Most factor VIII B domain missense mutations are unlikely to be causative mutations for severe hemophilia A: implications for genotyping

J Thromb Haemost. 2011 Jun;9(6):1183-90. doi: 10.1111/j.1538-7836.2011.04268.x.


Background & objective: The factor VIII (FVIII) B domain shares very little amino acid homology with other known proteins and is not directly necessary for procoagulant activity. Despite this, missense mutations within the B domain have been reported in patients with hemophilia A. Given that the B domain is dispensable for secretion and function of FVIII, we hypothesized that these mutations should not be causative of hemophilia A in these patients.

Methods: Plasmid vectors containing B domain missense mutations that were reported to be associated with moderate/severe hemophilia A (T751S, D826E, V993L, H1047Y, T1353A, N1441K, L1462P, E1579D, A1591S, P1641L and S1669L) were analyzed for their effect on synthesis and secretion compared with FVIII wild-type (WT) following transient transfection into COS-1 and CHO cells in vitro. Further, H1047Y, N1441K and E1579D mutants were expressed in vivo in a hemophilia A mouse model by hydrodynamic tail-vein injection.

Results: FVIII activity and antigen levels for all mutants expressed into the conditioned media of COS-1 and CHO cells were similar to FVIII WT. Also, plasma expression of these mutants was similar to FVIII WT in hemophilia A mice. An in vivo tail clip bleeding assay also demonstrated that blood loss from hemophilia A mice expressing FVIII WT, H1047Y, N1441K and E1579D was similar.

Conclusions: We conclude that most missense mutations within the FVIII B domain would be unlikely to lead to severe hemophilia A and that the majority of such missense mutations represent polymorphisms or non-pathologic mutations.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CHO Cells
  • COS Cells
  • Chlorocebus aethiops
  • Cricetinae
  • Cricetulus
  • Factor VIII / genetics*
  • Genotype
  • Hemophilia A / etiology*
  • Hemophilia A / genetics
  • Humans
  • Mice
  • Mutation, Missense*
  • Protein Structure, Tertiary / genetics
  • Transfection


  • Factor VIII