The use of a single von Willebrand factor‐containing,plasma‐derived FVIII product in hemophilia A immune tolerance induction: the US experience

Summary. Background: Inhibitors are a serious complication for patients with severe hemophilia A. Immune tolerance induction (ITI) is the primary method for eradicating these inhibitors. The role of type of concentrate and in particular the use of von Willebrand factor‐containing, plasma‐derived factor VIII (VWF/pd‐FVIII) concentrate in primary or rescue ITI remains unclear. Objectives: To report retrospective collection of data on the use of a single VWF/pd‐FVIII concentrate in primary and rescue ITI. Methods: Retrospective chart review of hemophilia A inhibitor patients at 11 US institutions who received VWF/pd‐FVIII concentrate in primary or rescue ITI. Results: Primary ITI was carried out in eight inhibitor patients with a 75% complete and partial success. Secondary ITI was carried out in 25 inhibitor patients, with 52% attaining complete or partial success. Conclusions: This report represents the largest group of primarily pediatric, high‐titer inhibitor patients treated with a single VWF/pd‐FVIII concentrate. It adds retrospective data to the use of VWF‐containing plasma‐derived factor VIII concentrate in primary and rescue ITI, particularly in those patients with characteristics of poor response to ITI.     

Induction of tolerance to factor VIII by transient co‐administration with rapamycin

See also Miao CH. Tilt balance towards regulation: evolving new strategy for treatment of hemophilia inhibitors. This issue, pp 1521–3.DOI: 10.1111/j.1538‐7836.2011.04351.x . Summary. Background: Formation of inhibitory antibodies is a frequent and serious complication of factor (F) VIII replacement therapy for the X‐linked bleeding disorder hemophilia A. Similarly, hemophilia A mice develop high‐titer inhibitors to recombinant human FVIII after a few intravenous injections. Objective: Using the murine model, the study sought to develop a short regimen capable of inducing tolerance to FVIII. Methods: A 1‐month immunomodulatory protocol, consisting of FVIII administration combined with oral delivery of rapamycin, was developed. Results: The protocol effectively prevented formation of inhibitors to FVIII upon subsequent intravenous treatment (weekly for 3.5 months). Control mice formed high‐titer inhibitors and had CD4⁺ T effector cell responses characterized by expression of IL‐2, IL‐4 and IL‐6. Tolerized mice instead had a CD4⁺CD25⁺FoxP3⁺ T cell response to FVIII that suppressed antibody formation upon adoptive transfer, indicating a shift from Th2 to Treg if FVIII antigen was introduced to T cells during inhibition with rapamycin. CD4⁺ T cells from tolerized mice also expressed TGF‐β1 and CTLA4, but not IL‐10. The presence of FVIII antigen during the time of rapamycin administration was required for specific tolerance induction. Conclusions: The study shows that a prophylactic immune tolerance protocol for FVIII can be developed using rapamycin, a drug that is already widely in clinical application. Immune suppression with rapamycin was mild and highly transient, as the mice regained immune competence within a few weeks.     

Induction of factor VIII‐specific unresponsiveness by intrathymic factor VIII injection in murine hemophilia A

Summary. Background: Hemophilia A is a congenital bleeding disorder caused by a deficiency of coagulation factor VIII. Approximately 30% of hemophilia A patients develop inhibitors against FVIII following replacement therapy. We have reported that neonatal exposure of FVIII antigen can induce antigen‐specific immune tolerance by interferon‐γ (IFN‐γ)‐dependent T‐cell anergy in hemophilia A mice. Objective: The thymus plays crucial roles in self‐tolerance, with negative selection of self‐reactive effector T cells and positive selection of self‐reactive regulatory T cells. We investigated the possibility of the induction of antigen‐specific immune tolerance by intrathymic injection of FVIII in hemophilia A mice. Methods: Hemophilia A mice were injected with recombinant FVIII into the thymus under real‐time high‐resolution image guidance. Results: Anti‐FVIII inhibitory antibody titers in mice challenged with intravenous administration of FVIII were significantly lower in mice (n = 22) that had received thymic FVIII injection than in mice (n = 18) without thymic injection (9.4 ± 2.3 vs. 122.5 ± 27.6 BU mL^?1, respectively, P = 0.00078). The CD4⁺ T cells from thymic‐injected mice could not proliferate or produce interleukin (IL)‐2, IL‐12 and IFN‐γ in response to FVIII. The CD4⁺CD25⁺ T cells generated from thymic‐treated mice but not from naïve mice efficiently suppressed the in vitro proliferative response of CD4⁺ T cells and blocked the in vivo development of anti‐FVIII antibodies in the adoptive transfer. Conclusion: These data suggest that intrathymic administration of FVIII could result in immune tolerance by induction of FVIII‐specific regulatory T cells.     

