WO2016005564A2

WO2016005564A2 - Antibodies against activated factor v

Info

Publication number: WO2016005564A2
Application number: PCT/EP2015/065848
Authority: WO
Inventors: Heidi Lindgreen Holmberg; Gustav RØDER; Kristoffer Winther BALLING
Original assignee: Novo Nordisk A/S
Priority date: 2014-07-11
Filing date: 2015-07-10
Publication date: 2016-01-14
Also published as: AR101185A1; WO2016005564A3; TW201617371A

Abstract

The present invention relates to monoclonal antibodies or fragments thereof, that bind to activated Factor V (FVa), that display pro-coagulant effect and therapeutic uses in thereof.

Description

ANTIBODIES AGAINST ACTIVATED FACTOR V

FIELD OF THE INVENTION

The present invention relates to antibodies, and compositions thereof, that are capable of binding to coagulation Factor V complex. BACKGROUND

In subjects with a coagulopathy, such as in human beings with haemophilia A and B, various steps of the coagulation cascade are rendered dysfunctional due to, for example, the absence or insufficient presence of a coagulation factor. Such dysfunction of one part of the coagulation cascade results in insufficient blood coagulation and potentially life-threatening bleeding, or damage to internal organs, such as the joints. Subjects such as human beings with haemophilia A and B may receive coagulation factor replacement therapy such as exogenous Factor VIII (FVIII) or Factor IX (FIX), respectively. However, such patients are at risk of developing "inhibitors" (neutralising anti-drug antibodies) to such exogenous factors, rendering formerly efficient therapy ineffective. Furthermore, exogenous coagulation factors may only be administered intravenously, which is of considerable inconvenience and discomfort to patients. For example, infants and toddlers may have to have intravenous catheters surgically inserted into a chest vein, in order for venous access to be guaranteed. This leaves them at great risk of developing bacterial infections. Subjects with a

coagulopathy may only receive therapy after a bleed has commenced, rather than as a precautionary measure, which often impinges upon their general quality of life.

There are thus still many unmet medical needs in the haemophilia community, in particular, and in subjects with coagulopathies, in general.

Activation of the blood coagulation system relies on a complex cascade of biological reactions. When a vessel wall is injured, tissue factor (TF) is exposed to the contents of circulating blood and TF forms a complex with Factor VI l/activated Factor VII (FVII/FVIIa) on the surface of TF-bearing cells. This leads to the activation of Factor X (FX) to FXa which together with activated Factor V (FVa) generates a limited amount of thrombin (Fl la). Small amounts of thrombin activate platelets, which results in surface exposure of phospholipids (PL) that supports the binding of the tenase complex consisting of activated FVIII/FIX

(FVIIIa/FIXa). Generation of a thrombin burst is central for a stable clot formation after injury to the vessel wall. Key to the production of thrombin is FXa and its cofactor FVa. When FXa forms a complex with FVa, the prothrombin turnover is enhanced approximately 10⁵-fold. The FVa:FXa complex generates both the initial small amount of thrombin required for the first activation of platelets during the initiation phase of the coagulation process and the thrombin burst on the activated platelets during the coagulation propagation phase where large amounts of FXa are generated by the complex the tenase complex. In patients with haemophilia A or B the propagation phase cannot take place and consequently insufficient thrombin is generated to form a stable clot.

Despite being downstream of the tenase complex which is deficient in haemophilia, several in vivo studies in either FVIII or FIX knock-out mice have demonstrated a significant ameliorating effect of increased FVa levels (Schlachterman A et al., (2005) J Throm

Haemost, 3:2730).

Factor V (FV)

Factor V is a pro-cofactor in the coagulation system. In it's activated form it has a procoagulant action that is associated with cofactor activity for FXa in the prothrombinase complex. FV is activated by thrombin or FXa, where it is spliced in two chains which are noncovalently bound to each other by metal ions. FVa is a cofactor of the prothrombinase complex and catalyses the conversion of prothrombin to thrombin on a phospholipid (PL) surface.

Blood coagulation factor V (FV) is a single-chain 330 kDa glycoprotein produced by hepatocytes and present in both plasma and platelets (Chesney, C. et al. (1981 ) Natl Acad Sci USA 78:5180). It is composed of six domains A1 (AA 29-329), A2 (AA 348-684), B (AA 692-1573), A3 (AA 1578-1907), C1 (AA 1907-2061 ), and C2 (AA2066-2221 ) (SEQ ID NO: 36). The A and C domains of the two chains are approximately 40% homologous with the equivalent domains of FVIII, but the B domains are not conserved. Following cleavage of the 28 amino acid signal peptide, the mature Factor V molecule consists of 2196 residues and has a domain structure that comprises three A-domains, one B-domain and two C-domains organized as follows: NH2-A1 -A2-B-A3-C1-C2-COOH.

Activation of FV is catalysed by thrombin or activated factor X (FXa) and occurs via cleavages at residues within and flanking the B-domain (Arg 709, Arg 1018 and Arg 1545) (Tans, G. et al. (1994) J Biol Chem 269:15969). Activation of FV to FVa causes dissociation of the B-domain from the remaining FVa heavy chain (HC, A1-A2, SEQ ID NO 37) and light chain (LC, A3-C1-C2, SEQ ID 38). The two chains of the FVa hetero-dimer remain associated in a bivalent metal-ions dependent fashion (Hibbard, L. and Mann, K. (1980) J Biol Chem 255:638).

Following activation, FVa is a co-factor to the enzyme FXa in the activation of prothrombin to thrombin. The prothrombinase complex (FVa:FXa complex) is responsible for the thrombin burst which occurs during normal haemostasis. Complete deficiency of either FV or FX has been shown to cause fatal embryonic or postnatal bleeding in mice (Cui J et al.

(1996) Nature 384:66, Dewerchin M et al. (2000) Thromb Haemost. 83:185).

Down-regulation of prothrombinase activity occurs via FVa inactivation catalysed by activated protein C (APC) (Walker, F. et al. (1979) Biochim Biophys Acta 571 :333). APC is generated from activation of protein C by thrombin in complex with thrombomodulin. APC inactivates FVa via two pathways: one involves a kinetically favoured cleavage at Arg 506 in FVa, which yields a partially active FVa intermediate that is subsequently inactivated completely by cleavage at Arg 306 which leads to dissociation of the A2 domain from the rest of the molecule; the alternative inactivation pathway involves initial cleavage at Arg 306 in FVa (Nicolaes, GA et al. (1995) J Biol Chem 270:21 158).

Naturally occurring mutations in either of the two APC cleavage sites in FVa have been described: FV Leiden (Arg 506 Gin substitution), FV Hong-Kong (Arg 306 Gly substitution) and FV Cambridge (Arg 306 Thr) (Bertina, R. et al. (1994) Nature 369:64, Chan, WP. et al. (1998) Blood 91 :1 135, Williamson, D. et al. (1998) Blood 91 : 1 140). FV Leiden is a defect in the rapid phase of FVa inactivation, causes APC resistance and is a risk factor for venous thrombosis. In contrast, FV Cambridge and FV Hong Kong are not associated with APC resistance or an increased risk of venous thrombosis. Furthermore, FV Leiden also shows impaired anti-coagulant co-factor activity in APC-mediated inactivation of FVIIIa (Thorelli E. et al. (1998) J Biol Chem 273:16140). Similar features to the FV Leiden gene mutation have been described for FV Nara gene mutation (Trp 1920 Arg substitution) (Nogami et al., (2014) Blood 123:2420).

The FV Leiden mutation has been shown to cause increased thrombin generation in normal plasma in which the APC system was activated either by the including thrombomodulin or by direct addition of APC (Castoldi et al. (2004) Blood 103:4173).

Furthermore, FV Leiden has been shown to improve the haemophilia A and B phenotype in mice, especially in the microcirculation (Schlachterman, A et al. (2005) J Thromb Haemost 3:2730). In addition to the increase FVa levels by direct supplementation of exogenous FVa (Schlachterman A et al., (2005) J Throm Haemost, 3:2730), it has also been suggested that prevention of the APC-mediated inactivation of FVa by using an antibody directed towards APC (Patent application number: 200901 10683) would improve the haemophilia A and B phenotypes.

Approaches pursued to increase FVa levels include direct supplementation of exogenous FVa (Schlachterman A et al., (2005) J Throm Haemost, 3:2730), FVa variants (von Drygalski, A. et al., (2012) Blood 120, 17) or interference with FVa inactivation by activated protein C (APC; Patent application number: WO200901 10683).

