Acute Efficacy of a Novel Anti-Tissue Factor Pathway Inhibitor Antibody BAY 1093884 in Hemophilia A Mouse Severe Tail Bleeding

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3500-3500 ◽  
Author(s):  
Yifan Xu ◽  
Maria Koellenberger ◽  
Volker Laux ◽  
Katalin Kauser ◽  
Derek Sim
Keyword(s):  
Blood Loss ◽  
Tissue Factor ◽  
Tissue Factor Pathway Inhibitor ◽  
Hemophilia A ◽  
Human Monoclonal Antibody ◽  
Factor Vii ◽  
Tissue Factor Pathway ◽  
Median Blood Loss

Abstract Tissue factor pathway inhibitor (TFPI) is the major inhibitor of the tissue factor initiated extrinsic coagulation pathway in blood and is intact in patients with hemophilia. The inhibition of TFPI may restore hemostasis in patients with hemophilia. BAY 1093884 is a fully human monoclonal antibody against TFPI developed as a bypass agent for hemophilia patients with or without inhibitors. It restores thrombin burst for stable clot formation in hemophilic conditions in vitro. The goal of these studies was to determine the in vivo acute efficacy of BAY 1093884 in the hemophilia A (HemA) mouse. In the first study, the acute efficacy of BAY 1093884 (3−100 mg/kg) was demonstrated and compared with full-length recombinant factor VIII (rFVIII; 10−100 IU/kg) by a HemA mouse tail clip model, in which blood loss from a severed tail tip was measured over 45 minutes after injury (n=12−27 mice/group). Naive C57/BL6 and HemA mice were used as positive and negative controls, respectively. Whereas isotype control antibody−treated HemA mice had median blood loss of 870 μL, increasing doses of BAY 1093884 to 50 and 100 mg/kg significantly reduced blood loss to a median of 55 and 5 μL. The dose required to reduce blood loss by 50% was 18 mg/kg, approximately equivalent to the efficacy of 20 IU/kg rFVIII. In a second study, we characterized the combined action of BAY 1093884 and activated recombinant factor VII (rFVIIa; n=10−25 mice/group). Low doses of BAY 1093884 (2.5 mg/kg) and rFVIIa (0.5 and 1.0 mg/kg) with minimal efficacies were tested. Untreated HemA mice had median blood loss of 860 μL. As stand-alone treatments, 2.5 mg/kg BAY 1093884, 0.5 mg/kg rFVIIa, and 1 mg/kg rFVIIa provided minimal blood loss protection, with bleeding volume reduced to 675, 830, and 770 μL, respectively. In comparison, the combination of 2.5 mg/kg BAY 1093884 with 0.5 mg/kg rFVIIa or 1.0 mg/kg rFVIIa reduced median blood loss to 215 and 35 μL, respectively. These results showed a combination effect of BAY 1093884 and rFVIIa in this severe acute efficacy model. These studies demonstrate that BAY 1093884 could potently reduce acute blood loss in HemA mice and may offer a new treatment option for hemophilia patients. Disclosures Xu: Bayer HealthCare LLC: Employment. Koellenberger:Bayer Pharma AG: Employment. Laux:Bayer Pharma AG: Employment. Kauser:Bayer HealthCare LLC: Employment. Sim:Bayer HealthCare LLC: Employment.

scholarly journals In Vitro and In Vivo Characterization of Marstacimab, an Anti-TFPI Antibody

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2391-2391
Author(s):  
Chuenlei Parng ◽  
Macy Jin ◽  
Matthew Holsti ◽  
Susan Benard ◽  
Swapnil Rakhe ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Tissue Factor Pathway Inhibitor ◽  
Human Monoclonal Antibody ◽  
Subcutaneous Administration ◽  
The Other ◽  
Pharmacokinetic Parameters ◽  
Human Pharmacokinetics ◽  
Tissue Factor Pathway

