Kinetics of protein synthesis inactivation by ricin-anti-Thy 1.1 monoclonal antibody hybrids. Role of the ricin B subunit demonstrated by reconstitution.

Antibody-toxin conjugates of different compositions have been reported to have varying differential toxicities between target and non-target cells in uitro. In this report, we compare the kinetics of protein synthesis inhibition of several different types of anti-Thy 1.1-ricin hybrids. Utilizing two monoclonal antibodies which vary in affinity for the Thy 1.1 antigen by >IO3, thioether-linked hybrids have been made with intact ricin and disulfide-linked hybrids made with ricin A chain. Protein synthesis inactivation kinetics of ricin A chain-S-S-anti-Thy 1.1 IgG is first order and the maximal rate occurs when the Thy 1.1 receptors of AKR cells are saturated. The concentration of the high afiinity anti-Thy 1.1 ricin A chain hybrid required to inhibit protein synthesis 90% after 20 h was lO-‘-fold lower than the low affinity hybrid and lO-’-fold lower than ricin. However, the maximal rate of protein synthesis inhibition by high affinity Thy 1.1-ricin A chain is only one-eighth the rate obtained with ricin. This rate of A chain transport to the cytosol via the Thy 1.1 antigen can be increased 6-fold by adding excess ricin B chain to the cells with no change in the amount of A chain bound to cells. Hybrids between intact ricin and low affinity anti-Thy 1.1 antibodies in the presence of lactose have up to 19-fold faster rates of protein synthesis inhibition than corresponding ricin A chain-antibody hybrids; however, target cell specificity is reduced. We conclude that the ricin B chain facilitates entry of the ricin A chain to the cytosol compartment by a process which is independent of the amount of A chain bound to the surface membrane. These results also show the importance of using kinetic studies to elucidate the variables which affect hybrid-toxin efficacy.