Induction of partial immune tolerance to factor VIII through prior mucosal exposure to the factor VIII C2 domain

Summary. Background: The development of anti‐factor VIII (FVIII) neutralizing antibodies (inhibitors) is a significant obstacle to FVIII replacement therapy. Objective: As mucosal administration of an antigen may induce immune tolerance we have evaluated the efficacy of mucosal antigen exposure to achieve tolerance to FVIII. Methods: We investigated the effects of oral and nasal administration of the purified FVIII C2 domain (FVIII‐C2) to FVIII‐deficient BALB/c mice prior to FVIII protein challenge. Mice received oral or nasal doses of FVIII‐C2, followed by a subcutaneous challenge of either FVIII‐C2 or FVIII. The development of anti‐FVIII inhibitors, cytokine production by splenocytes in vitro, and adoptive transfer assays were analyzed. Results and Conclusions: Mucosal administration of FVIII‐C2 decreases the titer of anti‐FVIII‐C2 inhibitors after FVIII‐C2 challenge, and decreases the percentage of FVIII‐C2 specific antibodies after challenge with full‐length FVIII. Tolerance induction to FVIII‐C2 is associated with increased IL‐10 production by splenocytes in vitro, and can be adoptively transferred to naïve mice. This study is the first to demonstrate that tolerance to the FVIII‐C2 domain can be induced via the mucosal route. Based on these results, the potential use of FVIII‐specific mucosal tolerance induction as an immunotherapy treatment for anti‐FVIII inhibitor development warrants further investigation.     

T‐cell responses in two unrelated hemophilia A inhibitor subjects include an epitope at the factor VIII R593C missense site

Summary. Background: Development of neutralizing anti‐factor (F)VIII antibodies (‘inhibitors’) is a serious clinical problem in hemophilia A. Increased inhibitor risk has been associated with certain FVIII missense substitutions, including R593C in the A2 domain. Objectives: The aim of the present study was to identify T‐cell epitopes in FVIII and characterize T‐cell responses in two unrelated hemophilia A subjects sharing F8‐R593C and HLA‐DRB1*1101 genotypes. We hypothesized that the hemophilic substitution site coincides with an important T‐cell epitope. Patients/methods: The binding affinities of peptides for recombinant HLA‐DR proteins were measured and compared with epitope prediction results. CD4+ T cells were stimulated using peptides and stained with fluorescent, peptide‐loaded tetramers. Results: The inhibitor subjects, but not HLA‐matched controls, had high‐avidity HLA‐DRB1*1101‐restricted T‐cell responses against FVIII_589–608, which contains the hemophilic missense site. Antigen‐specific T cells secreted Th1 and Th2 cytokines and proliferated in response to FVIII and FVIII_592–603. FVIII_589–608 bound with physiologically relevant (micromolar) IC₅₀ values to recombinant DR0101, DR1101 and DR1501 proteins. Conclusions: Hemophilia A patients with R593C missense substitutions and these HLA haplotypes had an increased incidence of inhibitors in our cohorts, supporting a paradigm in which presentation of FVIII epitopes containing the wild‐type R593 influences inhibitor risk in this hemophilia A sub‐population.     