Protein C

Protein C, also known as autoprothrombin IIA and blood coagulation factor XIV, is a vitamin K-dependent glycoprotein structurally similar to other vitamin K-dependent proteins affecting blood clotting, such as prothrombin, FVII, FIX and FX. The activation of protein C is strongly promoted by thrombomodulin and endothelial protein C receptor (EPCR), the latter of which is found primarily on endothelial cells (cells on the inside of blood vessels). The presence of thrombomodulin accelerates activation by several orders of magnitude, and EPCR speeds up activation by a factor of 20. On the endothelium, APC performs a major role in regulating blood clotting, inflammation, and cell death (apoptosis). APC proteolytically inactivates proteins FVa and FVIIIa. APC inactivates FVa by catayzing three cleavages (Arg³⁰⁶, Arg⁵⁰⁶, Arg⁶⁷⁹). The cleavages at both Arg³⁰⁶ and Arg⁵⁰⁶ diminish the molecule's affinity to FXa, and though the first of these sites is cleaved at the slowest rate, it is entirely necessary to the functioning of FV. Protein S aids this process by catalyzing the proteolysis at Arg³⁰⁶, in which the A2 domain of FV dissociates from the rest of the protein. Protein S also binds to FXa, inhibiting the latter from diminishing APC's inactivation of Factor Va. Protein S

Protein S is a vitamin K-dependent plasma glycoprotein with a molecular weight of approximately 70 kDa synthesized predominantly within the liver. However a significant amount is also synthesized in endothelial cells. Protein S is a cofactor for APC. Mature Protein S comprises 5 distinct structural domains, including an N-terminal gamma- carboxylation (Gla) domain and aromatic stack, a so-called "thrombin-sensitive region" , 4 EGF-like domains and EGF-4, and a large C-terminal region of 393 amino acids referred to as a sex-hormone binding globulin (SHBG)-like domain the structure of which represents two laminin G-type domains. The plasma concentration of Protein S is -350 nM and roughly 60% is bound to the complement 4 binding protein (C4b-BP), while the remaining fraction circulates as "free" Protein S. The complex bound Protein S has approximately 40% anti-coagulant activity compared to that of free Protein S. The half-life in plasma is 48-60 hours. Factor VIII

Factor VIII (FVIII) is a large, complex glycoprotein that is primarily produced by hepatocytes. FVIII consists of 2351 amino acids, including a signal peptide, and contains several distinct domains as defined by homology. There are three A-domains, a unique B- domain, and two C-domains. The domain order can be listed as NH2-A1 -A2-B-A3-C1 -C2- COOH. FVIII circulates in plasma as two chains, separated at the B-A3 border. The chains are connected by bivalent metal ion-bindings. The A1-A2-B chain is termed the heavy chain (HC) while the A3-C1 -C2 is termed the light chain (LC). Small acidic regions C-terminal of the A1 (the a1 region) and A2 (the a2 region) and N-terminal of the A3 domain (the a3 region) play important roles in its interaction with other coagulation proteins, including thrombin and von Willebrand factor (vWF or VWF), the carrier protein for FVIII.

Endogenous FVIII molecules circulate in vivo as a pool of molecules with B domains of various sizes, the shortest having C-terminal at position 740, i.e. at the C-terminal of A2- a2. These FVIII molecules with B-domains of different length all have full procoagulant activity. Upon activation with thrombin, FVIII is cleaved C-terminal of A1 -a1 at position 372, C-terminal of A2-a2 at position 740, and between a3 and A3 at position 1689, the latter cleavage releasing the a3 region with concomitant loss of affinity for VWF. The activated FVIII molecule is termed FVIIIa. The activation allows interaction of FVIIIa with phospholipid surfaces like activated platelets and activated factor IX (FIXa), i.e. the tenase complex is formed, allowing efficient activation of factor X (FX).

The B domain is cleaved at several different positions, generating large

heterogeneity in circulating plasma FVIII molecules. The exact function of the heavily glycosylated B domain is unknown. Thrombin and prothrombin

Thrombin, also known as activated blood-coagulation factor II, is a serine protease that in humans is encoded by the F2 gene.

Thrombin is produced by the enzymatic cleavage of two sites on prothrombin by activated Factor X (Xa). The activity of factor Xa is greatly enhanced by binding to activated Factor V (Va), termed the prothrombinase complex. Prothrombin is produced in the liver and is post-translationally modified in a vitamin K-dependent reaction that converts ten glutamic acids on prothrombin to gamma-carboxyglutamic acid (Gla). In the presence of calcium, the Gla residues promote the binding of prothrombin to phospholipid bilayers

Prothrombin (coagulation factor II) is proteolytically cleaved to form thrombin in the coagulation cascade, which ultimately results in the reduction of blood loss. Thrombin in turn acts as a serine protease that converts soluble fibrinogen into insoluble strands of fibrin, as well as catalyzing many other coagulation-related reactions. Protrombinase

The prothrombinase complex consists of the serine protease, Factor Xa, and the protein cofactor, Factor Va. The complex assembles on negatively charged phospholipid membranes in the presence of calcium ions. The prothrombinase complex catalyzes the conversion of prothrombin (Factor II), an inactive zymogen, to thrombin (Factor lla), an active serine protease. The activation of thrombin is a critical reaction in the coagulation cascade, which functions to regulate hemostasis in the body. To produce thrombin, the

prothrombinase complex cleaves two peptide bonds in prothrombin, one after Arg271 and the other after Arg320. Although it has been shown that Factor Xa can activate prothrombin when unassociated with the prothrombinase complex, the rate of thrombin formation is severely decreased under such circumstances. The prothrombinase complex can catalyze the activation of prothrombin at a rate 10⁵-fold faster than can Factor Xa alone. Thus, the prothrombinase complex is required for the efficient production of activated thrombin and also for adequate hemostasis.

SUMMARY

The present invention relates to antibodies or fragments thereof, that bind to activated Factor V (FVa). The invention describes a group of anti-FVa antibodies that display a pro-coagulant effect. The effect has been demonstrated in various setups to show that the antibodies are capable of increasing thrombin generation compared to set-ups where the antibody is not included. Different parameters in the thrombin generation test have been measured showing that antibodies of the invention increase or stimulate one or more of; peak thrombin, velocity index and/or endogenous thrombin potential.

One aspect of this invention relates to a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa, that when bound to FVa protects it from inactivation by activated protein C (APC).

In further embodiments the antibodies of the invention display the pro-coagulant effect also in the presence of activated protein C (APC).

In further embodiments the antibodies of the invention display the pro-coagulant effect in normal plasma.

In further embodiments the antibodies of the invention display the pro-coagulant effect in haemophilia plasma, such as in, haemophilia A plasma which is factor VIII deficient.

In further embodiments the antibodies of the invention display the pro-coagulant effect under haemophilia A like conditions, which may also include the ability to reverse the inhibitory effect of thrombomodulin on thrombin In further embodiments the antibodies of the invention display the pro-coagulant effect in haemophilic platelet poor plasma (PRP) in the presence of thrombomodulin.

In a further embodiment the antibodies of the invention display the pro-coagulant effect by stimulation of the thromboelastographic response, such as by increasing the elastic properties of blood during thrombus formation measured by thrombelastography using a TEG® hemostasis analyzer. In one embodiment the antibody dosage dependency decrease Clot time (R) in the absence of APC. In one embodiment the antibody dosage dependency decrease Clot time (R) in the presence of APC. In one embodiment the antibody dosage dependency increase MTG in the absence of APC. In one embodiment the antibody dosage dependency increase MTG in the presence of APC.

Another aspect of this invention relates to a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa, that when bound to FVa improves or stabilizes FVa co-factor activity to the enzyme FXa.

Another aspect of this invention relates to a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa that displays pro-coagulant effect in haemophilia.

One aspect of this invention regards a use of a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa as a pro-coagulant in prophylactic treatment of haemophilia.

Another aspect of this invention relates to the use of a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa, for the prophylactic treatment of haemophilia, such as haemophilia A and B and patients with inhibitors. One aspect of this invention relates to a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa for use in a method of treatment. Said treatment may be prophylactic treatment of haemophilia, such as haemophilia A. BRIEF DESCRIPTION OF THE SEQUENCE LISTING

SEQ ID NO: 1 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0161 monoclonal antibody. SEQ ID NO: 2 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0161 monoclonal antibody.

SEQ ID NO: 3 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0006 monoclonal antibody.

SEQ ID NO: 4 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0006 monoclonal antibody.

SEQ ID NO: 5 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0005 monoclonal antibody.

SEQ ID NO: 6 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0005 monoclonal antibody. SEQ ID NO: 7 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0028 monoclonal antibody.

SEQ ID NO: 8 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0028 monoclonal antibody.

SEQ ID NO: 9 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0128 monoclonal antibody. SEQ ID NO: 10 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0128 monoclonal antibody.

SEQ ID NO: 1 1 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0302 monoclonal antibody.

SEQ ID NO: 12 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0302 monoclonal antibody. SEQ ID NO: 13 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0337 monoclonal antibody.

SEQ ID NO: 14 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0337 monoclonal antibody.

SEQ ID NO: 15 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0293 monoclonal antibody.

SEQ ID NO: 16 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0293 monoclonal antibody.

