The Abnormal Factor IX of Hemophilia B+ Variants

1978 ◽  
Vol 40 (02) ◽  
pp. 335-349 ◽  
Author(s):  
Rogier M Bertina ◽  
Jan J Veltkamp
Keyword(s):  
Prothrombin Time ◽  
Electrophoretic Mobility ◽  
Human Factor ◽  
Binding Capacity ◽  
Factor Ix ◽  
Hemophilia B ◽  
Human Factor Ix

SummaryA rather large proportion of the hemophilia B patients can be characterized as hemophilia B+ because of the presence in their plasma of a protein which is immunologically identical with human factor IX. In a group of 33 hemophilia B patients we found 14 cases of hemophilia B+ belonging to 11 independent pedigrees. The variant factor IX molecules of these families have been compared with respect to the following properties: 1) factor IX activity and its dependence on phospholipid concentration; 2) factor IX antigen; 3) prolongation of prothrombin time with an ox brain thromboplastin; 4) electrophoretic mobility; 5) Ca2+ binding capacity; 6) affinity for binding to heparin and 7) susceptibility of the factor IX antigen to contact-induced activation. In the study of these parameters the use of a precipitating antibody against highly purified human factor IX showed to be of great value. According to our criteria at least 7 different factor IX variants were present in the 11 families with hemophilia B+ studied. Because of this rather high heterogeneity a suitable nomenclature for subclassification of hemophilia B+ variants is proposed.

scholarly journals Hemophilia B: Genetic Variants and Carrier Detection

1977 ◽  
Author(s):  
C.K. Kasper ◽  
B. Østerud ◽  
S.I. Rapaport
Keyword(s):  
Prothrombin Time ◽  
Genetic Variants ◽  
Human Factor ◽  
Factor Ix ◽  
Hemophilia B ◽  
Carrier Detection ◽  
Severe Hemophilia ◽  
Human Factor Ix ◽  
Normal Women

In 92 males with hemophilia B from 71 kindreds, we measured factor IX activity, prothrombin time using bovine thromboplastin (bovine tpln time), and factor IX antigen both by inhibitor neutralization using a human factor IX inhibitor and by electroimmunoassay using a precipitating rabbit anti-human-factor IX antiserum. Eighty patients with 3% or less factor IX activity could be divided into 4 groups:(1) 7 patients with greatly prolonged bovine tpln times and normal levels of factor IX antigen;(2) 17 patients with mildly prolonged bovine tpln times and factor IX antigen levels between about 25% and normal;(3) 8 patients with normal bovine tpln times and antigen levels between about 25% and normal; (4) 48 patients with normal bovine tpln times and no measureable antigen excess. Some of the latter group were also tested with a chicken anti-human-factor IX antiserum and no antigen was found. None of 12 patients with mild hemophilia B (factor IX activity of 4 to 22%) had a prolonged bovine tpln time although 4 patients had excess factor IX antigen over activity. Thus, about 1/3 of these 92 hemophilia B patients had evidence of an abnormal factor IX molecule. Factor IX activity was also measured in 48 normal women and in 51 definite carriers of severe hemophilia B. Probability curves were derived to estimate the chance of a woman being a carrier based upon her factor IX level and her degree of kinship to a definite carrier. The relation between factor IX activity and antigen was also delineated for normal women and for carriers. In kindreds in which affected males had excess antigen, some carriers could be distinguished from normal women on the basis of excess antigen over activity. In appropriate kindreds, prolonged bovine tpln times helped distinguish some carriers.

scholarly journals Production of human factor IX in animals by genetically modified skin fibroblasts: potential therapy for hemophilia B

Blood ◽  
1989 ◽  
Vol 73 (2) ◽  
pp. 438-445
Author(s):  
TD Palmer ◽  
AR Thompson ◽  
AD Miller
Keyword(s):  
Human Factor ◽  
Normal Skin ◽  
Human Fibroblasts ◽  
Retroviral Vectors ◽  
Factor Ix ◽  
Skin Fibroblasts ◽  
Hemophilia B ◽  
Clotting Factor ◽  
Human Factor Ix

Inherited diseases might be treated by introducing normal genes into a patient's somatic tissues to correct the genetic defects. In the case of hemophilia resulting from a missing clotting factor, the required gene could be introduced into any cell as long as active factor reached the circulation. We previously showed that retroviral vectors can efficiently transfer genes into normal skin fibroblasts and that the infected cells can produce high levels of a therapeutic product in vitro. In the current study, we examined the ability of skin fibroblasts to secrete active clotting factor after infection with different retroviral vectors encoding human clotting factor IX. Normal human fibroblasts infected with one vector secreted greater than 3 micrograms factor IX/10(6) cells/24 h. Of this protein, greater than 70% was structurally and functionally indistinguishable from human factor IX derived from normal plasma. This suggests that infected autologous fibroblasts might provide therapeutic levels of factor IX if transplanted into patients suffering from hemophilia B. By transplanting normal diploid fibroblasts infected with the factor IX vectors, we showed that human factor IX can be produced and is circulated at readily detectable levels in rats and mice.

scholarly journals Development of Monospecific Antibodies to Human Factor IX

1977 ◽  
Author(s):  
Cheryl Y. Tiarks ◽  
Chin-Hai Chang ◽  
Liberto Pechet
Keyword(s):  
Neutralizing Antibodies ◽  
Human Factor ◽  
Human Albumin ◽  
Factor Ix ◽  
Hemophilia B ◽  
Red Cross ◽  
Factor X ◽  
Normal Plasma ◽  
Human Factor Ix ◽  
Monospecific Antibodies

The purpose of this research was to develop neutralizing and precipitating antibodies to factor IX. Human factor IX, purified by the method of Rosenberg et.al. (J. Biol. Chem. 250:8883, 1975), was electrophoresed on acrylamide gel. Two major bands migrating adjacently were eluted. They contained factor IX activity only. The eluates and their homogenized gel segments 7 and 8 were injected separately into two rabbits, Rl and R2, respectively. On immunodiffusion the antiserum Rl showed one precipitating line with normal plasma. It neutralized human factor IX (20 Bethesda units) and also slightly neutralized factor X. It had no effect on factors II and VII. Following absorption of this antiserum with purified factor X it neutralized factor IX only. With continuous immunization, however, this antiserum revealed two new precipitating contaminants. The antiserum R2 neutralized only factor IX; it reached 220 Bethesda inhibitory units. On immunodiffusion it showed two precipitating lines, one of which disappeared after absorption with human albumin. On immunodiffusion and Laurell immunoelectrophoresis, the albumin-absorbed R2 antiserum showed one precipitin line of identity, or one rocket, with normal plasma, a Red Cross factor IX preparation (rich in factors IX, II and X), the original eluates 7 and 8, and a Hemophilia-B antigen-positive plasma. No line or rocket developed with normal plasma absorbed with aluminum hydroxide or with antigen-negative Hemophilia-B plasma. We conclude that the antisera Rl and R2 contain factor IX neutralizing antibodies and that albumin-absorbed R2 has monospecific precipitating antibodies to human non-activated factor IX.

scholarly journals Permanent partial phenotypic correction and tolerance in a mouse model of hemophilia B by stem cell gene delivery of human factor IX

Gene Therapy ◽  
2005 ◽  
Vol 13 (2) ◽  
pp. 117-126 ◽  
Author(s):  
B W Bigger ◽  
E K Siapati ◽  
A Mistry ◽  
S N Waddington ◽  
M S Nivsarkar ◽  
...  
Keyword(s):  
Stem Cell ◽  
Gene Delivery ◽  
Mouse Model ◽  
Human Factor ◽  
Factor Ix ◽  
Hemophilia B ◽  
Human Factor Ix ◽  
Cell Gene ◽  
Stem Cell Gene

scholarly journals Use of a BamHI polymorphism in the factor IX gene for the determination of hemophilia B carrier status

Blood ◽  
1986 ◽  
Vol 67 (5) ◽  
pp. 1508-1511 ◽  
Author(s):  
CW Hay ◽  
KA Robertson ◽  
SL Yong ◽  
AR Thompson ◽  
GH Growe ◽  
...  
Keyword(s):  
Human Factor ◽  
Factor Ix ◽  
Hemophilia B ◽  
Carrier Status ◽  
X Chromosomes ◽  
Human Factor Ix ◽  
History Of

Abstract A BamHI polymorphism has been identified in the human factor IX gene. This polymorphism, which occurs in approximately 6% of X chromosomes, has been used to determine the carrier status of a female in a family with a history of hemophilia B. This family was uninformative for the previously reported TaqI and Xmnl polymorphisms in the factor IX gene.

scholarly journals Delivery of human factor IX in mice by encapsulated recombinant myoblasts: a novel approach towards allogeneic gene therapy of hemophilia B

Blood ◽  
1996 ◽  
Vol 87 (12) ◽  
pp. 5095-5103 ◽  
Author(s):  
G Hortelano ◽  
A Al-Hendy ◽  
FA Ofosu ◽  
PL Chang
Keyword(s):  
Gene Therapy ◽  
Human Factor ◽  
Ex Vivo ◽  
Cost Effective ◽  
Factor Ix ◽  
Hemophilia B ◽  
Therapy Strategy ◽  
Human Factor Ix

A potentially cost-effective strategy for gene therapy of hemophilia B is to create universal factor IX-secreting cell lines suitable for implantation into different patients. To avoid graft rejection, the implanted cells are enclosed in alginate-polylysine-alginate microcapsules that are permeable to factor IX diffusion, but impermeable to the hosts' immune mediators. This nonautologous approach was assessed by implanting encapsulated mouse myoblasts secreting human factor IX into allogeneic mice. Human factor IX was detected in the mouse plasma for up to 14 days maximally at approximately 4 ng/mL. Antibodies to human factor IX were detected after 3 weeks at escalating levels, which were sustained throughout the entire experiment (213 days). The antibodies accelerated the clearance of human factor IX from the circulation of the implanted mice and inhibited the detection of human factor IX in the mice plasma in vitro. The encapsulated myoblasts retrieved periodically from the implanted mice up to 213 days postimplantation were viable and continued to secrete human factor IX ex vivo at undiminished rates, hence suggesting continued factor IX gene expression in vivo. Thus, this allogeneic gene therapy strategy represents a potentially feasible alternative to autologous approaches for the treatment of hemophilia B.

scholarly journals Production of human factor IX in animals by genetically modified skin fibroblasts: potential therapy for hemophilia B

Blood ◽  
1989 ◽  
Vol 73 (2) ◽  
pp. 438-445 ◽  
Author(s):  
TD Palmer ◽  
AR Thompson ◽  
AD Miller
Keyword(s):  
Human Factor ◽  
Normal Skin ◽  
Human Fibroblasts ◽  
Retroviral Vectors ◽  
Factor Ix ◽  
Skin Fibroblasts ◽  
Hemophilia B ◽  
Clotting Factor ◽  
Human Factor Ix

Abstract Inherited diseases might be treated by introducing normal genes into a patient's somatic tissues to correct the genetic defects. In the case of hemophilia resulting from a missing clotting factor, the required gene could be introduced into any cell as long as active factor reached the circulation. We previously showed that retroviral vectors can efficiently transfer genes into normal skin fibroblasts and that the infected cells can produce high levels of a therapeutic product in vitro. In the current study, we examined the ability of skin fibroblasts to secrete active clotting factor after infection with different retroviral vectors encoding human clotting factor IX. Normal human fibroblasts infected with one vector secreted greater than 3 micrograms factor IX/10(6) cells/24 h. Of this protein, greater than 70% was structurally and functionally indistinguishable from human factor IX derived from normal plasma. This suggests that infected autologous fibroblasts might provide therapeutic levels of factor IX if transplanted into patients suffering from hemophilia B. By transplanting normal diploid fibroblasts infected with the factor IX vectors, we showed that human factor IX can be produced and is circulated at readily detectable levels in rats and mice.

Sustained and therapeutic levels of human factor IX in hemophilia B mice implanted with microcapsules: key role of encapsulated cells

2006 ◽  
Vol 8 (3) ◽  
pp. 362-369 ◽  
Author(s):  
Jianping Wen ◽  
Andrew Gómez Vargas ◽  
Frederick A. Ofosu ◽  
Gonzalo Hortelano
Keyword(s):  
Human Factor ◽  
Factor Ix ◽  
Hemophilia B ◽  
Encapsulated Cells ◽  
Human Factor Ix ◽  
B Mice

scholarly journals Self-complementary adeno-associated virus vectors containing a novel liver-specific human factor IX expression cassette enable highly efficient transduction of murine and nonhuman primate liver

Blood ◽  
2006 ◽  
Vol 107 (7) ◽  
pp. 2653-2661 ◽  
Author(s):  
Amit C. Nathwani ◽  
John T. Gray ◽  
Catherine Y. C. Ng ◽  
Junfang Zhou ◽  
Yunyu Spence ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Human Factor ◽  
Factor Ix ◽  
Expression Cassette ◽  
Hemophilia B ◽  
Adeno Associated Virus ◽  
Knock Out ◽  
Human Factor Ix ◽  
Murine Liver ◽  
Knock Out Mice

AbstractTransduction with recombinant adeno-associated virus (AAV) vectors is limited by the need to convert its single-stranded (ss) genome to transcriptionally active double-stranded (ds) forms. For AAV-mediated hemophilia B (HB) gene therapy, we have overcome this obstacle by constructing a liver-restricted mini–human factor IX (hFIX) expression cassette that can be packaged as complementary dimers within individual AAV particles. Molecular analysis of murine liver transduced with these self-complementary (sc) vectors demonstrated rapid formation of active ds-linear genomes that persisted stably as concatamers or monomeric circles. This unique property resulted in a 20-fold improvement in hFIX expression in mice over comparable ssAAV vectors. Administration of only 1 × 1010 scAAV particles led to expression of hFIX at supraphysiologic levels (8I U/mL) and correction of the bleeding diathesis in FIX knock-out mice. Of importance, therapeutic levels of hFIX (3%-30% of normal) were achieved in nonhuman primates using a significantly lower dose of scAAV than required with ssAAV. Furthermore, AAV5-pseudotyped scAAV vectors mediated successful transduction in macaques with pre-existing immunity to AAV8. Hence, this novel vector represents an important advance for hemophilia B gene therapy.

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