Plasma-derived FVIII Products Research Articles

Ethical Decision Making Framework in Previously Untreated Severe Hemophilia A Patients

Treatment recommendations and decision making for severe hemophilia A previously untreated patients (PUPs) have recently become a significant debate with an ethical valence. Primary prophylaxis, initiation of factor replacement prior to the second joint bleed and before the age of three, has been the recommended standard of care in resource rich countries for the last two decades. Since the widespread availability of recombinant factor VIII (rFVIII) products, the choice has been fairly straightforward for PUPs, whereby transmission of blood borne pathogens associated with plasma-derived FVIII (pdFVIII) products were avoided. The Survey of Inhibitors in Plasma-Products Exposed Toddlers (SIPPET) randomized controlled trial demonstrated a higher incidence of FVIII alloantibody inhibitors in PUPs started on rFVIII products compared to pdFVIII products (Peyvandi F, et al., 2016). This finding has had a tremendous controversial impact on the decision making of product choice for PUPs despite the supportive Medical and Scientific Advisory Council recommendations to consider starting a PUP on a pdFVIII product (MASAC, 2016).For this patient population, we developed a modified utilitarian framework of decision making that employs clinical, public health, and research ethics. The framework recognizes that there are many stakeholders involved in the provision of FVIII products and health care of those with severe hemophilia A and provides guidance when there is concern for bias. Utilizing the premise of shared decision making, the framework takes a patient and family centered rather than a paternalistic approach. In discussions about care, it is recommended that the provider learn about relevant patient and family values. Doing so would better enable providers to inform the patient and family about the available treatments (including the risks, benefits and alternatives) and disclose any conflicts of interest and biases. Using shared decision making would serve to maximize understanding, minimize bias, respect informed consent or dissent, and provide care that aligns with patient values when medically and practically feasible.The framework has three tiers. First, it evaluates whether resources are scarce or abundant, which is important in equitably allocating resources. Where life-saving FVIII products are scarce, there could be a direct impact on patient care and clinical research. Healthcare providers would be at risk of compromising not only standard of care but also their fiduciary responsibilities to patient care in bedside rationing. If FVIII products are scarce, we recommend developing a central supply for emergency use and then evaluating the needs of the severe hemophilia A patients. Prioritization of who receives the factor products would be decided by a designated team based on the availability of the factor products and clinical scenarios, with no preference given to those on research trials. However, if resources are abundant, treatment for acute bleeding and standard of care prophylaxis measures, including primary prophylaxis, could continue.The second tier takes into account whether there is a new infectious epidemic or concern where the pathogen cannot be eliminated. If not, pdFVIII and rFVIII products are to be equally presented. If there is an epidemic, healthcare and public health workers may limit the use of pdFVIII products.The third tier evaluates the clinical scenario and if there is an acute bleeding emergency or not. If there is an acute bleeding event, the immediately available resource should be recommended, along with bypassing and/or adjuvant resources as needed until the bleeding has resolved or improved. To align with patient and family preferences, attempts to have both pdFVIII and rFVIII products available at similar costs in institutions would be ideal.The ethical framework we developed endeavors to balance autonomy, beneficence, nonmaleficence and justice. As treatment decisions are complex, the framework may help guide discussions among providers, PUPs with severe hemophilia A, and their families.Disclaimer:The findings and conclusions in this abstract are those of the author(s) and do not necessarily represent the official position of the Centers for Disease Control and Prevention, Emory University, or Children's Healthcare of Atlanta. DisclosuresSidonio:Bioverativ: Consultancy; CSL Behring: Consultancy; Grifols: Research Funding; Bioverativ: Research Funding; Shire: Consultancy, Research Funding; Novo Nordisk: Consultancy; Bayer: Consultancy.

Immune Responses to Plasma-Derived Versus Recombinant FVIII Products.

The most severe side effect of hemophilia treatment is the inhibitor development occurring in 30% of patients, during the earliest stages of treatment with factor (F)VIII concentrates. These catastrophic immune responses rapidly inactivate the infused FVIII, rendering the treatment ineffective. This complication is associated with a substantial morbidity and mortality. The risk factors involved in the onset of the inhibitors are both genetic and environmental. The source of FVIII products, i.e. plasma-derived or recombinant FVIII products, is considered one of the most relevant factors for inhibitor development. Numerous studies in the literature report conflicting data on the different immunogenicity of the products. The SIPPET randomized trial showed an increased in the inhibitor rate in patients using recombinant FVIII products than those receiving plasma-derived products in the first exposure days. The SIPPET randomized trial showed an increase in the inhibitor rate in patients using recombinant FVIII products compared to those treated with plasma-derived products in the first days of exposure. The potential increase in the immunogenicity of recombinant products can be attributed to several factors such as: the different post-translational modification in different cell lines, the presence of protein aggregates, and the role played by the chaperon protein of FVIII, the von Willebrand factor, which modulates the uptake of FVIII by antigen presenting cells (APCs). Furthermore, the presence of non-neutralizing antibodies against FVIII has shown to be in increased inhibitor development as demonstrated in a sub-analysis of the SIPPET study. In addition, the presence of the specific subclasses of the immunoglobulins may also be an important biomarker to indicate whether the inhibitor will evolve into a persistent neutralizing antibody or a transient one that would disappear without any specific treatment. Recently, the availability of novel non-replacement therapies as well as emicizumab, administered by weekly subcutaneous infusion, have significantly changed the quality of life of patients with inhibitors showing a considerable reduction of the annual bleeding rate and in most patients the absence of bleeding. Although, these novel drugs improve patients' quality of life, they do not abolish the need to infuse FVIII during acute bleeding or surgery. Therefore, the issue of immunogenicity against FVIII still remains an important side effect of hemophilia treatment.