Immune tolerance induction for the eradication of inhibitors in patients with hemophilia A

Background: Both genetic and environmental factors contribute to the formation of alloantibodies that bind to functional domains on the Factor VIII (FVIII) molecule and inhibit its function. Patients with hemophilia A who develop high-titer inhibitors are at increased risk for serious hemorrhage and disability, particularly arthropathy, because bleeding events do not respond to standard therapy. Immune tolerance induction (ITI) is usually attempted to eradicate newly diagnosed inhibitors, restore replacement FVIII pharmacokinetics, and improve bleed management and quality of life. Objective: This paper summarizes regimens used for ITI, predictors of success and failure, and adjunctive therapies for patients failing ITI therapy. Methods: This is a systematic review of published reports on ITI regimens, data from registries capturing response rates and predictors of success, and reports of adjunctive treatments used to enhance ITI therapy. Results/conclusion: Many issues remain unresolved, chief among them optimal dose and dosing regimen, choice of FVIII product, and the role of adjunctive therapy. Resolution of these issues, as well as new approaches to inhibitor management, may come from ongoing basic science research and clinical trials.     

B cell–activating factor modulates the factor VIII immune response in hemophilia A

Inhibitors of factor VIII (FVIII) remain the most challenging complication of FVIII protein replacement therapy in hemophilia A (HA). Understanding the mechanisms that guide FVIII-specific B cell development could help identify therapeutic targets. The B cell–activating factor (BAFF) cytokine family is a key regulator of B cell differentiation in normal homeostasis and immune disorders. Thus, we used patient samples and mouse models to investigate the potential role of BAFF in modulating FVIII inhibitors. BAFF levels were elevated in pediatric and adult HA inhibitor patients and decreased to levels similar to those of noninhibitor controls after successful immune tolerance induction (ITI). Moreover, elevations in BAFF levels were seen in patients who failed to achieve FVIII tolerance with anti-CD20 antibody–mediated B cell depletion. In naive HA mice, prophylactic anti-BAFF antibody therapy prior to FVIII immunization prevented inhibitor formation and this tolerance was maintained despite FVIII exposure after immune reconstitution. In preimmunized HA mice, combination therapy with anti-CD20 and anti-BAFF antibodies dramatically reduced FVIII inhibitors via inhibition of FVIII-specific plasma cells. Our data suggest that BAFF may regulate the generation and maintenance of FVIII inhibitors and/or anti-FVIII B cells. Finally, anti-CD20/anti-BAFF combination therapy may be clinically useful for ITI.     

Bortezomib delays the onset of factor VIII inhibitors in experimental hemophilia A,but fails to eliminate established anti‐factor VIII IgG‐producing cells

Summary. Background: Replacement therapy with exogenous factor VIII to treat hemorrhages induces inhibitory anti‐FVIII antibodies in up to 30% of patients with hemophilia A. Current approaches to eradicate FVIII inhibitors using high‐dose FVIII injection protocols (immune tolerance induction) or anti‐CD20 depleting antibodies (Rituximab) demonstrate limited efficacy; they are extremely expensive and/or require stringent compliance from the patients. Objectives: To investigate whether the proteasome inhibitor bortezomib, which depletes plasmocytes, modulates the anti‐FVIII immune response in FVIII‐deficient mice. Methods and results: Preventive 4‐week treatment of naïve mice with bortezomib at the time of FVIII administration delayed the development of inhibitory anti‐FVIII IgG, and depleted plasma cells as well as different lymphoid cell subsets. Conversely, curative treatment of inhibitor‐positive mice for 10 weeks, along with FVIII administration, failed to eradicate FVIII inhibitors to extents that would be clinically relevant if achieved in patients. Accordingly, bortezomib did not eradicate anti‐FVIII IgG‐secreting plasmocytes that had homed to survival niches in the bone marrow, despite significant elimination of total plasma cells. Conclusions: The data suggest that strategies for the efficient reduction of anti‐FVIII IgG titers in patients with hemophilia A should rely on competition for survival niches for plasmocytes in the bone marrow rather than the mere use of proteasome inhibitors.     

