Recombinant Coagulation Factor VII Research Articles

Effect of the intraoperative infusion of recombinant activated coagulation factor VII on short-term prognosis and thoracic complications after acute aortic coarctation

Purpose: To investigate the impact of intraoperative infusion of recombinant human-activated coagulation factor VII (rFVIIa) on postoperative outcomes in patients with acute aortic dissection (AAD). Methods: This study involved 120 individuals who underwent aortic coarctation surgery between January 1, 2020, and December 31, 2022 in the First Affiliated Hospital of Xinjiang Medical University, Xinjiang, China. The participants were divided into control (receiving only conventional treatment) and study (receiving rFVIIa infusion in addition to conventional treatment) groups. Various parameters, including patient status, postoperative measures, and complications were evaluated. Results: There was no significant difference in age, gender distribution, duration of myocardial block, onset time, or surgery duration (p > 0.05) between both groups. However, the study group exhibited longer extracorporeal circulation duration (p < 0.05). The study group also demonstrated lower levels of chest drainage compared to the control group 12 and 24 h after surgery (p < 0.05). Moreover, fewer concentrated red blood cells and fresh frozen plasma transfusions were administered to study group (p < 0.05). Despite having longer stays in the ICU (p < 0.05), the study group required less postoperative mechanical ventilation (p < 0.05). There was a significant difference in the incidence of postoperative thrombosis cases, positive gram-negative bacilli sputum culture results, and deaths within the study group (p < 0.05). Conclusion: Intraoperative rFVIIа infusion in AAD patients enhances short-term prognosis, with elevated risk of chest complications. However, further studies with larger sample sizes is essential to comprehensively assess clinical advantages and drawbacks.

Design and construction of a phage-displayed Camelid nanobody library using a simple bioinformatics method

BackgroundRational design of synthetic phage-displayed libraries requires the identification of the most appropriate positions for randomization using defined amino acid sets to recapitulate the natural occurrence. The present study uses position-specific scoring matrixes (PSSMs) for identifying and randomizing Camelidae nanobody (VHH) CDR3. The functionality of a synthetic VHH repertoire designed by this method was tested for discovering new VHH binders to recombinant coagulation factor VII (rfVII). MethodsBased on PSSM analysis, the CDR3 of cAbBCII10 VHH framework was identified, and a set of amino acids for the substitution of each PSSM-CDR3 position was defined. Using the Rosetta design SwiftLib tool, the final repertoire was back-translated to a degenerate nucleotide sequence. A synthetic phage-displayed library was constructed based on this repertoire and screened for anti-rfVII binders. ResultsA synthetic phage-displayed VHH library with 1 × 108 variants was constructed. Three VHH binders to rfVII were isolated from this library with estimated dissociation constants (KD) of 1 × 10−8 M, 5.8 × 10−8 M and 2.6 × 10-7 M. ConclusionPSSM analysis is a simple and efficient way to design synthetic phage-displayed libraries.

Rare factor deficiencies in children: A review of 23 cases from a single center

Objective: The prevalence of rare factor deficiency (RFD) is one in 500.000-2.000.000 in the general population. Different symptoms may occur from mild or moderate bleeding to severe and life-threatening bleeding. This study aimed to evaluate children with RFD in a single Turkish center.
 Materials and Methods: The records of children with RFD (Factor I, V, VII, X, XIII deficiency) were evaluated retrospectively.
 Results: Twenty-three cases (70% female) were reviewed. The mean age of patients was 9.52 years at review, and mean follow-up was 66.3 months. The most common factor (F) deficiencies were FVII (35%) and FX (35%). Parental consanguinity was present in 65%. The most common symptoms were mucocutaneous bleeding and epistaxis. Regarding treatment, fresh frozen plasma (FFP) was given to two patients, FXIII concentrate was given to one patient, and prothrombin complex concentrate (PCC) was given to two patients. Prophylaxis was started in patients with recurrent bleeding. Of the 16 receiving prophylaxis, three received FFP, seven received recombinant coagulation factor VIIa, and six received PCC.
 Conclusion: Treatment was given to a fifth of patients while nearly three-quarters received prophylaxis. As parental consanguinity was present in most of these patients, obtaining a detailed family history may aid in diagnosis.