Acquired Haemophilia A: An Intriguing Disease.

. Acquired Haemophilia A is a rare acquired bleeding disorder caused by Factor VIII autoantibodies, which neutralise FVIII activity. These inhibitors differ from alloantibodies against FVIII, which can occur in congenital Haemophilia A after repeated exposures to plasma-derived or recombinant FVIII products. In most cases, the disease occurs suddenly in subjects without a personal or familiar history of bleedings, with symptoms that may be mild, moderate, or severe. However, only laboratory alterations are present in ~ 30% of patients. The incidence varies from 1 to 4 cases per million/year; more than 80% of patients are elderly, males and females are similarly affected. There is a small peak of incidence related to pregnancy in young women aged 20–40 years. The disease may be underdiagnosed in the elderly. The diagnostic algorithm is based on an isolated prolonged activated partial thromboplastin time, normal thrombin time, absence of Lupus Anticoagulant, and a mixing test that reveals the presence of an inhibitor: the finding of reduced FVIII activity and the detection of neutralising autoantibodies against FVIII lead to the diagnosis. The disease is idiopathic in 44%–63% of cases, while in the others etiological factors are present. Bleeding prevention and treatment are based on therapeutic tools as by-passing agents, recombinant porcine FVIII concentrate or, in a limited number of cases, FVIII concentrates and desmopressin. As soon as the diagnosis has been made, immunosuppressive therapy must be started to eradicate the inhibitor. Better knowledge of the disease, optimal management of bleeding and eradication of the inhibitor have significantly reduced morbidity and mortality in most patients.

Status of Recombinant Factor VIII Concentrate Treatment for Hemophilia a in Italy: Characteristics and Clinical Benefits.

The current interest in recombinant factor VIII (rFVIII) products stems from the fact that they offer a technological solution to prolonging the half-life of and reducing the risk of formation of alloantibodies (inhibitors) against FVIII in treated patients with hemophilia A (HA). The Italian health care system has authorized the use of a wide range of rFVIII concentrates of the first, second, and third generation, as well as new innovative rFVIII preparates with an extended half-life (EHL) (Kogenate FS®-Bayer, belonging to the second generation and replaced since 2017 by a product consisting of the same modified molecule; because it is only available until the end of the current year, it will not be considered in this review). Some of these products have unique pharmacodynamic and pharmacokinetic (PK) profiles, including an EHL. The first-generation full-length rFVIII (FL-rFVIII), octocog alfa (Recombinate® Baxter/BIOVIIIx), although the oldest rFVIII product, has several desirable features. Third-generation products include two modified octocog alfa molecules (Advate®, Shire; Kovaltry®, Bayer) as well as the B domain-deleted rFVIII (BDD-rFVIII) moroctocog alfa (ReFacto®-Pfizer). The B domain-truncated (BDT-rFVIII) turoctocog alfa (NovoEight®, Novo Nordisk), the BDD-rFVIII simoctocog alfa (Nuwiq®, Kedrion), the single-chain BDT-rVIII lonoctocog alfa (Afstyla®, CSL Behring), and the BDD-rFVIIIFc efmoroctocog alfa (Elocta®, Sobi-Biogen) are new, innovative products. Simoctocog alfa, because its peculiarities, is considered a fourth-generation rFVIII concentrate. Turoctocog alfa, simoctocog alfa, and lonoctocog alfa have a high affinity for von Willebrand factor (vWF) that reduces renal clearance and prolongs the half-life of rFVIII. Efmoroctocog alfa, a first-in-class rFVIII-Fc fusion protein (rFVIIIFc), has a half-life 1.5–1.8 times longer than that of conventional plasma-derived FVIII (pd-rFVIII) and other rFVIII products. Clinical studies have evaluated the efficacy, safety, and inhibitor development of all these innovative concentrates in both previously treated (PTPs) and untreated patients (PUPs). This review considers the rFVIII products that are indicated for the treatment of patients with severe HA, focusing on those that are commercially available in Italy. Their PK characteristics, immunogenicity, and clinical benefits are discussed and compared.

Inhibitor Development with Simoctocog Alfa in Previously Untreated Patients with Severe Haemophilia a: Final Results of the Nuprotect Study

