scholarly journals Phospholipid-Independent Activity of Fviiia Mimetic Bispecific Antibodies in Plasma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 2461-2461
Author(s):  
Maria M Aleman ◽  
Siddharth Jindal ◽  
Nina Leksa ◽  
Robert Peters ◽  
Joe Salas
Keyword(s):  
Thrombin Generation ◽  
Hemophilia A ◽  
Bispecific Antibodies ◽  
Molar Ratio ◽  
Phospholipid Vesicles ◽  
Employment Equity ◽  
One Stage ◽  
Equity Ownership ◽  
Substrate Cleavage

Abstract Introduction: An important coagulation regulatory mechanism is localization of clotting complexes to exposed phosphatidylserine (PS) on cell surfaces. All components of the intrinsic tenase complex (factor (F)VIIIa, FIXa, and FX) bind to PS. FVIIIa mimetic bispecific antibodies are drugs in development for hemophilia A that aim to mimic the cofactor function of FVIIIa by bringing together FIXa and FX to generate FXa. However, these antibodies differ from FVIII in many ways including no requirement of activation and a lack of direct PS binding. Emicizumab is a bispecific antibody currently on the market for hemophilia A patients with inhibitors. It binds to factor FIX, FIXa, FX, and FXa with micromolar affinities in solution. Previously, we have shown that in-house preparations of sequence-identical emicizumab (SI-Emi) showed similar weak affinities to its antigens and similar in vitro activity to published emicizumab results by one-stage clotting, chromogenic FXa generation, and thrombin generation. However, in chromogenic FXa generation using antibody concentrations in the range of the mean steady state plasma concentration of patients on emicizumab prophylaxis [~360 nM, (Oldenburg, et al., NEJM 2017)], SI-Emi maintained 28% of its activity even in the absence of PS-containing phospholipid vesicles. Another FVIIIa mimetic antibody, BS-027125, was discovered by our group and binds with low nanomolar affinity to FIX, FIXa and FX, with no detectable binding to FXa. In one-stage clotting, BS-027125 achieved clot times similar to physiological levels of FVIII, but had poor activity in thrombin generation at these concentrations. Furthermore, it too maintained small amounts of phospholipid-independent activity in chromogenic FXa generation. Given the artificial nature of the chromogenic FXa generation assay, and that activity of prothrombinase is PS-dependent thereby precluding omission of phospholipids from thrombin generation assays, we developed an assay to detect FXa generation in a plasma milieu by FVIIIa mimetic antibodies or FVIII with and without phospholipid vesicles. Methods: FVIIIa mimetic antibodies or recombinant FVIII (rFVIII) were incubated with thrombin for 5 minutes, quenched with hirudin, then spiked into platelet-free congenital hemophilia A plasma treated with additional hirudin. FXIa (to generate FIXa in situ) with and without PC:PE:PS (40:40:20 molar ratio) phospholipid vesicles was added and reactions were triggered with a solution of CaCl2 and fluorogenic FXa substrate (Mes-D-LGR-ANSN(C2H5)2). Substrate cleavage was monitored kinetically on a fluorescent plate reader. Substrate cleavage by FXIa could not be detected, yet another unknown plasma peptidase did cleave substrate at a constant low rate that was background subtracted. Results: In the absence of phospholipid vesicles, SI-Emi maintained 51±3.7% of its FXa generation activity at all concentrations tested (3.8±0.4 versus 8.0±1.1 RFU/min at 333 nM). BS-027125 showed very low activity (0.43±0.12 RFU/min at 50 nM) in the presence of phospholipid vesicles, however, in the absence of phospholipid vesicles, BS-027125 activity was not detectable above baseline. Nearly all rFVIII activity (>99%) was lost in the absence of phospholipid vesicles (0.14±0.04 versus 15.1±1.8 RFU/min at 0.3 IU/mL). Addition of annexin V was sufficient to block all rFVIII activity in the presence or absence of phospholipid vesicles, but could not block SI-Emi activity. Furthermore, addition of rivaroxaban, a direct FXa inhibitor, confirmed that detection of substrate cleavage was due to FXa activity. Conclusions: In the absence of phosphatidylserine-containing phospholipid vesicles, SI-Emi promoted the generation of FXa in plasma triggered with FXIa. The activity of BS-027125 was too low in this assay to clearly determine its phospholipid-independent activity. These results suggest SI-Emi has mis-regulated (PS-independent) procoagulant activity due to a lack of phospholipid localization of the antibody-FIXa-FX complex. Given the weak affinity of SI-Emi for its antigens, the exact mechanism enabling this activity is unclear. Further study of this phenomenon and its relevance to overall thrombin generation and in vivo activity are needed. Disclosures Aleman: Bioverativ, a Sanofi company: Employment. Jindal:Bioverativ, a Sanofi company: Employment. Leksa:Bioverativ a Sanofi company: Employment. Peters:Bioverativ a Sanofi company: Employment, Equity Ownership. Salas:Bioverativ a Sanofi company: Employment, Equity Ownership.

scholarly journals Thrombin Generation Assay in Plasma from Hemophilia a Patients with or without Inhibitors on Concizumab Prophylaxis: Spiking with rFVIIa or aPCC

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3639-3639
Author(s):  
Marianne Kjalke ◽  
Mads Kjelgaard-Hansen ◽  
Søren Andersen ◽  
Ida Hilden
Keyword(s):  
Thrombin Generation ◽  
Significant Contribution ◽  
Hemophilia A ◽  
Employment Equity ◽  
Breakthrough Bleeding ◽  
Thrombin Generation Assay ◽  
Bleeding Episodes ◽  
Equity Ownership ◽  
Bypassing Agents

