title: A recombinant bispecific single-chain Fv antibody against HLA class II and FcgammaRIII (CD16) triggers effective lysis of lymphoma cells.
abstract: Bispecific antibodies offer the possibility of improving effector-cell recruitment for antibody therapy. For this purpose, a recombinant bispecific single-chain Fv antibody (bsscFv), directed against FcgammaRIII (CD16) and human leucocyte antigen (HLA) class II, was constructed and tested in functional assays. RNA from the hybridomas 3G8 and F3.3, reacting with CD16 and HLA class II, respectively, was used to generate phage display libraries. From these libraries, reactive phages were isolated and the bsscFv was constructed by connecting both single-chain Fv components through a 20 amino acid flexible linker. After expression in SF21 insect cells and chromatographic purification, the bsscFv bound specifically and simultaneously to both antigens. The affinities of the anti-CD16 and the anti-HLA class II scFv components of the bsscFv were 8.6 x 10(-8) mol/l and 13.7 x 10(-8) mol/l, respectively, which was approximately sevenfold lower than the F(ab) fragments of the parental antibodies. In antibody-dependent cellular cytotoxicity experiments with human mononuclear cells as effectors, the bsscFv-mediated specific lysis of both HLA class II-positive, malignant human B-lymphoid cell lines and primary cells from patients with chronic B-cell lymphocytic leukaemia. Optimal lysis was obtained at bsscFv concentrations of approximately 400 ng/ml, similar to the concentration required for maximum lysis by the corresponding chemically linked bispecific antibody. Thus, this recombinant bsscFv-antibody is an efficient molecule for effector-cell mediated lysis of malignant human B-lymphoid cells.
authors.affiliation: University of Erlangen-Nuremburg, Staudtstrasse 5, D-91058 Erlangen, Germany.
authors.forename: Joerg
authors.initials: J
authors.lastname: Bruenke
authors.forename: Barbara
authors.initials: B
authors.lastname: Fischer
authors.forename: Karin
authors.initials: K
authors.lastname: Barbin
authors.forename: Katja
authors.initials: K
authors.lastname: Schreiter
authors.forename: Yvonne
authors.initials: Y
authors.lastname: Wachter
authors.forename: Kerstin
authors.initials: K
authors.lastname: Mahr
authors.forename: Fritz
authors.initials: F
authors.lastname: Titgemeyer
authors.forename: Michael
authors.initials: M
authors.lastname: Niederweis
authors.forename: Matthias
authors.initials: M
authors.lastname: Peipp
authors.forename: Susan J
authors.initials: SJ
authors.lastname: Zunino
authors.forename: Roland
authors.initials: R
authors.lastname: Repp
authors.forename: Thomas
authors.initials: T
authors.lastname: Valerius
authors.forename: Georg H
authors.initials: GH
authors.lastname: Fey
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000276:immunology*; D002478:Cells, Cultured; D006224:Cricetinae; D000949:Histocompatibility Antigens Class II / Q000276:immunology*; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology; D016393:Lymphoma, B-Cell / Q000276:immunology* / Q000628:therapy; D017452:Receptors, IgG / Q000276:immunology*
chemical_list: D018033:Antibodies, Bispecific; D000949:Histocompatibility Antigens Class II; D017452:Receptors, IgG