Impact of Early Intravenous Haemostatic Drugs on Brain Haemorrhage Patients and Their Image Segmentation Based on RGB-D Images.

Cerebral haemorrhage is a serious subtype of stroke, with most patients experiencing short-term haematoma enlargement leading to worsening neurological symptoms and death. The main hemostatic agents currently used for cerebral haemorrhage are antifibrinolytics and recombinant coagulation factor VIIa. However, there is no clinical evidence that patients with cerebral haemorrhage can benefit from hemostatic treatment. We provide an overview of the mechanisms of haematoma expansion in cerebral haemorrhage and the progress of research on commonly used hemostatic drugs. To improve the semantic segmentation accuracy of cerebral haemorrhage, a segmentation method based on RGB-D images is proposed. Firstly, the parallax map was obtained based on a semiglobal stereo matching algorithm and fused with RGB images to form a four-channel RGB-D image to build a sample library. Secondly, the networks were trained with 2 different learning rate adjustment strategies for 2 different structures of convolutional neural networks. Finally, the trained networks were tested and compared for analysis. The 146 head CT images from the Chinese intracranial haemorrhage image database were divided into a training set and a test set using the random number table method. The validation set was divided into four methods: manual segmentation, algorithmic segmentation, the exact Tada formula, and the traditional Tada formula to measure the haematoma volume. The manual segmentation was used as the “gold standard,” and the other three algorithms were tested for consistency. The results showed that the algorithmic segmentation had the lowest percentage error of 15.54 (8.41, 23.18) % compared to the Tada formula method.

Transfusion management of abdominal delivery in pregnant women at high risk of bleeding with hemostasis control

Objective of the study . Optimization of transfusion support of abdominal delivery in pregnant women at risk of obstetric haemorrhage using monitored management of haemostasis disorders. Study materials. The study group consisted of 24 women of 23-47 years old with abnormal placental invasion (API) and bleeding during caesarean section at a gestational age of 32-36 weeks. Group 1 consisted of 14 patients, who were injected with tranexamic acid as part of combined therapy; Group 2 consisted of 10 women, who were injected with aminomethylbenzoic acid as part of combined therapy. Results. All patients received instrumental autoerythrocyte reinfusion. Three patients of Group 1 (21.4%) with refractory bleeding received recombinant coagulation factor VIIa (Coagil, Russia) at a dose of 60 pg/kg body weight (average dose 6.6 ± 1.4 mg), a decrease in bleeding was observed, the operative treatment was completed in the form of metroplasty. In the early stages of bleeding, eight patients were injected with a prothrombin complex concentrate of 600-1,200 IU before administration of fresh-frozen plasma (FFP), which allowed to reduce the volume of donor FFP in these patients. The study of hemostasis showed a very high inverse relationship between extent of blood loss and fibrinogen level in the groups of patients. Conclusion. The use of factor VIIa in refractory major obstetric haemorrhage (MOH) in patients with API allows to reduce the blood loss, perform metroplasty, and preserve the woman’s fertility. Patient blood management (PCM) in pregnant women with API makes it possible to conduct organ-preserving treatment in 91.7% of women.

HEK293 Cells Overexpressing Nuclear Factor E2-Related Factor-2 Improve Expression of Recombinant Coagulation Factor VII.