Introduction FVIII inhibitor development is the greatest challenge when treating previously untreated patients (PUPs) with hemophilia A (HA). The SIPPET study reported a cumulative inhibitor incidence of 44.5% (28.4% high-titre) in PUPs treated with recombinant FVIII (rFVIII) products produced in hamster cell lines and 26.8% (18.6% high-titre) with plasma-derived FVIII products containing von Willebrand factor (pdFVIII/VWF). Simoctocog alfa (Nuwiq®) is a 4th generation rFVIII produced in a human cell line without chemical modification or protein fusion. The NuProtect study assessed the immunogenicity, efficacy and safety of simoctocog alfa in PUPs with severe HA. A prespecified interim analysis was published with data up to 20 and 50 EDs (Haemophilia 2018; 24:211) and here we report the final results. Methods NuProtect was a prospective, multinational, open-label, non-controlled, phase III study. True PUPs (no prior FVIII treatment) with severe HA of any age and ethnicity were to be enrolled and treated for 100 exposure days (EDs) or a maximum of 5 years with simoctocog alfa for prophylaxis, on-demand treatment, treatment of breakthrough bleeding episodes (BEs) and surgical prophylaxis. Type of treatment and dose were determined by the investigator. Inhibitor screening (modified Bethesda assay) was performed at screening, every 3-4 EDs until ED20, then every 10-12 EDs or at least every 3 months, at completion, and if inhibitor development was suspected. Inhibitor levels of ≥0.6 to <5 Bethesda units [BU]/mL were defined as low-titer and ≥5 BU/mL as high-titer. Cumulative inhibitor incidence (primary endpoint) and 95% confidence intervals (CIs) were calculated (Kaplan-Meier). Efficacy endpoints (inhibitor-free periods) included the annualized bleeding rate (ABR) during prophylaxis and efficacy in treating BEs/surgical prophylaxis (4-point objective scales: excellent, good [successful]; moderate or none). Adverse events (AEs) were monitored throughout the study. Results Of 108 subjects consented, 105 PUPs, median age of 12 months (range 0-146) at ED1 were evaluable for inhibitor development. They were treated for a median of 101 EDs (range 1-1164), with 96 patients treated for ≥100 EDs (or until inhibitor development, including 5 patients with 97-99 EDs). The majority of patients with available genetic data had null F8 gene mutations (90/102 [88.2%]) and 13 (12.0%) had a family history of inhibitors. Cumulative inhibitor incidence was 17.6% (95% CI: 10.0%, 25.3%) for high-titre inhibitors and 27.9% (95% CI: 19.1%, 36.7%) for all inhibitors (Figure 1). No PUPs with non-null F8 mutations developed inhibitors. In 50 PUPs on continuous prophylaxis for ≥6 months, the mean (SD) ABR was 0.54 (1.07) [median 0] for spontaneous BEs and 3.61 (3.82) [median 2.53] for all BEs. The treatment of BEs was successful in 92.9% (747/804) of rated BEs in 85 patients with treated BEs and 91.9% of BEs were controlled with 1 or 2 infusions. Surgical prophylaxis was successful for 94.7% (18/19) of rated procedures and moderate for 5.3% (1/19). Excluding inhibitors, only one (0.9%) patient had an AE classified as serious by the investigator (hospitalization due to a mild rash that resolved with anti-histamine treatment). Conclusions Simoctocog alfa had a similar inhibitor incidence in PUPs with severe HA as pdFVIII/VWF products in SIPPET. No inhibitors occurred in PUPs with non-null F8 mutations. Simoctocog alfa had a median spontaneous ABR of 0 during prophylaxis and was successful in the treatment of 92.9% of BEs and in 94.7% of surgical procedures. These results complement results in previously treated patients (PTPs) and support the use of simoctocog alfa in the prevention and treatment of BEs in PUPs and PTPs. References Liesner R, et al. Haemophilia 2018; 24: 211-20. Disclosures Liesner: Octapharma, Bayer, Takeda, Novo Nordisk, CSL Behring, Roche: Research Funding; Octapharma, SOBI, Novo Nordisk: Speakers Bureau; Octapharma, Bayer, Takeda: Consultancy. Neufeld:Octapharma, Shire Pharmaceuticals (Baxalta), Novo Nordisk, Celgene, NHLBI/NIH: Research Funding; Octapharma, Agios, Acceleron, Grifols, Pfizer, CSL Behring, Shire Pharmaceuticals (Baxalta), Novo Nordisk, ApoPharma, Genentech, Novartis, Bayer Healthcare: Consultancy; Octapharma: Other: study investigator, NuProtect study (Octapharma-sponsored).

Immune tolerance induction in paediatric patients with haemophilia A and inhibitors receiving emicizumab prophylaxis

The formation of neutralizing antifactor VIII (fVIII) antibodies, called inhibitors, is the most common complication in modern haemophilia A care. Novel non-factor replacement therapies, such as emicizumab, have sought to address the limitations of bypassing agents for bleeding management in patients with inhibitors. However, immune tolerance induction (ITI) remains the primary method for eradicating inhibitors and restoring the hemostatic response to fVIII. The aim of this study was to review a case series of paediatric patients with haemophilia A and inhibitors who have received ITI for inhibitor eradication concomitantly with emicizumab prophylaxis for bleeding prevention. Single institution retrospective chart review of paediatric patients with severe haemophilia A receiving ITI and emicizumab. Seven patients were included in this cohort. Six patients used three different recombinant fVIII products at 100IU/kg three times per week, and one patient used a plasma-derived fVIII product at an initial dose of 50IU/kg three times per week. Three patients achieved a negative inhibitor titre <0.6 Chromogenic Bethesda Units per mL (CBU/mL), and two patients achieved a normal fVIII recovery ≥66%. There were nine bleeding events in four patients, but no thrombotic events. All patients remained on ITI and emicizumab. Immune tolerance induction while on emicizumab prophylaxis is a feasible approach in paediatric haemophilia A patients with inhibitors. This is the first report of the concomitant use of ITI in patients receiving emicizumab prophylaxis described as the 'Atlanta Protocol'.

