- Use of thrombin generation assay to personalize treatment of breakthrough bleeds in a patient with hemophilia and inhibitors receiving prophylaxis with emicizumab. [Journal Article]
- HHaematologica 2018 Feb 22
- Emicizumab is a recombinant, humanized, bispecific, monoclonal antibody that bridges activated factor IX and factor X to restore the function of deficient factor VIII. Treatment of bleeding in patien...
Emicizumab is a recombinant, humanized, bispecific, monoclonal antibody that bridges activated factor IX and factor X to restore the function of deficient factor VIII. Treatment of bleeding in patients with hemophilia and inhibitors involves the use of bypassing agents (BPA).These molecules are also used for the management of breakthrough bleeds in patients on prophylaxis with emicizumab, with increased concerns about the risks of combining two procoagulant drugs. Using thrombin generation assay, we tailored the dosage of activated prothrombin complex concentrate (APCC) and successfully treated an acute spontaneous arterial bleeding in a patient on prophylaxis with emicizumab 1.5mg.kg-1. Low dosages of APCC 15-25U.kg-1 were sufficient to normalize thrombin generation. The patient safely recovered with no thrombotic complications despite multiple infusions of APCC given during a 3-week period. We also showed that recombinant factor IX concentrates might represent an alternative to BPA in treating breakthrough bleeds in patients receiving prophylaxis with emicizumab.
- Factor XIa-triggered thrombin generation in severe haemophilia A. [Letter]
- BJBr J Haematol 2018 Feb 21
- Effects of pre-analytical heat treatment in factor VIII (FVIII) inhibitor assays on FVIII antibody levels. [Journal Article]
- HHaemophilia 2018 Feb 20
- CONCLUSIONS: The optimal temperature for PHT in the FVIII NBA is 56°C. Higher temperatures may lead to loss of inhibitory antibodies.
- Pharmacokinetic Modelling to Predict FVIII:C Response to Desmopressin and Its Reproducibility in Nonsevere Haemophilia A Patients. [Journal Article]
- THThromb Haemost 2018 Feb 19
- CONCLUSIONS: FVIII:C response to desmopressin in nonsevere HA patients was adequately described by a population PK model. Large variability in FVIII:C response was observed, which could only partially be explained by FVIII-recent.
- Hemlibra's Remarkable Efficacy A Beacon For Hemophilia Patients. [Journal Article]
- MCManag Care 2018; 27(2):30-32
- Hemlibra demonstrates how far antibody science has progressed. Genentech's drug, approved late last year, connects two clotting factors to prevent the devastating bleeds in hemophilia patients with i...
Hemlibra demonstrates how far antibody science has progressed. Genentech's drug, approved late last year, connects two clotting factors to prevent the devastating bleeds in hemophilia patients with inhibitors. The high price may be offset by avoided costs in patients with factor VIII inhibitors.
- Optimization of prophylaxis for hemophilia A. [Journal Article]
- PlosPLoS One 2018; 13(2):e0192783
- CONCLUSIONS: The methods described here can be used to identify optimal, person-specific prophylaxis regimens for children with hemophilia A.
- What´s new in Gene Therapy of Hemophilia [Journal Article]
- CGCurr Gene Ther 2018 02 14
- CONCLUSIONS: Achieving a safe and efficient remedy for hemophilia A and B by means of GT vector engineering needs further improvement. No randomized or quasi-randomized clinical trials of GT for hemophilia have been found. Given it is in its incipient period, there is need for well-designed clinical trials to evaluate the long-term practicability, efficacy and risks of GT for PWH.
- Laboratory testing for factor VIII and IX inhibitors in haemophilia: A review. [Review]
- HHaemophilia 2018 Feb 15
- Inhibitors are antibodies directed against haemophilia treatment products which interfere with their function. Factor VIII (FVIII) inhibitors in haemophilia A and factor IX (FIX) inhibitors in haemop...
Inhibitors are antibodies directed against haemophilia treatment products which interfere with their function. Factor VIII (FVIII) inhibitors in haemophilia A and factor IX (FIX) inhibitors in haemophilia B are significant clinically when they require a change in a patient's treatment regimen. Their persistence may increase morbidity and mortality. Multiple laboratory tests are now available for detecting and understanding inhibitors in haemophilia. Inhibitors are traditionally measured by their interference in clotting or chromogenic factor assays. They may also be detected using immunologic assays, such as enzyme-linked immunosorbent assay or fluorescence immunoassay. Anti-FVIII or anti-FIX antibodies of IgG4subclass best correlate with the presence of functional inhibitors. Improvements in inhibitor measurement have been recently introduced. Preanalytical heat treatment of patient specimens allows testing of patients without delaying treatment. Use of chromogenic and immunologic assays may aid in identification of false-positive results, which are frequent among low-titre inhibitors. Validated reagent substitutions can be used to reduce assay cost. New methods for defining assay positivity and reporting low-titre inhibitors have been suggested. Challenges remain in the areas of quality control, assay standardization, monitoring of patients undergoing immune tolerance induction therapy and testing in the presence of modified and novel treatment products.