The mammalian expression system plays a key central role in the production of therapeutic recombinant proteins. Conspicuously, any improvements in the expression system which lead to a higher expression level would have an impact especially in bio-pharmaceutical industries. In the current study, to take steps toward the improvement of expression of recombinant protein, first, we established a stable HEK293 cell line to overexpress a well-known cytoprotective and antioxidant gene, Nrf2. Next, we transiently expressed human recombinant coagulation factor VII, as an example of human recombinant protein, in the engineered-HEK293 cell line. Our results revealed that the established cells had a higher growth rate and were able to endure to UV-induced oxidative stress. Furthermore, within our expectation, our results revealed that the expression level of recombinant FVII in Nrf2-engineered HEK293 cells (315ng/ml) was higher than the HEK293 (198ng/ml) cells and it was functional in a coagulation test assay. Moreover, our new cell line could be a suitable cell to express other recombinant proteins especially for large-scale production of recombinant protein under other culture condition such as lower serum and suspension culture that imposed advantages especially in terms of cost benefits in bio-pharmaceutical industries.

Methodologies for data collection in congenital haemophilia with inhibitors (CHwI): critical assessment of the literature and lessons learned from recombinant factor VIIa.

To systematically review the effectiveness of on-demand treatment with recombinant coagulation factor VIIa (rFVIIa) in congenital haemophilia with inhibitors and, if feasible, perform a meta-analysis of the data. Publications from Embase® , MEDLINE® , MEDLINE® In-Process and the Cochrane Central Register of Controlled Trials were searched. Selected publications were reviewed for inclusion by two independent expert reviewers. Discrepancies were reconciled by a third independent reviewer. Data from selected studies were extracted using a predefined grid to ensure uniform and comparable results were captured. A systematic search (cut-off date of 2 May 2016) identified 20 studies (13 observational; seven randomized controlled trials). All studies were of sufficient quality to include in this analysis and comprised 1221 participants, with 5981 bleeds in 746 individuals treated with rFVIIa. Haemostatic overall effectiveness of the individual studies identified ranged from 68% to 100% at ≤12hours, 86% to 96% at 13-24hours and 76% to 99% at 24-48hours with rFVIIa <100μg/kg, with similar rates reported for the ≥250μg/kg dose. However, heterogeneity between the studies precluded pooling of results. Data from the individual studies confirmed that rFVIIa is an effective therapy for the on-demand treatment of bleeds in congenital haemophilia with inhibitors. However, the high levels of heterogeneity between studies precluded pooling of results for a valid, reliable or precise summary measure. There remains a need to implement standardized clinical definitions and measurements for the effectiveness and safety of haemophilia therapies in future clinical trials.

The application of recombinant human coagulation factor VIIa during perioperative period of congenital heart disease in children

Background Refractory hemorrhage is a fatal complication during peri-operative period of congenital heart disease(CHD) in children, which has great influence on the prognosis of patients. Recombinant human coagulation factor Ⅶa(rFⅦa) has been widely used to treat refractory bleeding, but there is no well controlled clinical trial with randomized selected cases or wide accepted guidance to guide clinical application. The application methods and evaluation still vary among different medical centers about the dosage, therapeutic efficacy and adverse effects of rFⅦa in refractory bleeding treatment. Objective This article aims to address the role of rFⅦa played in refractory bleeding treatment to guide clinical applications of rFⅦa. Content This article reviews the index of using rFⅦa, dosage, efficacy and adverse reactions of rFⅦa during the perioperative period of CHD. Trend As more studies are carried out, the use of rFⅦa will be standard and safe. Key words: Recombinant human coagulation factor Ⅶa; Refractory bleeding; Congenital heart disease

Role of agents for reversing the effects of target-specific oral anticoagulants.

