Measurement of the rate of copper(II) exchange for 64Cu complexes of bifunctional chelators

Measurement of the rate of loss of 64Cu from the chelators used in the preparation of 64Cu-labeled proteins is critical to the development of effective 64Cu-labeled radiopharmaceuticals. Typically, however, this assessment has been made indirectly, by comparison of the relative uptake of the labeled proteins in the liver, or under non-physiological conditions such as hot 5 M HCl. In the present study, we measured the rate of loss of 64Cu from a series of chelators (DOTA, TETA, and NOTA) that are commonly used to label proteins as a function of [Cu2+] (1, 2, and 5 μM) and pH (7.5, 6.0. and 5.0). We found that 64Cu-NOTA is somewhat more kinetically stable than 64Cu-TETA and that both 64Cu-NOTA and 64Cu-TETA are much more kinetically stable than 64Cu-DOTA. Furthermore, the rate of loss of 64Cu from Cu-DOTA increased with higher [Cu2+] and pH while the rate of loss of 64Cu from Cu-TETA and Cu-NOTA was minimally dependent on [Cu2+] and pH. These results suggest that NOTA is preferable to TETA and DOTA for the preparation of 64Cu-labeled proteins.