Immunogenicity of Two Plasma-Derived FVIII Products and Simoctocog Alfa in Previously Untreated Patients According to F8 Mutation Type

Background: The SIPPET study investigated inhibitor development in 251 previously untreated patients (PUPs) treated with either plasma-derived FVIII products containing von Willebrand factor (pdFVIII/VWF; n = 125), or recombinant FVIII (rFVIII; n = 126) from hamster cell lines. Amongst PUPs with non-null F8 mutations, none developed inhibitors when treated with pdFVIII/VWF while the cumulative inhibitor incidence was 43% in those treated with hamster-cell-derived rFVIII. In patients with null F8 mutations, the cumulative inhibitor incidences were 31% and 47% in patients treated with pdFVIII/VWF and rFVIII, respectively. In patients with null mutations the cumulative incidences of high-titre inhibitors were 22% and 30% with pdFVIII/VWF and rFVIII, respectively.Aim: To investigate the relationship between inhibitor development and F8 mutation type in PUPs with severe hemophilia A treated with either a rFVIII from a human cell line (Nuwiq®; simoctocog alfa) or either of two pdFVIII/VWF products, one with a VWF/FVIII ratio of 0.4 (octanate®) the other with a VWF/FVIII ratio of 1.0 (wilate®).Materials and Methods: Data from completed multicenter, prospective trials with octanate® and wilate® and interim data from the NuProtect study with Nuwiq® were analyzed. Data on F8 mutation type were available for 50/51, 27/28 and 58/66 patients in each of the studies. All patients in the three studies had no previous treatment with FVIII concentrates or other blood products containing FVIII.Results: In the three studies, 18% (9/50), 7.4% (2/27) and 19%.0% (11/58) of patients had non-null mutations. None of the patients with non-null mutations developed inhibitors with octanate®, wilate® or Nuwiq®. In patients with null mutations, 9.8% (4/41), 12.0% (3/25), and 17.0% (8/47) developed high-titre inhibitors.Conclusions: PUPs with non-null F8 mutations did not develop inhibitors when treated with octanate®, wilate® or Nuwiq®. Whilst the different studies are not directly comparable, the findings with these products, two pdFVIII/VWF and a rFVIII from a human cell line, show similar behavior to the SIPPET trial where no patients with non-null mutations treated with pdFVIII/VWF products developed inhibitors. DisclosuresLiesner:Bayer: Consultancy, Research Funding; Baxalta: Consultancy, Research Funding; Novo Nordisk: Research Funding, Speakers Bureau; Roche: Research Funding; Sobi: Speakers Bureau; Octapharma: Consultancy, Other: Clinical study investigator for NuProtect Study (Octapharma sponsored), Research Funding, Speakers Bureau. Versteden:Octapharma AG: Employment. Lowndes:Octapharma AG: Employment. Belyanskaya:Octapharma AG: Employment. Oldenburg:Grifols: Honoraria, Research Funding; Novo Nordisk: Honoraria, Research Funding; Biotest: Honoraria, Research Funding; Biogen: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Roche: Honoraria, Research Funding; Octapharma: Honoraria, Research Funding; Shire: Honoraria, Research Funding; CSL Behring: Honoraria, Research Funding; Bayer: Consultancy, Honoraria, Research Funding; Chugai: Honoraria, Research Funding; Swedish Orphan Biovitrum: Honoraria, Research Funding. Pavlova:Novo Nordisk: Honoraria; Octapharma: Honoraria.

Potential role of a new PEGylated recombinant factor VIII for hemophilia A.

Hemophilia A, a deficiency in the activity of coagulation factor (F) VIII, is an X-linked bleeding disorder with an approximate incidence of one in 5,000 male infants. Bleeding-related complications often result in greater severity of disease, poor quality of life, surgical interventions for severe joint destruction, and shortened life span. With the availability of plasma-derived and recombinant FVIII products, the benefits of primary prophylaxis were demonstrated and is now the standard of care for patients with severe factor deficiencies. Current hemophilia research is focusing on the creation of new factor replacement therapies with longer half-lives; accessing alternative mechanisms to achieve desired hemostasis and enhance bypassing activity; and limiting the immunogenicity of the protein. PEGylation involves the covalent attachment of polyethylene glycol (PEG) to a protein, peptide, or a small molecule drug. PEG effectively increases the molecular weight and size of the protein by creating a hydrophilic cloud around the molecule. This molecular change may reduce susceptibility of the molecule to proteolytic activity and degradation. It is also believed that PEGylation changes the surface charge of the protein that ultimately interferes with some receptor-mediated clearance processes. The half-life of PEGylated factor is more prolonged when compared to non-PEGylated full-length recombinant FVIII. The dawn of a new era in the care of hemophilia patients is upon us with the release of recombinant FVIII products with extended half-lives, and products with even more extended half-life will become available in a very short time. With all the promise of these new agents, many questions still remain.

