Gene deletions in patients with haemophilia B and anti-Factor IX antibodies

1987 ◽  
Author(s):  
B M Ludwig ◽  
R Schwaab ◽  
H H Brackmann ◽  
H Egli ◽  
K Olek

Deficiency or functional abnormality of factor IX protein leads to the X-linked recessive haemorrhagic disorder known as haemophilia B. Using the factor IX cDNA probe pTG 397 we have studied DNA purified from 40 patients afflicted with haemophilia B. Restriction analysis was carried out using Eco RI, Taq I and Xmn I which give result to DNA fragments representing 99 % of the whole factor IX gene. Mapping the gene this way four patients were shown to have structural alterations in the factor IX gene. To obtain more precise data concerning these mutations we additionally used restriction enzymes Bgl II and Hind III.The results listed below indicate more patients have to be examined to conclude a strong correlation between inhibitors to factor IX and the presence of gross gene deletions. This view is supported by the findings that two further inhibitor forming patients had no sizable deletions.


Nature ◽  
1983 ◽  
Vol 303 (5913) ◽  
pp. 181-182 ◽  
Author(s):  
F. Giannelli ◽  
K. H. Choo ◽  
D. J. G. Rees ◽  
Y. Boyd ◽  
C. R. Rizza ◽  
...  

1987 ◽  
Author(s):  
B F Giannelli

Haemophilia B, an X-linked recessive disease with an incidence of 1/30,000 newborn males, is due to defects in the gene for coagulation factor IX, which is on the long am of the X chromosome at band Xq27.1. This gene consists of approximately 34 Kb and contains 8 exons which specify a mRtfc of 2803 residues coding for a protein of 415 aa preceded by a prepro signal peptide of 46 aa. Coripanson of the functional domains of the factor IX protein with the exon structure of the gene supports the exon/protein domain hypothesis of gene evolution. The factor IX gene seems to be formed by a number of functionally and evolutionally independent modules. The signal peptide and the gla (γcarboxy-glutamic) region encoded in the first three exons are homologous to those of factor X, protein C and prothrombin. Thevfourth and fifth exons which code for the connecting peptide are homologous to one another and to the epidermal growth factor, a module that has been used in the construction of a great variety of proteins including different members of the coagulation and fibrinolytic pathways. The sixth exon encodes the activation peptide region, while the catalytic region of factor IX is coded by the seventh and eighth exon. This is at variance with other serine protease genes that have different exons for the segments containing the cardinal ami no-acids of the active centre (histidine, aspartic acid and serine).Natural selection acts against detrimental mutations of the factor IX gene and at each generation a proportion of haemophilia B genes is eliminated, as a significant number of patients does not reproduce. There appears to be no selective advantage to the heterozygote and therefore haemophilia B is maintained in the population by new mutations. Consequently, a significant proportion of patients should be born to non-carrier mothers, and unrelated patients should carry different gene defects, as recently verified by detailed analysis of individual haemophilia B genes.The defects of factor IX described so far comprise both point mutations and gene deletions. The latter affect either part or the whole of the gene and are often associated with the development of antibodies against therapeutically adninistered factor IX (the inhibitor complication). Since gene deletions may result in the complete absenceof factor IX synthesis or in the production of an extremely abnormal product, it has been suggested that mutationspreventing the synthesis of a factor IX gene product capable of inducing immune tolerance to normal factor IX is important in predisposing to the inhibitor complication.Among the point mutations described so far, those affecting the signal peptide are of particular interest. Substitutions of the arginine at positions -4 and -1 cause failure of propeptide cleavage. Thus they indicate that the propeptide consists of 18 aa an(lthat lts excision is necessary for factor IX function. It appears also that the propeptide contains a signal for γcarboxylation which has been conserved during the evolution of different γcarboxylated proteins.In spite of coagulant treatment, haemophilia B is a serious disease and one for which genetic counselling is required. Paramount for this is the detection of carriers and the diagnosis ofaffected male fetuses. DNA probes derived from the cloned factor IX gene have been used for this purpose. Carrier and first or second trimester prenatal diagnoses have been done using factors IX gene markers to follow the transmission of haemophilia B genes. Six sequence variations causing restriction fragment length polymorphisms (RFLP) in the factor IX gene have been detected and used as markers for such indirect diagnoses The efficiency of the above markers is reduced by linkage disequilibrium but, nevertheless, they offer definite carrier and nremtal diagnoses in 75-80% of the relatives of familial cases of haemophilia B.The indirect detection of gene defects is of modest help in the counselling of individuals from the families of isolated patients, but new methods for the direct detection of gene mutations promise better results in such families and also the attainment of % diagnostic success in relatives of familial cases.Finally the successful expression of recombinant factor IX genes in tissue culture and transgenic mammals raises hopes of therapeutic advances.