SEQ ID NO: 17 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0327 monoclonal antibody. SEQ ID NO: 18 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0327 monoclonal antibody.

SEQ ID NO: 19 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0296 monoclonal antibody.

SEQ ID NO: 20 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0296 monoclonal antibody.

SEQ ID NO: 21 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0426 monoclonal antibody. SEQ ID NO: 22 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0426 monoclonal antibody. SEQ ID NO: 23 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-01 10 monoclonal antibody.

SEQ ID NO: 24 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-01 10 monoclonal antibody.

SEQ ID NO: 25 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0222 monoclonal antibody.

SEQ ID NO: 26 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0222 monoclonal antibody.

SEQ ID NO: 27 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0143 monoclonal antibody. SEQ ID NO: 28 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0143 monoclonal antibody.

SEQ ID NO: 29 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0271 monoclonal antibody.

SEQ ID NO: 30 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0271 monoclonal antibody.

SEQ ID NO: 31 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0263 monoclonal antibody.

SEQ ID NO: 32 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0263 monoclonal antibody. SEQ ID NO: 33 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0270 monoclonal antibody.

SEQ ID NO: 34 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0270 monoclonal antibody.

SEQ ID NO: 35 - gives the amino acid sequences of human coagulation factor V precursor y.

SEQ ID NO: 36 - gives the amino acid sequences of human coagulation factor V.

SEQ ID NO: 37 - gives the amino acid sequences of the heavy chain of activated human coagulation factor V.

SEQ ID NO: 38 - gives the amino acid sequences of the light chain of activated human coagulation factor V.

SEQ ID NO: 39 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-000-0008 monoclonal antibody. SEQ ID NO: 40 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-000-0008 monoclonal antibody.

SEQ ID NO: 41 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0418 monoclonal antibody.

SEQ ID NO: 42 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0418 monoclonal antibody.

SEQ ID NO: 43 - gives the amino acid sequences of the light chain variable domain (VL) of the NNC 0233-0000-0422 monoclonal antibody.

SEQ ID NO: 44 - gives the amino acid sequences of the heavy chain variable domain (VH) of the NNC 0233-0000-0422 monoclonal antibody. DESCRIPTION

Coagulopathy/Haemophilia and treatment Currently, the gold standard in treatment of haemophilia is prophylactic replacement therapy, wherein treatment has to be administered intravenously 2-3 times weekly causing a significant burden to the patient. Furthermore, approximately 30% of the patients treated with factor VIII develop inhibitors which reduce the possibilities for an effective prophylactic treatment.

Antibodies according to the present invention or a pharmaceutical formulation comprising such antibodies may be used to treat a subject with a coagulopathy.

The term "subject", as used herein, includes any human patient, or non-human vertebrate.

The term "coagulopathy", as used herein, refers to an increased haemorrhagic tendency which may be caused by any qualitative or quantitative deficiency of any pro- coagulative component of the normal coagulation cascade, or any up-regulation of fibrinolysis. Such coagulopathies may be congenital and/or acquired and/or iatrogenic and are identified by a person skilled in the art.

Non-limiting examples of congenital hypocoagulopathies are haemophilia A, haemophilia B, Factor VII deficiency, Factor X deficiency, Factor XI deficiency, von

Willebrand's disease and thrombocytopenias such as Glanzmann's thombasthenia and Bernard-Soulier syndrome. Said haemophilia A or B may be severe, moderate or mild. The clinical severity of haemophilia is determined by the concentration of functional units of FIX/FVI 11 in the blood and is classified as mild, moderate, or severe. Severe haemophilia is defined by a clotting factor level of <0.01 U/ml corresponding to <1 % of the normal level, while moderate and mild patients have levels from 1-5% and >5%, respectively. Haemophilia A with "inhibitors" (that is, neutralizing allo-antibodies against factor VIII) and haemophilia B with "inhibitors" (that is, neutralizing allo-antibodies against factor IX) are non-limiting examples of coagulopathies that are partly congenital and partly acquired.

The term "treatment", as used herein, refers to the medical therapy of any human or other animal subject in need thereof. Said subject is expected to have undergone physical examination by a medical practitioner, who has given a tentative or definitive diagnosis which would indicate that the use of said specific treatment is beneficial to the health of said human or other animal subject. The timing and purpose of said treatment may vary from one individual to another, according to the status quo of the subject's health. Thus, said treatment may be prophylactic, palliative and/or symptomatic. In terms of the present invention, prophylactic, palliative and symptomatic may represent separate aspects of the invention.

A non-limiting example of an acquired coagulopathy is serine protease deficiency caused by vitamin K deficiency; such vitamin K-deficiency may be caused by administration of a vitamin K antagonist, such as warfarin. Acquired coagulopathy may also occur following extensive trauma. In this case otherwise known as the "bloody vicious cycle", it is

characterised by haemodilution (dilutional thrombocytopaenia and dilution of clotting factors), hypothermia, consumption of clotting factors and metabolic derangements (acidosis). Fluid therapy and increased fibrinolysis may exacerbate this situation. Said haemorrhage may be from any part of the body.

A non-limiting example of an iatrogenic coagulopathy is an over dosage of anticoagulant medication - such as heparin, aspirin, warfarin and other platelet aggregation inhibitors - that may be prescribed to treat thromboembolic disease. A second, non-limiting example of iatrogenic coagulopathy is that which is induced by excessive and/or

inappropriate fluid therapy, such as that which may be induced by a blood transfusion.

In one embodiment of the current invention, haemorrhage is associated with haemophilia A or B. In another embodiment, haemorrhage is associated with haemophilia A or B with acquired inhibitors. In another embodiment, haemorrhage is associated with thrombocytopenia. In another embodiment, haemorrhage is associated with von Willebrand's disease. In another embodiment, haemorrhage is associated with severe tissue damage. In another embodiment, haemorrhage is associated with severe trauma. In another

embodiment, haemorrhage is associated with surgery. In another embodiment, haemorrhage is associated with haemorrhagic gastritis and/or enteritis. In another embodiment, the haemorrhage is profuse uterine bleeding, such as in placental abruption. In another embodiment, haemorrhage occurs in organs with a limited possibility for mechanical haemostasis, such as intracranially, intraaurally or intraocularly. In another embodiment, haemorrhage is associated with anticoagulant therapy.

Antibodies

The term "antibody" herein refers to a protein, derived from an immunoglobulin sequence, which is capable of specifically binding to an antigen or a portion thereof. The term antibody includes, but is not limited to, full length antibodies of any class (or isotype), that is, IgA, IgD, IgE, IgG, IgM, and/or IgY. The term may also include one or more antigen- binding fragments of full length antibodies. An antibody that specifically binds to an antigen, or a portion thereof, may bind exclusively to that antigen, or portion thereof, or it may bind to a limited number of homologous antigens, or portions thereof.

Natural full-length antibodies usually comprise at least four polypeptide chains: two heavy (H) chains and two light (L) chains that are connected by disulfide bonds. In some cases, natural antibodies comprise less than four chains, as in the case of the heavy chain only antibodies found in camelids (V_HH fragments) and the IgNARs found in Chondrichthyes. One class of immunoglobulins of particular pharmaceutical interest is the IgGs. In humans, the IgG class may be sub-divided into four sub-classes lgG1 , lgG2, lgG3 and lgG4, based on the sequence of their heavy chain constant regions. The light chains can be divided into two types, kappa and lambda chains based on differences in their sequence composition. IgG molecules are composed of two heavy chains, interlinked by two or more disulfide bonds, and two light chains, each attached to a heavy chain by a disulfide bond. An IgG heavy chain may comprise a heavy chain variable region (VH) and up to three heavy chain constant (CH) regions: CH1 , CH2 and CH3. A light chain may comprise a light chain variable region (VL) and a light chain constant region (CL). VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDRs) or hypervariable regions (HvRs), interspersed with regions that are more conserved, termed framework regions (FR). VH and VL regions are typically composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1 , CDR1 , FR2, CDR2, FR3, CDR3, FR4. The variable domains with the hypervariable regions of the heavy and light chains form a domain that is capable of interacting with an antigen, whilst the constant region of an antibody may mediate binding of the immunoglobulin to host tissues or factors, including, but not limited to various cells of the immune system (effector cells), Fc receptors and the first component (C1 q) of the C1 complex of the classical complement system.

Antibodies of the invention may be monoclonal antibodies, in the sense that they represent a set of unique heavy and light chain variable domain sequences as expressed from a single B-cell or by a clonal population of B cells. Antibodies of the invention may be produced and purified using various methods that are known to the person skilled in the art. For example, antibodies may be produced from hybridoma cells. Antibodies may be produced by B-cell expansion. Antibodies or fragments thereof may be recombinantly expressed in mammalian or microbial expression systems, or by in-vitro translation.

