In vitro evaluation of the monoclonal antibody 64Cu-IgG M75 against human carbonic anhydrase IX and its in vivo imaging

Highlights: • Labelling of the monoclonal antibody IgG M75 with ⁶⁴Cu resulting using a new chelator. • Specific binding of ⁶⁴Cu-IgG M75 to the HT-29 cells was found to be 45 ± 2.8% in in vitro experiment. • The K_D value of the ⁶⁴Cu-IgG M75 was measured and found to be 9.2 nM. • In vivo imaging of the mouse inoculated with the HT-29 cells using ⁶⁴Cu-IgG M75 indicated increasing tumour-to-background ratio in time. - Abstract: Specific oncology diagnostics requires new types of the selective radiopharmaceuticals, particularly those suitable for the molecular PET imaging. The aim of this work is to present a new, specific PET-immunodiagnostic radiopharmaceutical based on the monoclonal antibody IgG M75 targeting human carbonic anhydrase IX labelled with ⁶⁴Cu (T_½ = 12.70 h) and its in vitro and in vivo evaluation. The antibody IgG M75 was conjugated with a non-commercial copper-specific chelator "phosphinate" and then labelled with the positron emitter ⁶⁴Cu. Stability of the labelled conjugated was tested in human serum. The immunoreactivity of the labelled conjugate was evaluated in vitro on a suitable cell cultures of the colorectal carcinoma (HT-29) and its imaging properties were estimated in vivo on a mouse model with inoculated colorectal carcinoma HT-29 imaged on a µPET/CT. The tested radioimmunoconjugate was obtained in a specific activity of 0.25–0.5 MBq/µg. In vitro uptake experiments revealed specific binding to the HT-29 cells (45 ± 2.8% of the total added activity) and the measured K_D value was found to be 9.2 nM. Imaging clearly demonstrated significant uptake of the labelled monoclonal antibody in the tumour at 18 h post administration. The radioimmunoconjugate ⁶⁴Cu-PS-IgG M75 seems to be a suitable candidate for PET diagnostics of hypoxic tumours expressing human carbonic anhydrase IX.