Factor VIII gene (F8) mutations as predictors of outcome in immune tolerance induction of hemophilia A patients with high-responding inhibitors

Summary. Background: Immune tolerance induction (ITI) is the only therapeutic approach that can eradicate factor VIII (FVIII) inhibitors in patients with hemophilia A. Predictors of ITI outcome are still debated, and the role of F8 gene mutations in this is not well established. Objectives: To investigate the relationship between F8 genotype and ITI outcome in patients with severe hemophilia A and high‐responding inhibitors. Patients and Methods:F8 mutations were identified in 86 patients recruited as part of the Italian ITI registry (the PROFIT study). ITI outcome was centrally reviewed according to the following definitions: success (undetectable inhibitor and normal FVIII pharmacokinetics), partial success (inhibitor titer < 5 BU mL^?1 and/or abnormal FVIII pharmacokinetics), and failure. Results:F8 mutations known to be associated with a high risk of inhibitor development (large deletions, inversions, nonsense mutations and splice site mutations) were found in 70 patients (81%); among these, the intron 22 inversion was present in 49 patients (57%). In 16 patients (19%) lower‐risk F8 defects (small insertions/deletions and missense mutations) were identified. The latter group of patients showed a significantly higher ITI success rate than those carrying high‐risk mutations [13/16 (81%) vs. 33/70 (47%); risk ratio 1.7, 95% confidence interval (CI) 1.1–2.1, P = 0.01]. On multivariate analysis, the mutation risk class remained a significant predictor of success [adjusted odds ratio (OR) 6.2, 95% CI 1.1–36.0, P = 0.04], as were inhibitor titer at ITI start (< 5 BU mL^?1, OR 11.8, 95% CI 3.5–40.2, P < 0.001), and peak titer during ITI (< 100 BU mL^?1, OR 11.4, 95% CI 3.2–40.8, P < 0.001). Conclusions: ITI success is influenced by F8 genotype. This knowledge should contribute to the stratification of prognosis, and to the clinical choices made for ITI in patients with high‐responding inhibitors.     

Surgery and inhibitor development in hemophilia A: a systematic review

Summary. Background: Although the association between intensive treatment and the formation of inhibiting antibodies towards factor VIII (FVIII) in hemophilia A has been demonstrated, the contributing effect of surgery is presently unclear. The release of immunological danger signals resulting from tissue damage during surgery in the presence of a high FVIII antigen load may elicit the formation of FVIII antibodies. The aim of this systematic review was to investigate the role of surgery in the inhibitor risk associated with intensive treatment as compared with treatment for bleeding and prophylactic administration of FVIII. Methods: A comprehensive literature search was performed that identified four cohort studies and three case control studies, comprising 342 inhibitor patients among a total of 957 hemophilia A patients. Results: Intensive treatment increased the inhibitor risk, most pronounced with intensive treatment of ≥ 5 exposure days (EDs) compared with < 3 EDs (OR, 4.1; 95% confidence interval, 2.6–6.5). Pooled odds ratio for inhibitor development in severe hemophilia patients that received intensive treatment for surgery at first exposure was 4.1 (95% confidence interval, 2.0–8.4) compared with treatment for bleeding or prophylaxis. Information on continuous infusion, previously treated patients and non‐severe hemophilia A was insufficient for valid meta‐analyses. Conclusions: Intensive FVIII treatment for surgery at first exposure leads to a higher inhibitor risk in hemophilia A patients compared with intensive treatment for bleeding.     

Suppression of FVIII Inhibitor Formation in Hemophilic Mice by Delivery of Transgene Modified Apoptotic Fibroblasts

The development of inhibitory antibodies to factor VIII (FVIII) is currently the most significant complication of FVIII replacement therapy in the management of patients with severe hemophilia A. Immune tolerance protocols for the eradication of inhibitors require daily delivery of intravenous FVIII for at least 6 months and are unsuccessful in 20–40% of treated patients. We hypothesize that tolerance can be induced more efficiently and reliably by delivery of FVIII antigen within autologous apoptotic cells (ACs). In this study, we demonstrated suppression of the T cell and inhibitor responses to FVIII by infusion of FVIII expression vector modified apoptotic syngeneic fibroblasts in both naive and preimmunized hemophilia A mice. ACs without FVIII antigen exerted modest generalized immune suppression mediated by anti-inflammatory signals. However, FVIII expressing apoptotic syngeneic fibroblasts produced much stronger antigen-specific immune suppression. Mice treated with these fibroblasts generated CD4⁺ T cells that suppressed the immune response to FVIII after adoptive transfer into naive recipients and antigen-specific CD4⁺CD25⁺ regulatory T cells (Tregs) that inhibited the proliferation of FVIII responsive effector T cells in vitro. These preclinical results demonstrate the potential for using FVIII vector modified autologous ACs to treat high-titer inhibitors in patients with hemophilia A.     