Antibodies or fragments thereof may also be recombinantly expressed as cell surface bound molecules, by means of e.g. phage display, bacterial display, yeast display, mammalian cell display or ribosome or mRNA display. Once produced, antibodies may be screened for binding to activated Factor V (FVa).

Antibodies of the current invention may be isolated. The term "isolated antibody" refers to an antibody that has been separated and/or recovered from (an)other component(s) in the environment in which it was produced and/or that has been purified from a mixture of components present in the environment in which it was produced.

Certain antigen-binding fragments of antibodies may be suitable in the context of the current invention, as it has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. The term "antigen-binding fragment" of an antibody refers to one or more fragment(s) of an antibody that retain(s) the ability to specifically bind to or recognise an antigen, such as activated Factor V (FVa) or another target molecule, as described herein. Examples of antigen-binding fragments include Fab, Fab', Fab₂, Fab'₂, FabS, Fv (typically the VL and VH domains of a single arm of an antibody), single-chain Fv (scFv); see e.g. Bird et al. Science 1988; 242:423-426; and Huston et al.

PNAS 1988; 85:5879-5883), dsFv, Fd (typically the VH and CH1 domain), and dAb (typically a VH domain) fragments; VH, VL, VhH, and V-NAR domains; monovalent molecules comprising a single VH and a single VL chain; minibodies, diabodies, triabodies, tetrabodies, and kappa bodies (see, e.g. Ill et al. Protein Eng 1997; 10:949-57); camel IgG; IgNAR; as well as one or more isolated CDRs or a functional paratope, where the isolated CDRs or antigen-binding residues or polypeptides can be associated or linked together so as to form a functional antibody fragment. Various types of antibody fragments have been described or reviewed in, e.g., Holliger and Hudson, Nat Biotechnol. 2005; 23:1 126-1 136;

WO2005040219, and published U.S. Patent Applications 20050238646 and 20020161201. These antibody fragments may be obtained using conventional techniques known to those of skill in the art, and the fragments may be screened for utility in the same manner as intact antibodies.

"Fab fragments" of an antibody, including "Fab" and "Fab'₂" fragments, can be derived from said antibody by cleavage of the heavy chain in the hinge region on the N- terminal or C-terminal side of the hinge cysteine residues connecting the heavy chains of the antibody. A "Fab" fragment includes the variable and constant domains of the light chain and the variable domain and the first constant domain (CH1 ) of the heavy chain. "Fab'₂" fragments comprise a pair of "Fab"' fragments that are generally covalently linked by their hinge cysteines. A Fab' is formally derived from a Fab'₂ fragment by cleavage of the hinge disulfide bonds connecting the heavy chains in the Fab'₂. Other chemical couplings than disulfide linkages of antibody fragments are also known in the art. A Fab fragment retains the ability of the parent antibody to bind to its antigen, potentially with a lower affinity. Fab'₂ fragments are capable of divalent binding, whereas Fab and Fab' fragments can bind monovalently. Generally, Fab fragments lack the constant CH2 and CH3 domains, i.e. the Fc part, where interaction with the Fc receptors would occur. Thus, Fab fragments are in general devoid of effector functions. Fab fragments may be produced by methods known in the art, either by enzymatic cleavage of an antibody, e.g. using papain to obtain the Fab or pepsin to obtain the Fab'₂, Fab fragments including Fab, Fab', Fab'₂ may be produced recombinantly using techniques that are well known to the person skilled in the art.

An "Fv" fragment is an antibody fragment that contains a complete antigen recognition and binding site, and generally comprises a dimer of one heavy and one light chain variable domain in association that can be covalent in nature, for example in a single chain variable domain fragment (scFv). It is in this configuration that the three hypervariable regions of each variable domain interact to define an antigen-binding site on the surface of the VH-VL dimer. Collectively, the six hypervariable regions or a subset thereof confer antigen binding specificity to the antibody. However, even a single variable domain comprising only three hypervariable regions specific for an antigen can retain the ability to recognize and bind antigen, although usually at a lower affinity than the entire binding site (Cai & Garen, PNAS 1996; 93:6280-6285). For example, naturally occurring camelid antibodies that only have a heavy chain variable domain (VHH) can bind antigen (Desmyter et al. J. Biol. Chem. 2002 277:23645-23650; Bond et al. J. Mol. Biol. 2003; 332:643-655).

"Single-chain Fv" or "scFv" antibody fragments comprise the VH and VL domains of antibody, where these domains are present in a single polypeptide chain. Generally, the Fv polypeptide further comprises a polypeptide linker between the VH and VL domains that enables the scFv to form the desired structure for antigen binding. For a review of scFv, see Pluckthun, 1994, In: The Pharmacology of Monoclonal Antibodies, Vol. 1 13, Rosenburg and Moore eds. Springer-Verlag, New York, pp. 269-315.

The term "diabodies" refers to small antibody fragments with two antigen-binding sites, in which fragments comprise a heavy chain variable domain (VH) connected to a light chain variable domain (VL) in the same polypeptide chain (VH and VL). By using a linker that is too short to allow pairing between the two variable domains on the same chain, the variable domains are forced to pair with complementary domains of another chain, creating two antigen-binding sites. Diabodies are described more fully, for example, in EP 404,097; WO 93/1 1 161 ; and Hollinger et al. PNAS 1993; 90:6444-6448. The expression "linear antibodies" refers to antibodies as described in Zapata et al. Protein Eng. 1995; 8(10): 1057-1062. Briefly, these antibodies contain a pair of tandem Fd segments (VH-CH1 -VH-CH1 ) that, together with complementary light chain polypeptides, form a pair of antigen binding regions. Linear antibodies can be bispecific or monospecific.

The term "monobody" as used herein, refers to an antigen binding molecule with a heavy chain variable domain and no light chain variable domain. A monobody can bind to an antigen in the absence of light chains and typically has three hypervariable regions, for example CDRs designated CDRH 1 , CDRH2, and CDRH3. A heavy chain IgG monobody has two heavy chain antigen binding molecules connected by a disulfide bond. The heavy chain variable domain comprises one or more hypervariable regions, preferably a CDRH3 or HVL- H3 region.

Antibody fragments may be obtained using conventional recombinant or protein engineering techniques and the fragments can be screened for binding to factor V or activated factor V, or another function, in the same manner as intact antibodies.

Antibody fragments of the invention may be made by truncation, e.g. by removal of one or more amino acids from the N and/or C-terminal ends of a polypeptide. Fragments may also be generated by one or more internal deletions.

An antibody of the invention may be, or may comprise, an antigen binding portion of one of these antibodies, or variants thereof. For example, the antibody of the invention may be a Fab fragment of one of these antibodies or variants thereof, or it may be a single chain antibody derived from one of these antibodies, or a variant thereof.

The term "binding affinity" is herein used as a measure of the strength of a non- covalent interaction between two molecules, e.g. an antibody, or fragment thereof, and an antigen. The term "binding affinity" is used to describe monovalent interactions (intrinsic activity).

Binding affinity between two molecules, e.g. an antibody, or fragment thereof, and an antigen, through a monovalent interaction may be quantified by determining the equilibrium dissociation constant (KD). In turn, KD can be determined by measurement of the kinetics of complex formation and dissociation, e.g. by the SPR method. The rate constants corresponding to the association and the dissociation of a monovalent complex are referred to as the association rate constant k_a (or k_on) and dissociation rate constant k_d (or k_off), respectively. K_D is related to ka and kd through the equation K_D = k_d / k_a.

Following the above definition, binding affinities associated with different molecular interactions, such as comparison of the binding affinity of different antibodies for a given antigen, may be compared by comparison of the KD values for the individual antibody/antigen complexes.

An antibody according to the current invention may be able to compete with another molecule, such as a naturally occurring ligand or receptor or another antibody, for binding to FVa or FV. Therefore, an antibody according to the current invention may be able to bind FVa or FV with a greater affinity that that of another molecule also capable of binding FVa or FV. The ability of an antibody to compete with a natural ligand/receptor for binding to an antigen may be assessed by determining and comparing the KD value for the interactions of interest, such as a specific interaction between an antibody and an antigen, with that of the KD value of an interaction not of interest. Typically, the KD for the antibody with respect to the target will be 2-fold, preferably 5-fold, and more preferably 10-fold less than KD with respect to the other, non-target molecule such as unrelated material or accompanying material in the environment. More preferably, the KD will be 50-fold less, such as 100-fold less, or 200-fold less; even more preferably 500-fold less, such as 1 ,000-fold less, or 10,000- fold less. The value of this dissociation constant can be determined directly by well-known methods. Standard assays to evaluate the binding ability of ligands such as antibodies towards targets are known in the art and include, for example, ELISA, Western blot, RIA, and flow cytometry analysis. The binding kinetics and binding affinity of the antibody also can be assessed by standard assays known in the art, such as SPR.

A competitive binding assay can be conducted in which the binding of the antibody to the target is compared to the binding of the target by another ligand of that target, such as another antibody.