The available clinical data on target-specific oral anticoagulant (TSOAC) reversal agents that are currently in development or have been approved by the Food and Drug Administration (FDA) are reviewed. The development of TSOACs such as dabigatran, rivaroxaban, edoxaban, and apixaban has presented benefits and new challenges. One of the main challenges associated with the use of TSOACs is the lack of suitable agent-specific reversal agents. Several treatment options for the management of life-threatening bleeding events associated with TSOAC use, such as fresh frozen plasma, prothrombin complex concentrates, and recombinant coagulation factor VIIa, have been used, with inconsistent results. Currently, two potential reversal agents for oral direct factor Xa inhibitors (andexanet alfa and ciraparantag) are at various stages of clinical development. Idarucizumab, a reversal agent for the oral direct thrombin inhibitor dabigatran, was approved by FDA in October 2015. Idarucizumab and andexanet alfa have been reported to produce anticoagulation reversal effects within minutes of administration. Ciraparantag was demonstrated to decrease whole blood clotting time to within 10% of baseline values in 10 minutes or less, with a return to baseline hemostasis in 10-30 minutes. TSOAC reversal agents have been generally well tolerated in clinical trials. Idarucizumab and other TSOAC reversal agents, such as andexanet alfa and ciraparantag, present the potential for consistent and effective treatment and management options when life-threatening or uncontrolled TSOAC-associated bleeding occurs or when emergency surgery is warranted in patients using TSOACs.

Enhanced recombinant factor VII expression in Chinese hamster ovary cells by optimizing signal peptides and fed-batch medium

ABSTRACTSignal peptides play an important role in directing and efficiently transporting secretory proteins to their proper locations in the endoplasmic reticulum of mammalian cells. The aim of this study was to enhance the expression of recombinant coagulation factor VII (rFVII) in CHO cells by optimizing the signal peptides and type of fed-batch culture medium used. Five sub-clones (O2, I3, H3, G2 and M3) with different signal peptide were selected by western blot (WB) analysis and used for suspension culture. We compared rFVII expression levels of 5 sub-clones and found that the highest rFVII expression level was obtained with the IgK signal peptide instead of Ori, the native signal peptide of rFVII. The high protein expression of rFVII with signal peptide IgK was mirrored by a high transcription level during suspension culture. After analyzing culture and feed media, the combination of M4 and F4 media yielded the highest rFVII expression of 20 mg/L during a 10-day suspension culture. After analyzing cell density and cell cycle, CHO cells feeding by F4 had a similar percentage of cells in G0/G1 and a higher cell density compared to F2 and F3. This may be the reason for high rFVII expression in M4+F4. In summary, rFVII expression was successfully enhanced by optimizing the signal peptide and fed-batch medium used in CHO suspension culture. Our data may be used to improve the production of other therapeutic proteins in fed-batch culture.

Absence of Vitamin K-Dependent γ-Carboxylation in Human Periostin Extracted from Fibrotic Lung or Secreted from a Cell Line Engineered to Optimize γ-Carboxylation.

Periostin (PN, gene name POSTN) is an extracellular matrix protein that is up-regulated in bronchial epithelial cells and lung fibroblasts by TH-2 cytokines. Its paralog, TGF-β-induced protein (βig-h3, gene name TGFBI), is also expressed in the lung and up-regulated in bronchial myofibroblasts by TGF-β. PN and βig-h3 contain fasciclin 1 modules that harbor putative recognition sequences for γ-glutamyl carboxylase and are annotated in UniProt as undergoing vitamin K-dependent γ-carboxylation of multiple glutamic acid residues. γ-carboxylation profoundly alters activities of other proteins subject to the modification, e.g., blood coagulation factors, and would be expected to alter the structure and function of PN and βig-h3. To analyze for the presence of γ-carboxylation, proteins extracted from fibrotic lung were reacted with monoclonal antibodies specific for PN, βig-h3, or modification with γ-carboxyglutamic acid (Gla). In Western blots of 1-dimensional gels, bands stained with anti-PN or -βig-h3 did not match those stained with anti-Gla. In 2-dimensional gels, anti-PN-positive spots had pIs of 7.0 to >8, as expected for the unmodified protein, and there was no overlap between anti-PN-positive and anti-Gla-positive spots. Recombinant PN and blood coagulation factor VII were produced in HEK293 cells that had been transfected with vitamin K 2, 3-epoxide reductase C1 to optimize γ-carboxylation. Recombinant PN secreted from these cells did not react with anti-Gla antibody and had pIs similar to that found in extracts of fibrotic lung whereas secreted factor VII reacted strongly with anti-Gla antibody. Over 67% coverage of recombinant PN was achieved by mass spectrometry, including peptides with 19 of the 24 glutamates considered targets of γ-carboxylation, but analysis revealed no modification. Over 86% sequence coverage and three modified glutamic acid residues were identified in recombinant fVII. These data indicate that PN and βig-h3 are not subject to vitamin K-dependent γ-carboxylation.