The pharmacokinetics of a B‐domain truncated recombinant factor VIII, turoctocog alfa (NovoEight®), in patients with hemophilia A

Turoctocog alfa (NovoEight(®)) is a human recombinant coagulation factor VIII (rFVIII) for the treatment of patients with hemophilia A. To evaluate the pharmacokinetics of turoctocog alfa in all age groups across clinical trials. Data from previously treated patients with severe hemophilia A (FVIII activity level of ≤ 1%) with no history of FVIII inhibitors, in a non-bleeding state, were included. The pharmacokinetics were assessed following a wash-out period and a subsequent single intravenous 50 IU kg(-1) dose of turoctocog alfa. Blood was sampled during a 48-h period postdose. Standard pharmacokinetic (PK) parameters were estimated on the basis of plasma FVIII activity vs. time (PK profiles) with non-compartmental methods. Furthermore, a population PK analysis was conducted. Data from 76 patients (aged 1-60 years) enrolled globally across six clinical trials were included, totaling 105 turoctocog alfa PK profiles. Single-dose PK results 3-6 months after the first dose of turoctocog alfa were comparable with the results obtained after the first dose. Similar PK characteristics were shown for different lots and strengths of the drug product. Overall, area under the plasma concentration (activity) curve from administration to infinity (AUC) and t1(/2) tended to increase with increasing age, with lower AUC and shorter t(1/2) being seen in children than in adolescents and adults. The PK profiles of turoctocog alfa and other commercially available plasma-derived FVIII and rFVIII products were similar in all age groups. The PK characteristics of turoctocog alfa have been thoroughly studied, and shown to be consistent over time, reproducible between different lots and strengths of drug product, and similar to those observed for other FVIII products.

Turoctocog alfa (recombinant factor VIII)

Turoctocog alfa (NovoEight®) is a new recombinant factor VIII (rFVIII) with a truncated B domain and a high degree of tyrosine sulphation, similar to plasma-derived FVIII products. The manufacturing process includes double nanofiltration with a 20-nm pore size and immunoaffinity chromatography with monoclonal F25 anti-FVIII antibodies. Treatment with turoctocog alfa can be monitored with both one-stage and chromogenic substrate assays without a product-specific laboratory standard. In total, 213 previously-treated patients with severe haemophilia A participated in the pivotal part of the clinical trial programme guardianTM. The median annualised bleeding rate during turoctocog alfa prophylaxis was 3.7 and 3.0 in adolescents/adults and children, respectively, with marked differences between participating countries. The success rate for the treatment of breakthrough bleeds was 85% (adults/adolescents) and 94% (children). A total of 41 surgical procedures (15 major, 26 minor) were performed in 33 patients, with a successful haemostatic response reported in all cases. No patient developed confirmed inhibitors in any of the trials.

The Rodin (Research Of Determinants of INhibitor Development among PUPs with haemophilia) study: the clinical conundrum from the perspective of haemophilia treaters

The Rodin study, recently published in the New England Journal of Medicine, has begun to provide some very important answers to several questions pertinent to the quality and safety of replacement therapy to individuals with haemophilia [1]. Probably the most serious complication of modern haemophilia care is currently the development of alloantibody neutralizing inhibitors and considerable recent literature until the Rodin trial had speculated that the recombinant factor (rFVIII) replacement products were more likely than the plasma-derived products to produce this. The Rodin trial has extended our thoughts on this issue because it is a robustly populated prospective observational study, in contrast to previously published results, which have been generated from smaller, retrospective, mostly uncontrolled heterogeneous population studies. Based on the results of 574 previously untreated severe haemophilia A patients (FVIII activity, <0.01 IU mL ), the observations in Rodin indicated that there was no difference in the incidence of alloantibody inhibitors whether patients received plasma-derived or recombinant full-length FVIII products (adjusted hazards ratio = 0.96). Furthermore, among those who developed alloantibody inhibitors while on plasma-derived FVIII concentrates, the content of von Willebrand factor (VWF) did not influence the risk of inhibitor formation (adjusted hazards ratio = 0.90). Lastly, the Rodin population study indicated that switching from a plasma-derived FVIII product (irrespective of relative VWF content) to a full-length rFVIII product did not increase the risk of inhibitor development. These conclusions are extremely cogent to patients and their physicians alike as the possibility that rFVIII products were more immunogenic, that the high content of VWF protein in plasma-derived FVIII products could protect against alloantibody formation, and that product switching would be harmful all were used as rationale to determine the choice of replacement product for previously untreated, slightly previously treated, and even those previously treated individuals with over 150 exposure days. This published report notwithstanding the possibility that it may be underpowered to conduct most of the above comparisons due to the relatively low number of patients treated with plasma-derived molecules has added to the cumulative published data, which tend to discount these concerns when determining product choice for previously untreated patients (PUPs) and others. In fact, several of the haemophilia treaters and haemophilia treatment centres (HTC), who participated in Rodin, had expressed publicly prior to this publication that these same concerns influenced their decision to initiate their PUPs considered at higher risk for inhibitor development on plasma-derived products. The Rodin trial was observational, that is NOT a randomized controlled study, and allowed each HTC to determine independently how it was to treat its PUPs. This approach could have led to an imbalance in the baseline prognostic characteristics of the groups being compared in Rodin and this potential bias could have introduced a significant biostatical flaw into the study design [2].

Reduced Antigenicity of Recombinant Ovine Factor VIII in Hemophilia A Inhibitor Patient Plasmas

