Analysis of large structural changes of the factor VIII gene, involving intron 1 and 22, in severe hemophilia A

Institute of Enzymology, Hungarian Academy of Sciences, Budapeŝto, Budapest, Hungary
Haematologica (Impact Factor: 5.81). 08/2003; 88(7):778-84.
Source: PubMed


Hemophilia A (HA), the deficiency of coagulation factor VIII (FVIII), is the most common, sex-linked inherited bleeding disorder. The disease is caused by FVIII gene intron 22 inversion in approximately 50% of the patients, and by intron 1 inversion in 5% of the patients with severe HA. Both inversions occur as a result of intrachromosomal recombination between homologous regions, in intron 1 or 22, and their extragenic copy located telomeric to the FVIII gene. The goal of the present study was to analyze the presence of large structural changes in the FVIII gene in patients with severe hemophilia A.
We studied 104 unrelated, severe HA-patients or obligate carriers for the presence of intron 22 and intron 1 inversions by Southern blotting, long-distance polymerase chain reaction (PCR), and simple PCR.
We found altered intron 22 restriction profiles by Southern analyses in 58 cases: 43 type 1, 11 type 2 inversions and 4 unusual patterns. Upon further examination of the last 4 cases, large deletions involving intron 22 were demonstrated in two cases. In the remaining two patients extra homologous regions were detected by Southern analysis, and long-distance PCR showed the presence of unaltered intra- and extragenic copies together with one inversion-affected copy, suggesting that an additional intronic fragment participated in the inversion process and was inserted in the genome. During screening for intron 1 inversion among 43 patients, who were intron 22 inversion negative, we identified only wild type individuals.
The relatively large proportion of unusual patterns further supports the observation that the structure of FVIII intron 22 represents a hot spot for large gene rearrangements with various mechanisms, while intron 1 inversion seems to be not common in Hungary.

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    • "Among the identified intron 22 inversion types, the type I inversion was the most frequent in our patients (families 10, 11 and 12), which correlates with what previously has been reported in the literature [16]. The type II inversion was identified in one patient (family 9) and a novel atypical pattern (family 8) with 3 bands (18 kb, 15.5 kb and 14 kb), since the amplification of exons 22 and 23 gave the expected size this atypical pattern may be explained by a deletion of 2 kb in the INT22-h1 or in the INT22-h3 homologous regions (Table 1). "
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    • "It demonstrates that the chromosomal instability in Xq28 associated with the intron 22 homology regions may not only involve deletions but even additional insertions. Several independent reports of unusual patterns in diagnostic Southern blots or long-range PCR [summarized in (Andrikovics et al., 2003)] have led to the assumption that a number of patients may carry both an inversion and a deletion in F8 ( " rare inversion type " ). In a pilot study of DNA samples from 32 patients with severe hemophilia A carrying large deletions, we identified three cases with similar patterns (C. "
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    • "A mutant DNA sam ple was obtained by Dr. Cemal Un (Boaziçi University, Istanbul, Turkey) from a patient and used as a positive control in every amplification (Fig. 1). Thus far, the intron 1 inversion of the FVIII gene has not been investigated in West Anatolian HA patients, whereas this inversion has been detected in Italian (1.7-5%), in Spanish (5%), in the Czech Republic (4.3%), in the UK (1.8-3.8%), in Iranian (1.5%) and in Indian (3.8%) HA patients67891011. Although the number of HA patients is low, the results of this study provide the first data from the West Anatolian Turkish population. "
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