Introduction: Concizumab is a humanized monoclonal antibody that inhibits tissue factor pathway inhibitor (TFPI). Concizumab is currently in clinical development as a subcutaneous prophylactic therapy for hemophilia A and B patients with and without inhibitors. Breakthrough bleeding episodes experienced by inhibitor patients while on concizumab prophylaxis may be treated with the bypassing agents recombinant activated factor VII (rFVIIa; NovoSeven®) or activated prothrombin complex concentrate (aPCC; FEIBA®). Aim: To investigate the in vitro effect of rFVIIa and aPCC on hemophilia A plasma containing concizumab using a thrombin generation assay and pooled plasma spiked with concizumab or samples from patients treated prophylactically with concizumab. Methods: Pooled hemophilia A plasma was spiked with concizumab at 1, 3 and 10 nM and patient plasma samples from explorer4 (n=16; hemophilia with inhibitors; NCT03196284) and explorer5 (n=30; hemophilia A; NCT03196297) before and during concizumab prophylaxis at steady state exposure levels were assessed. Samples were spiked with rFVIIa (25 or 75 nM) or aPCC (0.25, 0.5 or 1 U/mL), and analyzed using a thrombin generation assay initiated with tissue factor (PPP-Low, Thrombinoscope). The effects of rFVIIa or aPCC in the absence or presence of concizumab were compared using ANOVA methodology. Results: Addition of rFVIIa or aPCC to hemophilia A plasma with or without inhibitors increased peak thrombin generation both in the absence and presence of concizumab. A significant additional effect of rFVIIa and aPCC was observed for all concizumab concentrations spiked to the plasma pool. Overall, the effects of the combination of concizumab and rFVIIa or aPCC were mainly additive; however, a small but statistically significant drug-drug interaction was observed for rFVIIa (25 nM or 75 nM) and aPCC (0.5 U/ml or 1 U/mL) in the presence of 10 nM concizumab. At this concizumab concentration, the additive effect of aPCC corresponded to 68% of the total observed effect and the additive effect of rFVIIa to 85% of the total observed effect. At lower concizumab concentrations (1 and 3 nM), statistically significant drug-drug effects were only observed in combination with aPCC. No excessive thrombin generation above the level obtained with 1 IU/mL recombinant factor VIII (rFVIII) was observed at 1 nM concizumab combined with either rFVIIa (25 and 75 nM) or aPCC 0.5 U/mL. However, addition of 1 U/mL aPCC to 1 nM concizumab resulted in a thrombin peak modestly above the upper 95% confidence interval of the rFVIII range. In the experiments using plasma from patients treated with concizumab, the increase in thrombin peak upon addition of rFVIIa was within or below the range observed by spiking with 1 IU/mL rFVIII. The increase in thrombin peak upon addition of aPCC was within or above the rFVIII range. The effects of concizumab and rFVIIa or aPCC were mainly additive; however, a small, statistically significant contribution caused by drug-drug interaction was observed for concizumab and rFVIIa (75 nM) in both plasma from patients with and without inhibitors, and for 1 U/mL aPCC in plasma from patients with inhibitors. The additive effects of concizumab and rFVIIa corresponded to between 60% (25 nM rFVIIa, plasma without inhibitors) and 75% (75 nM rFVIIa, inhibitor plasma), and the additive effects of concizumab and 1 U/mL aPCC corresponded to 77% of the total observed effects. Conclusions: Addition of rFVIIa or aPCC to hemophilia A plasma with or without inhibitors increased peak thrombin generation as expected both in the absence and presence of concizumab. Thus, the bypassing agents function as expected in plasma containing concizumab. The effects of concizumab and rFVIIa or aPCC were mainly additive. A small but statistically significant contribution was synergistic in accordance with the concizumab mechanism of action (Hilden I et al, Blood, 2012). These in vitro results support the concomitant use of bypassing agents to treat breakthrough bleeding episodes in hemophilia with inhibitor patients on concizumab prophylactic treatment. Disclosures Kjalke: Novo Nordisk A/S: Employment, Honoraria. Kjelgaard-Hansen:Novo Nordisk A/S: Employment, Equity Ownership. Andersen:Novo Nordisk A/S: Employment, Equity Ownership, Honoraria. Hilden:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties.

Antithrombin Reduction Corrected Thrombin Generation in Samples from Hemophilia A and B Patients with Inhibitors

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 552-552 ◽  
Author(s):  
Gili Kenet ◽  
Tami Livnat ◽  
Emma Fosbury ◽  
Pratima Chowdary ◽  
Alfica Sehgal ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Research Funding ◽  
Thrombin Generation ◽  
Hemophilia A ◽  
Severe Hemophilia ◽  
Employment Equity ◽  
Advisory Committees ◽  
Advisory Boards ◽  
Equity Ownership

Abstract Background: Severe hemophilia A and B patients with inhibitors experience serious musculoskeletal hemorrhage as well as high risk of limb and life threatening bleeds. However, lack of effect of FVIII or FIX substitution therapy and short functional half-life of by-passing agents, leave these patients with very limited bleed preventive treatment options. ALN-AT3 (Alnylam Pharmaceuticals, Cambridge, MA, USA), a subcutaneously administered investigational RNAi therapeutic targeting reduction of antithrombin for potential treatment of hemophilia is currently in phase 1 clinical development in hemophilia A and B patients without inhibitors. Initial data from that ongoing study in 12 patients suggest an AT KD dependent correction of thrombin generation. This study aims to assess changes in peak thrombin generation in samples from patients with severe hemophilia A and B with inhibitors following in vitro reduction of antithrombin. Materials and methods: Citrated plasma samples were obtained from patients with severe hemophilia A and B with high responding inhibitors. Samples were spiked in vitro with isotype specific control IgG or a monoclonal antibody (Haemtech Inc, Essex Junction, VT, USA) targeting antithrombin knockdown of 50% and 90%. Dynamic formation of thrombin was measured by calibrated automated thrombin generation using 1pM tissue factor PPP reagent and 4μM phospholipid (Thrombinoscope, Maastricht, The Nederlands). The primary effect measure was peak thrombin (nM). Data were tested by a 1-way ANOVA and p<0.05 was considered statistically significant. Results: A total of 12 inhibitor hemophilia samples were investigated; 9 hemophilia A and 3 hemophilia B. All the control samples demonstrated a profound defect in thrombin generation with a median peak thrombin of 19.9 nM (range 6.7 - 42.4). Patients with severe hemophilia A and inhibitors had a median peak thrombin generation of 19.7 nM (range 6.7 - 42.4), whereas patients with severe hemophilia B and inhibitors had a median peak thrombin generation of 19.2nM (range 19.4 - 38.1). An AT reduction dependent improvement in peak thrombin generation was observed in all 12 tested plasma samples (Figure 1). In the first 12 subjects, peak thrombin generation was increased up to 363% from a mean of 22nM (control) to 39 nM (50% AT reduction) and 80nM (90% AT reduction) (p<0.05); levels comparable to thrombin generation observed in healthy male volunteers and in hemophilia patients treated with ALN-AT3. Conclusions: These in vitro data suggest that reduction of AT is a promising approach for restoring hemostatic balance and correcting thrombin generation in hemophilia patients with inhibitors. Furthermore, the present laboratory data compare well with clinical data generated with ALN-AT3 administered to patients with hemophilia A or B. Thus, both laboratory and emerging clinical data suggest that targeting antithrombin could be a promising approach for restoring hemostatic balance in hemophilia. The potential for low volume subcutaneous administration, infrequent dosing, and applicability to persons with hemophilia who have inhibitors, make ALN-AT3 a particularly encouraging investigational therapy. Figure 1. Figure 1. Disclosures Kenet: Bayer, Novo Nordisk: Other: Advisory Boards, Speakers Bureau; Opko Biologics: Consultancy, Other: Advisory Boards; BPL; Baxelta: Research Funding; Pfizer: Honoraria. Off Label Use: ALN-AT3 is an investigational RNAi therapeutic targeting the endogenous anticoagulant antithrombin.. Chowdary:Sobi: Membership on an entity's Board of Directors or advisory committees; CSL Behring: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novo Nordisk: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Baxalta: Membership on an entity's Board of Directors or advisory committees; Biogen: Membership on an entity's Board of Directors or advisory committees. Sehgal:Alnylam Pharmaceuticals: Employment, Equity Ownership. Akinc:Alnylam Pharmaceuticals: Employment, Equity Ownership. Sorensen:Alnylam Pharmaceuticals: Employment, Equity Ownership.