AbstractAbstract 1124Hemophilia A is an X-linked recessive disorder caused by deficiencies or functional defects in coagulation factor VIII (fVIII). Approximately 20–30% of patients with severe hemophilia A develop antibodies against fVIII (inhibitors) following fVIII replacement therapy, which presents significant complication to the control of subsequent bleeding episodes. State of the art treatment options for patients with inhibitors include fVIII-bypassing agents such as recombinant factor VIIa or activated prothrombin-complex concentrate. Previously, plasma-derived porcine fVIII was a treatment option for inhibitor patients and was effective due to the reduced antigenicity of porcine fVIII toward anti-human fVIII inhibitors. However due to concerns regarding viral contamination, the plasma-derived porcine fVIII products were discontinued and no alternative fVIII products have been made available to patients with inhibitors. Presently, a recombinant porcine fVIII product (OBI-1, Inspiration Biopharmaceuticals) is being investigated in two phase 3 clinical trials for congenital and acquired hemophilia A. Rationale for the development of such a product consists of the prior success of plasma-derived porcine fVIII and the concept that the most effective and lowest risk treatment for fVIII deficiency, even in the presence of inhibitors, remains a fVIII product. Recently, a line of hemophilia A sheep was reestablished from banked frozen sperm and the ovine fVIII (ofVIII) gene, causal mutation, and protein were genetically and biochemically characterized. B-domain deleted (BDD) ovine fVIII shares 86% identity to human fVIII at the amino acid level and confers phenotypic correction, in vivo, to hemophilia A mice using a tail transaction bleeding model. Recombinant ofVIII was expressed in baby-hamster kidney cells and purified to > 95% homogeneity using a two-step ion exchange chromatography procedure. Highly purified ofVIII displays a specific activity of 18,300 units/mg, which is approximately twice that of recombinant BDD human fVIII. Furthermore, the decay of ofVIII activity following thrombin activation is slower than BDD human fVIII suggesting prolonged activity in vivo. Lastly, ofVIII demonstrates equivalent binding to human von Willebrand factor at physiological concentrations in vitro. A translational aim of the present study was to test the hypothesis that unique sequences within ofVIII confer differential antigenicity compared to human and/or porcine fVIII in congenital and acquired inhibitor patient plasmas. To address this hypothesis, the reactivity of 28 samples (22 congenital patient samples designated 1–22, and 6 acquired hemophilia A patient samples designated A1-A6) from the Emory IRB approved inhibitor bank towards recombinant BDD human, porcine, and ovine fVIII were assessed by enzyme-linked immunosorbant assay (Figure 1). When normalized to the reactivity towards human fVIII, the data revealed reduced reactivity towards ofVIII in 27 of 28 total samples. In only one patient was the reactivity towards ofVIII greater than that towards human fVIII and, in this sample, the reactivity towards porcine fVIII also was greater than 100%. Furthermore, plasma reactivity to ovine fVIII was significantly reduced compared to porcine fVIII (P = 0.025; Mann-Whitney U Test). Median values of the relative cross reactivity towards porcine and ovine fVIII were 54 and 38%, respectively. Preliminary inhibitor analysis (Bethesda assay) of three samples shown to contain titers against human fVIII of 25, 19, and 68 BU/ml, revealed undetectable inhibitor titers towards ofVIII in 2 samples, and a titer of 5 BU/ml in the third, respectively. These results suggest that additional orthologous recombinant fVIII molecules may be enabling to the treatment of patients harboring pathogenic inhibitors to human fVIII. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Kinetics of the interaction between anti‐FVIII antibodies and FVIII from therapeutic concentrates, with and without von Willebrand factor, assessed by surface plasmon resonance

The presence of VWF in plasma-derived FVIII (pdFVIII/VWF) products has been pointed out as a key difference with recombinant FVIII (rFVIII) products with regard to immunogenicity. A Surface Plasmon Resonance (SPR) study was designed to characterize in detail the interaction between anti-FVIII (IgGs) from a severe haemophilia A patient, and FVIII from concentrates of different sources. Full-length rFVIII (preincubated or not with purified VWF), B domain-deleted (BDD)-rFVIII and pdFVIII/VWF were analysed. To ensure reproducible conditions for accurate determination of kinetic constants, a capture-based assay format was developed using protein G surfaces for specific and reversible coupling of endogenous anti-FVIII antibodies. Concentration ranges (nm) of FVIII products tested were 9-0.03 (rFVIII) and 6-0.024 (pdFVIII/VWF). The association with antibodies was monitored for 3-5 min, whereas dissociation of the complex was followed for 5-20-240 min. A strong interaction of rFVIII and BDD-rFVIII with patient's IgG was detected with the K (D) values in the low picomolar range (5.9 ± 3.0 and 12.7 ± 6.9 pm, respectively) and very slow dissociation rates, while pdFVIII/VWF showed only marginal binding signals. The VWF complexed rFVIII displayed reduced binding signals compared with uncomplexed rFVIII, but the K (D) was still in the picomolar range (4.1 ± 1.9 pm) indicating insufficient complex formation. rFVIII, alone or bound to exogenously added VWF, showed high affinity for anti-FVIII IgGs from a severe haemophilia A patient whereas pdFVIII/VWF did not. These results are in agreement with those studies that point towards rFVIII concentrates to be more immunogenic than pdFVIII concentrates.

Efficacy and safety of recombinant factor VIII products in patients with hemophilia A

The introduction of recombinant factor VIII (rFVIII) clotting factor concentrates nearly 20 years ago represented a significant advance in the treatment of hemophilia A. The major advantage of rFVIII products compared with plasma-derived FVIII products is related to product safety, with rFVIII products virtually eliminating bloodborne pathogen transmission. The most challenging aspect of hemophilia A management today is the development of FVIII inhibitors; previously untreated patients are at the highest risk for inhibitor formation. Presented in this article are results of clinical trials in previously treated and untreated patients and postmarketing surveillance studies for the four commercially available rFVIII products (Recombinate, ReFacto, Kogenate FS/Kogenate Bayer and Advate). Recombinant FVIII therapies are highly efficacious when used ondemand and prophylactically, and they have excellent safety profiles; there have been no reports of viral- or prion-based disease transmission associated with rFVIII administration. The incidence rate of inhibitors in previously untreated patients ranges from 15% to approximately 30%. Because rFVIII concentrates have proven efficacy and safety profiles, a number of hemophilia treatment groups recommend rFVIII products as first-line therapy in the management of hemophilia A.

