Heptadentate chelates for ⁸⁹Zr-radiolabelling of monoclonal antibodies

Herein, we report the synthesis of three new bifunctional heptadentate metal ion binding chelates derived from desferrioxamine B (DFO) linked to a tripeptide unit that comprises of a glutamic acid and two glycine residues. The three DFO derivatives were also functionalised with a photoactivatable aryl azide unit for light-triggered labelling of proteins. The chelates were obtained in 3 synthetic steps in good overall yields by using solid phase peptide synthesis (SPPS). Density Functional Theory (DFT) calculations were used to estimate thermodynamic formation constants (log β) of the corresponding Zr⁴⁺ complexes. Quantitative zirconium-89 radiolabelling (>95%) was obtained in <5 min at room temperature, and the stability of the radioconjugates toward different competitors (human serum, EDTA and Fe³⁺) was assessed in vitro. One-pot ⁸⁹Zr-photoradiosynthesis produced [⁸⁹Zr]Zr-2-onartuzumab directly from the formulated, clinical-grade sample MetMAb™, without pre-purifying the monoclonal antibody (mAb) component, with an isolated decay-corrected radiochemical yield of 36.4 ± 2.4%. PET imaging and biodistribution studies were performed in female athymic nude mice bearing subcutaneous xenografts derived from the MKN-45 human gastric cancer cell line to assess the pharmacokinetic profile and tumour binding of [⁸⁹Zr]Zr-2-onartuzumab. Specific tumour uptake of [⁸⁹Zr]Zr-2-onartuzumab was confirmed by using competitive inhibition (blocking) studies and bone uptake was significantly reduced compared to the parent DFO analogue.