Hemophilia A and B are X-linked genetic disorders resulting from functional deficiencies of the intrinsic coagulation plasma proteins Factor VIII (FVIII) or Factor IX (FIX), respectively. An approach to achieving hemostatic pharmacology in hemophilia is to augment the extrinsic cascade is to neutralize Tissue Factor Pathway Inhibitor (TFPI). TFPI is a multi-Kunitz (K) domain inhibitor which binds to and inhibits Factor Xa via the K2 domain and Factor VIIa/Tissue Factor activity (K1 domain). Marstacimab is a fully human monoclonal antibody that binds to and neutralizes TPFI activity and is under development for treatment of hemophilia. To support lead identification during late discovery, five anti-tissue factor pathway inhibitor (TFPI) antibodies (EC50<5nM) were evaluated in an in vivo rabbit pharmacokinetics/pharmacodynamics (PK/PD) study and in vitro using rabbit and human hemostatic assays. Marstacimab displayed potent in vitro activities, and potent pharmacology in shortening the dilute prothrombin time (dPT) assay (1.3 nM) and in the thrombin generation assay (TGA) (2.8 nM). Following an intravenous administration in rabbits, marstacimab and the other anti-TFPI antibodies exhibited target-mediated drug disposition (TMDD) with a half-life less than 2 days, when compared to a control IgG. The pharmacokinetics of anti-TFPI antibody can be described using a Michaelis-Menten mechanistic model in which in vivo Km and Vmax were estimated for the observed nonlinear clearance. The estimated pharmacokinetic parameters for marstacimab was 45 and 38 mL/kg for the central and peripheral volume of distribution, 23 nM for Km, 131 nM/kg/hr for Vmax. In addition, the correlation between in vitro Kd, in vivo Km and in vitro/in vivo potency were assessed. Compared to other anti-TFPI antibodies, marstacimab exhibited reduced clearance and a good in vitro-in vivo relationship. Second, the binding epitopes were further characterized using competitive surface plasmon resonance (SPR) and marstacimab has distinct binding epitopes from the other antibodies. Lastly, the potential human pharmacokinetics and efficacy were evaluated using allometry scaling and in vitro potency, the results predict a minimal target occupancy of 76% following a weekly subcutaneous administration of marstacimab. These studies indicate that the differential influence of the TFPI-binding epitopes on the pharmacokinetics, but not on the pharmacodynamics. The selection strategy could be applicable to the other antibody discovery and development. Disclosures Parng: Pfizer: Employment. Jin:Pfizer: Employment. Holsti:Pfizer: Employment. Benard:pfizer: Employment. Rakhe:Pfizer Inc.: Employment. Patel-Hett:Pfizer: Employment. Joyce:Pfizer: Employment. Webster:pfizer: Employment. Pittman:Pfizer Inc.: Employment.

Effect of Depolymerized Holothurian Glycosaminoglycan (DHG) on Tissue Factor Pathway Inhibitor: In Vitro and In Vivo Studies

1997 ◽  
Vol 78 (02) ◽  
pp. 864-870 ◽  
Author(s):  
Hideki Nagase ◽  
Kei-ichi Enjyoji ◽  
Yu-ichi Kamikubo ◽  
Keiko T Kitazato ◽  
Kenji Kitazato ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Tissue Factor Pathway Inhibitor ◽  
Factor Xa ◽  
Free Form ◽  
Initial Reaction ◽  
Extrinsic Pathway ◽  
Factor Xa Inhibition ◽  
Tissue Factor Pathway

SummaryDepolymerized holothurian glycosaminoglycan (DHG) is a glycosaminoglycan extracted from the sea cucumber Stichopus japonicusSelenka. In previous studies, we demonstrated that DHG has antithrombotic and anticoagulant activities that are distinguishable from those of heparin and dermatan sulfate. In the present study, we examined the effect of DHG on the tissue factor pathway inhibitor (TFPI), which inhibits the initial reaction of the tissue factor (TF)-mediated coagulation pathway. We first examined the effect of DHG on factor Xa inhibition by TFPI and the inhibition of TF-factor Vila by TFPI-factor Xa in in vitro experiments using human purified proteins. DHG increased the rate of factor Xa inhibition by TFPI, which was abolished either with a synthetic C-terminal peptide or with a synthetic K3 domain peptide of TFPI. In contrast, DHG reduced the rate of TF-factor Vila inhibition by TFPI-factor Xa. Therefore, the effect of DHG on in vitroactivity of TFPI appears to be contradictory. We then examined the effect of DHG on TFPI in cynomolgus monkeys and compared it with that of unfractionated heparin. DHG induced an increase in the circulating level of free-form TFPI in plasma about 20-fold when administered i.v. at 1 mg/kg. The prothrombin time (PT) in monkey plasma after DHG administration was longer than that estimated from the plasma concentrations of DHG. Therefore, free-form TFPI released by DHG seems to play an additive role in the anticoagulant mechanisms of DHG through the extrinsic pathway in vivo. From the results shown in the present work and in previous studies, we conclude that DHG shows anticoagulant activity at various stages of coagulation reactions, i.e., by inhibiting the initial reaction of the extrinsic pathway, by inhibiting the intrinsic Xase, and by inhibiting thrombin.