Repeated oral administration of chitosan/DNA nanoparticles delivers functional FVIII with the absence of antibodies in hemophilia A mice

Summary. Background: Current treatment of hemophilia A is expensive and involves regular infusions of factor (F)VIII concentrates. The supply of functional FVIII is further compromised by the generation of neutralizing antibodies. Thus, the development of an alternative safe, cost effective, non‐invasive treatment that circumvents immune response induction is desirable. Objectives: To evaluate the feasibility of oral administration of chitosan nanoparticles containing FVIII DNA to provide sustainable FVIII activity in hemophilia A mice. Methods: Nanoparticles were characterized for morphology, DNA protection and transfection efficiency. Oral administration of nanoparticles containing canine FVIII in C57Bl/6 FVIII^?/? hemophilia A mice was evaluated for biodistribution, plasma FVIII activity and phenotypic correction. Sustainable FVIII expression was elucidated after repeated nanoparticle administration. Immune responses to repeated oral nanoparticle administration were also investigated. Results: Chitosan nanoparticles had a particle size range of 200–400 nm and protected DNA from endonuclease and pH degradation. In addition, nanoparticles transfected HEK 293 cells resulted in expression of eGFP, luciferase and FVIII. Hemophilia A mice that ingested chitosan nanoparticles demonstrated transient canine FVIII expression reaching > 100 mU 1 day after treatment, together with partial phenotypic correction. The delivered FVIII plasmid DNA was detected in the intestine and, to a lesser extent, in the liver. Importantly, repeated weekly administrations restored FVIII activity. Furthermore, inhibitors and non‐neutralizing FVIII antibodies were not detectable. Conclusions: Repeat oral administration of FVIII DNA formulated in chitosan nanoparticles resulted in sustained FVIII activity in hemophilic mice, and thus may provide a non‐invasive alternative treatment for hemophilia A.     

Immune tolerance therapy for factor VIII inhibitors: moving from empiricism to an evidence-based approach

Summary.  Currently, the only proven strategy for achieving antigen-specific tolerance to factor VIII (FVIII) is immune tolerance induction (ITI) therapy. This paper discusses our current knowledge of the host and treatment factors, as well as supportive care initiatives, known or suspected to influence the outcome of ITI in the treatment of inhibitors arising in patients with severe hemophilia A. Among these, questions surrounding the choice of therapeutic product and/or dosing regimen generate the most controversy, given the lack of a definitive evidence-based approach to either. Furthermore, the potential for central venous access device (CVAD) and intercurrent bleeding complications to impact the ultimate success of ITI remains unclear. The ongoing clinical trials designed to further clarify several of these polarizing issues are reviewed. This paper also explores the current and future role of immune modulation in possible salvage, ancillary or primary alternative tolerance induction strategies. The special cases of low titer/ responding inhibitors and inhibitors developing in mild hemophilia A patients are considered. Finally, this paper summarizes the currently recommended approach to ITI and makes the case for a move from empiric therapeutics to a risk-stratified evidence-based approach to FVIII inhibitor eradication.     

