As part of a coordinated program to develop effective, specific, antibody-ricin (ricini A-chain) conjugates for use in immunotherapy, the studies described in this proposal seek to: (a) more fully understand the process of toxin and immunotoxin internalization and degradation by hematopoietic cells; (b) to characterize further the effects of lysosomotropic amines and proton ionophores on the cellular processing of toxins and conjugates; and (c) to produce whole-ricin immunotoxins with modified ricin moieties which no longer recognize cell-surface terminal galactose or N-acetylgalactosamine residues. More specifically we plan to study the effects of conjugation on antibody binding and affinity, to characterize the cellular metabolic products of the toxin and antibody moieties both in the presence and absence of lysosomotropic amines, proton ionophores, and other potentiating reagents, to characterize the functional domains of the ricin A- and B-chain polypeptides using anti-peptide antisera, and to chemically modify the ricin B-chain such that it retains only its translocation properties. This work is both timely and important. Timely because of the currently intense evaluation of immunotoxins as immunotherapeutic reagents, and important because of the as yet poorly understood characteristics which comprise an effective immunotoxin. Only when the significance of these characteristics is fully appreciated will truly effective immunotoxins be available for use in chemotherapeutic protocols.
