A possible mechanism for Inv22-related F8 large deletions in severe hemophilia A patients with high responding factor VIII inhibitors.
BACKGROUND
Intron 22 inversion (Inv22) of the coagulation factor (F)VIII gene (F8) is a frequent cause of severe hemophilia A. In addition to Inv22, a variety of F8 mutations (1492 unique mutations) causing hemophilia A have been reported, of which 171 involve deletions of over 50 bp (HAMSTeRs database; http://hadb.org.uk/). However, only 10% of these large deletions have been fully characterized at the nucleotide level.


METHODS
We investigated gene abnormalities in three unrelated severe hemophilia A patients with high titer FVIII inhibitors. They had previously been shown to carry large deletions of the F8, but the precise gene abnormalities remain to be elucidated.


RESULTS
Inverse shifting-PCR (IS-PCR) Inv22 diagnostic tests revealed that these patients carried either type I or II Inv22. However, they showed a wild-type (WT) pattern in the IS-PCR Inv22 complementary tests. We further analyzed their X chromosomes to account for the puzzling results, and found that they had different centromeric breakpoints in the Inv22 X chromosomes, adjacent to the palindromic regions containing int22h-2 or -3, and their spacer region, respectively. The connections appeared to be shifted towards the telomere of the WT F8 Xq28, resulting in a new telomere with an additional intact int22h copy.


CONCLUSIONS
These gene rearrangements might result from double-strand breaks in the most distal regions of the long arms of the Inv22 X chromosomes, followed by DNA restorations using the WT F8 Xq28 by non-homologous end joining or break-induced replication; thus leading to large F8 deletions in severe hemophilia A patients.
Department of Pathophysiological Laboratory Sciences, Nagoya University Graduate School of Medicine, Nagoya Japan.
J
J
Fujita
Y
Y
Miyawaki
A
A
Suzuki
A
A
Maki
E
E
Okuyama
M
M
Murata
A
A
Takagi
T
T
Murate
N
N
Suzuki
T
T
Matsushita
H
H
Saito
T
T
Kojima
D000293:Adolescent; D000328:Adult; D001323:Autoantibodies / Q000097:blood*; D001483:Base Sequence; D056905:Chromosome Breakpoints; D002874:Chromosome Mapping; D041321:Chromosomes, Human, X; D053903:DNA Breaks, Double-Stranded; D004252:DNA Mutational Analysis; D004260:DNA Repair; D005169:Factor VIII / Q000235:genetics* / Q000276:immunology*; D015321:Gene Rearrangement; D020022:Genetic Predisposition to Disease; D006467:Hemophilia A / Q000097:blood / Q000235:genetics* / Q000276:immunology*; D006801:Humans; D007438:Introns; D055029:Inverted Repeat Sequences; D008297:Male; D010641:Phenotype; D016133:Polymerase Chain Reaction; D017384:Sequence Deletion; D057345:Sequence Inversion; D012720:Severity of Illness Index; D016615:Telomere / Q000235:genetics
D001323:Autoantibodies; C078147:F8 protein, human; D005169:Factor VIII