Effect of activated recombinant factor VII versus tranexamic acid infusion on bleeding during spine surgery, randomized, controlled, double blinded trial

BackgroundAntifibrinolytic drugs, such as tranexamic acid are medications that facilitate hemostasis and decrease blood loss and the need for blood transfusion during major surgery. Activated recombinant coagulation factor VII is a novel hemostatic agent, studies revealed that it is helpful hemostatic in disorders with impaired hemostasis, as well as in patients with normal hemostatic function to minimize perioperative blood loss. This study aimed to compare the efficacy of activated recombinant factor VII with tranexamic acid in reducing the perioperative blood loss.Methods50 patients undergoing spine surgery were enrolled in this study. Group A patients were given activated recombinant factor VII and group T patients given tranexamic acid. In both groups, anesthesia was induced using fentanyl 3 μg/kg and propofol 2 mg/kg; muscle relaxation was initiated using cisatracurium 0.2 mg/kg. Transfusion of blood and its products was done according to a value guide. The primary outcome variable of the study was the total volume of blood loss in the perioperative period. Secondary outcome variables include perioperative transfusion requirement, and the number of patients who needed transfusion, as well as time of operation. A P-value less than 0.05 is considered statistically significant.ResultsThe current study showed that the total perioperative blood loss in group A was significantly lower than group T. None of the patients required ICU admission, as well as reentry to operating theater. Furthermore, no significant difference was detected in the number of patients needed blood transfusion. Intraoperative bleeding was associated with a slight decrease of hemoglobin in group A.ConclusionThe present study concluded that administration of activated recombinant factor VII in spine surgery reduces the total perioperative blood loss and the total volume of intraoperative blood transfusion compared with tranexamic acid, with no evidence of adverse effects.

A Case of Successful Endoscopic Submucosal Dissection of Gastric Adenoma in a Patient with Congenital Factor VII Deficiency

Congenital factor VII deficiency is a rare hemorrhagic disorder, and invasive procedures are likely to cause excessive bleeding in these patients. Endoscopic submucosal dissection (ESD) has been accepted as a curative treatment modality for gastric adenoma, early gastric cancer (EGC) and any other mucosal and submucosal tumors. The most important complications of ESD are bleeding and perforation. The use of antiplatelet agents or coagulopathies are risk factors for these complications. There are only few reports of successful ESD with coagulation disorders. We report a case of a 70-year-old female patient who was diagnosed with a gastric adenoma and factor VII deficiency. The patient was successfully treated with ESD. Before ESD, recombinant Coagulation factor VIIa was injected, and the procedure was performed successfully without any complications. In conclusion, ESD can be performed successfully in patients with factor VII deficiency, when recombinant human factor VIIa is administered properly. (Korean J Helicobacter Up Gastrointest Res 2015;15:49-52)

Safety and pharmacokinetics of a recombinant fusion protein-linking coagulation factor VIIa with albumin (rVIIa-FP) in healthy volunteers

Safety and pharmacokinetics of a recombinant fusion protein-linking coagulation factor VIIa with albumin (rVIIa-FP) in healthy volunteers

