Immunogenicity, efficacy and safety of Nuwiq® (human-cl rhFVIII) in previously untreated patients with severe haemophilia A-Interim results from the NuProtect Study.

Nuwiq® (Human-cl rhFVIII) is a fourth generation recombinant FVIII. This is an ongoing, prospective, multicentre, multinational, open-label, non-controlled, phase III study (NuProtect study) of 110 male previously untreated patients, which aimed to assess immuno-genicity, efficacy and safety of Nuwiq in severe Hemophilia A patients. Here, they report data from a subgroup of 66 patients treated with Nuwiq for up to 100 exposure days (EDs) or 5 years.
Patients received Nuwiq for standard prophylaxis or on-demand treatment, and for treatment of breakthrough bleeds or surgical prophylaxis, as required. The recommended dose for standard prophylaxis was 20-50 U FVIII/kg. The dosage and duration for treating breakthrough bleeds, on-demand treatment and during surgical prophylaxis depended on the location and extent of bleeding, and the clinical condition of the patient. The primary objective is to assess the immunogenicity of Nuwiq (inhibitor activity ≥0.6 BU) using the Nijmegen-modified Bethesda assay at a central laboratory.
High-titre (HT) inhibitors developed in 8 of 66 patients after a median of 11.5 EDs (range 6-24) and low-titre inhibitors in 5 patients. Cumulative incidence was 12.8% for HT inhibitors and 20.8% for all inhibitors. Efficacy was rated as “excellent” or “good” in treating 91.8% of bleeds and for 8 (89%) procedures. Nuwiq was generally well tolerated. The majority of AEs were mild.
Interim analysis included only the 66 patients who were treated for ≥20 EDs with Nuwiq. As almost all patients enrolled across 14 countries in this interim analysis were Caucasian, no data were collected for patients of African descent who are assumed to be at higher risk of inhibitor development.
These interim data showed that Nuwiq was well tolerated, and had good efficacy in Hemophilia A. Cumulative incidence of inhibitors was 12.8% for High titre and 20.8% for all inhibitors.

Plasminogen replacement therapy for the treatment of children and adults with congenital plasminogen deficiency

Congenital plasminogen deficiency is an exceedingly rare autosomal recessive disorder characterized by pseudomembranous lesions on mucous membranes throughout the body, abnormal wound healing and infertility. With the highest severity in infants and children, it is caused by mutations in PLG and is estimated to affect 1.6 individuals per million. Treatment is usually surgical excision of the lesions, although plasminogen concentrates purified from human plasma are being developed as an IV replacement therapy.
This is an open label study of human plasminogen given every 2-4 days based on pharmacokinetic (PK) studies, to achieve a trough plasminogen level of 10% above the patient's baseline. The primary outcome was the ability to obtain this PK endpoint; secondary outcomes were overall clinical success in both the number and size of lesions or change in organ function. Clinical examination of visible lesions and medical imaging of non-visible lesions were used to quantify change.
14 patients, including 5 children, were enrolled. All achieved the primary endpoint, with three patients having transient falls in their trough levels associated with interruption of the infusion schedule. PK studies reveal an approximate 2-week interval to achieve steady state levels. Clinical endpoints also had a 100% success rate, with resolution or improvement of all visible and non-visible manifestations assessed at baseline. The therapy was well tolerated in all patients.
Because of the very small sample size, there were no power calculations or statistical determinations, which the authors acknowledged a priori.
Replacement therapy with human derived plasminogen effectively increased plasminogen activity and improved clinical symptoms. This is an important advance given the limitations of other current treatment options.

