Non-Inversion Factor VIII Mutations in 80 Hemophilia A Families Including 24 with Alloimmune Responses

Summary

Heteroduplex screening identified 74 small mutations in the factor VIII genes of 72 families with hemophilia A. In addition, patients from 3 families with high titer inhibitors had partial gene deletions and 5 unrelated families that were negative for heteroduplex formation had a mutation on direct sequencing. The latter had mild hemophilia A with an inhibitor, and sequencing their exon 23 fragments found a transition predicting a recurrent Arg²¹⁵⁰ to His. Of 69 distinct mutations (including the 3 partial gene deletions), 47 are novel. Of small mutations, 51 were missense (one possibly a normal variant and two that could also alter splicing) at 39 sites, 13 were small deletions or insertions (3 inframe and one a normal variant in an intron), 13 were nonsense at 12 sites and 2 altered intron splice junctions. In 24 families, at least one affected member had evidence for an alloimmune response to factor VIII; of these, 11 were associated with missense mutations. In 14 families, de novo origin was demonstrated.

Keywords

Hemophilia A - factor VIII - mutations - inhibitors

• References

• 1 Lakich D, Kazazian HH, Antonarakis SE, Gitschier J. Inversions disrupting the factor VIII gene are a common cause of severe haemophilia A. Nat Genet 1993; 05: 236-41.
• 2 Naylor JA, Brinke A, Hassock S, Green PM, Giannelli F. Characteristic mRNA abnormality found in half the patients with severe hemophilia A is due to large DNA inversions. Hum Mol Genet 1993; 02: 1773-8.
• 3 Kemball-Cook G, Tuddenham EGD, Wacey AI. The factor VIII Structure and Mutation Resource Site: HAMSTeRS version 4. Nucl Acids Res 1998; 26: 216-219 http://europium.csc.mrc.ac.uk/ (10/00 update).
• 4 Hay CRM, Ludlam CA, Colvin BT, Hill FGH, Preston FE, Wasseem N, Bagnall R, Peake IR, Berntorp E, Bunschoten EPM, Fijnvandraat K, Kasper CK, White G, Santagostino E. Factor VIII inhibitors in mild and moderateseverity haemophilia A. Thromb Haemost 1998; 79: 762-6.
• 5 Liu M, Murphy ME, Thompson AR. A domain mutations in 65 haemophilia A families and molecular modeling of dysfunctional factor VIII proteins. Br J Haematol 1998; 103: 1051-60.
• 6 Liu ML, Shen BW, Nakaya S, Pratt KP, Fujikawa K, Davie EW, Stoddard BL, Thompson AR. Modeling hemophilic factor VIII (F.VIII) C1 and C2 domain mutations into the 1.5 A human C2 domain crystal structure. Blood 2000; 96: 979-87.
• 7 Nakaya S, Liu M-L, Thompson AR. Some factor VIII exon 14 frameshift mutations cause phenotypically moderately severe hemophilia A. Br J Haematol. 2001 in press..
• 8 Weinmann AF, Schoof JM, Thompson AR. Clinical correlates among 49 families with hemophilia A and factor VIII gene inversions. Am J Hematol 1996; 51: 192-9.
• 9 Liu ML, Thompson AR. Factor VIII gene inversions and an XbaI polymorphism: nonradioactive detection and clinical usage. Haemophilia 1999; 05: 26-31.
• 10 Donath M-JSH, de Laaf RTM, Biessels PTM, Lenting PJ, van de Loo J-W, van Mourik JA, Voorberg J, Mertens K. Characterization of des-(741-1668)-factor VIII, a single-chain factor VIII variant with a fusion site susceptible to proteolysis by thrombin and factor Xa. Biochem J 1995; 312: 49-55.
• 11 Gitschier J, Wood WI, Goralka TM, Wion KL, Chen EY, Eaton DH, Vehar GA, Capon DJ, Lawn RM. Characterization of the human factor VIII gene. Nature 1984; 312: 326-30.
• 12 Wood WI, Capon DJ, Simonsen CC. et al. Expression of active human factor VIII from recombinant DNA clones. Nature 1984; 312: 330-7.
• 13 Pemberton S, Lindley P, Zaitsev V, Card G, Tuddenham EGD, Kemball-Cook G. A molecular model for the triplicated A domains of human factor VIII based on the crystal structure of human ceruloplasmin. Blood 1997; 89: 2413-21.
• 14 Thompson AR, Murphy MEP, Liu ML, Saenko EL, Healey JF, Lollar P, Scandella D. Loss of tolerance to exogenous and endogenous factor VIII in a mild hemophilia A patient with an Arg593 to Cys mutation. Blood 1997; 90: 1902-10.
• 15 Goodeve AC, Williams I, Bray GL, Peake IR. Relationship between factor VIII mutation type and inhibitor development in a cohort of previously untreated patients treated with recombinant factor VIII (Recombinate). Recombinate PUP Study Group. Thromb Haemost 2000; 83: 844-8.
• 16 Fijnvandraat K, Turenhout EA, van den Brink EN, ten Cate JW, van Mourik JA, Peters M, Voorberg J. The missense mutation Arg593-Cys is related to antibody formation in a patient with mild hemophilia A. Blood 1997; 89: 4371-7.
• 17 Knobe KE, Villoutreix BO, Tengborn L, Petrini P, Ljung RCR. Factor VIII inhibitors in two families with mild haemophilia A: Structural analysis of the mutations. Haemostasis 2000; 30: 268-79.
• 18 Gilles JGG, Arnout J, Vermylen J, Saint-Remy JM. Anti-factor VIII antibodies of hemophilic patients are frequently directed towards nonfunctional determinants and do not exhibit isotype restriction. Blood 1993; 82: 2452-61.
• 19 Vidal F, Farssac E, Altisent C, Pulg L, Gallardo D. Rapid hemophilia A molecular diagnosis by a simple DNA sequencing procedure: identification of 14 novel mutations. Thromb Haemost 2001; 85: 580-3.
• 20 Weinmann AF, Murphy MEP, Thompson AR. Consequences of factor IX mutations in 26 families with haemophilia B. Br J Haematol 1998; 100: 58-61.