scholarly journals Mim8 - a Next-Generation FVIII Mimetic Bi-Specific Antibody - Potently Restores the Hemostatic Capacity in Hemophilia a Settings in Vitro and In Vivo

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 96-96 ◽  
Author(s):  
Stine L Kjellev ◽  
Henrik Østergaard ◽  
Per Jr Greisen ◽  
Mette B Hermit ◽  
Karina Thorn ◽  
...  
Keyword(s):  
Thrombin Generation ◽  
Bispecific Antibody ◽  
Hemophilia A ◽  
Next Generation ◽  
Employment Equity ◽  
Thrombin Generation Assay ◽  
Equity Ownership ◽  
Target Binding

The treatment of hemophilia A (HA) is primarily based on replacement of factor VIII (FVIII), and in people with HA with inhibitors (HAwI) on the use of by-passing agents. Recently, a FVIII mimetic bispecific antibody emicizumab (Hemlibra®) was approved for treatment of HA and HAwI, offering a subcutaneous, prophylactic treatment opportunity with potential for significantly reducing the treatment burden. We describe the development and pre-clinical characterization of Mim8, a novel, next-generation FVIII mimetic human bispecific antibody. Mim8 is a highly potent molecule bridging factor IXa (FIXa) and factor X (FX) in development for subcutaneous treatment of people with HA and HAwI. Development of Mim8 utilized the Duobody® platform to initially screen for compatible anti-FIXa and anti-FX antibodies followed by several iterations of systematic mutational optimization. In total, more than 30,000 bispecific antibodies were analyzed. The optimization process aimed for efficient Mim8-mediated activation of FX by FIXa in the presence of procoagulant membrane, low target binding in solution, low immunogenicity risk, and for desirable biophysical parameters such as low viscosity. In vitro characterization demonstrated that Mim8 efficiently localizes FIXa and FX to the phospholipid surface and enhances FXa activation. The monovalent anti-FIXa arm alone stimulates the proteolytic activity of FIXa in the range of 15,000-fold and is an important contributor to the activity of the bispecific antibody. The dissociation constants (Kd) of Mim8 for FIXa and FX is in the micromolar range, minimizing target binding in the blood. Using thrombin generation assay in congenital HA plasma and thrombelastography (TEG) in whole blood from healthy volunteers spiked with anti-FVIII antibodies, Mim8 was capable of normalizing thrombin generation and blood clot formation, respectively, with approximately 15 times greater potency than emicizumab (Figure 1). A similar potency improvement was demonstrated in a tail vein transection bleeding model in FVIII-deficient mice co-dosed with human FIX and FX to circumvent lack of Mim8 cross reactivity to murine FIX and FX. The terminal half-life of Mim8 was estimated to 14 days (range 10-17 days) in cynomolgus monkeys and the subcutaneous bioavailability to 97%. In conclusion, Mim8 is a novel, next-generation FVIII mimetic bispecific antibody with anti-FIXa and anti-FX arms that potently stimulates FX activation resulting in efficacious haemostasis in vitro and in vivo. Mim8 has a high potency allowing for administration of small volumes in a pen device, good PK parameters, minimal target binding in the blood, and good biophysical properties. Collectively, these properties support clinical development of Mim8 as a potentially improved next-generation FVIII-mimetic prophylactic treatment option for persons with hemophilia A regardless of inhibitor status. Figure 1: Left: FXI-triggered thrombin generation assay in congenital HA plasma (mean and SD of n = 5). Right: thromboelastography in whole blood from healthy donors spiked with polyclonal anti-FVIII antibody (mean and SD of n = 3). Coagulation was triggered with low concentration (∼30 fM) of tissue factor (Innovin® 1:200,000). Shaded areas: standard deviation of controls. Blue circles: Mim8. Grey squares: a sequence identical analogue (SIA) to emicizumab (comparable data were obtained with a commercially available batch of Hemlibra®). Disclosures Kjellev: Novo Nordisk A/S: Employment, Equity Ownership. Østergaard:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Greisen:Novo Nordisk A/S: Equity Ownership, Patents & Royalties: Patents. Hermit:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Thorn:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Hansen:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Zhou:Novo Nordisk A/S: Equity Ownership, Other: Previous employment, Patents & Royalties: Patents. Bjelke:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Kjalke:Novo Nordisk A/S: Employment, Honoraria. Lund:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Holm:Novo Nordisk A/S: Equity Ownership, Other: Previous employment. Ley:Novo Nordisk A/S: Employment, Equity Ownership. Elenius:Novo Nordisk A/S: Equity Ownership, Other: Previous employment; Leo Pharma A/S: Employment, Equity Ownership. Thygesen:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Loftager:Novo Nordisk A/S: Employment, Equity Ownership. Rasch:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Lorenzen:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Gandhi:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Lamberth:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Egebjerg:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Lund:Novo Nordisk A/S: Employment, Equity Ownership. Henriksen:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Rahbek-Nielsen:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Yang:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties: Patents. Hilden:Novo Nordisk A/S: Employment, Equity Ownership, Patents & Royalties.

scholarly journals The Combination of Marzeptacog Alfa (Activated) or Eptacog Alfa (Activated) with Emicizumab Appears Comparable As Assessed By the Thrombin Generation Test in Hemophilia a Plasma

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1112-1112
Author(s):  
Tom Knudsen ◽  
Brajesh Kumar ◽  
Frank Del Greco ◽  
Linda Neuman ◽  
Howard Levy ◽  
...  
Keyword(s):  
Thrombin Generation ◽  
Hemophilia A ◽  
Statistical Significance ◽  
Plasma Concentrations ◽  
Unmet Need ◽  
Surrogate Marker ◽  
Employment Equity ◽  
Thrombin Generation Assay ◽  
Equity Ownership