In non‐severe hemophilia A the risk of inhibitor after intensive factor treatment is greater in older patients: a case–control study

Summary. Background: Twenty‐five percent of new anti‐factor VIII (FVIII) antibodies (inhibitors) that complicate hemophilia A occur in those with mild and moderate disease. Although intensive FVIII treatment has long been considered a risk factor for inhibitor development in those with non‐severe disease, its strength of association and the influence of other factors have remained undefined. Objective: To evaluate risk factors for inhibitor development in patients with non‐severe hemophilia A. Methods: Information on clinical and demographic variables and FVIII genotype was collected on 36 subjects with mild or moderate hemophilia A and an inhibitor and 62 controls also with mild or moderate hemophilia A but without an inhibitor. Results: Treatment with FVIII for six or more consecutive days during the prior year was more strongly associated with inhibitor development in those ≥ 30 years of age compared with those < 30 years of age [adjusted odds ratio (OR) 12.62; 95% confidence interval (CI), 2.76–57.81 vs. OR 2.54; 95% CI, 0.61–10.68]. Having previously received < 50 days of FVIII was also not statistically associated with inhibitor development on univariate or multivariate analysis. Conclusions: These findings suggest that inhibitor development in mild and moderate hemophilia A varies with age, but does not vary significantly with lifetime FVIII exposure days: two features distinct from severe hemophilia A.     

Most factor VIII B domain missense mutations are unlikely to be causative mutations for severe hemophilia A: implications for genotyping

Summary. Background & Objective: The factor VIII (FVIII) B domain shares very little amino acid homology with other known proteins and is not directly necessary for procoagulant activity. Despite this, missense mutations within the B domain have been reported in patients with hemophilia A. Given that the B domain is dispensable for secretion and function of FVIII, we hypothesized that these mutations should not be causative of hemophilia A in these patients. Methods: Plasmid vectors containing B domain missense mutations that were reported to be associated with moderate/severe hemophilia A (T751S, D826E, V993L, H1047Y, T1353A, N1441K, L1462P, E1579D, A1591S, P1641L and S1669L) were analyzed for their effect on synthesis and secretion compared with FVIII wild‐type (WT) following transient transfection into COS‐1 and CHO cells in vitro. Further, H1047Y, N1441K and E1579D mutants were expressed in vivo in a hemophilia A mouse model by hydrodynamic tail‐vein injection. Results: FVIII activity and antigen levels for all mutants expressed into the conditioned media of COS‐1 and CHO cells were similar to FVIII WT. Also, plasma expression of these mutants was similar to FVIII WT in hemophilia A mice. An in vivo tail clip bleeding assay also demonstrated that blood loss from hemophilia A mice expressing FVIII WT, H1047Y, N1441K and E1579D was similar. Conclusions: We conclude that most missense mutations within the FVIII B domain would be unlikely to lead to severe hemophilia A and that the majority of such missense mutations represent polymorphisms or non‐pathologic mutations.     

Reduction of the immune response to factor VIII mediated through tolerogenic factor VIII presentation by immature dendritic cells

Summary. Background: The development of neutralizing antibodies to factor FVIII (FVIII) represents the most serious complication in the treatment of hemophilia A. Objective: We have explored the potential of using immature dendritic cells (iDCs) to present FVIII in a tolerogenic manner to T cells. Methods: The iDCs were isolated from hemophilic murine bone marrow and pulsed with canine cFVIII (cFVIII‐iDCs) in the presence or absence of the NFκB pathway blocking compound Andrographolide (Andro‐cFVIII‐iDCs). Three weekly intravenous infusions of one million cFVIII pulsed‐iDCs were administered to a group of five hemophilic Balb/c mice. Anti‐FVIII antibody levels were monitored by functional Bethesda assay after four weekly intravenous challenges with 2 IU of cFVIII. Results: We have shown that cFVIII in the presence or absence of Andro is efficiently taken up by iDCs and that this process does not result in the maturation of DCs or the activation of co‐cultured T cells. Following repeated infusion of the cFVIII‐iDCs and Andro‐cFVIII‐iDCs into hemophilic mice, which were subsequently challenged with cFVIII, long‐term reductions of FVIII inhibitors of 25% and 40%, respectively, were documented. Studies of cytokine release and T‐cell phenotypes indicate that the mechanisms responsible for reducing immunologic responsiveness to cFVIII appear to involve an expansion of Foxp3 T regulatory cells in the case of cFVIII‐iDC infusion and the elaboration of the immunosuppressive cytokines IL‐10 and TGF‐β following andrographolide‐treated cFVIII‐iDCs. Conclusions: This study shows that tolerogenic presentation of cFVIII to the immune system can significantly reduce immunogenicity of the protein.     