Gene Therapy in Patients with Transfusion-Dependent Beta-Thalassemia

This study evaluates the safety and efficacy of a lentiviral beta-globin gene therapy compared to long-term red-cell transfusions in patients with beta-thalassemia. This study reports interim results from two companion phase I and II clinical studies, specifically evaluating the LentiGlobin BB305 vector using a multi-center, non-randomized, open-label single-dose approach.
Patients between the ages of 5 and 35 years with any genotype of transfusion-dependent B-thalassemia were eligible. Patients had hematopoietic stem cells harvested by apheresis. CD34+ cells were transduced with BB305. Conditioning occurred with myeloablative IV busulfan followed by infusion of the LentiGlobin vector-manipulated stem cells.
22 patients were treated on the study. Treatment with the BB305 vector-manipulated stem cells reduced or eliminated the need for transfusion therapy in all 22 patients. In patients 12/13 patients with a non-β0/β0 genotype and 3/9 with a β0/β0 genotype became transfusion-independent. In those who continued to require transfusions, transfusion volume was decreased by 73%. There were no major adverse events, apart from those typically associated with autologous stem cell transplantation.
Small study in 22 patients. The treatment requires autologous stem cell transplantation and a specific lentiviral product that is not yet widely available. Safety has not yet been fully evaluated as long-term or rare side effects can not be determined from only 22 patients.
This study presents an exciting alternative to allogenic hematopoietic stem cell transplantation in patients with transfusion-dependent beta-thalassemia. Further research is required to better characterize the safety profile.

Targeting anticoagulant protein S to improve hemostasis in hemophilia

Current therapy for hemophilia involves replacement of the missing coagulation factor, which has numerous drawbacks including frequent IV infusions and risk of inhibitor formation. Newer approaches have explored gene therapy, antibody-based mimetic therapy, and targeting of coagulation inhibitors to restore thrombin generation. Protein S is a co-factor to activated protein C and tissue factor pathway inhibitor (TFPI), and is therefore a key regulator of thrombin generation.
A protein S (ProS) knockout mouse model was used to evaluate the potential role of protein S inhibition in management of hemophilia. This model was interbred with Factor 8 (F8) and Factor 9 (F9) knockout mice to generate mice which were co-deficient in either F8 or F9 and ProS. These mice were then studied in several bleeding models, including tail clipping and acute hemarthrosis, as well as a model of tissue factor-induced pulmonary embolism.
The model demonstrated that the loss of thrombin generation from F8 or F9 deficiency rescued the ProS deficient mice from thromboembolic complications, although it did not prevent induced pulmonary embolism. Tail bleeding was limited but not completely abrogated in F8 and ProS deficient mice, although ProS deficient mice with F8 or F9 deficiency were protected from acute hemarthrosis. Subsequent studies demonstrated that both ProS and TFPI are expressed in the synovium of both mice and human patients, which may explain this effect. As a final human proof of concept, the research group demonstrated that ProS inhibition in plasma restored thrombin generation in patients with F8 deficiency.
As an animal study, this requires further evaluation in human clinical trials.
Targeting ProS can improve hemophilia in a mouse model, both as an improved bleeding phenotype and protection against hemarthrosis. This may lead to the development of novel therapies for patients with hemophilia.

Gene therapy with adeno-associated virus vector 5-human factor IX in adults with hemophilia B

Gene therapy for hemophilia B is an area of active investigation. The first trial using an adeno-associated virus (AAV) - 8 vector demonstrated durable Factor IX (F9) expression, with reduction in bleeding rates and usage of factor replacement products. However, T-cell activation in response to the viral vector led to loss of transgene, which was temporally associated with transaminitis. The AAV-5 vector has a preferential immune profile, with lower prevalence of neutralizing antibodies and an apparent lack of cellular immune response. As such, it may be a better vector for F9 gene therapy.
10 adults with Hemophilia B either on prophylactic therapy or bleeding manifestations were given a single IV infusion of AMT-060 - a codon-optimized wild-type F9 within an AAV-5 vector. Safety outcome measures included neutralizing antibodies, F9 antibodies, and treatment-related adverse events. Efficacy outcomes included use of F9 concentrates, F9 plasma levels, bleed data, and joint health scores.
6 participants experienced a total of 14 adverse events, most of which were mild, and three had mild elevations of liver transaminases. No patients developed any neutralizing antibodies to AAV-5 or F9. F9 activity rose in all participants and remained stable - patients in the lower-dose cohort achieved mean levels of > 2 U/dL and those in the higher-dose cohort achieved levels of >5 U/dL. Eight of the 9 patients on prophylaxis were able to stop, with a total reduction in F9 use of 79% across all participants.
This is a small cohort with a limited duration of follow up. Long term durability of the transgene remains to be determined.
Utilization of the AAV-5 vector for F9 gene transfer was not associated with vector specific T-cell responses or loss of F9 activity, suggesting it may be a preferable gene delivery system.

Immune tolerance induction: What have we learned over time?

Good review on immune tolerance induction including different protocols to tackle inhibitors in hemophilia.

