Flow cytometric observations on the in vivo use of Fab fragments of a chimaeric monoclonal antibody to platelet glycoprotein IIb-IIIa.
Fluorescence flow cytometry (FC) was employed to monitor the platelet surface in humans receiving intravenous infusions of Fab fragments of a chimaeric (murine/human) construct of monoclonal antibody 7E3, which binds to the fibrinogen receptor (glycoprotein IIb-IIIa) and inhibits platelet aggregation. Platelet surface-bound chimaeric 7E3-Fab (c7E3-Fab) was measured using a fluorescein-conjugated polyclonal anti-7E3 antibody and residual c7E3-Fab binding capacity was measured using fluorescein-conjugated c7E3-Fab. Turbidometrically measured platelet aggregation response to ADP was shown to be a linear function (r = 0.9) of the logarithm of residual free binding sites for c7E3-Fab. The distribution of c7E3-Fab in the platelet population was unimodal at all time points following the infusion, demonstrating in vivo transfer of antibody to newly released circulating platelets. Clearance of platelet surface-bound antibody followed an exponential model but all circulating platelets were bearing c7E3-Fab at time points well beyond the lifespan of the platelets exposed to c7E3-Fab during the infusion. Ex vivo and in vitro mixing experiments showed that c7E3-Fab transfer between platelets was possible and suggested that differences in the in vivo kinetics between monovalent and bivalent forms of monoclonal antibodies might be relevant in their therapeutic application.
Christopoulos
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C
Department of Haematology, University College and Middlesex School of Medicine, London, UK.
Mackie
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I
Lahiri
A
A
Machin
S
S
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000494:pharmacology; D001666:Binding Sites, Antibody; D001792:Blood Platelets / Q000276:immunology* / Q000502:physiology; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000032:analysis* / Q000494:pharmacology; D051379:Mice; D009391:Nephelometry and Turbidimetry; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000032:analysis / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000276:immunology* / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D010975:Platelet Aggregation Inhibitors; D010980:Platelet Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D000244:Adenosine Diphosphate