title: Non-classical anti-factor VIII C2 domain antibodies are pathogenic in a murine in vivo bleeding model.
abstract: OBJECTIVE
  The pathogenicity of anti-human factor (F) VIII monoclonal antibodies (MAbs) was tested in a murine bleeding model.
  
  
  METHODS
  MAbs were injected into the tail veins of hemophilia A mice to a peak plasma concentration of 60 nm, followed by injection of human B domain-deleted FVIII at 180 U kg(-1), producing peak plasma concentrations of approximately 2 nm. At 2 h, blood loss following a 4-mm tail snip was measured. The following MAbs were tested: (i) 4A4, a type I anti-A2 FVIII inhibitor, (ii) I54 and 1B5, classical type I anti-C2 inhibitors, (iii) 2-77 and B45, non-classical type II anti-C2 inhibitors, and (iv) 2-117, a non-classical anti-C2 MAb with inhibitory activity less than 0.4 Bethesda Units per mg IgG.
  
  
  RESULTS
  All MAbs except 2-117 produced similar amounts of blood loss that were significantly greater than control mice injected with FVIII alone. Increasing the dose of FVIII to 360 U kg(-1) overcame the bleeding diathesis produced by the type II MAbs 2-77 and B45, but not the type I antibodies, 4A4, I54, and 1B5. These results were consistent with the in vitro Bethesda assay in which 4A4 completely inhibited both 1 U mL(-1) and 3 U mL(-1) FVIII, while there was 40% residual activity at saturating concentrations of 2-77 at either concentration of FVIII.
  
  
  CONCLUSIONS
  For patients with an inhibitor response dominated by non-classical anti-C2 antibodies both the in vivo and in vitro results suggest that treatment with high-dose FVIII rather than bypassing agents may be warranted.
authors.affiliation: Department of Pediatrics, Aflac Cancer Center and Blood Disorders Service, Children's Healthcare of Atlanta, Emory University, Atlanta, GA 30322, USA.
authors.forename: S L
authors.initials: SL
authors.lastname: Meeks
authors.forename: J F
authors.initials: JF
authors.lastname: Healey
authors.forename: E T
authors.initials: ET
authors.lastname: Parker
authors.forename: R T
authors.initials: RT
authors.lastname: Barrow
authors.forename: P
authors.initials: P
authors.lastname: Lollar
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000008:administration & dosage* / Q000009:adverse effects / Q000097:blood; D004195:Disease Models, Animal; D000939:Epitopes / Q000276:immunology*; D005169:Factor VIII / Q000008:administration & dosage / Q000276:immunology*; D006467:Hemophilia A / Q000276:immunology*; D006470:Hemorrhage / Q000276:immunology*; D006801:Humans; D051379:Mice
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D005169:Factor VIII