title: Developing recombinant HPA-1a-specific antibodies with abrogated Fcgamma receptor binding for the treatment of fetomaternal alloimmune thrombocytopenia.
abstract: Fetomaternal alloimmune thrombocytopenia (FMAIT) is caused by maternal generation of antibodies specific for paternal platelet antigens and can lead to fetal intracranial hemorrhage. A SNP in the gene encoding integrin beta3 causes a clinically important maternal-paternal antigenic difference; Leu33 generates the human platelet antigen 1a (HPA-1a), whereas Pro33 generates HPA-1b. As a potential treatment to prevent fetal intracranial hemorrhage in HPA-1a alloimmunized pregnancies, we generated an antibody that blocks the binding of maternal HPA-1a-specific antibodies to fetal HPA-1a1b platelets by combining a high-affinity human HPA-1a-specific scFv (B2) with an IgG1 constant region modified to minimize Fcgamma receptor-dependent platelet destruction (G1Deltanab). B2G1Deltanab saturated HPA-1a+ platelets and substantially inhibited binding of clinical HPA-1a-specific sera to HPA-1a+ platelets. The response of monocytes to B2G1Deltanab-sensitized platelets was substantially less than their response to unmodified B2G1, as measured by chemiluminescence. In addition, B2G1Deltanab inhibited chemiluminescence induced by B2G1 and HPA-1a-specific sera. In a chimeric mouse model, B2G1 and polyclonal Ig preparations from clinical HPA-1a-specific sera reduced circulating HPA-1a+ platelets, concomitant with transient thrombocytopenia. As the Deltanab constant region is uninformative in mice, F(ab')2 B2G1 was used as a proof of principle blocking antibody and prevented the in vivo platelet destruction seen with B2G1 and polyclonal HPA-1a-specific antibodies. These results provide rationale for human clinical studies.
authors.affiliation: NHS Blood and Transplant, Department of Haematology, University of Cambridge, Cambridge, United Kingdom. cedric.ghevaert@nhsbt.nhs.uk
authors.forename: Cedric
authors.initials: C
authors.lastname: Ghevaert
authors.forename: David A
authors.initials: DA
authors.lastname: Wilcox
authors.forename: Juan
authors.initials: J
authors.lastname: Fang
authors.forename: Kathryn L
authors.initials: KL
authors.lastname: Armour
authors.forename: Mike R
authors.initials: MR
authors.lastname: Clark
authors.forename: Willem H
authors.initials: WH
authors.lastname: Ouwehand
authors.forename: Lorna M
authors.initials: LM
authors.lastname: Williamson
mesh_terms: D000906:Antibodies / Q000276:immunology* / Q000378:metabolism; D016824:Antigens, Human Platelet / Q000235:genetics / Q000276:immunology*; D001792:Blood Platelets / Q000276:immunology / Q000378:metabolism; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry / Q000235:genetics; D007135:Immunoglobulin Variable Region / Q000276:immunology / Q000378:metabolism; D007231:Infant, Newborn; D039661:Integrin beta3; D008958:Models, Molecular; D009154:Mutation; D010976:Platelet Count; D011247:Pregnancy; D011485:Protein Binding; D017452:Receptors, IgG / Q000276:immunology / Q000378:metabolism; D011994:Recombinant Proteins / Q000276:immunology; D054098:Thrombocytopenia, Neonatal Alloimmune / Q000209:etiology / Q000628:therapy*
chemical_list: D000906:Antibodies; D016824:Antigens, Human Platelet; C061248:ITGB3 protein, human; D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region; D039661:Integrin beta3; D017452:Receptors, IgG; D011994:Recombinant Proteins