1324

Introduction: Strategies to reverse the anticoagulant effect of rivaroxaban include prothrombin complex concentrate, fresh frozen plasma, and recombinant coagulation factor VIIa (rFVIIa). However, the best method of emergent reversal is uncertain. An 81 year old female with a history of atrial fibrillation on rivaroxaban and coronary artery disease on aspirin presented after fall with a Glasgow coma score (GCS) of 15. A CT scan of the head showed acute subarachnoid hemorrhage (SAH). Patient also had melena, hypotension, and acute kidney injury. Laboratory results at that time were: prothrombin time (PT) 17.5 s, international normalized ratio (INR) 1.5, activated partial thromboplastin time (aPTT) 26.8 s, hemoglobin (Hgb) 6.9 g/dL, and serum creatinine 3.3 mg/dL. After 2 hours the patient's GCS decreased to 9 and she was intubated. The patient was given rFVIIa 6000mcg (92mcg/kg), phytonadione 5mg by mouth, 4 units packed red blood cells, and 1 pool of platelets. Laboratory results 4 hours later were: PT 8.9 s, INR 0.9, aPTT 27.8 s, Hgb 12.7 g/dL. Repeat CT showed no significant change in SAH. The troponin level upon admission was 0.7 ng/mL and post rFVIIa dose increased to 2.73 ng/mL, peaking at 5.23 ng/mL. There were no significant electrocardiogram changes. Despite improvements in her mental status, the patient ultimately expired as a result of respiratory failure in accordance with her expressed wishes. Although the routine use of coagulation assays does not appear to be necessary with rivaroxaban, it has a more reliable effect on the PT than the aPTT. It is unknown whether administration of any procoagulant factor will merely reverse laboratory parameters or will truly impact the clinical outcome of an actively bleeding patient. There is a black box warning for rFVIIa because of an association with thromboembolic events. This warning must be weighed against the potential clinical benefit of anticoagulation reversal, particularly in patients with life-threatening hemorrhage. Consideration of need for reversal and pharmacokinetic parameters, including anticoagulant half-life and route of elimination, may assist in balancing the risks vs. benefit of procoagulant administration.

抜歯後の止血困難をきっかけに発見された後天性血友病Aの1例

Acquired hemophilia A is a rare disease. It is caused by the acquirement of factor VIII inhibitor. We report a case of acquired hemophilia A incidentally diagnosed on bleeding after tooth extraction. A 75-yearold woman referred to our hospital because of bleeding after tooth extraction. Hematological examination revealed a prolonged activated partial thromboplastin time (APTT) and decreased coagulation activity of factor VIII. In addition, a high level of factor VIII inhibitor was detected. Therefore, our patient was given a diagnosis of acquired hemophilia A. The patient was admitted to our hospital and received corticosteroids and bypass hemostasis using recombinant coagulation factor VII. On hospital day 42, APTT and factor VIII activity returned to the normal range. Moreover, factor VIII inhibitor disappeared. The patient was discharged and is being treated at a local general hospital.

Treatment of Dabigatran-Associated Bleeding

Dabigatran etexilate is a competitive, direct thrombin inhibitor that works in the coagulation cascade to ultimately prevent thrombus formation. It is recommended by the 2012 American College of Chest Physicians evidence-based clinical practice guidelines as first-line therapy over vitamin k antagonists for long-term antithrombotic therapy in patients with paroxysmal or persistent nonrheumatic atrial fibrillation who are at intermediate to high risk of stroke and systemic embolism (grade 2B). However, serious postmarketing events involving life-threatening bleeding are emerging with no antidote for reversal of the anticoagulant effect being available for use. Potential reversal agents are being used in clinical practice with questionable efficacy and safety profiles. We report a case involving an 84-year-old male with acute kidney injury who developed life-threatening gastrointestinal and surgical site bleeding secondary to dabigatran accumulation. Use of the Naranjo probability scale indicated a probable cause between the bleeding event and dabigatran use. After discontinuation of drug therapy, fresh frozen plasma, recombinant coagulation factor VIIa, and cryoprecipitate were administered as potential reversal agents with negligible benefit. However, this patient appeared to slowly benefit with administration of continuous venovenous hemodialysis. Based upon our experience with this patient and literature review, the most effective treatment algorithm for dabigatran-associated bleeding may be to utilize hemodialysis initially.

What Causes Hematoma Enlargement in Lobar Intracerebral Hemorrhage?

