SITE-SPECIFIC CONJUGATION OF ETHIDIUM HOMODIMER TO B72.3 MAB - RETENTION OF IMMUNOREACTIVITY, ISOELECTRIC POINT AND IN-VIVO TUMOR TARGETING

In our initial studies, we developed a lysine-directed conjugation of EHD to B72.3 in which the protein-bound EHD was able to intercalate wi th DNA successfully. Compared with the native antibody, the immunoconj ugate retained full immunoreactivity. However, it exhibited an altered isoelectric point and poor tumor targeting. In the work reported here in, a site-specific conjugate was prepared by generating aldehydes in the Fc region of the antibody, conjugating cysteine-containing 14-mer peptide, and reacting this moiety with maleimido-EHD. The monomeric co njugate contained 6 EHD per antibody molecule, and the bound EHD inter calated fully with DNA as indicated by the fluorescence titration curv es. The conjugate not only retained its original isoelectric point and immunoreactivity but also showed substantial in vivo tumor targeting (similar to 21% ID/g).