An antibody of the invention may have a KD for its target of 1 x 10^"7M or less, 1 x 10^" ⁸M or less, or 1 x 10^"9M or less, or 1 x 10^"10M or less, 1 x 10^"11M or less, or 1 x 10^"12M or less. The KD of an antibody of the current invention may be less than 5 nM, such as less than 4 nM, such as less than 3 nM, such as less than 2 nM, such as less than 1 nM, such as less than 0.8 nM, such as less than 0.7 nM, such as less than 0.6 nM, such as less than 0.5 nM, such as less than 0.4 nM, such as less than 0.3 nM, such as less than 0.2 nM, such as less than 0.1 nM, such as less than 0.05 nM, such as less than 0.025 nM, such as less than 0.015 nM, such as between 0.015 nM and 0 nM.

An antibody of the invention may be a human antibody or a humanised antibody. The term "human antibody", as used herein, is intended to include antibodies having variable regions in which at least a portion of a framework region and/or at least a portion of a CDR region are derived from human germline immunoglobulin sequences. (For example, a human antibody may have variable regions in which both the framework and CDR regions are derived from human germline immunoglobulin sequences.) Furthermore, if the antibody contains a constant region, the constant region or a portion thereof is also derived from human germline immunoglobulin sequences. The human antibodies of the invention may include amino acid residues not encoded by human germline immunoglobulin sequences {e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo).

Such a human antibody may be a human monoclonal antibody. Such a human monoclonal antibody may be produced by a hybridoma which includes a B cell obtained from a transgenic nonhuman animal, e.g., a transgenic mouse, having a genome comprising human immunoglobulin heavy and light chain gene segments repertoires, fused to an immortalised cell.

Human antibodies may be isolated from sequence libraries built on selections of human germline sequences, further diversified with natural and synthetic sequence diversity.

Human antibodies may be prepared by in vitro immunisation of human lymphocytes followed by transformation of the lymphocytes with Epstein-Barr virus.

The term "human antibody derivative" refers to any modified form of the human antibody, such as a conjugate of the antibody and another agent or antibody.

The term "humanised antibody", as used herein, refers to a human/non-human chimeric antibody that contains a sequence (CDR regions or parts thereof) derived from a non-human immunoglobulin. A humanised antibody is, thus, a human immunoglobulin

(recipient antibody) in which at least residues from a hypervariable region of the recipient are replaced by residues from a hypervariable region of an antibody from a non-human species (donor antibody) such as from a mouse, rat, rabbit or non-human primate, which have the desired specificity, affinity, sequence composition and functionality. In some instances, framework (FR) residues of the human immunoglobulin are replaced by corresponding non- human residues. An example of such a modification is the introduction of one or more so- called back-mutations, which are typically amino acid residues derived from the donor antibody. Humanisation of an antibody may be carried out using recombinant techniques known to the person skilled in the art (see, e.g., Antibody Engineering, Methods in Molecular Biology, vol. 248, edited by Benny K. Lo). A suitable human recipient framework for both the light and heavy chain variable domain may be identified by, for example, sequence or structural homology. Alternatively, fixed recipient frameworks may be used, e.g., based on knowledge of structure, biophysical and biochemical properties. The recipient frameworks can be germline derived or derived from a mature antibody sequence. CDR regions from the donor antibody can be transferred by CDR grafting. The CDR grafted humanised antibody can be further optimised for e.g. affinity, functionality and biophysical properties by identification of critical framework positions where re-introduction (back mutation) of the amino acid residue from the donor antibody has beneficial impact on the properties of the humanised antibody. In addition to donor antibody derived back mutations, the humanised antibody can be engineered by introduction of germline residues in the CDR or framework regions, elimination of immunogenic epitopes, site-directed mutagenesis, affinity maturation, etc.

Furthermore, humanised antibodies may comprise residues that are not found in the recipient antibody or in the donor antibody. These modifications are made to further refine antibody performance. In general, a humanised antibody will comprise at least one - typically two - variable domains, in which all or substantially all of the CDR regions correspond to those of a non-human immunoglobulin and in which all or substantially all of the FR residues are those of a human immunoglobulin sequence. The humanised antibody can, optionally, also comprise at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin.

The term "humanised antibody derivative" refers to any modified form of the humanised antibody, such as a conjugate of the antibody and another chemical agent or antibody or antibody fragment or polypeptide.

The term "chimeric antibody", as used herein, refers to an antibody whose light and heavy chain genes have been constructed, typically by genetic engineering, from

immunoglobulin variable and constant region genes that originate from different species. For example, the variable segments of genes from a mouse monoclonal antibody may be joined to human constant regions.

The fragment crystallisable region ("Fc region'V'Fc domain") of an antibody is the C- terminal region of an antibody, which comprises the constant CH2 and CH3 domains. The Fc domain may interact with cell surface receptors called Fc receptors, as well as some proteins of the complement system. The Fc region enables antibodies to interact with the immune system. In one aspect of the invention, antibodies may be engineered to include

modifications within the Fc region, typically to alter one or more of its functional properties, such as plasma half-life, complement fixation, Fc-receptor binding, protein stability and/or antigen-dependent cellular cytotoxicity, or lack thereof, among others. Furthermore, an antibody of the invention may be chemically modified (e.g., one or more chemical moieties can be attached to the antibody) or be modified to alter its glycosylation, again to alter one or more functional properties of the antibody. An lgG1 antibody may carry a modified Fc domain comprising one or more, and perhaps all of the following mutations that will result in decreased affinity to certain Fc receptors (L234A, L235E, and G237A) and in reduced C1 q- mediated complement fixation (A330S and P331 S), respectively (residue numbering according to the EU index).

The isotype of an antibody of the invention may be IgG, such as lgG1 , such as lgG2, such as lgG4. If desired, the class of an antibody may be "switched" by known techniques. For example, an antibody that was originally produced as an IgM molecule may be class switched to an IgG antibody. Class switching techniques also may be used to convert one IgG subclass to another, for example: from lgG1 to lgG2 or lgG4; from lgG2 to lgG1 or lgG4; or from lgG4 to lgG1 or lgG2. Engineering of antibodies to generate constant region chimeric molecules, by combination of regions from different IgG subclasses, can also be performed.

In one embodiment, the hinge region of CH1 is modified such that the number of cysteine residues in the hinge region is altered, e.g., increased or decreased. This approach is described further for instance in U.S. Patent No. 5,677,425 by Bodmer et al.

The constant region may be modified to stabilise the antibody, e.g., to reduce the risk of a bivalent antibody separating into two monovalent VH-VL fragments. For example, in an lgG4 constant region, residue S228 (according to the EU numbering index and S241 according to Kabat) may be mutated to a proline (P) residue to stabilise inter heavy chain disulphide bridge formation at the hinge (see, e.g., Angal et al. Mol Immunol. 1993; 30:105- 8).

Antibodies or fragments thereof may be defined in terms of their complementarity- determining regions (CDRs). The term "complementarity-determining region" or

"hypervariable region", when used herein, refers to the regions of an antibody in which amino acid residues involved in antigen-binding are situated. The region of hypervariability or CDRs can be identified as the regions with the highest variability in amino acid alignments of antibody variable domains. Databases can be used for CDR identification such as the Kabat database, the CDRs e.g. being defined as comprising amino acid residues 24-34 (L1 ), 50-56 (L2) and 89-97 (L3) of the light-chain variable domain and 31 -35 (H1 ), 50-65 (H2) and 95- 102 (H3) in the heavy-chain variable domain; (Kabat et al. 1991 ; Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242) Alternatively CDRs can be defined as those residues from a "hypervariable loop" (residues 26-33 (L1 ), 50-52 (L2) and 91-96 (L3) in the light-chain variable domain and 26-32 (H1 ), 53-55 (H2) and 96-101 (H3) in the heavy-chain variable domain; Chothia and Lesk, J. Mol. Biol. 1987; 196:901 -917). Typically, the numbering of amino acid residues in this region is performed by the method described in Kabat et al. supra. Phrases such as "Kabat position", "Kabat residue", and "according to Kabat" herein refer to this numbering system for heavy chain variable domains or light chain variable domains. Using the Kabat numbering system, the actual linear amino acid sequence of a peptide may contain fewer or additional amino acids corresponding to a shortening of, or insertion into, a framework (FR) or CDR of the variable domain. For example, a heavy chain variable domain may include amino acid insertions (residue 52a, 52b and 52c according to Kabat) after residue 52 of CDR H2 and inserted residues (e.g. residues 82a, 82b, and 82c, etc. according to Kabat) after heavy chain FR residue 82. The Kabat numbering of residues may be determined for a given antibody by alignment at regions of homology of the sequence of the antibody with a "standard" Kabat numbered sequence.

The term "framework region" or "FR" residues refer to those VH or VL amino acid residues that are not within the CDRs, as defined herein.

