A novel cause of mild/moderate hemophilia A: mutations scattered in the factor VIIIC1 domain reduce factor VIII binding to von Willebrand factor

The mechanisms responsible for the low factor VIII (fVIII) activity in the plasma of patients with mild/moderate hemophilia A are poorly understood. I n such patients, we Rave identified a series of fVIII mutations (IIe2098Ser , Ser2119Tyr, Asn2129Ser, Arg2150His, and Pro2153Gln) clustered In the C1 d omain and associated with reduced binding of fVIII to von Willebrand factor (vWf), For each patient plasma, the specific activity of mutated fVIII was close to that of normal fVIII, Scatchard analysis showed that the affinity for vWf of recombinant IIe2098Ser, Ser2119Tyr, and Arg2150His fVIII mutant s was reduced 8-fold, 80-fold, and 3-fold, respectively, when compared with normal fVIII, Given the importance of vWf for the stability of fVIII in pl asma, these findings suggested that the reduction of fVIII binding to vWf r esulting from the above-mentioned mutations could contribute to patients' l ow fVIII plasma levels. We, therefore, analyzed the effect of vWf on fVIII production by Chinese hamster ovary (CHO) cells transfected with expression vectors for recombinant B domain-deleted normal, IIe2098Ser, Ser2119Tyr, a nd ArgP1B0His fVIII, These 3 mutations impaired the vWf dependent accumulat ion of functional fVIII in culture medium. Analysis of fVIII production by transiently transfected CHO cells indicated that, in addition to the impair ed stabilization by vWf, the secretion of functional IIe2098Ser and ArgP1B0 His fVIII was reduced about 2-fold and 6-fold, respectively, by comparison to Ser2119Tyr and normal fVIII, These findings indicate that C1-domain muta tions resulting in reduced fVIII binding to vWf are an important cause of m ild/moderate hemophilia A. (C) 2000 by The American Society of Hematology.