title: Anti-Human Platelet Antigen-1a Immunoglobulin G Preparation Intended to Prevent Fetal and Neonatal Alloimmune Thrombocytopenia.
abstract: Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is a severe disease that is caused by maternal alloantibodies generated during pregnancy or at delivery as a result of incompatibility between maternal and fetal human platelet antigens (HPAs) inherited from the father. Antibody-mediated immune suppression using anti-HPA-1a immunoglobulins is thought to be able to prevent FNAIT caused by HPA-1a. A fractionation process to prepare anti-HPA-1a immunoglobulin (Ig) G (IgG) from human plasma was therefore developed. Anti-HPA-1a plasma was obtained from volunteer mothers who underwent alloimmunization against HPA-1a during a previous pregnancy. Plasma was cryoprecipitated and the supernatant treated with caprylic acid and solvent/detergent (S/D), purified by chromatography, nanofiltered, concentrated, and sterile-filtered. The anti-HPA-1a immunoglobulin fraction was characterized for purity and safety. PAK12 and quantitative monoclonal antibody immobilization of platelet antigen (MAIPA) assays were used to detect anti-HPA-1a IgG. Hepatitis C virus (HCV) removal during nanofiltration was assessed by spiking experiments, using cell culture-derived reporter HCV and luciferase analysis. The caprylic acid treatment precipitated non-Ig proteins yielding a 90% pure Ig supernatant. S-HyperCel chromatography of the S/D-treated supernatant followed by HyperCel STAR AX provided high IgG recovery (>80%) and purity (>99.5%), and efficient IgA and IgM removal. Concentrations of complement factors C3 and C4 were < 0.5 and < 0.4 mg/dL, respectively. The final IgG could be nanofiltered on Planova 20N under conditions removing more than 3 log HCV infectivity to baseline mock infection level, and concentrated to ca. 30 g/L. Proteolytic activity and thrombin generation were low in the final fraction. The Pak12 and MAIPA assays showed good recovery of anti-HPA-1a throughout the process. Clinical-grade HPA-1a IgG can be prepared using a process compliant with current quality requirements opening perspectives for the prevention of FNAIT.
authors.affiliation: Graduate Institute of Biomedical Materials and Tissue Engineering, College of Biomedical Engineering, Taipei Medical University, Taipei, Taiwan.
authors.forename: Ying-Jan
authors.initials: YJ
authors.lastname: Weng
authors.affiliation: Department of Medical Biology, UiT The Arctic University of Norway, Tromsø, Norway.
authors.forename: Anne
authors.initials: A
authors.lastname: Husebekk
authors.affiliation: Department of Medical Biology, UiT The Arctic University of Norway, Tromsø, Norway.|Department of Laboratory Medicine, University Hospital North Norway, Tromsø, Norway.
authors.forename: Björn
authors.initials: B
authors.lastname: Skogen
authors.affiliation: Department of Laboratory Medicine, University Hospital North Norway, Tromsø, Norway.|Finnmark Hospital Trust, Hammerfest, Norway.
authors.forename: Mette
authors.initials: M
authors.lastname: Kjaer
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.|Department of Microbiology and Immunology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Liang-Tzung
authors.initials: LT
authors.lastname: Lin
authors.affiliation: Graduate Institute of Biomedical Materials and Tissue Engineering, College of Biomedical Engineering, Taipei Medical University, Taipei, Taiwan.
authors.forename: Thierry
authors.initials: T
authors.lastname: Burnouf
mesh_terms: D016824:Antigens, Human Platelet / Q000276:immunology / Q000302:isolation & purification / Q000627:therapeutic use*; D005591:Chemical Fractionation; D003720:Densitometry; D004591:Electrophoresis, Polyacrylamide Gel; D016174:Hepacivirus; D006801:Humans; D007231:Infant, Newborn / Q000276:immunology; D039661:Integrin beta3; D013917:Thrombin / Q000378:metabolism; D054098:Thrombocytopenia, Neonatal Alloimmune / Q000517:prevention & control*; D014462:Ultrafiltration / Q000379:methods
chemical_list: D016824:Antigens, Human Platelet; C061248:ITGB3 protein, human; D039661:Integrin beta3; D013917:Thrombin