1987 ◽  
Author(s):  
R J Matthews ◽  
D S Anson ◽  
I R Peake ◽  
A L Bloom

Previous studies have indicated that the majority of haemophilia B patients who produce anti-factor IX inhibitors (antibodies) have some kind of deletion of the factor IX gene. We have analysed DNA from nine haemophilia B inhibitor patients using the Southern blotting method and hybridisation with (i) factor IX cDNA and intragenomic probes (ii) probes originating from flanking sequences up to 60kb 5' and 170kb 3' to the factor IX gene that have been isolated by gene walking experiments (D.S.Anson and G.G.Brownlee, unpublished observations).Two patients who are brothers (haemophilia B (Chicago I)) were shown to have a presumably identical complex rearrangement of the factor IX gene involving two separate deletions. The first deletion is approx. 5.0kb and removes exon e. The second deletion is between 9 and 29kb and removes exons g and h but leaves exon f intact. An abnormal TaqI fragment at one end of the deletions junctions acted as a marker for the inheritance of haemophilia B in the patients' family. Furthermore, an abnormal llkb Bglll fragment (detected with an intragenomic probe containing exon f) in DNA from both patients and their mother acted as a marker for the presence of both deletions. Since the patients' grandmother only showed the normal 12kb Bgl II fragment then both deletions appear to have arisen at the same time. We believe that haemophilia B (Chicago 1) is the first observation of a natural gene rearrangement involving two separate deletions within the same gene.Patient haemophilia B (Jersey 1) was revealed to have a deletion of at least 170kb including the entire factor IX gene and > 60kb of 5' flanking sequence. The 3' breakpoint of this deletion was mapped to between 80 and 140kb 3' to the factor IX gene. One further patient, haemophilia B (Boston I) was shown to have a deletion of > 230kb including the factor IX gene, > 60kb of 5' flanking sequence and >140kb of 3' flanking sequence. Five other inhibitor patients had a structurally intact gene as detected by this method.Although all nine haemophilia B inhibitor patients studied did not have a detectable plasma factor IX only in four of them is this absence due to a large deletion of the factor IX gene.


Haemophilia ◽  
1999 ◽  
Vol 5 (2) ◽  
pp. 101-105 ◽  
Author(s):  
E. C. THORLAND ◽  
J. B. DROST ◽  
J. M. LUSHER ◽  
I. WARRIER ◽  
A. SHAPIRO ◽  
...  

1975 ◽  
Vol 33 (03) ◽  
pp. 547-552 ◽  
Author(s):  
L Meunier ◽  
J. P Allain ◽  
D Frommel

SummaryA mixture of adsorbed normal human plasma and chicken plasma was prepared as reagent for factor IX measurement using a one-stage method. The substrate was found to be specific for factor IX. Its performances tested on samples displaying factor IX activity ranging from <l%–2,500% compared favorably with those obtained when using the plasma of severe haemophilia B patients as substrate.


1976 ◽  
Vol 35 (03) ◽  
pp. 510-521 ◽  
Author(s):  
Inga Marie Nilsson

SummaryThe incidence of living haemophiliacs in Sweden (total population 8.1 millions) is about 1:15,000 males and about 1:30,000 of the entire population. The number of haemophiliacs born in Sweden in 5-year periods between 1931-1975 (June) has remained almost unchanged. The total number of haemophilia families in Sweden is 284 (77% haemophilia A, 23% haemophilia B) with altogether 557 (436 with A and 121 with B) living haemophiliacs. Of the haemophilia A patients 40 % have severe, 18 % moderate, and 42 % mild, haemophilia. The distribution of the haemophilia B patients is about the same. Inhibitors have been demonstrated in 8% of the patients with severe haemophilia A and in 10% of those with severe haemophilia B.There are 2 main Haemophilia Centres (Stockholm, Malmo) to which haemophiliacs from the whole of Sweden are admitted for diagnosis, follow-up and treatment for severe bleedings, joint defects and surgery. Minor bleedings are treated at local hospitals in cooperation with the Haemophilia Centres. The concentrates available for treatment in haemophilia A are human fraction 1-0 (AHF-Kabi), cryoprecipitate, Antihaemophilic Factor (Hyland 4) and Kryobulin (Immuno, Wien). AHF-Kabi is the most commonly used preparation. The concentrates available for treatment in haemophilia B are Preconativ (Kabi) and Prothromplex (Immuno). Sufficient amounts of concentrates are available. In Sweden 3.2 million units of factor VIII and 1.0 million units of factor IX are given per year. Treatment is free of charge.Only 5 patients receive domiciliary treatment, but since 1958 we in Sweden have practised prophylactic treatment of boys (4–18 years old) with severe haemophilia A. At about 5-10 days interval they receive AHF in amounts sufficient to raise the AHF level to 40–50%. This regimen has reduced severe haemophilia to moderate. The joint score is identical with that found in moderate haemophilia in the same age groups. For treatment of patients with haemophilia A and haemophilia B complicated by inhibitors we have used a large dose of antigen (factor VIII or factor IX) combined with cyclophosphamide. In most cases this treatment produced satisfactory haemostasis for 5 to 30 days and prevented the secondary antibody rise.


1961 ◽  
Vol 6 (02) ◽  
pp. 224-234 ◽  
Author(s):  
E. T Yin ◽  
F Duckert

Summary1. The role of two clot promoting fractions isolated from either plasma or serum is studied in a purified system for the generation of intermediate product I in which the serum is replaced by factor X and the investigated fractions.2. Optimal generation of intermediate product I is possible in the purified system utilizing fractions devoid of factor IX one-stage activity. Prothrombin and thrombin are not necessary in this system.3. The fraction containing factor IX or its precursor, no measurable activity by the one-stage assay method, controls the yield of intermediate product I. No similar fraction can be isolated from haemophilia B plasma or serum.4. The Hageman factor — PTA fraction shortens the lag phase of intermediate product I formation and has no influence on the yield. This fraction can also be prepared from haemophilia B plasma or serum.


Haemophilia ◽  
2021 ◽  
Author(s):  
Akihiro Tamura ◽  
Keiko Shinozawa ◽  
Suguru Uemura ◽  
Sayaka Nakamura ◽  
Takahiro Fujiwara ◽  
...  

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