title: Phenotypes of allo- and autoimmune antibody responses to FVIII characterized by surface plasmon resonance.
abstract: Evidence of antibody isotype/subtype switching may provide prognostic value regarding the state of immune responses to therapeutic proteins, e.g. anti-factor VIII (FVIII) antibodies that develop in many hemophilia A patients, clinically termed "inhibitors". A sensitive, high- information-content surface plasmon resonance (SPR) assay has been developed to quantify IgG subtype distributions and the domain specificity of anti-drug antibodies. Plasma samples from 22 subjects with an allo- or auto-immune reaction to FVIII were analyzed. Pre-analytical treatment protocols were developed to minimize non-specific binding and specific matrix interference due to von Willebrand factor-FVIII interactions. The dynamic range for IgG quantification was 0.2-5 µg/ml (∼1-33 nM), allowing characterization of inhibitor-positive samples. Subtype-specific monoclonal antibodies were used to quantify the IgG subtype distribution of FVIII-specific antibodies. Most samples obtained from multiply-infused inhibitor subjects contained IgG₄ antibodies. Several distinct phenotypes were assigned based on the IgG subtype distribution: IgG₁, IgG₄, IgG₁ & IgG₄, and IgG₁, IgG₂ & IgG₄. An IgG₁-only response was found in mild/moderate HA subjects during early FVIII infusions, and analysis of serial samples followed antibody class switching as several subjects' immune responses developed. Competition studies utilizing a recombinant FVIII-C2 domain indicated 40-80% of FVIII-specific antibodies in most samples were directed against this domain.
authors.affiliation: Puget Sound Blood Center Research Institute, Seattle, Washington, United States of America.
authors.forename: Kenneth B
authors.initials: KB
authors.lastname: Lewis
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Hughes
authors.forename: Melinda S
authors.initials: MS
authors.lastname: Epstein
authors.forename: Neil C
authors.initials: NC
authors.lastname: Josephson
authors.forename: Christine L
authors.initials: CL
authors.lastname: Kempton
authors.forename: Craig M
authors.initials: CM
authors.lastname: Kessler
authors.forename: Nigel S
authors.initials: NS
authors.lastname: Key
authors.forename: Tom E
authors.initials: TE
authors.lastname: Howard
authors.forename: Rebecca
authors.initials: R
authors.lastname: Kruse-Jarres
authors.forename: Jeanne M
authors.initials: JM
authors.lastname: Lusher
authors.forename: Christopher E
authors.initials: CE
authors.lastname: Walsh
authors.forename: Raymond G
authors.initials: RG
authors.lastname: Watts
authors.forename: Ruth A
authors.initials: RA
authors.lastname: Ettinger
authors.forename: Kathleen P
authors.initials: KP
authors.lastname: Pratt
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000917:Antibody Formation / Q000276:immunology*; D005169:Factor VIII / Q000276:immunology*; D006467:Hemophilia A / Q000097:blood / Q000276:immunology*; D006801:Humans; D007074:Immunoglobulin G / Q000145:classification / Q000276:immunology*; D051379:Mice; D010641:Phenotype; D020349:Surface Plasmon Resonance / Q000379:methods*
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; C078147:F8 protein, human; D005169:Factor VIII