Generation of low-toxicity interleukin-2 fusion proteins devoid of vasopermeability activity.
Because of its key role in immunity, interleukin-2 (IL-2) has been studied extensively for the adoptive immunotherapy of cancer. Although systemic administration of IL-2 has been shown to stimulate antitumor responses in vivo, its efficacy in the clinic has been limited by the development of serious side effects, including the induction of vascular leak syndrome. Previously, we have identified a small peptide fragment of IL-2 that was found to contain the entire vasopermeability activity of the cytokine. The identification of the location of this potentially undesirable property of IL-2 enabled us to focus on the generation of mutant derivatives that might be lacking vasopermeability activity but that retain cytokine functionality. In addition to this discovery, our laboratory has constructed monoclonal antibody/IL-2 fusion proteins that can target this potent cytokine directly to tumor for the immunotherapy of both solid and lymphoid malignancies. Using this fusion protein technology, we have constructed a series of point mutations in the newly identified vasopermeability region of IL-2 for the purpose of deleting this activity. Fusion proteins showing reduced or deleted vasopermeability activity were then tested for their cytokine potency by several methods, including their binding to IL-2 receptors, T-cell proliferation assays, the induction of secondary cytokines, dose-escalating toxicity, and finally their ability to treat established solid tumors in syngeneic immunocompetent mice. The results of these studies clearly show that the vasopermeability activity of IL-2 can be substantially deleted by single point mutations such as Arg38Trp without grossly affecting the immune function of the cytokine.
Hu
Peisheng
P
Department of Pathology, Keck School of Medicine, University of Southern California, Los Angeles 90033, USA.
Mizokami
Myra
M
Ruoff
Gina
G
Khawli
Leslie A
LA
Epstein
Alan L
AL
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics; D001483:Base Sequence; D002199:Capillary Permeability / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D016219:Immunotherapy, Adoptive; D007371:Interferon-gamma / Q000096:biosynthesis; D007375:Interleukin-1 / Q000096:biosynthesis; D007376:Interleukin-2 / Q000235:genetics* / Q000494:pharmacology* / Q000633:toxicity; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects; D016399:Lymphoma, T-Cell / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D016296:Mutagenesis; D009369:Neoplasms / Q000628:therapy; D017354:Point Mutation; D015375:Receptors, Interleukin-2 / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000494:pharmacology* / Q000627:therapeutic use / Q000633:toxicity; D013601:T-Lymphocytes / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
D000911:Antibodies, Monoclonal; D007375:Interleukin-1; D007376:Interleukin-2; D015375:Receptors, Interleukin-2; D011993:Recombinant Fusion Proteins; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma