Immune response against serial infusion of factor VIII antigen through an implantable venous-access device system in haemophilia A mice

Haemophilia A is a life long bleeding disorder caused by an inherited deficiency of factor VIII (FVIII). About 30% of haemophilia A patients develop neutralizing antibodies as a consequence of treatment with FVIII concentrates. Immune tolerance protocols for the eradication of inhibitors require daily delivery of intravenous FVIII. We evaluated the immune responses to serial intravenous administration of FVIII in preimmunized haemophilia A mice. We introduced an implantable venous-access device (iVAD) system into haemophilia A mice to facilitate sequential infusion of FVIII. After preimmunization with FVIII, the haemophilia A mice were subjected to serial intravenous administration of FVIII through the iVAD system. In all mice with serial infusion of FVIII, high titers of anti-FVIII inhibitory antibodies developed at 10 exposure days (EDs). However, the anti-FVIII IgG titers were decreased after 150 EDs of sequential low-dose infusion of FVIII [0.05 U g(-1) body weight (BW) five times per week]. Proliferative response to ex vivo FVIII stimulation was significantly suppressed in splenic CD4(+) T cells from mice with serial low-dose FVIII infusion compared with those from mice with high-dose FVIII infusion (0.5 U g(-1) BW five times per week) or preimmunized mice. Moreover, splenic CD4(+) T cells from mice with serial low-dose infusion of FVIII failed to produce interleukin-2 and interferon-γ. These data suggest that serial infusion of FVIII could induce T-cell anergy in haemophilia A mice with inhibitor antibodies.     

Combined prednisolone and intravenous immunoglobulin treatment for acquired factor VIII inhibitors: a 2-year review

Acquired inhibitors to factor VIII (FVIII) are rare, but life-threatening in up to 22% of cases. The optimal therapy for suppression of these inhibitors remains unclear. Prednisolone is the mainstay of therapy, producing responses in approximately 30% of cases. Intravenous immunoglobulin (IVIg) has a similar response rate, but a more rapid effect. We report the results of prednisolone 1 mg kg(-1) combined with IVIg 2 g kg(-1) in divided doses as first-line therapy in seven consecutive patients with acquired FVIII inhibitors. All patients were bleeding at the time of diagnosis with prolonged activated partial thromboplastin time. There were four complete responses, one partial response, one nonresponse and one with an inadequate follow-up for assessment of response, giving an overall response rate of 71%. In all complete responders the inhibitor declined rapidly and was undetectable by day 21 from start of treatment. Therapy was well tolerated and responses have been maintained off treatment for 2-8 months. This is a safe, well-tolerated rapidly acting regimen with good response rates.     

Spinal epidural haematoma in haemophilia A with inhibitors – efficacy of recombinant factor VIIa concentrate

We report the case of a 21-year-old man with severe haemophilia A and factor VIII inhibitors who presented with an extensive spinal epidural haematoma (C2-T12), probably induced by sit-up exercises. The bleed was defined by magnetic resonance imaging of the cervical and thoracic spine and prompt treatment with recombinant factor VIIa concentrate led to complete resolution at 4 weeks. Neurological sequelae were averted and surgical decompression was not necessary. We discuss the difficulties in diagnosis and management of such a case.     

Management of oral bleedings with recombinant factor VIIa in children with haemophilia A and inhibitor

Dental extraction in patients with haemophilia A and high-titre inhibitor is always a high-risk procedure, which often presents a lot of problems associated with bleeding. Prothrombin complex concentrates or recombinant activated factor VII (rFVIIa) has been used to control bleeding. rFVIIa was administered to five boys with severe haemophilia A complicated with inhibitor, who underwent seven dental extractions. The age of the patients ranged between 8 and 13 years (median 10 years). The concentrate was administered in doses of 90-100 microg kg(-1) body weight. Duration in the therapy and intervals between rFVIIa doses depended on the severity of bleeding. rFVIIa was proven to be highly effective and no side-effects of the product were observed.     

The prevalence of factor VIII inhibitors and genetic aspects of inhibitor development in Chinese patients with haemophilia A

Summary. The prevalence of inhibitors in Chinese haemophiliacs has not yet been reported. The aim of this study was to identify the prevalence of factor VIII (FVIII) inhibitors among haemophiliacs who are treated only with plasma‐derived FVIII (pdFVIII), cryoprecipitate or fresh frozen plasma (FFP), and tried to explore the relationship between the generation of inhibitors and particular FVIII deficiency genotypes. Clinical information and blood samples of 1435 patients with haemophilia A (HA) were collected by six haemophilia centres in China. The Nijmegen modification of the Bethesda assay was used to detect inhibitors. Multiplex PCR, long‐range PCR and direct sequencing were performed for genotyping. The overall prevalence of inhibitors in Chinese HA patients was 3.9% and the prevalence of severe haemophiliacs was 4.3%; 18 of the 56 patients with inhibitors had high titres. A total of 38 different mutations were identified in the 55 patients with inhibitors, including 15 intron 22 and 3 intron 1 inversions, seven large deletions, 14 small deletion/insertions, seven nonsense mutations, one splice site mutations and eight missense mutations. Of 38 mutations, 28 were novel. Patients with large deletions and nonsense mutations were prone to have high titre inhibitors, with incidence rates of 57.1% (4/7) and 42.9% (3/7), respectively. In conclusion, the prevalence of inhibitors in Chinese HA patients is much lower than that reported for other ethnic groups and the large deletion and nonsense mutations are high risk factors for high titre inhibitor development.     

Use of recombinant activated factor VII for treatment of a retropharyngeal hemorrhage in a hemophilic patient with a high titer inhibitor

Treatment options during an acute hemorrhage for a hemophilic patient who has an inhibitor (antibody) to factor VIII (FVIII) are limited. If the inhibitor titer is high, even massive doses of FVIII are not sufficient to neutralize the antibody. Likewise, immunoadsorption techniques or plasmapheresis cannot remove enough antibody to permit treatment with FVIII. Allergic reactions and cross-reacting inhibitors complicate therapy with porcine FVIII. Prothrombin complex concentrates (PCC) may be effective but the mechanism is unclear. The theory that activated factor VII (FVIIa) is the active principle in PCC prompted our treatment of a patient with a recombinant FVIIa (rFVIIa) product (NOVO Industries). The patient presented with a large retropharyngeal hemorrhage and an initial inhibitor titer of 129 Bethesda units (BU). Despite Autoplex therapy (100U/kg), tracheal compression by the hematoma increased and asphyxiation was imminent. RFVIIa therapy (60 micrograms/kg) was substituted for Autoplex and nine doses were given without complication. The hemorrhage was controlled. By 18 hr breathing was normal and swallowing and speech were greatly improved. Clinically, the patient dramatically responded to rFVIIa. In addition, the purity, the lack of known infectious agents, and the ease of administration make rFVIIa a potentially attractive new therapy. Use of this product also promises to further our understanding of in vivo hemostasis.     

A modified chromogenic assay for the measurement of very low levels of factor VIII activity (FVIII:C)

A precise and sensitive chromogenic assay for the measurement of very low levels of factor VIII (FVIII) in plasma has been developed. The assay is based on modifications of a commercially available chromogenic assay. The modifications include reduction of sample final dilution factor and prolongation of the development period. The modified assay allows accurate and precise measurement of FVIII in the range of 0.001-0.02 IU mL(-1). The detection limit is 0.0005 IU mL(-1) and the quantitation limit is 0.0015 IU mL(-1). This assay can be used in research and to study the clinical efficacy of low circulating levels of FVIII in haemophilia A patients.     

Unresponsiveness to factor VIII inhibitor bypassing agents during haemostatic treatment for life-threatening massive bleeding in a patient with haemophilia A and a high responding inhibitor

We report a case of haemophilia A with a high responding inhibitor of factor VIII (FVIII) who had a serious retroperitoneal haematoma caused by penetration of a duodenal ulcer. Inhibitor-bypassing therapy was commenced immediately on admission. On the 17th day of treatment with activated prothrombin complex concentrate (APCC; FEIBA, re-bleeding occurred and thrombelastography (TEG) demonstrated resistance to therapy. Treatment was changed to recombinant activated factor VII (rFVIIa; NovoSeven and resulted in clinical improvement together with an improvement in TEG parameters. On the 10th day of continuous infusion with NovoSeven, however, TEG again showed resistance to therapy. FEIBA infusions were re-introduced and TEG results remained satisfactory for 7 days. On day 34, however, further retroperitoneal bleeding was evident and a decline in the haemostatic efficiency of FEIBA was recorded by TEG. NovoSeven was again successfully administered for 7 days. There were no laboratory findings to indicate disseminated intravascular coagulation (DIC), hypercoagulability or abnormal fibrinolysis. The plasma-based clotting tests did not show any additional prolongation on the occasions when the TEG demonstrated unresponsiveness to FEIBA or NovoSeven. These findings suggested that some component of whole blood, other than plasma might have governed the TEG data. The long-term use of APCC such as FEIBA or rFVIIa, requires careful monitoring in terms of FVIII inhibitor bypassing activity as well as the tendency to DIC.     

Acquired factor VIII inhibitor associated with chronic interferon-alpha therapy in a patient with haemophilia A

SUMMARY. Both the development of factor VIII inhibitors and infection by hepatitis C virus are serious complications of haemophilia A. We describe the first reported case of the subsequent development of a factor VIII inhibitor in a patient with haemophilia A after treatment with interferon-alpha for chronic active hepatitis C.     

Breastfeeding does not influence the development of inhibitors in haemophilia

Our aim was to test the hypothesis that breastfeeding may reduce development of inhibitors in male infants with haemophilia by inducing an oral immune tolerance to factor VIII. To achieve that goal, we performed a structured epidemiological survey comprising all males born with severe haemophilia A (in all 100 patients, 19 with inhibitors) or haemophilia B (in all 16 patients, six with inhibitors) in Sweden in 1980-99. Our results show no protective effect of breastfeeding.     

Successful treatment of acute subdural haemorrhage with continuous intravenous infusion of factor VIII in a 17 year old with haemophilia A

Intracranial haemorrhage is a common complication of haemophilia, occurring in 2–8% of sufferers [1]. Half of the cases give a history of trauma and despite developments in the management of acute bleeding the condition still carries a mortality of approximately 30% [2]. Surgery may be required, most commonly for evacuation of a subdural haematoma which carries a mortality of up to 40% [3]. We present the case of a 17-year-old haemophiliac with a traumatic subdural haemorrhage, who was treated with a continuous intravenous infusion of factor VIII and made a complete recovery without recourse to surgery. We have found no reference in the world literature of such treatment for acute subdural haemorrhage in a haemophiliac.     

Low response to high-dose intravenous immunoglobulin in the treatment of acquired factor VIII inhibitor

Prednisone is the classic first-line therapy to suppress an acquired factor VIII inhibitor and may achieve complete remission in about 30% of patients. More recently, promising results have been reported with high-dose intravenous immunoglobulin (IVIg). However, after an extensive review of the literature, we found only three complete remissions (12%) among the 26 assessable patients treated by IVIg. These data are in agreement with the low response to IVIg that we experienced in our series of patients. This study suggests that steroids should still be preferred to IVIg, an expansive therapy, to suppress an acquired factor VIII inhibitor.     

Mechanisms of action of immune tolerance induction against factor VIII in patients with congenital haemophilia A and factor VIII inhibitors

In its most severe form, haemophilia A is a life-threatening haemorrhagic bleeding disorder that is caused by mutations in the factor VIII (FVIII) gene. About 25% of patients who receive replacement therapy with intravenous FVIII products develop neutralising antibodies (FVIII inhibitors) that inhibit the function of substituted FVIII. Long-term application of high or low doses of FVIII has evolved as an effective strategy for eradicating antibodies and inducing long-lasting immune tolerance. Despite clinical experience with the therapy, little is known about the immunological mechanisms that cause the down modulation of FVIII-specific immune responses or the induction of long-lasting immune tolerance against FVIII. This review summarises current knowledge of the immunological mechanisms that might be involved in the induction of immune tolerance against FVIII in patients with haemophilia A who have FVIII inhibitors. In addition to data from patients with haemophilia A, data from patients who have had organ transplants or have immune-related disorders, such as autoimmune diseases, are considered as well as data from animal models.     

Pharmacokinetics and safety of OBI-1, a recombinant B domain-deleted porcine factor VIII, in subjects with haemophilia A

Summary. OBI‐1 is a recombinant B‐domain deleted porcine factor VIII (FVIII). FVIII treatment in those with haemophilia A may be complicated by the development of anti‐FVIII antibodies (inhibitors) leading to a failure to respond to treatment with human FVIII. To compare the pharmacokinetics and safety of a single dose of OBI‐1 with Hyate:C in subjects with haemophilia A and inhibitors, subjects were randomized to receive either Hyate:C followed by placebo or placebo followed by OBI‐1 in a double‐blind fashion. FVIII levels were assayed using both a one‐stage coagulation assay (OSCA) and chromogenic assay. Pharmacokinetic parameters for FVIII were calculated for 6/9 subjects randomized; in three subjects baseline anti‐porcine FVIII inhibitors led to a lack of measurable FVIII activity. Mean C_max appeared higher for OBI‐1 (OSCA: 176.00 U dL^?1, standard deviation ± 88.00; chromogenic: 151.00 ± 31.51 U dL^?1) than Hyate:C (OSCA: 82.3 ± 19.22 U dL^?1; chromogenic: 52.67 ± 13.8 U dL^?1). Mean AUC also appeared higher for OBI‐1 (OSCA: 2082.87 ± 1323.43 U h^?1dL^?1; chromogenic: 1817.28 ± 625.14 U h^?1dL^?1) than Hyate:C (OSCA: 1177.8 ± 469.49 U h^?1dL^?1; chromogenic: 707.61 ± 420.05 U h^?1dL^?1). Two infusion‐related events occurred: one with Hyate:C, one with placebo. Four of five subjects without anti‐porcine FVIII inhibitors at baseline remained porcine FVIII inhibitor negative 29 days after infusion. A single dose of OBI‐1 appears to have higher bioavailability than Hyate:C in subjects with haemophilia A without measurable anti‐porcine FVIII inhibitors, and is well tolerated. These results should be confirmed in a larger phase 2/3 study.     

Excision of pseudotumour in a patient with haemophilia A and inhibitor managed with recombinant factor VIIa

We describe a patient with haemophilia A and factor VIII inhibitor who underwent surgical excision of a large pseudotumour in the left femoral region. Haemostasis was successfully maintained with bolus doses of recombinant factor VIIa at 2-h intervals and anti-fibrinolytic therapy, and the pseudotumour was removed. Subsequently, the dose interval was gradually prolonged to 8 h for a total of 18 days. Although a spontaneous haemorrhage was observed on postoperative day 8, haemostasis was achieved by reducing the dosage interval. No adverse event occurred during the course of treatment.     

Characterization of inhibitors to FVIII with an ELISA in congenital and acquired haemophilia A

Different methods can be used for the detection and quantification of inhibitors or antibodies to coagulation factor VIII (FVIII). Traditionally, clotting assays have been used, in particular the Bethesda assay. These assays have, however, several shortcomings, due to the complex reaction kinetics of some inhibitors and a low sensitivity to low-titre antibodies. In addition, a universal FVIII inhibitor standard is lacking. Furthermore, clotting assays do not detect noninhibitory antibodies. Use of ELISAs has been described and FVIII from various commercially available FVIII concentrates has been used as target antigen in the assays. In the present study, we systematically explored the influence of different FVIII concentrates on the performance of an ELISA for detection of FVIII antibodies. The described ELISA was also used for further characterization of FVIII inhibitors in patients with acquired and congenital haemophilia A. We found that the source of FVIII had a substantial impact on the frequency of antibody detection. Albumin-free recombinant FVIII as target antigen gave the highest sensitivity for the assay, whereas plasma-derived concentrates containing a high level of von Willebrand factor (vWF) gave the lowest sensitivity. Presumably vWF interferes with the binding of antibodies to FVIII. We suggest that albumin-free recombinant FVIII should be used as target antigen when ELISAs are used for detection of FVIII antibodies.