CHARACTERIZATION OF A RICIN FUSION TOXIN TARGETED TO THE INTERLEUKIN-2 RECEPTOR

Fusion toxins are hybrid proteins consisting of peptide ligands linked through amide bonds to polypeptide toxins. The ligand directs the mol ecule to the surface of target cells and the toxin enters the cytosol and induces cell death, Ricin is an excellent candidate for use in fus ion toxins because of its extreme potency, the extensive knowledge of its atomic structure and the lack of prior immunological exposure in p atients. We synthesized a baculovirus transfer vector with the polyhed rin promoter followed sequentially from the 5' end with DNA encoding t he gp67A leader sequence, the tripeptide ADP, IL-2 (interleukin-2), an other ADP tripeptide and RTB (ricin toxin B chain) with lectin-site mu tations W37S and Y248H, Recombinant baculovirus was generated in Sf9 i nsect cells and used to infect Sf9 cells, Recombinant IL-2-RTB[W37S/Y2 48H] protein (fusion protein of IL-2 with modifications W37S and Y238H ) was recovered at high yields from day 6 insect cell supernatants, pa rtially purified by affinity chromatography and reassociated with RTA (ricin toxin A chain), The fusion toxin was soluble. immunoreactive wi th antibodies to RTB, IL-2 and RTA and had a molecular weight of 80 kD a by SDS-PAGE. The molecule reacted poorly with asialofetuin, but boun d strongly to IL-2 receptor based on selective cytotoxicity to IL-2 re ceptor bearing cells, The specific cytotoxicity could be blocked with IL-2 but not lactose, Thus, we report a novel targeted fusion toxin pr otein with full biological activity.