Efficacy and Safety of ReFacto AF for Surgical Hemostasis in Previously Treated Patients with Severe Hemophilia A.

ReFacto AF (Albumin-Free Cell Culture Process) is a BDDrFVIII manufactured by a modified process designed to enhance viral safety relative to currently licensed ReFacto. Efficacy and safety of ReFacto AF for management of surgical hemostasis has been evaluated in an ongoing open label study of at least 25 efficacy evaluable previously treated patients, age ≥12 years, with severe hemophilia A [FVIII:C ≤ 2%] undergoing elective major surgery. ReFacto AF was to be administered for at least 6 postoperative days by bolus injection (BI) or by continuous infusion (CI) at the investigator's discretion, and up to a maximum of 6 weeks following surgery. An interim analysis has been performed on the initial 22 patients. Fourteen (14) patients treated by BI received a total of 647 infusions (range 17 to 72 infusions per patient) for a cumulative total dose of 1,279,150 IU over 457 exposure days (ED), while 8 patients assigned to treatment by CI, including 1 patient who received only 1 dose for PK assessment, received a total dose of 348,618 IU over 140 total ED (range 1 to 64 ED per patient). Of the 21 patients who underwent major surgery, 18 were evaluable for efficacy and had the following procedures: 11 total knee replacements, 3 synovectomies, 1 left ulnar nerve transposition/release, 1 ventral hernia repair/scar revision, 1 knee arthroscopy, and 1 revision/debridement of the knee after total knee replacement. Investigator ratings of efficacy at the end of surgery and the initial postoperative period were all excellent or good [72% excellent (13/18) and 28% good (5/18) at the end of surgery; and 94% excellent (15/16) and 6% good (1/16) at the end of the initial postoperative period]. Blood loss was assessed for the intra- and post- operative periods. Sixteen (16) efficacy-evaluable patients had intraoperative blood loss; for all subjects, blood loss was rated normal. Ten (10) efficacy-evaluable patients had postoperative blood loss; in 9 cases the postoperative blood loss was rated normal, 1 case was rated abnormal due to hemorrhage following surgical trauma to the epigastric artery. Eleven (11) of 18 efficacy-evaluable patients were predicted to require transfusions during the intraoperative period; only 2 were transfused. No intraoperative transfusions were administered to the other 7 efficacy-evaluable patients. During the postoperative period 2 efficacy-evaluable patients received transfusions. One (1) of these had excessive hemorrhage following trauma to the epigastric artery during surgery, the other received 2 units of PRBCs in the postoperative period following normal blood loss. The most frequently reported treatment emergent adverse events (AEs) were fever, anemia and pain, not unexpected in the perioperative setting. Only one AE was related to ReFacto AF: a clinically silent low titer inhibitor was detected in one patient during a routine protocol-specified surveillance test before his surgery, after several injections of a plasma derived FVIII product that were preceded by only 1 dose of ReFacto AF. This patient had a left knee synovectomy, both of his hemostatic efficacy assessments were excellent, and he had less intraoperative blood loss than predicted. These interim data demonstrate that ReFacto AF is effective and safe when used for surgical prophylaxis in hemophilia A patients undergoing major surgery.

ReFacto®1 and Advate®2: a single‐dose, randomized, two‐period crossover pharmacokinetics study in subjects with haemophilia A

ReFacto is a recombinant B-domain-deleted, monoclonal antibody-purified, solvent-detergent-treated factor VIII (BDDrFVIII) with no albumin added to the final formulation. Although ReFacto has been shown to be bioequivalent to a plasma-derived FVIII product (Hemophil-M) in a randomized, crossover pharmacokinetic (PK) study, the comparability of ReFacto with the full-length (complete sequence) recombinant FVIII (FLrFVIII, Advate) product has not been previously examined in this manner. The primary objective of this study was to compare the PKs of ReFacto with those of Advate in patients with severe haemophilia A. This was a third-party unblinded, randomized, multicentre, two-period crossover PKs study of ReFacto and Advate in subjects with severe haemophilia A (FVIII:C < or =1%). Blood samples were collected over a 48-h period after i.v. administration of each of the FVIII products. FVIII:C was determined using the chromogenic substrate assay (CSA) in a central laboratory. The plasma FVIII:C PK parameters of ReFacto and Advate were determined using non-compartmental analysis. Bioequivalence was assessed on maximum plasma concentration (C(max)) and the area under the plasma concentration vs. time curves (AUCs) using an anova. The two products were judged to be equivalent if the 90% confidence limits of the ratio of the geometric mean values of C(max) and AUCs fell within the interval of 80-125%. Results from this PKs comparison of two different rFVIII products, using chromogenic substrate assay to measure FVIII:C, showed that ReFacto and Advate are bioequivalent to each other.

