IL2-ricin fusion toxin is selectively cytotoxic in vitro to IL2 receptor-bearing tumor cells.
Fusion toxins consist of peptide ligands linked through amide bonds to polypeptide toxins. The ligand directs the molecule to the surface of target cells and the toxin enters the cytosol and induces cell death. Ricin toxin is an excellent candidate for use in fusion toxins because of its extreme potency, the extensive knowledge of its atomic structure, and the years of experience with RTA chemical conjugates in clinical trials. We synthesized a baculovirus transfer vector with the polyhedrin promoter followed sequentially from the 5' end with DNA encoding the gp67A leader sequence, the tripeptide ADP, IL2, another ADP tripeptide, and RTB. Recombinant baculovirus was generated in Sf9 insect cells and used to infect Sf9 cells. Recombinant IL2-RTB protein was recovered at high yields from day 5 insect cell supernatants, partially purified by affinity chromatography, and characterized. The recombinant product was soluble and immunoreactive with antibodies to RTB and IL2, bound asialofetuin and lactose, and reassociated with RTA. In the presence of lactose to block galactose-binding sites on RTB, the IL2-RTB-RTA heterodimer was selectively cytotoxic to IL2 receptor, bearing cells. Specific cytotoxicity could be blocked with IL2. Thus, we report a novel targeted plant toxin fusion protein with full biological activity.
Frankel
A
A
Department of Medicine, Medical University of South Carolina, Charleston 29425, USA.
Tagge
E
E
Chandler
J
J
Burbage
C
C
Hancock
G
G
Vesely
J
J
Willingham
M
M
D000818:Animals; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D001212:Asialoglycoproteins / Q000378:metabolism; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D005260:Female; D060748:Fetuins; D006801:Humans; D015151:Immunoblotting; D007376:Interleukin-2 / Q000627:therapeutic use*; D007785:Lactose / Q000378:metabolism; D015458:Leukemia, T-Cell / Q000188:drug therapy; D008969:Molecular Sequence Data; D010051:Ovarian Neoplasms / Q000188:drug therapy; D015375:Receptors, Interleukin-2 / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000627:therapeutic use*; D012276:Ricin / Q000627:therapeutic use*; D018411:Spodoptera / Q000378:metabolism; D014407:Tumor Cells, Cultured; D000509:alpha-Fetoproteins / Q000378:metabolism
D000970:Antineoplastic Agents; D001212:Asialoglycoproteins; D060748:Fetuins; D007376:Interleukin-2; D015375:Receptors, Interleukin-2; D011993:Recombinant Fusion Proteins; D000509:alpha-Fetoproteins; C017029:asialofetuin; D012276:Ricin; D007785:Lactose