The term "antigen" (Ag) refers to the molecular entity used for immunisation of an immunocompetent vertebrate to produce the antibody (Ab) that recognizes the Ag. Herein, Ag is termed more broadly and is generally intended to include target molecules that are specifically recognized by the Ab, thus including fragments or mimics of the molecule used in the immunisation process, or other process, e.g. phage display, used for generating the Ab.

The term "epitope", as used herein, is defined in the context of a molecular interaction between an "antigen binding polypeptide", such as an antibody (Ab), and its corresponding antigen (Ag). Generally, "epitope" refers to the area or region on an Ag to which an Ab specifically binds, i.e. the area or region in physical contact with the Ab.

Physical contact may be defined using various criteria (e.g. a distance cut-off of 2-6A, such as 3A, such as 4 A, such as 5A; or solvent accessibility) for atoms in the Ab and Ag molecules. A protein epitope may comprise amino acid residues in the Ag that are directly involved in binding to a Ab (also called the immunodominant component of the epitope) and other amino acid residues, which are not directly involved in binding, such as amino acid residues of the Ag which are effectively blocked by the Ab, i.e. amino acid residues within the "solvent-excluded surface" and/or the "footprint" of the Ab.

The term epitope herein comprises both types of binding region in any particular region of FVa that specifically binds to an anti-FVa antibody, or another FVa specific agent according to the invention, unless otherwise stated. FVa may comprise a number of different epitopes, which may include, without limitation, (1 ) linear peptide epitopes (2) conformational epitopes which consist of one or more non-contiguous amino acids located near each other in the mature FVa conformation; and (3) post-translational epitopes which consist, either in whole or part, of molecular structures covalently attached to Fva, such as carbohydrate groups.

The epitope for a given antibody (Ab)/antigen (Ag) pair can be described and characterized at different levels of detail using a variety of experimental and computational epitope mapping methods. The experimental methods include mutagenesis, X-ray crystallography, Nuclear Magnetic Resonance (NMR) spectroscopy, Hydrogen deuterium exchange Mass Spectrometry (HX-MS) and various competition binding methods; methods that are known in the art. As each method relies on a unique principle, the description of an epitope is intimately linked to the method by which it has been determined. Thus, depending on the epitope mapping method employed, the epitope for a given Ab/Ag pair may be described differently.

At its most detailed level, the epitope for the interaction between the Ag and the Ab can be described by the spatial coordinates defining the atomic contacts present in the Ag- Ab interaction, as well as information about their relative contributions to the binding thermodynamics. At a less detailed level, the epitope can be characterized by the spatial coordinates defining the atomic contacts between the Ag and Ab. At an even less detailed level the epitope can be characterized by the amino acid residues that it comprises as defined by a specific criteria such as the distance between or solvent accessibility of atoms in the Ab:Ag complex. At a further less detailed level the epitope can be characterized through function, e.g. by competition binding with other Abs. The epitope can also be defined more generically as comprising amino acid residues for which substitution by another amino acid will alter the characteristics of the interaction between the Ab and Ag.

In the context of an X-ray derived crystal structure defined by spatial coordinates of a complex between an Ab, e.g. a Fab fragment, and its Ag, the term epitope is herein, unless otherwise specified or contradicted by context, specifically defined as activated Factor V (FVa) residues characterized by having a heavy atom (i.e. a non-hydrogen atom) within a distance of 4 A, from a heavy atom in the Ab.

The definition of the term "paratope" is derived from the above definition of "epitope" by reversing the perspective. Thus, the term "paratope" refers to the area or region on the Ab to which an Ag specifically binds, i.e. with which it makes physical contact to the Ag.

In the context of an X-ray derived crystal structure, defined by spatial coordinates of a complex between an Ab, such as a Fab fragment, and its Ag, the term paratope is herein, unless otherwise specified or contradicted by context, specifically defined as Ag residues characterized by having a heavy atom (i.e. a non-hydrogen atom) within a distance of 4 A from a heavy atom in activated Factor V (FVa). The epitope and paratope for a given antibody (Ab)/antigen (Ag) pair may be identified by routine methods. For example, the general location of an epitope may be determined by assessing the ability of an antibody to bind to different fragments or variant FVa polypeptides. The specific amino acids within FVa that make contact with an antibody (epitope) and the specific amino acids in an antibody that make contact with FVa (paratope) may also be determined using routine methods. For example, the antibody and target molecule may be combined and the Ab:Ag complex may be crystallised. The crystal structure of the complex may be determined and used to identify specific sites of interaction between the antibody and its target.

Antibodies that bind to the same antigen can be characterised with respect to their ability to bind to their common antigen simultaneously and may be subjected to "competition binding'Vbinning". In the present context, the term "binning" refers to a method of grouping antibodies that bind to the same antigen. "Binning" of antibodies may be based on competition binding of two antibodies to their common antigen in assays based on standard techniques such as surface plasmon resonance (SPR), ELISA or flow cytometry.

An antibody's "bin" is defined using a reference antibody. If a second antibody is unable to bind to an antigen at the same time as the reference antibody, the second antibody is said to belong to the same "bin" as the reference antibody. In this case, the reference and the second antibody competitively bind the same part of an antigen and are coined

"competing antibodies". If a second antibody is capable of binding to an antigen at the same time as the reference antibody, the second antibody is said to belong to a separate "bin". In this case, the reference and the second antibody do not competitively bind the same part of an antigen and are coined "non-competing antibodies".

Antibody "binning" does not provide direct information about the epitope. Competing antibodies, i.e. antibodies belonging to the same "bin" may have identical epitopes, overlapping epitopes or even separate epitopes. The latter is the case if the reference antibody bound to its epitope on the antigen takes up the space required for the second antibody to contact its epitope on the antigen ("steric hindrance"). Non-competing antibodies generally have separate epitopes.

Improved utilization of endogenous FV

The problem addressed in the present invention relates to methods for improved utilization of the endogenous FV pool by a method of increasing the level and/or improving the co-factor activity of FVa by a monoclonal antibody and/or fragments hereof. Furthermore the problem addressed in the present invention relates to methods for improved utilization of the endogenous FV pool by a method of increasing the level and/or improving the co-factor activity of FVa by a monoclonal antibody binding to FV and FVa and/or fragments hereof. Thus, the present invention relates to antibodies that bind to Factor Va. In particular, the invention relates to antibodies that modulate the function of FVa. In particular, the invention relates to antibodies capable of improving the co-cofactor function of FVa. A key functionality of FVa is in the generation of thrombin. As mentioned above FVa function is to stimulate thrombin generation working as a cofactor for FXa in the

prothrombinase and as described in the Examples herein anti-FVa antibodies capable of increasing thrombin generation has been identified.

In particular, the invention relates to monoclonal antibodies and/or fragments thereof capable of reducing inactivation of FVa.

The invention also relates to uses for such antibodies and fragments thereof, such as therapeutic and pharmaceutical uses.

An antibody according to the invention against FVa and/or FV may offer a prophylactic treatment option for haemophilia patients with inhibitors. The present invention also provides a method for treatment of haemophilia patients in a FVIII and FIX independent manner. It has been found that monoclonal antibodies of the invention raised against human FVa and/or FV significantly improve the thrombin generation in haemophilic plasma as the addition of the antibody increases thrombin generation compared to the assay performed in the absence of the antibody e.g. the pro-coagulant effect of the antibody is to stimulate thrombin generation beyond the thrombin generation of FVa alone (e.g. in the absence of antibody).

For example, an antibody of the invention may have the ability to improve the peak thrombin generation in human FVII l-deficient plasma (see example 3) or to reduce time to clot as measured in a thromboelastography (TEG) analysis of human FVII l-deficient whole blood (see example 4). Furthermore, the antibodies may provide a prophylactic treatment for patients with haemophilia. Anti FVa Antibodies

The present invention relates to antibodies or fragments thereof, that bind to activated Factor V (FVa) and/or Factor V also refered to as anti-activated Factor V (FVa) and/or anti FV antibodies.

The present invention relates to antibodies, or fragments thereof, that bind to Factor V and activated Factor V (FVa). The invention describes anti-FV and/or anti-FVa antibodies that display a pro-coagulant effect and in particular antibodies that bind both Factor V and activated Factor V that display a pro-coagulant effect. The effect has been demonstrated in various setups to show that the antibodies are capable of increasing thrombin generation. Different parameters have been measured showing that antibodies of the invention increase one or more of peak thrombin, velocity index and/or endogenous thrombin potential. Likewise the thromboelastographic response has been measured by thrombelastography using a TEG® hemostasis analyzer.

In one embodiment the anti-FVa antibody or an antigen-binding fragment thereof increases thrombin generation in the presence of APC, such as in the presence of exogenously added APC in a human plasma-based thrombin generation assay.

