title: Use of uteroglobin for the engineering of polyvalent, polyspecific fusion proteins.
abstract: We report a novel strategy to engineer and express stable and soluble human recombinant polyvalent/polyspecific fusion proteins. The procedure is based on the use of a central skeleton of uteroglobin, a small and very soluble covalently linked homodimeric protein that is very resistant to proteolytic enzymes and to pH variations. Using a human recombinant antibody (scFv) specific for the angiogenesis marker domain B of fibronectin, interleukin 2, and an scFv able to neutralize tumor necrosis factor-alpha, we expressed various biologically active uteroglobin fusion proteins. The results demonstrate the possibility to generate monospecific divalent and tetravalent antibodies, immunocytokines, and dual specificity tetravalent antibodies. Furthermore, compared with similar fusion proteins in which uteroglobin was not used, the use of uteroglobin improved properties of solubility and stability. Indeed, in the reported cases it was possible to vacuum dry and reconstitute the proteins without any aggregation or loss in protein and biological activity.
authors.affiliation: Laboratory of Recombinant Therapeutic Proteins, Advanced Biotechnology Centre, Istituto G Gaslini, Genoa, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Ventura
authors.forename: Francesca
authors.initials: F
authors.lastname: Sassi
authors.forename: Sara
authors.initials: S
authors.lastname: Fossati
authors.forename: Arianna
authors.initials: A
authors.lastname: Parodi
authors.forename: William
authors.initials: W
authors.lastname: Blalock
authors.forename: Enrica
authors.initials: E
authors.lastname: Balza
authors.forename: Patrizia
authors.initials: P
authors.lastname: Castellani
authors.forename: Laura
authors.initials: L
authors.lastname: Borsi
authors.forename: Barbara
authors.initials: B
authors.lastname: Carnemolla
authors.forename: Luciano
authors.initials: L
authors.lastname: Zardi
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity / Q000276:immunology; D045744:Cell Line, Tumor; D003602:Cytotoxicity, Immunologic / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D007376:Interleukin-2 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008815:Mice, Inbred Strains; D008958:Models, Molecular; D009374:Neoplasms, Experimental / Q000276:immunology / Q000473:pathology; D010084:Oxidation-Reduction; D010957:Plasmids / Q000235:genetics; D055503:Protein Multimerization; D021382:Protein Sorting Signals / Q000235:genetics; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D014598:Uteroglobin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D007135:Immunoglobulin Variable Region; D007376:Interleukin-2; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D014409:Tumor Necrosis Factor-alpha; D014598:Uteroglobin