21] Preparation and application of antibodies coupled to the A chain of ricin

Abstract

This chapter describes the selective cytotoxicity of a variety of normal and neoplastic cells by toxic peptides that have been covalently coupled to specific antibodies. The synthesis and applications of such antibody–toxin conjugates is also discussed in the chapter. Ricin is an example of a toxin that has been used in this manner. Ricin is obtained from castor beans and is composed of two 3 x 10 4 dalton polypeptide chains bridged by a disulfide bond. One chain (the B chain) has binding specificity for galactose and is responsible for the binding of the toxin to the surface of ricin-sensitive cells. Once bound, the other peptide chain (A chain) enters the cytoplasm and catalytically and irreversibly inhibits protein synthesis. The covalent coupling of antibody to the A chain of ricin can be performed by a disulfide exchange reaction mediated by the heterobifunctional, thiol-containing cross-linker N-succinimidyl-3-(2-pyridyldithio) propionate (SPDP). The hybrid product mimics the structure of the native toxin in that the A chain is covalently linked through a disulfide bridge to a binding moiety. Covalent coupling is performed in three steps: (1) purification of ricin A chain; (2) introduction of 2-pyridyl disulfides to the primary amino groups on the antibody; and (3) ricin A chain substitution via disulfide exchange.