1P-398 : Development of an immunotoxin based on a full-length antibody via site-specific conjugation using a cysteine residue introduced to IgG and an unnatural amino acid

Immunotoxins consisting of a toxin from bacteria or plants, and a targeting module have been developed as potent anti-cancer therapeutics. The majority of them, especially those in preclinical or clinical testing stages, are based on antibody fragments, even though the advantage of using full-length antibodies has been well documented. Here, we generated an immunotoxin via site-specific conjugation using a cysteine (Cys) residue introduced to IgG and a bio-orthogonally reactive unnatural amino acid. A Her2-targeting IgG, trastuzumab, was engineered to have an unpaired Cys at position 425 in the heavy chain, and an unnatural amino acid having the azido group (azidophenylalanine) was incorporated into an engineered Pseudomonas exotoxin A (PE24). The two protein molecules were conjugated site-specifically using a bifunctional linker having dibenzocyclooctyne and maleimide groups. The resulting trastuzumab-PE24 conjugate was cytotoxic to Her2-overexpressing cell lines.