Monitoring the bioavailability of FEIBA with a thrombin generation assay

Summary. Background: Hemophilia A patients with inhibitors are generally treated with preparations containing activated coagulation factors to achieve hemostasis by bypassing factor (F)VIII. Objectives: We developed an assay for monitoring the kinetic of thrombin generation in human FVIII inhibitor plasma reconstituted in vitro with activated prothrombin complex concentrate, FEIBA, and in plasma samples from hemophilia A patients taken after FEIBA treatment. Patients and methods: For pharmacokinetic studies three patients with severe hemophilia A and with a high‐titer inhibitor received a single dose of FEIBA. Repeated FEIBA treatment was monitored in one patient with acquired hemophilia A. Coagulation was triggered in citrated plasma by adding a low concentration of tissue factor/phospholipid complex and CaCl₂ in the presence of a fluorogenic thrombin substrate. The intensity of the fluorescence signal (FU) was continuously monitored, and the rate of increase in the fluorescence signal for every time point, which reflects the actual thrombin concentrations, was calculated. Results: The maximum rate of substrate conversion, which indicates the highest thrombin concentration, was approximately 1900 FU min^?1 in a normal plasma pool. Practically no thrombin generation was observed in the FVIII inhibitor plasma, but when it was spiked with FEIBA, the rate and the peak of thrombin generation increased dose‐dependently to close to normal. Plasma samples from FVIII inhibitor patients treated with a single dose of FEIBA had an improved thrombin maximum within an hour after treatment, which gradually returned to baseline values with a half‐life of 4–7 h. Changes in the characteristic parameters of thrombin generation coincided with the repeated administration of FEIBA in a patient with acquired hemophilia A. Conclusions: This assay enables the pharmacodynamic and pharmacokinetic properties of bypassing therapies to be monitored, thus helping to optimize treatment.     

血友病A伴抑制物一例及其免疫耐受诱导治疗探讨

目的 探讨血友病A伴抑制物的免疫耐受诱导(immune tolerance induction,ITI)治疗,提高血友病A伴抑制物患者的诊疗水平.方法 对重型血友病A患者用APTT标准曲线一期法测定凝血因子Ⅷ(FⅧ)活性(FⅧ∶C);用Bethesda法定量测定FⅧ抗体;用长距离PCR方法检测内含子22倒位.结果 经检测发现患者为FⅧ基因22内含子倒位;ITI治疗3个月后,患者FⅧ抑制物滴度下降为0,输注FⅧ后回收率＞66%,治疗6个月后达到抑制物清除标准,停止治疗.结论 该患者是国内首例采用ITI成功治疗血友病A伴抑制物的病例,ITI是目前最有希望根除血友病抑制物的方法.     

Immune tolerance induction with high-dose FVIII and pulsed intravenous immunoglobulin

The development of neutralizing allo-antibodies against factor VIII (FVIII) or FVIII inhibitors is a severe complication in the treatment of haemophilia A. About 25% of the children with severe haemophilia A develop FVIII inhibitors. Here we report on a boy with severe haemophilia A and intron 22 inversion of the FVIII gene who was diagnosed at ten months of age. After 16 exposure days to FVIII (81 days after initial exposure) he developed a FVIII inhibitor (maximum: 9.76 BU/ml). Therapy: We started immune tolerance induction (ITI) according to the Bonn protocol with high dose plasma derived FVIII concentrate (100 IU per kg body weight) twice daily. For additional inhibitor elimination treatment the patient received intravenous immunoglobulin (ivIg) at a dose of 1-2 g/kg body weight every 4 to 6 weeks. After start of treatment a rapid decline of the inhibitor level was observed, nevertheless low FVIII inhibitor levels persisted (<5 BU/ml). Furthermore, the FVIII half-life was still accelerated. However, after every course of ivIg the inhibitor level declined and FVIII half-life was prolonged. Currently, the FVIII half-life is approaching normal values after more than seven months of ITI duration. Conclusion: Additional application of immunoglobulin is beneficial for immune tolerance induction.