Induction of fetal hemoglobin synthesis by CRISPR/Cas9-mediated editing of the human β-globin locus

Synthesis of fetal gamma-globin is usually silenced to a large degree after the first months of life. Upregulation of gamma-globin production was shown to mitigate the severity of beta-globin diseases like beta-thalassemia and sickle cell disease. This report investigated whether upregulation of fetal hemoglobin through genome editing might be beneficial in beta-globin diseases.
Large deletions in the beta-globin gene lead to hereditary persistence of fetal hemoglobin. Mutations in these locations were introduced with a CRISPR/Cas9 system in apheresed blood progenitor cells of patients with sickle cell disease or beta-thalassemia.
Of different deletions tested, a 13.6kb measuring deletion (largest) was the most effective with robust fetal hemoglobin production. The same was true for an inversion that spanned the same region. Half the edited cells and about 1/3 overall carried the biallelic rearrangement.
So far, this is a cell line experiment and results cannot directly be transferred to humans. It remains to be proven that these genetically modified cells do engraft in a recipient with sickle cell disease or thalassemia.
This is a promising new approach on how to treat beta-globin diseases with gene editing of patient-derived cells to increase hemoglobin F production. A clinical trial utilizing this approach is in the works and has already passed several regulatory hurdles.

Predictors of remission in children with newly diagnosed immune thrombocytopenia: Data from the Intercontinental Cooperative ITP Study Group Registry II participants

Immune thrombocytopenia (ITP) in childhood spontaneously remits in up to 80% of affected children. Predictors of remission are not well understood and identifying them could be useful for treatment decisions that impact outcomes.
Data from a large prospective cohort of pediatric patients with ITP (Registry II of Intercontinental Cooperative ITP Study Group [ICIS]) was analyzed to investigate factors that might predict remission. Included children were >4 months and 20 years of age with newly diagnosed ITP based on standard criteria. Therapy was classified as no systemic pharmacologic therapy, IVIG alone, corticosteroids alone, anti-D Ig alone, IVIG, and steroids in combination, and other.
1,239 patients were analyzed, 705 had follow-up data and came from 45 different institutions. Data collected through 12 months or through 12 and 24 months. More than half of the patients had no or mild bleeding. Patients were managed with observation alone (28%), IVIG alone (25%), corticosteroids alone (13%), and anti-D Ig (3%). Remission was achieved in 419/705 (59%) at 12 months and 211/383 (55%) at 24 months. Younger age (especially patients 1 year and 1 to 6 years) was associated with highest remission rates at both 12 and 24 months. Significant bleeding at diagnosis and pharmacologic treatment at diagnosis were statistically associated with remission at both 12 and 24 months. Highest remission rate at both 12 and 24 months occurred in the group treated with combination of corticosteroids and IVIG at diagnosis (76% and 77% respectively). Gender and platelet count at diagnosis were not significantly associated with ITP remission rates. Lower platelet count at diagnosis was not predictive of remission (contrary to previous reports).
Large numbers of patients were lost to follow-up, management decisions were at the discretion of the treating provider at multiple different institutions, and other lab studies that can impact disease course (such as ANA and red cell antibodies) were not collected.
Although this study found that combination treatment with IVIG and steroids may help prevent chronic disease, it would not be appropriate to recommend this upfront at this point.

Venous Thromboembolism in Pediatric Hematopoietic Cell Transplant: A Multicenter Cohort Study

This retrospective study examined the prevalence of venous thromboembolism (VTE) in pediatric patients undergoing a hematopoietic stem cell transplant. To date there has been no large retrospective study to examine both the prevalence and risk factors associated with VTE in this at-risk population.
Utilizing the pediatric health information system; a large database comprising information from 49 US pediatric tertiary care hospitals, ICD-9 codes were used to identify patients who underwent allogeneic or autologous stem cell transplant from January 2010 to September 2014. ICD-9 codes were used to identify VTE events, disease-specific indications for transplant and transplant complications including GVHD up to one-year post transplant.
4,158 eligible patients were identified that underwent a transplant. A total of 209 VTE events were identified, leading to a prevalence of 6.97%. 70% were DVTs, 13% Budd-Chiari syndrome, 9% Pulmonary embolism and 8% portal venous thrombosis. When divided between age groups, the highest prevalence was seen in those 1 month to 1-year-old (12.54%) and patients older than 21 years old (10.31%). Risk factors for VTE were identified as age >13yrs old (OR 1.38; 95%CI 1.08-1.77, p0.01) and allogeneic transplant (OR 1.60; 95%CI 1.19-2.15, p0.01). Finally, patients with VTE had increased length of stay in hospital (81 vs 54 days, p0.01), a median length of stay in the intensive care unit (18 vs 12 days, p0.01) and increased 1-year mortality (13.9% vs 5.9%, p0.01).
This is a retrospective study that depends on ICD-9 coding to correctly identify patients. Certain risk factors for VTE were not analyzed in the data including inherited thrombophilias, or prior history of VTE which would have been helpful information when analyzing the results.
The overall prevalence of VTE in patients undergoing hematopoietic stem cell was 6.97%, which is comparable to previous smaller pediatric studies. The identified age groups for VTE, and risk factors demonstrated by this study may increase awareness of this complication in the post-transplant period.

