PLoS ONE (Jan 2013)

Phenotypes of allo- and autoimmune antibody responses to FVIII characterized by surface plasmon resonance.

  • Kenneth B Lewis,
  • Richard J Hughes,
  • Melinda S Epstein,
  • Neil C Josephson,
  • Christine L Kempton,
  • Craig M Kessler,
  • Nigel S Key,
  • Tom E Howard,
  • Rebecca Kruse-Jarres,
  • Jeanne M Lusher,
  • Christopher E Walsh,
  • Raymond G Watts,
  • Ruth A Ettinger,
  • Kathleen P Pratt,
  • PATH (Personalized Alternative Therapies for Haemophilia) Study Investigators

DOI
https://doi.org/10.1371/journal.pone.0061120
Journal volume & issue
Vol. 8, no. 5
p. e61120

Abstract

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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.