Comparison of bifunctional chelates for 64Cu antibody imaging

Improved bifunctional chelates (BFCs) are needed to facilitate efficient Cu-64 radiolabeling of monoclonal antibodies (mAbs) under mild conditions and to yield stable, target-specific agents. The utility of two novel BFCs, 1-Oxa-4,7,10-triazacyclododecane-5-S-(4-isothiocyanatobenzyl)-4,7,10-triacetic acid (p-SCN-Bn-Oxo-DO3A) and 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-4-S-(4-isothiocyanatobenzyl)-3,6,9-triacetic acid (p-SCN-Bn-PCTA), for mAb imaging with Cu-64 were compared to the commonly used S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-tetraacetic acid (p-SCN-Bn-DOTA). The BFCs were conjugated to trastuzumab, which targets the HER2/neu receptor. Cu-64 radiolabeling of the conjugates was optimized. Receptor binding was analyzed using flow cytometry and radioassays. Finally, PET imaging and biodistribution studies were done in mice bearing either HER2/neu-positive or HER2/neu-negative tumors. Cu-64-Oxo-DO3A- and PCTA-trastuzumab were prepared at room temperature in > 95% radiochemical yield (RCY) in < 30 min, compared to only 88% RCY after 2 h for the preparation of Cu-64-DOTA-trastuzumab under the same conditions. Cell studies confirmed that the immunoreactivity of the mAb was retained for each of the bioconjugates. In vivo studies showed that Cu-64-Oxo-DO3A- and PCTA-trastuzumab had higher uptake than the Cu-64-DOTA-trastuzumab at 24 h in HER2/neu-positive tumors, resulting in higher tumor to background ratios and better tumor images. By 40 h all three of the Cu-64-BFC-trastuzumab conjugates allowed for clear visualization of the HER2/neu-positive tumors but not the negative control tumor. The antibody conjugates of PCTA and Oxo-DO3A were shown to have superior Cu-64 radiolabeling efficiency and stability compared to the analogous DOTA conjugate. In addition, Cu-64-PCTA and Oxo-DO3A antibody conjugates may facilitate earlier imaging with greater target to background ratios than the analogous Cu-64-DOTA antibody conjugates.