BSH Guideline: management of thrombotic and haemostatic issues in paediatric malignancy.

Very good guideline for the management of VTE and bleeding in pediatric malignancy.

Utility of the immature platelet fraction in pediatric immune thrombocytopenia: Differentiating from bone marrow failure and predicting bleeding risk

The immature platelet fraction (IPF) is an emerging laboratory test for measuring platelet production/turnover. It has been proposed that these immature platelets are more hemostatic and an increase in IPF more consistent with ITP, compared to BMF.
The two objectives were to examine the ability of IPF to help distinguish patients with BMF from ITP in their initial presentation of thrombocytopenia and to determine whether the IPF or AIPN could identify a subset of patients with ITP who were at increased risk of significant bleeding.
Retrospective chart review of thrombocytopenic patients, 2 months – 21 years old, with platelet counts 50 × 10e9/L at Children's Hospital of Philadelphia (CHOP) from November 1, 2013 to July 1, 2015. Charts reviewed for final diagnosis and bleeding symptoms. A bleeding severity score was retrospectively assigned.
272 patients were reviewed: 97 ITP, 11 BMF, 126 malignancy, 38 other. Cut-off IPF > 5.2% differentiated ITP from BMF with 93% sensitivity and 91% specificity. Absolute immature platelet number differed in all three bleeding severity groups (mild, moderate, severe). On multivariate analysis, an IPF 10.4% was confirmed as an independent predictor of bleeding risk at platelet counts 10 × 1e09/l in patients with ITP.
Single centre, retrospective, chart review (bleeding symptoms collected based on what was documented in chart), management of ITP not standardized (physician dependent if treatment provided or not thus potentially influencing bleeding symptoms), no blinding when assigning bleeding scores.
IPF measurement may have utility in diagnosis of ITP. In addition, IPF may be useful for prediction of relative future risk of bleeding in ITP patients, however there are limitations with the bleeding scores in this study.

Timing and severity of inhibitor development in recombinant versus plasma-derived factor VIII concentrates: a SIPPET analysis

Inhibitor development remains a significant problem in congenital hemophilia, with a multi-factorial etiology. The SIPPET trial demonstrated that the risk of inhibitors was nearly twice as high with recombinant products as compared to plasma-derived. While most inhibitors developed within the first 20 exposure days (ED), there is little data in the literature on the time course of inhibitor development.
SIPPET was a randomized trial comparing recombinant and plasma-derived (von Willebrand containing) factor replacement products for their risk of inhibitor development. This post-hoc analysis examined the results of frequent inhibitor testing done on the included patients. Inhibitors were assessed every 3-4 EDs for patients treating on demand and every 2 weeks for patients on prophylaxis, or as clinically indicated.
All inhibitors occurred within 39 ED and 90% within 20 ED (34 and 16 ED respectively for high-titre inhibitors). Inhibitors occurred earlier with recombinant products, peaked at a higher level, and persisted longer. In the first 5 EDs, the hazard ratio for recombinant products was 3.14 for all inhibitors and 4.19 for high-titre. Evaluation of the severity of the immunogenic effect also revealed an increasing hazard ratio for recombinant products with increasing Bethesda titres.
This is a post-hoc analysis of a relatively small sample pool. Because the comparison is between high-purity recombinant product and von Willebrand containing plasma derived product, the role of von Willebrand factor cannot be determined (which may be a relevant consideration in immunogenicity).
The difference in immunogenicity between recombinant and plasma-derived Factor VIII products is temporal, qualitative, and quantitative.