Inhibition of restenosis by tissue factor pathway inhibitor: in vivo and in vitro evidence for suppressed monocyte chemoattraction and reduced gelatinolytic activity

Blood ◽  
2004 ◽  
Vol 103 (5) ◽  
pp. 1653-1661 ◽  
Author(s):  
Christoph W. Kopp ◽  
Thomas Hölzenbein ◽  
Sabine Steiner ◽  
Rodrig Marculescu ◽  
Helga Bergmeister ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Tissue Factor Pathway Inhibitor ◽  
Early Stage ◽  
Thrombus Formation ◽  
Matrix Metalloproteinase 2 ◽  
Monocyte Chemoattractant Protein 1 ◽  
Markers Of Inflammation ◽  
Tissue Factor Pathway

AbstractActivation of inflammatory and procoagulant mechanisms is thought to contribute significantly to the initiation of restenosis, a common complication after balloon angioplasty of obstructed arteries. During this process, expression of tissue factor (TF) represents one of the major physiologic triggers of coagulation that results in thrombus formation and the generation of additional signals leading to vascular smooth muscle cell (VSMC) proliferation and migration. In this study, we have investigated the mechanisms by which inhibition of coagulation at an early stage through overexpression of tissue factor pathway inhibitor (TFPI), an endogenous inhibitor of TF, might reduce restenosis. In a rabbit femoral artery model, percutaneous delivery of TFPI using a recombinant adenoviral vector resulted in a significant reduction of the intimamedia ratio 21 days after injury. Investigating several markers of inflammation and coagulation, we found reduced neointimal expression of monocyte chemoattractant protein-1 (MCP-1), lesional monocyte infiltration, and expression of vascular TF, matrix metalloproteinase-2 (MMP-2), and MMP-9. Moreover, overexpression of TFPI suppressed the autocrine release of platelet-derived growth factor BB (PDGF-BB), MCP-1, and MMP-2 in response to factors VIIa and Xa from VSMCs in vitro and inhibited monocyte TF activity. These results suggest that TFPI exerts its action in vivo through not only thrombotic, but also nonthrombotic mechanisms.

scholarly journals Plasma Tissue Factor Pathway Inhibitor (TFPI) Levels in Healthy Subjects and Patients with Hemophilia A and B

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4672-4672 ◽  
Author(s):  
Jian-Ming Gu ◽  
Chandra Patel ◽  
Katalin Kauser
Keyword(s):  
Tissue Factor ◽  
Healthy Subjects ◽  
Tissue Factor Pathway Inhibitor ◽  
Hemophilia A ◽  
Hemophilia B ◽  
Severe Hemophilia ◽  
Capture Assay ◽  
Healthy Donors ◽  
Tissue Factor Pathway

