T-CELL-TARGETED IMMUNOFUSION PROTEINS FROM ESCHERICHIA-COLI

Fusion proteins between cell-targeting domains and cytotoxic proteins should be particularly effective therapeutic reagents. We constructed a family of immunofusion proteins linking humanized Fab, F(ab')(2), or single chain antibody forms of the H65 antibody (which recognizes the CD5 antigen on the surface of human T cells) with the plant ribosome- inactivating protein gelonin. We reasoned that such an immunofusion wo uld kill human target cells as efficiently as the previously described chemical conjugates of H65 and gelonin (Better M., Bernhard, S. L., F ishwild, D. M., Nolan, P. A., Bauer, R. J., Kung, A. H. C., and Carrol l, S. F. (1994) J. Biol. Chem. 269, 9644-9650) if both the recognition and catalytic domains remained active, and a proper linkage between d omains could be found. Immunofusion proteins were produced in Escheric hia coli as secreted proteins and were recovered directly from the bac terial culture supernatant in an active form. All of the immunofusion proteins were purified by a common process and were tested for cytotox icity toward antigen-positive human cells. A 20-60-fold range of cytot oxic activity was seen among the fusion family members, and several fu sion proteins were identified which are approximately as active as eff ective chemical conjugates. Based on these constructs, immunofusion av idity and potency can be controlled by appropriate selection of antibo dy domains and ribosome-inactivating protein.