In a further embodiment the FVa antibody has an EC₅₀ below 150 nM in the thrombin generation assay employing haemophilia A plasma and added APC as described in Example 2 herein. In one embodiment the EC50 is below 125 nM, such as below 100 nM, such as below 75 nM such as below 50 nM, such as below 40 nM.

In one embodiment the FVa antibody has EC₅₀ equal to or below the EC₅₀ for the 01 10 antibody (defined by SEQ ID NO 23 and 24) in the thrombin generation assay employing haemophilia A plasma and added APC as described in Example 2 herein. Equal here means that the EC50 ratio 01 10/FVa antibody is around 1 , such as 0.5-1 . 5, 0.6-1.4, 0.7-1.3, 0.8-1.2 or 0.9-1.1 .

In a further embodiment the peak height, in the thrombin generation assay employing haemophilia A plasma and added APC as described in Example 2 herein, is equal to the peak height of the 01 10 antibody. In one embodiment the peak height ratio for an FVa antibody/ the 01 10 antibody is at least 0.5, such as 0.6, such as 0.7 or such as 0.8.

In a series of further embodiments the antibodies of the invention display the pro- coagulant effect under haemophilia A like conditions such as in the presence of an anti-FVIII antibody. In one embodiment ability of the antibody to increase thrombin generation is measured by an ability to increase one or more of peak thrombin, the velocity index and/or endogenous thrombin potential as described in example 3 herein. The effect may be compared to the effect of Factor VIII.

In one embodiment the peak thrombin, the velocity index and/or endogenous thrombin potential is higher for 90 nM antibody than 10 % FVIII either in the presence or absence of APC.

The effect (peak thrombin, the velocity index and/or endogenous thrombin potential) may be compared to one or more of the antibodies of the present invention. In one embodiment the effect is equal to or higher than the effect of one or more of antibody 0005, 0028, 0159, 0056, 0008, 0150, 0161 and 01 10. In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) is equal to or higher than the effect of antibody 0005, 0028, 0159, 0056, 0008, 0150, 0161 or 01 10.

In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) is equal to or better than the effect of antibody 0005.

In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) is equal to or better than the effect of antibody 0028.

In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) is equal to or better than the effect of antibody 0159.

In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) ise equal to or better than the effect of antibody 0056.

In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) is equal to or better than the effect of antibody 0008.

In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) is equal to or better than the effect of antibody fragment 0150.

In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) is equal to or better than the effect of antibody 0161 .

In one embodiment the effect (one or more of peak thrombin, the velocity index and/or endogenous thrombin potential) is equal to or better than the effect of antibody 01 10.

Reference is made to tables 2-1 1 where the three parameters have been tested under various conditions such as in the presence of APC and different concentrations hereof, such as with different concentrations of antibody, such as in the presence or absence of thrombo-modulin. Based on table 5, the following embodiments can be derived covering antibodies with functionalities similar to 0005, 0028 and/or 01 10.

In one embodiment Peak Thrombin in haemophilia A patient plasma in the presence of APC (5 nM) as measured after Thrombinoscope PPP triggering of thrombin generation is at least 10 nM when using 100 nM anti-FVa antibody. In one further embodiment Peak Thrombin is at least 25 nM, such as at least 50nM, such as at least 75 nM when using 100 nM anti-FVa antibody. In one further embodiment Peak Thrombin is at least 40 when using 500 nM anti-FVa antibody. In one further embodiment Peak Thrombin is at leastl OO, such as at least 125 when using 500 nM anti-FVa antibody.

Based on table 6, the following embodiments can be derived covering antibodies with functionalities similar to 0005, 0028 and/or 01 10.

In one embodiment the Velocity Index, in haemophilia A patient plasma in the presence of APC (5 nM) as measured after Thrombinoscope PPP triggering of thrombin generation, is at least 1 .5 nM/min when using 20 nM anti-FVa antibody. In one further embodiment the Velocity Index is at least 2.5 nM/min, such as at least 15 nM/min, when using 50 nM anti-FVa antibody. In one further embodiment the Velocity Index is at least 3.5 nM/min, such as at least 10 nM/min, such as at least 25 nM/min when using 100 nM anti-FVa antibody. In one further embodiment the Velocity Index is at least 4 nM/min, such as at least 8, such as at least 50 nM/min when using 500 nM anti-FVa antibody.

Similar embodiment can be derived from other tables, such as an anti-FVa antibody with Velocity Index is at least 10 nM/min, such as at least 20, such as at least 25 nM/min or such as at least 40 nM/min when using 500 nM anti-FVa antibody in haemophilia A patient plasma in the presence of APC (5 nM) and measured after Thrombinoscope PPP triggering of thrombin generation which is based on table 8 and the data obtained for antibodies 0159, 0056, 0008, 01 10 (and Fab 0150) and 0161.

In one embodiment the antibody stimulates the thromboelastographic response, such as by increasing the elastic properties of blood during thrombus formation measured by thrombelastography using a TEG® hemostasis analyzer.

The effect of the antibody may be measured as the clot time or maximum rate of Thrombus Generation (MTG) measuring the elastic properties of blood during thrombus formation as described in Example 4 herein. In one embodiment the antibody dose dependency reduce clotting time. In one embodiment a similar effect is observed in the presence of APC. In a further embodiment the antibody is capable of reducing clotting time equal to recombinant FVIIa. In one embodiment the relative Clot time (R) is dosage dependency reduced by the anti-FVa antibody. It may be observed that the antibody clot time is increased by a low concentration of the antibody while increasing dosage reduces the clot time in a dosage dependent fashion.

In one embodiment the antibody increase the Maximum rate of Thrombus

Generation (MTG). In one embodiment the antibody dosage dependency increases MTG. In one such embodiment a similar effect is observed in the absence of APC. In one such further embodiment a similar effect is observed in the presence of APC.

In further embodiments the antibodies of the invention display the pro-coagulant effect under haemophilia A like conditions, which may also include the ability to reverse the inhibitory effect of thrombomodulin on thrombin generation. In further embodiments the antibodies of the invention display the pro-coagulant effect in haemophilic platelet rich plasma (PRP) in the presence of thrombomodulin. In a further embodiment the FVa antibody of the invention has a peak height above 20 % of the peak height for the 01 10 antibody in a thrombin generation assay in platelet rich plasma under haemophilia A like conditions in the presence of thrombomodulin as described in Example 5 herein.

In further such embodiments the peak height is above 20 % of the 01 10 antibody when the antibody concentration is 500 nM. In even further embodiments the peak height is above 40 %, such as above 60 % of the peak height for the 01 10 antibody when the antibody concentration is or 500 nM.

In further embodiments the peak height is above 20 % of the peak height for the 01 10 antibody when the antibody concentration is or 50 nM. In even further embodiments the peak height is above 40 %, such as above 60 % of the peak height for the 01 10 antibody when the antibody concentration is or 50 nM.

The examples, tables and embodiments herein together present various parameters that enable the persons skilled in the art to determine if an antibody with functionalities similar to the antibodies of the present invention has been identified.

One aspect of this invention regards a use of a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa (as a pro-coagulant) in prophylactic treatment of haemophilia. Another aspect of this invention relates to the use of a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa, for the prophylactic treatment of haemophilia.

One aspect of this invention relates to a monoclonal antibody or a fragment of said monoclonal antibody that binds to FVa for use in a method of treatment. Said treatment may be prophylactic treatment of haemophilia, such as haemophilia A.

As described the examples herein a series of antibodies binding FVa capable of enhancing thrombin generation in vitro has been identified.

The antibodies identified binds to various regions of FV, and the invention thus includes anti-FVa antibodies or an antigen-binding fragments thereof that binds to a) the A3 domain of the FVa light chain, b) the C2 domain of the FVa light chain, c) the A1 domain the FVa heavy chain or d) the A2 domain of the FVa heavy chain.

A series of the identified antibodies have been sequence and are defined in the sequence listing. The antibodies of the invention include one or more of an anti-FVa antibody or antigen-binding fragment thereof, wherein the CDRs of the light and heavy chain variable regions are as in SEQ ID NO 1 and SEQ ID NO 2, SEQ ID NO 3 and SEQ ID NO 4, SEQ ID NO 5 and SEQ ID NO 6, SEQ ID NO 7 and SEQ ID NO 8, SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 1 1 and SEQ ID NO 12, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 17 and SEQ ID NO 18, SEQ ID NO 19 and SEQ ID NO 20, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24,SEQ ID NO 25 and SEQ ID NO 26, SEQ ID NO 27 and SEQ ID NO 28, SEQ ID NO 29 and SEQ ID NO 30, SEQ ID NO 31 and SEQ ID NO 32, SEQ ID NO 33 and SEQ ID NO 34, SEQ ID NO 39 and SEQ ID NO 40, SEQ ID NO 41 and SEQ ID NO 42 or SEQ ID NO 43 and SEQ ID NO 44.

