DOTAGA-Trastuzumab. A New Antibody Conjugate Targeting HER2/Neu Antigen for Diagnostic Purposes

Improved bifunctional chelating agents (BFC) are required for indium-111 radiolabeling of monoclonal antibodies (mAbs) under mild conditions to yield stable, target-specific agents. 2,2',2 ''-(10-(2,6-Dioxotetrahydro-2H-pyran-3-yl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid (DOTAGA-anhydride) was evaluated for mAb conjugation and labeling with indium-111. The DOTA analogue was synthesized and conjugated to trastuzumab which targets the HER2/neu receptor in mild conditions (PBS pH 7.4, 25 degrees C, 30 min) and gave a mean degree of conjugation of 2.6 macrocycle per antibody. Labeling of this immunoconjugate with indium-111 was performed in 75% yield after 1 h at 37 degrees C, and the proportion of In-111-DOTAGA-trastuzumab reached 97% after purification. The affinity of DOTAGA-trastuzumab was 5.5 +/- 0.6 nM as evaluated by in vitro saturation assays using HCC1954 breast cancer cell line. SPECT/CT imaging and biodistribution studies were performed in mice bearing breast cancer BT-474 xenografts. BT-474 tumors were clearly visualized on SPECT images at 24, 48, and 72 h postinjection. The tumor uptake of [In-111-DOTAGA]-trastuzumab reached 65%ID/g 72 h postinjection. These results show that the DOTAGA BFC appears to be a valuable tool for biologics conjugation.