Background: Hemophilia patients treated prophylactically with emicizumab (Hemlibra®) may experience break-through bleeds or require additional hemostatic coverage for procedures or surgery. Currently available therapies including rFVIIa (Eptacog Alfa (Activated); NovoSeven®) and aPCC (FEIBA®) have been used with Hemlibra to treat bleeding or when additional coverage is required. While NovoSeven appears safe in combination with Hemlibra (HAVEN 1 to 4 clinical trials), thrombotic events have been observed with concurrent use of FEIBA and Hemlibra. While safe and efficacious when used as directed, NovoSeven must be infused intravenously. Ideally, for patients on subcutaneous (SC) prophylaxis with Hemlibra adjunct rFVIIa could be dosed SC too. Marzeptacog alfa (activated) (MarzAA) is a novel rFVIIa differentiated by increased potency and the ability to be administered SC to achieve pharmacologically relevant plasma concentrations. Thus, MarzAA provides a potential solution to address this unmet need in hemophilia therapy. Objective: Demonstrate the procoagulant potential of MarzAA, NovoSeven or FEIBA alone or in combination with Hemlibra using the thrombin generation assay in platelet poor hemophilia A (HA) plasma. The thrombin generation potential will therefore provide a surrogate marker to assess the potential safety and efficacy of SC MarzAA in combination with Hemlibra. Methods: A thrombin generation assay was performed using the PPP-Low Tissue Factor and phospholipid containing thrombin generation assay reagent (#TS31.00, Thrombinoscope, Stago). Citrated hemophilia A plasma was spiked with increasing concentrations (0, 25, 50, and 100 µg/mL) of Hemlibra together with various concentrations of each bypassing agent: MarzAA at 0, 0.1, 0.5, 1, 2.5, 5, and 10 µg/mL, NovoSeven at 1, 2.5, 5, 10, and 50 µg/mL or FEIBA at 0.25, and 0.50 IU/ml. Statistical significance was set at α = 0.05. Results: We assessed the relative potencies of MarzAA and NovoSeven in HA plasma. As expected, MarzAA demonstrated an approximate ten-fold increased potency vs NovoSeven. Both rFVIIa compounds increased peak thrombin generation in the HA plasma to the level of normal plasma and beyond. The effect of adding normalizing levels of MarzAA (1 µg/mL), NovoSeven (10 µg/mL) or FEIBA (0.5 IU/mL) to HA plasma containing clinically relevant concentrations of Hemlibra was evaluated (Fig 1). When correcting for the effect of Hemlibra alone, the increase in peak thrombin generation induced by FEIBA was significantly greater than that observed for both MarzAA and NovoSeven (P<0.002). In contrast, the observed increases in thrombin generation for MarzAA and NovoSeven in combination with Hemlibra were statistically indistinguishable. FEIBA was not tested at the highest clinically relevant concentration (2.0 IU/mL) as assay limitations were already approached at 0.5 IU/ml, corresponding to ~25% of the plasma concentration expected for a clinical FEIBA dose of 100 IU/kg. Furthermore, concentrations of MarzAA (5 µg/mL) or NovoSeven (50 µg/mL) 50-fold higher than expected after standard doses were required before peak thrombin generation became statistically indistinguishable from FEIBA at 0.5 IU/mL when all three compounds were evaluated in the presence of Hemlibra. Conclusion: As assessed by in vitro thrombin generation, equipotent concentrations of MarzAA and NovoSeven exhibit comparable characteristics when spiked into HA plasma containing Hemlibra at clinically relevant concentrations. Based on these data, MarzAA and NovoSeven are expected to behave similarly in combination with Hemlibra when dosed to achieve equipotent plasma concentrations. Figure 1 Disclosures Knudsen: Catalyst Biosciences: Employment, Equity Ownership. Kumar:Catalyst Biosciences: Employment, Equity Ownership. Del Greco:Catalyst Biosciences: Consultancy, Equity Ownership. Neuman:Catalyst Biosciences: Employment, Equity Ownership. Levy:Catalyst Biosciences: Employment, Equity Ownership. Blouse:Catalyst Biosciences: Employment, Equity Ownership.

scholarly journals Elucidating the Excessive Pro-Coagulant Effect of a Sequence Identical Analogue to ACE910 in Combination with Bypassing Agents

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 90-90
Author(s):  
Rudolf Hartmann ◽  
Tjerk Feenstra ◽  
Sabine Knappe ◽  
Michael Dockal ◽  
Friedrich Scheiflinger
Keyword(s):  
Whole Blood ◽  
Thrombin Generation ◽  
Hemophilia A ◽  
Thrombus Formation ◽  
Phase Iii ◽  
Hek293 Cells ◽  
Employment Equity ◽  
Equity Ownership ◽  
Bypassing Agents