Abstract BAY 1093884 is a fully human monoclonal antibody against tissue factor pathway inhibitor (TFPI) developed as a potential bypass agent for patients with hemophilia with or without inhibitors. It restores insufficient thrombin burst, leading to stable clot formation in hemophilic conditions in vitro, and effectively stops bleeding in vivo. TFPI is a potent inhibitor of factor Xa (FXa) and the factor VIIa tissue factor complex in the extrinsic pathway. The majority of TFPI is associated with vascular endothelial cells. The mean plasma TFPI concentration in healthy individuals is ~70 ng/mL (1.6 nM) and about 80% of the circulating TFPI is bound to lipoproteins [Dahm, et al. Blood. 2003;101(11):4387-4392; Broze,et al. Front Biosci. 2012;17:262-280]. Some reports indicate that patients with hemophilia B have lower free TFPI levels than patients with hemophilia A, irrespective of phenotypic severity (Tardy-Poncet, et al. Haemophilia 2011;17:312-313). The objective of this study is to determine the plasma TFPI concentration in healthy donors and patients with hemophilia by a newly developed functional TFPI capture assay and to evaluate this assay with inhibition of TFPI by anti-TFPI neutralizing antibody (BAY 1093884) in vitro. A quantitative enzyme-linked immunosorbent assay using FXa as capture agent was developed and validated to measure TFPI levels in human plasma. The assay shows very good precision, accuracy, and reproducibility and should capture all coagulation-relevant forms of TFPI from plasma. Plasma TFPI was determined in 30 healthy donors (15 males and 15 females) and 30 patients with severe hemophilia (hemophilia A [n=12], hemophilia A with inhibitors [n=9], hemophilia B [n=9]). The plasma TFPI levels (mean ± SD) in healthy individuals, patients with severe hemophilia A without and with inhibitors, and severe hemophilia B were 59.5±18.4 ng/mL, 62.9±14.6 ng/mL, 47.3±4.3 ng/mL, and 68.1±8.8 ng/mL, respectively (Table 1). No statistical differences were found based on sex or race (Hispanic, African American, white) in the healthy population and between patients with hemophilia with and without inhibitors. TFPI levels were also not affected by addition of corn trypsin inhibitor (CTI) in citrate plasma. Furthermore, the concentration that inhibits 50% of TFPI levels (IC50) of anti-TFPI antibody (BAY 1093884) was determined to be 4.76 nM in normal human plasma using this assay. In conclusion,plasma TFPI does not appear to be affected by sex or race in healthy subjects, or the deficiency of factor VIII or IX in patients with hemophilia. The functional TFPI capture assay could potentially be used as a pharmacodynamic marker for monitoring plasma TFPI levels after the administration of anti-TFPI antibody and guide dosing strategies. Table 1. Plasma TFPI Levels in Healthy Subjects and Patients With Severe Hemophilia A and B HealthyHuman Donors(n=30) SevereHem A(n=12) Severe Hem AWith inhibitors(n=9) SevereHem B(n=9) TFPI, ng/mL Mean ± SD 59.5±18.4 62.9±14.6 47.3±4.3 68.1±8.8 Hem=hemophilia; TFPI=tissue factor pathway inhibitor. Disclosures Gu: Bayer HealthCare: Employment. Patel:Bayer HealthCare: Employment. Kauser:Bayer HealthCare LLC: Employment.

Tissue Factor Reduction and Tissue Factor Pathway Inhibitor Release after Heparin Administration

1999 ◽  
Vol 81 (04) ◽  
pp. 589-593 ◽  
Author(s):  
A. M. Gori ◽  
G. Pepe ◽  
M. Attanasio ◽  
M. Falciani ◽  
R. Abbate ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Plasma Levels ◽  
Tissue Factor Pathway Inhibitor ◽  
Day Treatment ◽  
Procoagulant Activity ◽  
Cytokine Gene Expression ◽  
Heparin Administration ◽  
Tissue Factor Pathway

SummaryElevated plasma levels of tissue factor (TF) and tissue factor pathway inhibitor (TFPI) and large amounts of monocyte procoagulant activity (PCA) have been documented in unstable angina (UA) patients. In in vitro experiments heparin is able to blunt monocyte TF production by inhibiting TF and cytokine gene expression by stimulated cells and after in vivo administration it reduces adverse ischemic outcomes in UA patients. TF and TFPI plasma levels and monocyte PCA have been investigated in 28 refractory UA patients before and during anticoagulant subcutaneous heparin administration (thrice daily weight- and PTT-adjusted for 3 days) followed by 5000 IU × 3 for 5 days. After 2-day treatment, immediately prior to the heparin injection, TF and TFPI plasma levels [(median and range): 239 pg/ml, 130-385 pg/ ml and 120 ng/ml, 80-287 ng/ml] were lower in comparison to baseline samples (254.5 pg/ml, 134.6-380 pg/ml and 135.5 ng/ml, 74-306 ng/ml). Four h after the heparin injection TF furtherly decreased (176.5 pg/ml, 87.5-321 pg/ml; -32.5%, p<0.001) and TFPI increased (240.5 ng/ml, 140-450 ng/ml; +67%, p<0.0001).After 7-day treatment, before the injection of heparin, TF and TFPI plasma levels (200 pg/ml, 128-325 pg/ml and 115 ng/ml, 70-252 ng/ml) significantly decreased (p<0.05) in comparison to the pre-treatment values. On the morning of the 8th day, 4 h after the injection of heparin TF plasma levels and monocytes PCA significantly decreased (156.5 pg/ml, 74-259 pg/ml and from 180 U/105 monocytes, 109-582 U/105 monocytes to 86.1 U/105 monocytes, 28-320 U/105 monocytes; - 38% and -55% respectively) and TFPI increased (235.6 ng/ml, 152-423 ng/ ml; +70%, p<0.001). In conclusion, heparin treatment is associated with a decrease of high TF plasma levels and monocyte procoagulant activity in UA patients. These actions of heparin may play a role in determining the antithrombotic and antiinflammatory properties of this drug.

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