Characterization of a new monoclonal anti-Fc gamma RII antibody, AT10, and its incorporation into a bispecific F(ab')2 derivative for recruitment of cytotoxic effectors.
Fc gamma RII (CDw32) on monocytes is capable of triggering both phagocytosis and lysis of chick red blood cells (CRBC) coated with antibody of the appropriate isotype. In this report we describe the production and characterization of a mouse monoclonal IgG1 antibody specific for Fc gamma RII and compare its activity in binding studies, tissue distribution and redirected cellular cytotoxicity (RCC), with the previously identified anti-Fc gamma RII antibodies KB61 and IV.3. Immunohistochemical and flow cytometry analyses demonstrated that AT10 binds very strongly to Fc gamma RII on normal monocytes, but only weakly to that expressed on lymphocytes. This pattern does not correspond to the staining seen with either KB61 or IV.3, and appears to give an intermediate profile. The binding constant (Ka) for the Fab' fragment of AT10 was calculated at 5.3 x 10(8) M-1, four times higher than that for KB61 (1.4 x 10(8) M-1). Bispecific F(ab')2 antibodies were constructed from Fab' fragments of AT10 or KB61 thioether-linked to Fab' from an anti-CRBC monoclonal antibody. These bispecific derivatives directed monocyte cytotoxicity against CRBC as efficiently as either a monoclonal or polyclonal anti-chick erythrocyte antibody. The bispecific F(ab')2 antibodies had a distinct advantage over the conventional reagents, in that they were not blocked in the presence of human Fc gamma at 3.5 mg/ml (a concentration comparable with that provided by IgG in serum). Therefore, bispecific derivatives constructed with the high affinity anti-Fc gamma RII antibody, AT10, may be used as therapeutic reagents for targeting tumour cell lysis in vivo.
Greenman
J
J
Lymphoma Research Unit, Tenovus Laboratory, General Hospital, Southampton, U.K.
Tutt
A L
AL
George
A J
AJ
Pulford
K A
KA
Stevenson
G T
GT
Glennie
M J
MJ
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000943:Antigens, Differentiation / Q000276:immunology*; D002460:Cell Line; D003601:Cytotoxicity Tests, Immunologic; D003602:Cytotoxicity, Immunologic; D004306:Dose-Response Relationship, Immunologic; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D007140:Immunoglobulin Fab Fragments / Q000276:immunology; D007074:Immunoglobulin G / Q000096:biosynthesis; D008214:Lymphocytes / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C / Q000276:immunology*; D009000:Monocytes / Q000276:immunology; D011233:Precipitin Tests; D011961:Receptors, Fc / Q000276:immunology*; D017452:Receptors, IgG; D012397:Rosette Formation
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D000943:Antigens, Differentiation; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D011961:Receptors, Fc; D017452:Receptors, IgG