Abstract Introduction: Emicizumab (ACE910), an antibody to FIX(a) and FX(a), is currently under investigation for treatment of hemophilia with inhibitors. In a phase III trial, two thromboembolic complications and three cases of microangiopathy were reported in patients on ACE910 prophylaxis [Oldenburg et al. NEJM 2017], whose breakthrough bleeding was treated with activated prothrombin complex concentrate aPCC (FEIBA) or aPCC and rFVIIa. We generated a sequence identical analogue (SIA) to ACE910 and analyzed its synergistic interplay with bypassing agents. Aims: To monitor in vitro the pro-coagulant activity of SIA ACE910 in the presence of FEIBA and rFVIIa, and detect the source of excessive coagulation induced by SIA ACE910 combined with FEIBA. Methods: A sequence identical analogue (SIA) to ACE910 was expressed in HEK293 cells, purified as previously described [Sampei et al. PLoS One 2013], and analyzed in several global hemostatic assays at different concentrations and test conditions using plasma and whole blood assays. In thrombin generation (TG) experiments, platelet-poor plasma (PPP) from hemophilia A inhibitor patients and hemophilia A plasma reconstituted with platelets from 3 healthy donors (PRP) was used. A normal TG range was established in healthy donor plasma. Therapeutic concentrations of SIA ACE910 (20-600 nM) were tested alone and with FEIBA (0.05-1 U/mL) or rFVIIa (0.88-5.25 µg/mL). To measure FEIBA components' contribution to the synergistic effect with SIA ACE910, PPP was spiked with select FEIBA components at concentrations corresponding to 0.5 U/mL FEIBA in combination with the antibody. Thrombus formation was analyzed in FVIII-inhibited blood using rotational thromboelastometry (ROTEM) and Total Thrombus-formation Analysis System (T-TAS). Results: Normal peak thrombin was 47-144 nM for PPP and 88-231 nM for PRP. rFVIIa and FEIBA had an additive effect on TG in combination with SIA ACE910 in both plasma types. Combined with rFVIIa (0.88 µg/mL) or FEIBA (0.5 U/mL), SIA ACE910 (600 nM) induced a ~2- and ~16-fold increase over SIA ACE910 alone. SIA ACE910+rFVIIa did not reach the normal range, while SIA ACE910+FEIBA far exceeded it. Adding individual FEIBA components to PPP showed that FIX was, with a half-maximal effect, the main driver for enhanced TG, followed by FIXa. formation in FVIII-inhibited whole blood using ROTEM and T-TAS confirmed the excessive effect of SIA ACE910+FEIBA. In ROTEM, FEIBA and rFVIIa reduced clotting time to shorter than normal, whereas SIA ACE910 had only little effect. Moreover, adding SIA ACE910 to rFVIIa exerted no effect over rFVIIa alone. Conclusion: Combining SIA ACE910 at plasma concentrations observed in patients [Oldenburg et al. NEJM 2017] with FEIBA induced excessive thrombin generation and faster clot formation. In vitro, this effect is mainly mediated by FEIBA component FIX. ACE910 binds to FIX and FIXa to the same extent, and displays its pro-coagulant effect via an unregulated mechanism. Therefore, careful judgement is needed in treating breakthrough bleeds with FEIBA. Disclosures Hartmann: Shire: Employment. Feenstra: Shire: Employment. Knappe: Shire: Employment. Dockal: Baxalta: Patents & Royalties; Shire: Employment, Equity Ownership; Baxter: Equity Ownership, Patents & Royalties. Scheiflinger: Baxter: Equity Ownership; Shire: Employment, Equity Ownership.

Fitusiran, an Investigational RNAi Therapeutic Targeting Antithrombin for the Treatment of Hemophilia: Updated Results from a Phase 1 and Phase 1/2 Extension Study in Patients with Inhibitors

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1397-1397 ◽  
Author(s):  
K John Pasi ◽  
Pencho Georgiev ◽  
Tim Mant ◽  
Toshko Lissitchkov ◽  
Michael Desmond Creagh ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Research Funding ◽  
Thrombin Generation ◽  
Hemophilia A ◽  
Phase 1 ◽  
Extension Study ◽  
Severe Hemophilia ◽  
Employment Equity ◽  
Advisory Committees ◽  
Equity Ownership

Abstract Background: Development of inhibitory antibodies, also known as "inhibitors," to replacement factor is considered the most serious unmet need in hemophilia and occurs in up to 30% of persons with severe hemophilia A, and 3-5% of persons with severe hemophilia B. Once inhibitors are present in high titer, treatment or prevention of bleeding can become more difficult due to the decreased responsiveness to factor concentrates, requiring bypassing agents (BPA) for bleed management. Current BPAs have a short half-life and are sub-optimally effective. Fitusiran is a subcutaneously (SC) administered investigational RNA interference (RNAi) therapeutic targeting the endogenous anticoagulant antithrombin (AT) as a means to improve thrombin generation and promote hemostasis in patients with hemophilia. Preliminary data from an ongoing Phase 1, multi-center, study showed that fitusiran was generally well tolerated in patients with hemophilia A or B with and without inhibitors and that administration of once-monthly SC doses of fitusiran led to dose-dependent AT lowering, thrombin generation increase, and decrease in bleeding frequency (Pasi KJ, et al. Haemophilia 2016, 22[Suppl 4]). Here we report the updated safety, pharmacodynamic (PD) effect, and clinical activity of fitusiran in patients with hemophilia with inhibitors as well as long term data from the Phase 1/2 extension study. Methods: We are conducting a multi-center Phase 1, four part (Part A: healthy volunteers; Parts B and C: patients with moderate to severe hemophilia A or B; Part D: patients with hemophilia A or B with inhibitors) study (NCT02035605) followed by a multi-center Phase 1/2 extension study (NCT02554773). Primary endpoints include safety and tolerability; secondary endpoints include AT activity, thrombin generation and exploratory evaluation of bleed pattern. In Part D, patients with inhibitors received once-monthly SC fixed doses of 50 or 80mg fitusiran. After receiving 3 monthly doses in the Phase 1 study, all patients were eligible to continue monthly dosing in the Phase 1/2 extension study. Utilization of BPA for breakthrough bleed management was permissible in these patients. Results: Part D of the Phase 1 study included 12 hemophilia A or B patients with inhibitors in 2 dosing cohorts (50mg SC, qM dosing cohort, n=6; 80mg SC, qM dosing cohort, n=6). Within the 50mg dosing cohort there were five patients with severe hemophilia A with inhibitors and one patient with severe hemophilia B with inhibitors; mean age: 33 ± 7 years; mean weight: 73 ± 17kg. Previously reported safety data from the 50mg dosing cohort demonstrated fitusiran was generally well tolerated in hemophilia A or B patients with inhibitors and that there were no serious adverse events related to study drug and no thromboembolic events. Monthly administration of fitusiran at the 50mg dose led to a mean maximal AT lowering of 81 ± 2% and mean maximal thrombin generation increase of 368 ± 113%. A preliminary, post-hoc analysis suggested a 49-100% reduction in bleeding frequency at the lower dose of 50mg during initial follow-up in the Phase 1 study. As of July 2016, the 80mg dose cohort has been fully enrolled and includes 6 patients with hemophilia A with inhibitors; mean age: 39 ± 15 years; mean weight: 75 ± 19kg, and 5 of the 6 patients in the initial 50mg cohort have transitioned to the Phase 1/2 extension study. Follow-up in the Phase 1, 80mg cohort and Phase 1/2 extension study is ongoing. Updated safety, tolerability and clinical activity from the Phase 1 and Phase 1/2 extension studies among all 12 patients with inhibitors will be presented. Conclusions: Emerging clinical data suggest that targeting AT is generally safe and could be a promising approach for promoting hemostasis in patients with hemophilia with inhibitors. Furthermore, the potential for low volume SC administration, monthly dosing, and applicability to patients with hemophilia A and B with and without inhibitors make fitusiran a potentially encouraging investigational therapy. Disclosures Pasi: Biogen: Consultancy, Honoraria; Octapharma: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria; Genzyme: Consultancy, Honoraria; SOBI: Honoraria, Membership on an entity's Board of Directors or advisory committees. Georgiev:Alnylam Pharmaceuticals: Consultancy. Chowdary:Bayer: Honoraria; Baxalta: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sobi: Honoraria; Biogen: Honoraria, Membership on an entity's Board of Directors or advisory committees; CSL Behring: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novo Nordisk: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Ragni:Novo Nordisk: Research Funding; Biomarin: Consultancy; Biogen: Consultancy, Research Funding; Alnylam Pharmaceuticals: Consultancy, Research Funding; Genentech: Research Funding; SPARK: Research Funding; Baxalta: Research Funding; CSL Behring: Research Funding; Shire: Consultancy; Vascular Medicine Institute: Research Funding; Tacere Benitec: Consultancy; OPKO: Research Funding. Soh:Alnylam Pharmaceuticals: Employment, Equity Ownership. Akinc:Alnylam Pharmaceuticals: Employment, Equity Ownership. Partisano:Alnylam: Employment, Equity Ownership. Sorenson:Alnylam Pharmaceuticals: Employment, Equity Ownership.

