LYSINE-DIRECTED CONJUGATION OF ETHIDIUM HOMODIMER TO B72.3 ANTIBODY -RETENTION OF IMMUNOREACTIVITY BUT ALTERED TUMOR TARGETING

Ethidium homodimer (EHD) was conjugated to B72.3 monoclonal antibody u sing a method whereby 85-90% of the conjugated EHD remains available f or DNA intercalation. Antibody was thiopropionylated by reaction with N-succinimidyl 3-(2-pyridyldithio)propionate and reduction of pyridyld ithio groups with dithiothreitol. EHD was maleimido-functionalized wit h succinimidyl 4-(N-maleimidoethyl) cyclohexane-1-carboxylate and trea ted with thiopropionylated antibody to obtain a conjugate containing s imilar to 3.4 EHD per antibody molecule. For biologic studies, C-14-la beled EHD was synthesized by reductive amination and conjugated as abo ve. In vitro the conjugate maintained chemical integrity and immunorea ctivity, while in vivo its targeting of LS174T tumors was reduced comp ared with that of iodinated antibody. A decrease in isoelectric point of the immunoconjugate was also observed. (C) 1998 Elsevier Science In c.