Molecular Characterization of Hemophilia a in Southeast Bulgaria
The results of molecular characterization of Hemophilia A in 50 patients from Southeast Bulgaria are presented. Southern blot analysis for the detection of inversions in intron 22, and polymerase chain reaction (PCR) followed by single strand conformation polymorphism (SSCP) or de-naturing gradient gel electrophoresis (DGGE) for screening of the coding sequences of the Factor VIII (FVIII) gene were used. A molecular defect was found in 35 (70%), the most frequent being an inversion in intron 22, found in 19 (38%) patients; an intron 1 inversion was not detected. In one severely affected patient, an Alu insert was found, which disrupted exon 14 at codon 1224. Nucleotide substitutions were found in 15 (30%) patients, the most frequent being an Arg531→Cys missense mutation in exon 11. The same nonsense mutation (codon -5, CGA>TGA) was found in two patients with a severe phenotype. Seven missense mutations (Asn90→Thr, Arg 372→His, Glu456→Val, Tyr473→His, Arg1689→ Cys, Arg2159→Cys and Arg2163→His) were detected in isolated families. Two of these (Asn90→Thr and Glu456→ Val) are being reported for the first time.
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Bolton-Maggs PHB Pasi JK. Haemophilia A and B. Lancet 2003; 361(9371): 1801-1809.
Wood WI Capon DJ Simonsen CC Eaton DL Gitschier J Keyt B Seebork PH Smith DH Hollingshead P Wion KL Delwart E Tuddenham EGD Vehar GA Lawn RM. Expression of active human factor VIII from recombinant cDNA clones. Nature 1984; 312(5992): 330-337.
Saenko EL Ananyeva NM Tuddenham EGD Kemball-Cook G. Factor FVIII - novel insights into form and function. Br J Haematol 2002; 119(2): 323-331.
Lakich D Kazazian HH Jr Antonarakis SE Gitschier J. Inversions disrupting the factor VIII are a common cause for severe Haemophilia A. Nat Genet 1993; 5(3): 236-241.
Bagnall RD Waseem N Green PM Giannelli F. Recurrent inversion breaking intron 1 of the factor VIII gene is a frequent cause of severe hemophilia A. Blood 2002; 99(1): 168-174.
Kemball Cook G Tuddenham EGD Wacey AI. The factor VIII structure and mutation resource site HAMSTeRS version 4. Nucleic Acids Res 1998; 26(1): 216-219 (http://europium.csc.mrc.ac.uk) http://europium.csc.mrc.ac.uk
Antonarakis SE Kazazian HH Tuddenham EGD. Molecular etiology of factor VIII deficiency in hemophilia A. Hum Mutat 1995; 5(1): 1-22.
Efremov G D Dimovski A J Plaseska-Karanfilska D Simjanovsa L Sukarova E Koceva S Popovski Z. Laboratory Manual 2nd ed. 1998. ICGEB Affilated Center "Nucleic acid based methods in human and veterinary medicine." Skopje: Macedonian Academy of Sciences and Arts 1998.
Gitschier J Wood WI Tuddenham EGD Shuman MA Goralka TM Chen EY Lawn RM. Detection and sequence of mutations in the factor VIII gene of haemophiliacs. Nature 1985; 315(6018): 427-430.
David D Moreira I Lalloz MRA Rosa HAV Schwaab R Morais S Diniz MJ de Deus G Camops M Lavinha J Johnson D Tuddenham EGD. Analysis of the essential sequences of the human factor VIII gene in twelve haemophilia A patients with single strand conformation polymorphism. Blood Coagul Fibrinol 1992; 5(2): 257-264.
Diamond C Kogan S Levinson B Gitschier J. Amino acid substitutions in a conserved domains of factor VIII and related proteins: study of patients with mild and moderately severe hemophilia A. Hum Mutat 1992; 1(3): 248-257.
Antonarakis SE. and a consortium of more than 50 international authors. Factor VIII gene inversions in severe hemophilia A: results of an international consortium study. Blood 1995; 86(6): 2206-2212.
Rossiter JP Young M Kimberland ML Hutter P Ketterling RP Gitschier J Horst J Morris MA Schaid DJ de Moerloose P Sommer SS Kazazian HH Antonarakis SE. Factor VIII gene inversions causing severe hemophilia A originate almost exclusively in male germ cells. Hum Mol Genet 1994; 3(7): 1035-1039.
Schroder J El-Maarri O Schwaab R Muller CR Oldenburg J. Factor VIII intron-1 inversion: frequency and inhibitor prevalence. J Thromb Haemost 2006; 4(5): 1141-11413.
Andrikovics H Klein I Bors A Nemes L Marosi A Varadi A Tordai A. Analysis of large structural changes of the factor VIII gene involving intron 1 and 22 in severe hemophilia A. Haematologica 2003; 88(7): 778-784.
Sukarova E Dimovski AJ Tchacarova P Petkov GH Efremov GD. An Alu insert as the cause of a severe form of hemophilia A. Acta Haematol 2001 106(3): 126-129.