- FVIII proteins with a modified immunodominant T-cell epitope exhibit reduced immunogenicity and normal FVIII activity. [Journal Article]
- BABlood Adv 2018 Feb 27; 2(4):309-322
- Factor VIII (FVIII)-neutralizing antibodies (inhibitors) are a serious complication in hemophilia A (HA). The peptide FVIII2194-2213contains an immunodominant HLA-DRA*01-DRB1*01:01 (DRB1*01:01)-restr...
Factor VIII (FVIII)-neutralizing antibodies (inhibitors) are a serious complication in hemophilia A (HA). The peptide FVIII2194-2213contains an immunodominant HLA-DRA*01-DRB1*01:01 (DRB1*01:01)-restricted epitope recognized by CD4+T-effector cells from HA subjects. The aim of this study was to identify amino acid substitutions to deimmunize this epitope while retaining procoagulant function and expression levels comparable to those of wild-type (WT) FVIII proteins. The shortest DRB1*01:01-binding peptide was FVIII2194-2205, and residues important for affinity were identified as F2196, M2199, A2201, and S2204. T-cell proliferation experiments with Ala-substituted FVIII2194-2205peptides identified F2196A as a substitution that abrogated proliferation of clones specific for the WT sequence. T-cell clones that were stimulated by recombinant WT-FVIII-C2 (rWT-FVIII-C2) protein did not proliferate when cultured with rFVIII-C2-F2196A, indicating the immunogenic peptide includes a naturally processed T-cell epitope. Additional amino acid substitutions at F2196 and M2199 were evaluated by peptide-MHC class II (MHCII)-binding assays, T-cell proliferation assays, epitope prediction algorithms, and sequence homologies. Six B-domain-deleted (BDD)-FVIII proteins with substitutions F2196A, F2196L, F2196K, M2199A, M2199W, or M2199R were produced. Proliferation of T-cell clones and polyclonal lines in response to rBDD-FVIII-F2196K and rBDD-FVIII-M2199A was reduced compared with responses to WT-BDD-FVIII. The BDD-FVIII-F2196K sequence modification appears to be the most promising sequence variant tested here, due to its effectiveness at eliminating DRB1*01:01-restricted immunogenicity, low potential immunogenicity in the context of other MHCII alleles, expression level comparable to WT-BDD-FVIII, and retained procoagulant activity. These results provide proof of principle for the design of less immunogenic FVIII proteins targeted to specific subsets of HA patients.
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- Molecular mechanisms of missense mutations that generate ectopic N-glycosylation sites in coagulation factor VIII. [Journal Article]
- BJBiochem J 2018 Feb 14
- N-glycosylation is a common posttranslational modification of secreted and membrane proteins, catalyzed by the two enzymatic isoforms of the oligosaccharyltransferase, STT3A and STT3B. Missense mutat...
N-glycosylation is a common posttranslational modification of secreted and membrane proteins, catalyzed by the two enzymatic isoforms of the oligosaccharyltransferase, STT3A and STT3B. Missense mutations are the most common mutations in inherited diseases, however, missense mutations that generate extra, non-native N-glycosylation sites have not been well characterized. Coagulation factor VIII (FVIII) contains five consensus N-glycosylation sites outside its functionally dispensable B domain. We developed a computer program that identified hemophilia A mutations in FVIII that can potentially create ectopic glycosylation sites. We determined that eighteen of these ectopic sites indeed become N-glycosylated. These sites span the domains of FVIII, and are primarily associated with a severe disease phenotype. Using STT3A and STT3B knockout cells, we determined that ectopic glycosylation exhibited different degrees of dependence on STT3A and STT3B. By separating the effects of ectopic N-glycosylation from those due to underlying amino acid changes, we showed that ectopic glycans promote the secretion of some mutants but impair the secretion of others. However, ectopic glycans that enhanced secretion could not functionally replace a native N-glycan in the same domain. Secretion-deficient mutants, but not mutants with elevated secretion levels, show increased association with the ER chaperones BiP and calreticulin. Though secreted to different extents, all studied mutants exhibited lower relative activity than wild-type FVIII. Our results reveal differential impacts of ectopic N-glycosylation on FVIII folding, trafficking and activity which highlight complex disease-causing mechanisms of FVIII missense mutations. Our findings are relevant to other secreted and membrane proteins with mutations that generate ectopic N-glycans.