scholarly journals Platelet-Targeted rFVIIa-Xten Improves Thrombin Generation and Fibrin Formation Compared to Recombinant FVIIa

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2269-2269 ◽  
Author(s):  
Maria M. Aleman ◽  
Elena N. Kistanova ◽  
Nancy Moore ◽  
Volker Schellenberger ◽  
Robert Peters ◽  
...  
Keyword(s):  
Half Life ◽  
Thrombin Generation ◽  
Hemophilia A ◽  
Platelet Rich Plasma ◽  
Life Extension ◽  
Employment Equity ◽  
On Demand ◽  
Fibrin Formation ◽  
Fibrin Network ◽  
Equity Ownership

Abstract The development of inhibitors to replacement plasma factors in hemophilia is an ongoing clinical complication. Bypass therapies, such as recombinant factor VIIa (rFVIIa), have emerged as important alternative on-demand strategies for hemophilic patients with inhibitors to treat spontaneous bleeds and prevent bleeding during surgery. However, general prophylaxis strategies for hemophilia inhibitor patients are lacking. An attractive approach for effective prophylaxis and on-demand treatment includes engineering potent rFVIIa variants with prolonged pharmacokinetics. Because the primary mechanism of action of rFVIIa is thought to occur on the platelet surface, we combined rFVIIa with platelet-targeting and XTEN half-life extension technologies to improve its pharmacokinetic profile and procoagulant activity. Platelet-targeting was achieved by recombinant fusion of an antibody fragment which binds the human alpha IIb integrin. Half-life extension was achieved by fusion of an XTEN polypeptide which increases the hydrodynamic radius, and therefore half-life, of rFVIIa. We have shown that these rFVIIa modifications improve the pharmacokinetics and efficacy of rFVIIa in vivo in humanized alpha IIb transgenic hemophilia A mice. The goal of the current study was to evaluate and compare thrombin generation and fibrin formation kinetics in hemophilic platelet-rich plasma in the presence of platelet-targeted rFVIIa-XTEN or rFVIIa. To achieve this, platelet-rich plasma from normal human donors was treated with an inhibitory factor VIII antibody to model hemophilia A and spiked with doses of platelet-targeted rFVIIa-XTEN or rFVIIa. Reactions were triggered with low tissue factor and recalcification. Thrombin generation (n=9) was monitored by calibrated automated thrombography and fibrin formation (n=7) was monitored optically on a plate reader. Non-linear regression analysis of dose responses was used to determine EC50 values for each parameter for each donor. Platelet-targeted rFVIIa-XTEN increases the rate and peak of thrombin generation with 2- to 6-fold lower EC50 values (peak and rate, respectively) than rFVIIa. These data were confirmed by thrombin generation in platelet-rich plasma from 1 hemophilia A donor and 1 hemophilia B donor which demonstrated similar responses to platelet-targeted rFVIIa-XTEN with 2.5- to 5-fold lower EC50 values (peak and rate, respectively) than rFVIIa. Notably, platelet count-dependent changes in thrombin generation activity were similar between platelet-targeted rFVIIa-XTEN and rFVIIa. Fibrin formation analysis indicated platelet-targeted rFVIIa-XTEN increases the rate and time to plateau of fibrin formation with 5- to 13-fold lower EC50 values (rate and time to plateau, respectively) than rFVIIa. Analysis of fibrin network structure by confocal microscopy indicated platelet-targeted rFVIIa-XTEN increases fibrin network density in platelet-rich plasma clots. Together, these data show platelet-targeted rFVIIa-XTEN has more procoagulant activity than rFVIIa by supporting more thrombin generation and faster fibrin formation and suggest our approach has the potential to be an effective alternative for the treatment and prevention of bleeds in hemophilia patients with inhibitors. Disclosures Aleman: Biogen: Employment, Equity Ownership. Kistanova:Biogen: Employment, Equity Ownership. Moore:Biogen: Employment, Equity Ownership. Schellenberger:Amunix Operating Inc: Employment. Peters:Biogen: Employment. Salas:Biogen: Employment, Equity Ownership.

scholarly journals The Factor VIII Variant X5 Enhances Hemophilia a Gene Therapy Efficiency By Its Improved Secretion

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3356-3356
Author(s):  
Franziska Horling ◽  
Johannes Lengler ◽  
Wenjing Cao ◽  
Biao Dong ◽  
Bagirath Gangadharan ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Factor Viii ◽  
Liver Cell ◽  
Human Liver ◽  
Hemophilia A ◽  
Employment Equity ◽  
Fviii Activity ◽  
Human Liver Cell ◽  
Equity Ownership