Occurrence of inhibitors in previously untreated or minimally treated patients with haemophilia A after exposure to a plasma‐derived solvent‐detergent factor VIII concentrate

A solvent-detergent virus-inactivated plasma-derived FVIII concentrate (SD-pdFVIII) has been employed for treatment of Italian patients with haemophilia A for 15 years. This product is a non-monoclonally purified, high purity FVIII concentrate, containing large amounts of von Willebrand factor (VWF). A retrospective survey was carried out in Italy in order to evaluate the immunogenicity of SD-pdFVIII in previously untreated patients (PUPs) or in minimally treated patients (MTPs), i.e. previously exposed for up to 5 days only to other plasma-derived concentrates. The survey included 99 patients with ages ranging from 6 to 64 years (median=21.3) of whom 31 PUPs and 68 MTPs, the latter with a median of four exposure days (EDs; range 1-5) to other plasma products. Surveyed patients had been exposed to SD-pdFVIII for a median of 83 EDs (range 21-1580). Seven patients (three PUPs and four MTPs), all with severe haemophilia, had developed inhibitors [7.1%, 95%; confidence interval: 3-14%] after a median of 11 EDs (range 4-22). Of them, two were low responders (<or=5 BU mL-1) and five high responders. Two low responders and one high responder tolerated inhibitors spontaneously; one high responder underwent immunotolerance treatment with complete, long-standing response. These findings show that SD-pdFVIII is at low risk of inhibitor development in PUPs and MTPs with severe and moderate haemophilia, the risk of inhibitor development being similar to that previously reported for other plasma-derived VWF-containing FVIII products.

A postmarketing surveillance study of the safety and efficacy of ReFacto® (St Louis‐derived active substance) in patients with haemophilia A

This clinical trial evaluated the safety and efficacy of ReFacto (St Louis-derived active substance) in patients with severe or moderately severe haemophilia A over a period of 6 months or 50 exposure days (EDs), whichever occurred first. Sixty patients, 58 previously treated and two previously untreated, were enrolled into this study. This was an open-label, multicentre, postmarketing surveillance study in which patients received prophylaxis or on-demand treatment as determined by their doctor. Surgical prophylaxis was evaluated in seven patients requiring elective surgery. Thirty-two patients aged <1 to 66 years (median 19.5) received prophylaxis and 28 patients, aged 1-71 years (median 33.5), received on-demand treatment. The majority of patients had severe haemophilia A (FVIII:C < 2%): 84.4% in the prophylaxis group and 85.7% in the on-demand group. Prophylaxis with ReFacto was associated with a median of 6.7 bleeds per year (range: 0-37). The investigator's assessment of final outcome for prophylactic treatment was excellent or effective for 93.1% of patients. ReFacto resolved 92.8% of bleeds with one or two infusions. The investigator's assessment was excellent or good for 98.2% of bleeds treated with ReFacto. Haemostasis was achieved for all seven surgical cases and ReFacto gave an excellent or good response for each. The nature and incidence of adverse events was as expected and no new safety concerns emerged. One previously treated patient (PTP) developed a high-titre inhibitor (maximum 75 BU) and one minimally treated patient (MTP) developed a low-titre inhibitor while on the study but eventually achieved high titres (maximum 30 BU) after immune tolerance therapy was initiated with a plasma-derived FVIII product. One previously untreated patient (PUP) developed a transient low-titre inhibitor (0.4 BU). Other serious adverse events (SAEs) were unrelated to study treatment. There were no allergic events. The results of this study are consistent with the previously published ReFacto pivotal studies.

Validation of a quantitative SPR assay for recombinant FVIII.

Surface plasmon resonance was employed to establish a quantitative assay for recombinant FVIII (rFVIII) products using rFVIII as standard. The anti-FVIII monoclonal antibody ESH4 was immobilized onto a carboxymethyldextran surface. A range of rFVIII concentrations were injected over the surface and the binding response enhanced by the addition of a further monoclonal antibody ESH8. Validation using National Institute of Biological Standards and Controls (NIBSC) sixth International rFVIII concentrate standard gave inter- and intra-assay coefficient of variations (CVs) of 7.5 and 3.68% respectively for ESH4-rFVIII binding alone. Enhancement of the binding signal by secondary addition of ESH8 produced inter- and intra-assay CVs of 2.75 and 1.5%. The ESH4 immobilized chip was found to retain binding capacity following regeneration for at least 75 cycles. The assay was found to be unsuitable for quantitation of plasma derived FVIII product but may prove useful for monitoring of rFVIII production.

In vivo recovery and safety of human factor VIII product AAFACT® in patients with haemophilia A

AAFACT, a monoclonal purified, solvent/detergent treated human plasma-derived coagulation factor VIII concentrate obtained from plasma of voluntary, non-remunerated blood donors, is manufactured and marketed in the Netherlands by Sanquin Plasma Products since 1995. In a postmarketing surveillance study, 70 previously treated haemophilia A patients were included (73% severe, 14% moderate and 13% mild haemophilia A). Most of these patients were followed during 4 years for the appearance of adverse events, possible transmissions of blood-borne viruses and the occurrence of antibodies against FVIII. The efficacy of treatment was determined in each patient by the in vivo recovery of FVIII. During this study, only six adverse events, possibly related to the use of AAFACT, were reported. None of these were indicated as serious. Transmissions of HIV, HAV, HBV and HCV in the seronegative patients have not been observed. In none of the patients, inhibitors to FVIII were detected. The in vivo recovery of FVIII during this study was not different from the in vivo recovery observed in eight patients during the preregistration study. There was a correlation of in vivo recovery with age and body weight. From these results, we conclude that the clinical usage of this human plasma-derived FVIII product is efficient and safe.