Premium
A recombinant bispecific single‐chain Fv antibody against HLA class II and FcγRIII (CD16) triggers effective lysis of lymphoma cells
Author(s) -
Bruenke Joerg,
Fischer Barbara,
Barbin Karin,
Schreiter Katja,
Wachter Yvonne,
Mahr Kerstin,
Titgemeyer Fritz,
Niederweis Michael,
Peipp Matthias,
Zunino Susan J.,
Repp Roland,
Valerius Thomas,
Fey Georg H.
Publication year - 2004
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.2004.04893.x
Subject(s) - antibody , antigen , microbiology and biotechnology , cd16 , human leukocyte antigen , recombinant dna , immunoglobulin light chain , lysis , biology , fragment crystallizable region , cytolysis , monoclonal antibody , cytotoxicity , immunology , chemistry , in vitro , biochemistry , cd3 , cd8 , gene
Summary 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 Fc γ RIII (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 × 10 −8 mol/l and 13·7 × 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.