Introduction. Adeno-associated virus (AAV)-based factor VIII (FVIII) gene therapy holds great promise to provide clinical benefit in patients with hemophilia A. However, very high doses are currently needed to achieve therapeutic factor levels and the durability appears to be limited to a couple of years. Vector efficiency could be improved by employing more potent liver-specific promoters, but this might come at the price of overstraining the cellular protein folding capacity, causing FVIII to misfold in the lumen of the Endoplasmic Reticulum (ER). This event would in turn activate the unfolded protein response, cause oxidative stress, and if not resolved may even induce cell death. Aims. The objective of the presented study was to test whether the B-domain deleted (BDD)-FVIII-X5 variant can overcome the secretion challenge of high level FVIII expression in the context of hepatic gene therapy. Methods. The human FVIII variant BDD-FVIII-X5 harboring 5 amino acid exchanges in the A1 domain was previously isolated in a screen aimed at identifying those residues in porcine FVIII that are critical for efficient secretion. BDD-FVIII and BDD-FVIII-X5 were produced in Chinese Hamster Ovary (CHO) cells and purified to apparent homogeneity using standard procedures. The preparations were assayed for total protein by UV absorbance at 280 nm and FVIII activity by a chromogenic assay. Both FVIII variants were vectorized using AAV8 and tested in the human liver cell line HepG2 and FVIII knockout mice (E17) at various doses. Resulting samples were assayed for FVIII chromogenic activity. The potential immunogenic risk was evaluated in three hemophilic mouse strains (E17, human FVIII transgenic, humanized HLA-DRB1*1501). Results. A characterization of purified recombinant Refacto-like BDD-FVIII and the corresponding X5 variant revealed similarity of the two proteins and their specific activities in particular, indicating that introduction of the 5 amino acids from porcine FVIII did not alter functionality of human BDD-FVIII. In vitro expression of BDD-FVIII-X5 in a human liver cell line resulted in substantially increased FVIII activity levels in the supernatant compared with the non-modified BDD-FVIII, commensurate with enhanced secretion of the X5 variant. Intravenous delivery of liver-targeted AAV8 vectors carrying the BDD-FVIII-X5 transgene achieved substantial increases in plasma coagulation activity over BDD-FVIII in FVIII-deficient mice, even when highly efficient codon-optimized F8 nucleotide sequences were employed. Evaluation of the immunogenicity of the BDD-FVIII-X5 variant by an immunological risk assessment did not reveal any increased immunogenic risk compared to BDD-FVIII. Conclusions: The fully active BDD-FVIII-X5 variant demonstrated improved secretion in vitro and in vivo, resulting in substantially higher FVIII levels in a hemophilia A mouse model. No signs of enhanced immunogenicity were noted in a comparative immunogenicity study. The results obtained warrant further exploration of the BDD-FVIII-X5 variant for a next generation hemophilia A gene therapy. Disclosures Horling: Baxalta Innovations GmbH, a Takeda company: Employment. Lengler:Baxalta Innovations GmbH, a Takeda company: Employment. Gangadharan:Baxalta Innovations GmbH, a Takeda company: Employment. De La Rosa:Baxalta Innovations GmbH, a Takeda company: Employment, Equity Ownership. Hoellriegl:Baxalta Innovations GmbH, a Takeda company: Employment, Equity Ownership. Reipert:Baxalta Innovations GmbH, a Takeda company: Employment, Equity Ownership. Scheiflinger:Baxalta Innovations GmbH, a Takeda company: Employment, Equity Ownership. Xiao:Ivygen: Other: Patent application on FVIII-X5 has been submitted. Rottensteiner:Baxalta Innovations GmbH, a Takeda company: Employment, Equity Ownership.

A Subcutaneously Administered Investigational RNAi Therapeutic (ALN-AT3) Targeting Antithrombin for Treatment of Hemophilia: Interim Weekly and Monthly Dosing Results in Patients with Hemophilia A or B

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 551-551 ◽  
Author(s):  
K John Pasi ◽  
Pencho Georgiev ◽  
Tim Mant ◽  
Michael Desmond Creagh ◽  
Toshko Lissitchkov ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Research Funding ◽  
Thrombin Generation ◽  
Hemophilia A ◽  
Advisory Board ◽  
Employment Equity ◽  
Advisory Committees ◽  
Educational Support ◽  
Monthly Dosing ◽  
Equity Ownership

Abstract Background: Hemophilia A and B are bleeding disorders characterized by a profound defect in thrombin generation (TG). Furthermore, in the presence of normal levels of endogenous anticoagulants a deficiency in factor VIII and IX results in major hemostatic imbalance and a bleeding phenotype. ALN-AT3 is a subcutaneously administered investigational RNAi therapeutic targeting the endogenous anticoagulant antithrombin (AT) that aims to restore the hemostatic balance by increasing TG. Methods: We are conducting a phase 1 multi-center study (NCT02035605) in healthy volunteers and patients with moderate to severe hemophilia A or B. Part A of this study has been completed and assessed a single ascending dose study in healthy volunteers. Parts B and C are multiple ascending dose studies in patients with hemophilia who are receiving weekly or monthly dosing, respectively. Primary endpoints are safety and tolerability. Secondary endpoints include PK, AT knockdown; change in thrombin generation and whole blood clot formation as measured by Calibrated Automated Thrombin generation and ROTEM thromboelastometry. Exploratory endpoints include evaluations of bleed pattern and control. Results: Part A enrolled 4 healthy volunteers, randomized (3:1) to 30 mcg/kg ALN-AT3 or placebo; no serious adverse events (SAE) or injection site reactions were observed. A total of 12 patients with severe hemophilia (10 hemohilia A; 2 hemophilia B) were enrolled in Part B and received 3 weekly subcutaneous doses of ALN-AT3 at 15 (n=3), 45 (n=6), and 75 (n=3) mcg/kg. Similar to part A, weekly administration of ALT-AT3 was generally safe and well tolerated in patients with hemophilia; no SAEs, discontinuations, clinical thromboembolic events or clinically significant D-dimer increases were reported. In the 75 mcg/kg dosing cohort, the mean maximum AT knockdown was 59% (p<0.05, relative to baseline), with nadir levels achieved between days 28 and 42. Maximum plasma AT knockdown of 86% was achieved, resulting in thrombin generation increases that correlated with AT knockdown and a bleed-free period of 114 days in the patient achieving the highest level of AT knockdown. The association between AT KD and TG was assessed in a post hoc exploratory analysis in which AT KD was categorized into tertiles. Part C aims to enroll several cohorts (n=3 per cohort) and will assess a monthly dosing schedule (x3 doses) of ALN-AT3. Patients in cohort 1 and 2 were dosed at 225 and 450 mcg/kg, respectively. Up to 4 additional cohorts may be enrolled within Part C. Updated safety, PK, AT knockdown, TG results as well as bleed patterns from Parts B and C will be presented. Conclusions: Emerging clinical data suggest that targeting AT could be a promising approach for restoring hemostatic balance in hemophilia. The potential for low volume subcutaneous administration, monthly dosing, and applicability to patients with hemophilia A and B with and without inhibitors make ALN-AT3 a potentially encouraging investigational therapy. Disclosures Pasi: Octapharma: Research Funding; Biogen, Octapharma, Genzyme, and Pfizer: Consultancy, Honoraria. Off Label Use: ALN-AT3 is an investigational RNAi therapeutic targeting the endogenous anticoagulant antithrombin.. Mant:Quintiles: Employment, Equity Ownership. Creagh:Bayer Healthcare UK: Honoraria. Austin:SOBI: Other: member of advisory board and received educational support; Pfizer: Other: member of advisory board and received educational support; Novo Nordisk: Other: member of advisory board and received educational support; CSL Behring: Other: member of advisory board and received educational support; Bio Products Laboratory: Other: member of advisory board and received educational support; Bayer: Other: member of advisory board and received educational support; Baxter: Other: member of advisory board and received educational support. Brand:Alnylam: Honoraria. Chowdary:Bayer: Consultancy; Biogen Idec: Consultancy; Baxter: Consultancy; CSL Behring: Consultancy, Research Funding; Novo Nordisk: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; SOBI: Consultancy. Ragni:Tacere Benitec: Membership on an entity's Board of Directors or advisory committees; Alnylam: Research Funding; Bristol Myers Squibb: Research Funding; Biogen: Research Funding; Shire: Membership on an entity's Board of Directors or advisory committees, Research Funding; Dimension: Research Funding; Bayer: Research Funding; SPARK: Research Funding; Genentech Roche: Research Funding; Pfizer: Research Funding; Vascular Medicine Institute: Research Funding; Baxalta: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Biomarin: Research Funding; CSL Behring: Research Funding. Chen:Alnylam Pharmaceuticals: Employment, Equity Ownership. Akinc:Alnylam Pharmaceuticals: Employment, Equity Ownership. Sorensen:Alnylam Pharmaceuticals: Employment, Equity Ownership. Rangarajan:Octapharma: Other: Investigator.