See related article, p 1490. Intracerebral hemorrhage (ICH) accounts for 10% to 15% of all strokes worldwide.1 “Primary” ICH most often results from the rupture of a small penetrating artery, either associated with hypertensive arteriopathy or cerebral amyloid angiopathy (CAA). Whereas CAA-related ICH preferentially affects cortical–subcortical regions, hypertensive bleedings are located in “deep” areas of the brain such as the basal ganglia, thalamus, and brain stem. For both subtypes of ICH, hematoma size on initial brain scan and hematoma enlargement (HE) between the first and second brain imagings are strong and independent predictors of functional outcome.2,3 Hence, the prevention of HE by coagulation activation was recently evaluated as a treatment strategy in acute ICH in a large-scale randomized trial. Although recombinant coagulation factor VII reduced hematoma expansion, a positive effect on functional outcome after 3 months could not be demonstrated.4 One of the reasons for the lack of success of clinical trials so far might be our scarce knowledge regarding the pathophysiology that underlies hematoma growth. Observational studies reported HE to occur in approximately 30% to 40% of all ICH cases depending on the time spans to first and second brain imaging.5 Several predictors of HE have been identified so far, including clinical, laboratory, and radiological parameters.6,7 In this issue of Stroke , Brouwers et al8 identified the apolipoprotein E (APOE) genotype as a new risk factor for HE. APOE was initially characterized for its involvement in Alzheimer disease.9 It is a lipoprotein with …

Room-Temperature-Stable Recombinant Activated Coagulation Factor VII Recombinant: Chemical and Microbiologic Stability Over 24 Hours During Continuous In VitroInfusion

rFVIIa-25C is a recombinant coagulation Factor VIIa (rFVIIa) formulated to remain stable at room temperature (up to 25°C) for up to 2 hours. The bolus injection of rFVIIa-25C has been reported to be bioequivalent to that of rFVIIa. With alternative administration by continuous infusion (50 μg/kg/h) for surgical hemostatic coverage in hemophilia patients with inhibitors, rFVIIa has been reported to be stable. This research letter presents data from an in vitro study of the activity and chemical stability (rFVIIa content, clot activity, and degradation products), physical stability, and microbiologic stability of rFVIIa-25C administered as a continuous infusion. The findings suggest that rFVIIa-25C remained biochemically stable and aseptic during 24-hour continuous infusion in vitro at 19.3°C to 20.7°C, with no clinically significant changes in clot activity, solution constituents, or concentrations.

Poly-N-Acetylglucosamine Fibers Amplify the Effectiveness of Recombinant Factor VIIA on Clot Formation in Hemophilia B Canine Blood

Achieving hemostasis in anticoagulated patients is an increasingly important clinical issue. Poly-N-acetylglucosamine (pGlcNAc) nanofibers activate platelets by β3 subunit (CD61) and the von Willebrand receptor GP1b (CD42b) integrin signaling for generation of a prothrombotic surface membrane. Recombinant coagulation factor VIIa (rFVIIa) functions in hemophilia A and B by catalyzing formation of the Xa/Va complex on the surface of activated platelets. These observations suggest that pGlcNAc nanofibers may amplify the activity of rFVIIa in hemophilic blood. The activity of rFVIIa on platelets was tested by performing thromboelastographic analysis with blood from hemophilia B dogs in the presence of pGlcNAc nanofibers and increasing concentrations of rFVIIa. Mechanisms for hemostatic system activation were investigated with inhibitors of tissue factor, factor XIIa, and platelet function. Recombinant FVIIa was observed to partially restore the ability of the hemophiliac blood to form fibrin clots in a dose-dependent manner with thromboelastographic analysis. The addition of pGlcNAc nanofibers amplified the rFVIIa effect. The activity of rFVIIa and the amplification effect of pGlcNAc were dependent on platelet integrin function but independent of FXIIa and tissue factor activities. The pGlcNAc nanofibers amplify rFVIIa activity in hemophilia B canine blood by activating platelets through integrin-dependent mechanisms.