Spectrum of molecular defects and mutation detection rate in patients with mild and moderate hemophilia A

Abstract

The amount of residual F8 (FVIII:C) determines the clinical severity of hemophilia A. Recently, we showed that the mutation detection rate in severely affected male patients (FVIII:C<1% of normal) is virtually 100% when testing for the common intron 22-/intron 1- inversions and big deletions, followed by genomic sequencing of the F8 gene. Here we report on the spectrum of mutations and their distribution throughout the F8 gene sequence in 135 patients with moderate (n=23) or mild (n=112) hemophilia A. In contrast to the severe form of the disorder, analysis on the genomic level failed to detect the molecular defect in approximately 4% of the moderately and in approximately 12% of the mildly affected patients. A total of 36 of the mutations identified in this study are novel. The vast majority of the detected changes were missense. The newly detected amino acid substitutions were scored for potential distant or local conformational changes and influence on molecular stability for every single F8 domain with available structures, using homology modeling. Two molecular changes in the promoter region of the factor VIII gene (c.-112G>A and -219C>T), affecting the core segment (minimal promoter) were detected in two patients with mild hemophilia A. To our knowledge this is the first report on promoter mutations in the F8 gene.

Mutational spectrum of F8 gene and prothrombotic gene variants in haemophilia A patients from Southern Italy

“…In accordance to the previously published data [12], our study showed that the detection rate was higher in moderate cases than that observed in mild ones, although patients with moderate and mild HA formed a small group, possibly unrepresentative of all enrolled cases. Among severe patients, we found inv22 in 64 (59.2%) of them, a higher prevalence as compared with that of previous studies in North Italian and other populations [13,14].…”

“…This study revealed a marked heterogeneity of HA mutations in patients from Southern Italy, indicating that molecular analysis should include scanning of all exons [2,12]. Although molecular defects spread all over FVIII gene sequence, in severe patients, mutations were most frequently located in exon 14 encoding B domain, that has been shown to be a hot spot for small insertions/deletions resulting in frameshift and/or introduction of stop codon.…”

Characterization of duplication breakpoints in the factor VIII gene

“…Mutations in the FVIII gene ( F8 ), which spans 186 kb on chromosome Xq28 and consists of 26 exons, are the molecular basis of hemophilia A [3,4]. A great variety of missense and nonsense mutations distributed over the whole coding region of F8 have been published for patient cohorts of different ethnicity [5–12]. The most common mutation in severe hemophilia A patients is the inversion of intron 22, leading to a rearrangement of the F8 structure [13].…”

Pitfalls in the diagnosis of hemophilia severity: What to do?

“…The Worldwide Factor VIII Variant Database currently contains more than 2,000 F8 mutations in hemophilia A patients (http://www.factorviii-db.org). Nevertheless, in 2–18% of patients, no genetic abnormality is observed, depending on the type of mutational screening . Identical mutations may result in different FVIII:C baseline values, so DNA mutation analysis is not always conclusive for hemophilia severity.…”