate-Based Aza-Macrocycle Ligands for Low-Temperature, Stable Chelation of Medicinally Relevant Rare Earth Radiometals and Radiofluorination

Abstract

Radioisotopes of fluorine (¹⁸F), scandium (^43/44Sc, ⁴⁷Sc), lutetium (¹⁷⁷Lu), and yttrium (⁸⁶Y, ⁹⁰Y) have decay properties ideally suited for targeted nuclear imaging and therapy with small biologics, such as peptides and antibody fragments. However, a single-molecule strategy to introduce these radionuclides into radiopharmaceuticals under mild conditions to afford inert in vivo complexes is critically lacking. Here, we introduce H₄L2 and H₄L3, two small-cavity macrocyclic chelator structural isomers bearing a single phosphonate functional group. Potentiometry and spectrophotometry were employed to determine H₄L2 and H₄L3's ability to form a single [M(L)]^- species with metals of different sizes (Sc³⁺, Lu³⁺, and Y³⁺) under physiologically relevant conditions. NMR spectroscopy and density functional theory (DFT) calculations suggest modulation of H₄L2 and H₄L3's inner-sphere hydration across the Sc³⁺/Lu³⁺/Y³⁺ series. Radiochemical labeling experiments with ¹⁸F, ⁴⁴Sc, ¹⁷⁷Lu, and ⁸⁶Y reveal that H₄L2 selectively chelates radioscandium at room temperature with high apparent molar activity (AMA, 462 mCi/μmol), while radiofluorination remains inaccessible. In contrast, H₄L3 enables room temperature radiochelation ⁴⁴Sc, ¹⁷⁷Lu, and ⁸⁶Y (AMA: 96-275 mCi/μmol) and incorporates ¹⁸F via the Sc-¹⁸F methodology to form [¹⁸F][ScF(L3)]^2-. In vivo biodistribution analysis at 1 h postinjection confirms the broad utility of H₄L3: all four radiochemical complexes clear off-target organs and remain >98% intact in urine metabolite analyses. The scope of room temperature radiochemical labeling, paired with facile ¹⁸F incorporation, to afford in vivo compatible complexes exceeds the clinical gold standard chelator DOTA and previously reported acyclic chelators, rendering H₄L3 promising for prospective radiopharmaceutical applications.

Figure 1.

Chemical structures of M-chelates employed for the chelation of M = Sc³⁺, Lu³⁺, and Y³⁺ as discussed within the introduction section, including novel structures H₄L2 and H₄L3 introduced in this work. Structures do not reflect the inner-sphere hydration status of the corresponding complexes which varies from 0–1 in dependence of ionic radius.

Figure 2.

pH-dependent speciation diagrams of M(L2) and M(L3) coordination complexes. The purple box denotes the physiologically relevant pH range.

Figure 3.

¹H NMR of [M(L)]⁻; 500 MHz in H₂O (0.1 M KCl, pH 7.0–7.4) referenced to trimethylsilyl propanoic acid. Left (from bottom to top): H₄L2 pH 7.24 (black), [Y(L2)]⁻ pH 7.20 (teal), [Lu(L2)]⁻ pH 6.98 (purple), and [Sc(L2)]⁻ pH 7.08 (pink). Right (from bottom to top): H₄L3 pH 7.08 (black), [Y(L3)]⁻ pH 7.17 (teal), [Lu(L3)]⁻ pH 7.37 (purple), and [Sc(L2)]⁻ pH 7.24 (pink). Square and triangle symbols indicate peaks corresponding to the respective hydration species (q = 0 and 1, respectively), as indicated in the reaction equilibrium scheme above.

Figure 4.

DFT structures of Δ-[Sc(L2)]⁻ and Δ-C-[Sc(L3)]⁻, generated using olex2. Annotated bond angles are listed in Table 3, and additional structural information, including those for enantiomeric and diastereomeric structures, can be found in Supporting Information Section 6.

Figure 5.

Percent radiochemical conversion as a function of ligand quantity for H₄L2 and H₄L3 at 25 °C with (A) ⁴⁴Sc, (B) ¹⁷⁷Lu, and (C) ⁸⁶Y at 30 min, pH 5.5, determined by radioTLC in triplicate and validated with radioHPLC. Activity: 28–53 μCi/reaction. Volume: 40–116 μL.

Figure 6.

Top: Biodistribution analysis of free ions and H₄L2 and H₄L3 complexes, respectively. Bottom: Urine metabolite analysis of [M(L3)]⁻ radiochemical complex (M = ⁴⁴Sc, ¹⁷⁷Lu, ⁸⁶Y) at 60 min postinjection.

Figure 7.

Chemical structure (left), biodistribution analysis of [¹⁸F]ScF-acetate and the [¹⁸F][ScF(L3)]²⁻ radiochemical complex 60 min postinjection (center), with radiochromatographic characterization and metabolite analysis (right).