Fitusiran, an Investigational RNAi Therapeutic Targeting Antithrombin for the Treatment of Hemophilia: Updated Results from a Phase 1 and Phase 1/2 Extension Study in Patients without Inhibitors

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2572-2572 ◽  
Author(s):  
Margaret V. Ragni ◽  
Pencho Georgiev ◽  
Tim Mant ◽  
Michael Desmond Creagh ◽  
Toshko Lissitchkov ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Research Funding ◽  
Thrombin Generation ◽  
Hemophilia A ◽  
Phase 1 ◽  
Extension Study ◽  
Employment Equity ◽  
Advisory Committees ◽  
Part C ◽  
Equity Ownership

Abstract Background: Hemophilia is a bleeding disorder characterized by a profound defect in the ability to generate sufficient thrombin for effective hemostasis. Fitusiran is a subcutaneously (SC) administered investigational RNA interference (RNAi) therapeutic targeting the endogenous anticoagulant antithrombin (AT) as a means to improve thrombin generation and promote hemostasis in patients with hemophilia. Preliminary data from an ongoing Phase 1, four part (Part A: healthy volunteers; Parts B and C: patients with moderate to severe hemophilia A or B; Part D: patients with hemophilia A or B with inhibitors), multi-center, study showed that fitusiran was generally well tolerated and that administration of once-monthly SC doses of fitusiran led to dose-dependent AT lowering, thrombin generation increase, and decrease in bleeding frequency (Pasi KJ, et al. Haemophilia 2016, 22[Suppl 4]). The purpose of this abstract is to report the updated safety, pharmacodynamics (PD), and clinical activity of fitusiran in patients with hemophilia without inhibitors from the Phase 1 (Part C) as well as long term data from the Phase 1/2 extension study. Methods: We are conducting a multi-center Phase 1 study (NCT02035605) followed by a multi-center Phase 1/2 extension study (NCT02554773) in hemophilia A and B patients. Patients received SC administered, weekly (Part B) or once-monthly (Part C) weight-based or fixed doses of fitusiran. Primary endpoints include safety and tolerability; secondary endpoints include AT activity, thrombin generation and exploratory evaluation of bleed pattern. After receiving 3 monthly doses in the Phase 1 study, all patients were eligible to continue monthly dosing in the Phase 1/2 extension study. Standard replacement factor VIII or IX was utilized to manage any breakthrough bleeds. Results: Previously reported safety data demonstrated that fitusiran was generally well tolerated in hemophilia A or B patients without inhibitors (Phase 1, Parts B and C) and that there were no serious adverse events related to study drug and no thromboembolic events. In Phase 1, Part C, patients initially received weight-based doses ranging from 225 to 1800mcg/kg or a fixed dose of 80mg. Fitusiran dosing resulted in dose-dependent lowering of AT with a mean maximal AT lowering of 87 ± 1% at the 80mg fixed dose and mean thrombin generation increase of 289% relative to baseline with AT lowering >75%. Exploratory analysis of bleed events (Phase 1, Part C) showed a median annualized bleeding rate (ABR) of zero during the defined observation period. As of July 2016, a total of 25 non-inhibitor patients have been enrolled in the entire Phase 1 study and 16 of these patients have entered the ongoing Phase 1/2 extension study. All patients who enrolled in the Phase 1/2 extension study transitioned to a fixed monthly fitusiran dose of 50mg or 80mg. Updated safety, tolerability and clinical activity among all non-inhibitor patients will be presented. Conclusions: Emerging clinical data suggest that targeting AT is generally safe and could be a promising approach for promoting hemostasis in hemophilia. The potential for low volume SC administration, monthly dosing, and applicability to patients with hemophilia A and B with and without inhibitors make fitusiran a potentially encouraging investigational therapy. Disclosures Ragni: Tacere Benitec: Consultancy; Shire: Consultancy; Novo Nordisk: Research Funding; Genentech: Research Funding; CSL Behring: Research Funding; Biomarin: Consultancy; Biogen: Consultancy, Research Funding; Baxalta: Research Funding; Alnylam Pharmaceuticals: Consultancy, Research Funding; OPKO: Research Funding; Vascular Medicine Institute: Research Funding; SPARK: Research Funding. Georgiev:Alnylam Pharmaceuticals: Consultancy. Chowdary:Sobi: Honoraria; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novo Nordisk: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; CSL Behring: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Biogen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Baxalta: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bayer: Honoraria. Rangarajan:Novo Nordisk: Research Funding; Baxter: Research Funding; Baxalta, now part of Shire: Other: Investigator Clinical Studies, Research Funding; Biogen: Consultancy; Biotest: Research Funding; Pfizer: Research Funding; Grifols: Consultancy, Research Funding. Soh:Alnylam Pharmaceuticals: Employment, Equity Ownership. Akinc:Alnylam Pharmaceuticals: Employment, Equity Ownership. Partisano:Alnylam: Employment, Equity Ownership. Sorenson:Alnylam Pharmaceuticals: Employment, Equity Ownership. Pasi:Pfizer: Consultancy, Honoraria; Octapharma: Consultancy, Honoraria, Research Funding; SOBI: Honoraria, Membership on an entity's Board of Directors or advisory committees; Genzyme: Consultancy, Honoraria; Biogen: Consultancy, Honoraria.

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