In one embodiment the anti-FVa antibody or antigen-binding fragment thereof, comprises the CDRs of the light and heavy chain variable regions are as in the light chain and heavy chains set fort in SEQ ID NO 1 and SEQ ID NO 2, SEQ ID NO 7 and SEQ ID NO 8, SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 17 and SEQ ID NO 18, SEQ ID NO 19 and SEQ ID NO 20, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24,SEQ ID NO 25 and SEQ ID NO 26, SEQ ID NO 27 and SEQ ID NO 28, SEQ ID NO 29 and SEQ ID NO 30, SEQ ID NO 31 and SEQ ID NO 32, SEQ ID NO 33 and SEQ ID NO 34, SEQ ID NO 39 and SEQ ID NO 40, SEQ ID NO 41 and SEQ ID NO 42 or SEQ ID NO 43 and SEQ ID NO 44.

In one embodiment the anti-FVa antibody or antigen-binding fragment thereof, comprises the CDRs of the light and heavy chain variable regions are as in the light chain and heavy chains set fort in SEQ ID NO 5 and SEQ ID NO 6, SEQ ID NO 7 and SEQ ID NO 8, SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 17 and SEQ ID NO 18, SEQ ID NO 19 and SEQ ID NO 20, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24,SEQ ID NO 25 and SEQ ID NO 26, SEQ ID NO 27 and SEQ ID NO 28, SEQ ID NO 29 and SEQ ID NO 30, SEQ ID NO 31 and SEQ ID NO 32, SEQ ID NO 33 and SEQ ID NO 34, SEQ ID NO 39 and SEQ ID NO 40 or SEQ ID NO 41 and SEQ ID NO 42.

In one embodiment the anti-FVa antibody or antigen-binding fragment thereof, comprises the CDRs of the light and heavy chain variable regions are as in the light chain and heavy chains set fort in SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 17 and SEQ ID NO 18, SEQ ID NO 19 and SEQ ID NO 20, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24,SEQ ID NO 25 and SEQ ID NO 26, SEQ ID NO 27 and SEQ ID NO 28, SEQ ID NO 29 and SEQ ID NO 30, SEQ ID NO 31 and SEQ ID NO 32, SEQ ID NO 33 and SEQ ID NO 34, SEQ ID NO 39 and SEQ ID NO 40 or SEQ ID NO 41 and SEQ ID NO 42.

In one embodiment the anti-FVa antibody or antigen-binding fragment thereof, comprises the CDRs of the light and heavy chain variable regions are as in the light chain and heavy chains set fort in SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 17 and SEQ ID NO 18, SEQ ID NO 19 and SEQ ID NO 20, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24,SEQ ID NO 25 and SEQ ID NO 26, SEQ ID NO 27 and SEQ ID NO 28, SEQ ID NO 29 and SEQ ID NO 30, SEQ ID NO 31 and SEQ ID NO 32 or SEQ ID NO 41 and SEQ or ID NO 42.

In one embodiment the anti-FVa antibody or antigen-binding fragment thereof, comprises the CDRs of the light and heavy chain variable regions are as in the light chain and heavy chains set fort in SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 17 and SEQ ID NO 18, SEQ ID NO 19 and SEQ ID NO 20, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24, SEQ ID NO 29 and SEQ ID NO 30, SEQ ID NO 31 and SEQ ID NO 32 or SEQ ID NO 41 and SEQ or ID NO 42.

In one embodiment the anti-FVa antibody or antigen-binding fragment thereof, comprises the CDRs of the light and heavy chain variable regions are as in the light chain and heavy chains set fort in SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 17 and SEQ ID NO 18, SEQ ID NO 19 and SEQ ID NO 20, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24 or SEQ ID NO 41 and SEQ or ID NO 42.

In one embodiment the anti-FVa antibody or antigen-binding fragment thereof, comprises the CDRs of the light and heavy chain variable regions are as in the light chain and heavy chains set fort in SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24 or SEQ ID NO 41 and SEQ or ID NO 42.

In one embodiment the anti-FVa antibody or antigen-binding fragment thereof, comprises the CDRs of the light and heavy chain variable regions are as in the light chain and heavy chains set fort in SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24 or SEQ ID NO 41 and SEQ ID NO 42.

Antibodies of the invention include the human monoclonal antibodies 0233-0000- 0302, 0233-0000-0337, 0233-0000-0293, 0233-0000-0327, 0233-0000-0296, 0233-0000- 0418 and 0233-0000-0426. It further includes the preferred murine antibodies 0233-0000- 0028 and 0233-0000-0128. And current invention further includes the preferred rabbit antibodies 0233-0000-01 10, 0233-0000-0271 and 0233-0000-0270. Heavy and light CDRs of these antibodies are provided below and in tables 28-30.

An antibody of the invention may be, or may comprise, a fragment of the 0233-0000-

0008 antibody, the 0233-0000-0028 antibody or the 0233-0000-01 10 antibody, or a variant of any one of these antibodies.

In another embodiment of the invention, the antibody of the invention may be, or may comprise, a fragment of the 0233-0000-01 10 antibody, the 0233-0000-0128 antibody or the 0233-0000-0270 antibody, or a variant of any one of these antibodies.

In a further embodiment of the invention, the antibody of the invention may be, or may comprise, a fragment of the 0233-0000-0271 antibody, the 0233-0000-0293 antibody or the 0233-0000-0296 antibody, or a variant of any one of these antibodies. In yet another embodiment of the invention, the antibody of the invention may be, or may comprise, a fragment of the 0233-0000-0302 antibody, the 0233-0000-0327 antibody or the 0233-0000-0337 antibody, or a variant of any one of these antibodies.

In yet another embodiment of the invention, the antibody of the invention may be, or may comprise, a fragment of the 0233-0000-0418 antibody, the 0233-0000-0422 antibody or the 0233-0000-0426 antibody, or a variant of any one of these antibodies.

In a further embodiment of the invention, the antibody of the invention may be, or may comprise, a fragment of the 0233-0000-0005 antibody or the 0233-0000-0006 antibody, or a variant of any one of these antibodies.

An antibody of the invention may comprise a CDR region from one or more of the specific antibodies disclosed herein.

The light chain variable region (SEQ ID NO:1 1 ) of human monoclonal antibody 0233-0000-0302 contains the following CDRs; LC-CDR1 : "RASQDISHWLA" corresponding to Kabat residues 24-34 of SEQ ID NO 1 1 ; LC-CDR2: " IASTLQT" corresponding to Kabat residues 50-56 of SEQ ID NO 1 1 and LC-CDR3 " QQSNSFPLT" corresponding to Kabat residues (89-97) of SEQ ID NO 1 1. The heavy chain variable region (SEQ ID NO: 12) of human monoclonal antibody 0233-0000-0302 contains the following CDRs; HC-CDR1 "DYAMH" corresponding to Kabat residues 31-35 of SEQ ID NO 12; HC-CDR2

"GISWNSGAIGYADSVKG" corresponding to Kabat residues 50-66 of SEQ ID NO: 12 and HC-CDR3 "DA R WLV E E D YQYYG L D V" corresponding to Kabat residues 99-1 15 of SEQ ID NO:12. The light chain variable region (SEQ ID NO:13) of human monoclonal antibody

0233-0000-0337 contains the following CDRs: LC-CDR1 "RASQDISNWLA" corresponding to Kabat residues 24-34 of SEQ ID NO: 13, LC-CDR2 "ITSTLHI" corresponding to Kabat residues 50-56 of SEQ ID NO: 13, LC-CDR3 "QQANSFPFT" corresponding to Kabat residues 89-97 of SEQ ID NO:13. The heavy chain variable region (SEQ ID NO:14) of human monoclonal antibody 0233-0000-0337 contains the following CDRs: HC-CDR1 "DYAMH" corresponding to Kabat residues 31-35 of SEQ ID NO:14, HC-CDR2

"GISWNSGSTGYADSVQG" corresponding to Kabat residues 50-66 of SEQ ID NO: 14 and HC-CDR3 " D AR WLVE E D YYYYG M D V" corresponding to Kabat residues 90-1 15 of SEQ ID NO:14. The light chain variable region (SEQ ID NO:15) of human monoclonal antibody 0233-0000-0293 contains the following CDRs: LC-CDR1 " KSSQSVLYSSN N KN YLA" corresponding to Kabat residues 24-40 of SEQ ID NO:15, LC-CDR2 "WASTRES" corresponding to Kabat residues 56-62 of SEQ ID NO:15 and LC-CDR3 "QQYYSTPWT" corresponding to Kabat residues 95-103 of SEQ ID NO: 15. The heavy chain variable region (SEQ ID NO: 16) of human monoclonal antibody 0233-0000-0293 contains the following CDRs: HC-CDR1 "SYDIN" corresponding to Kabat residues 31 -35 of SEQ ID NO:16; HC- CDR2 "WMNPNSGNTGYALKFQG" corresponding to Kabat residues 50-66 of SEQ ID NO: 16 and HC-CDR3 "RTYYDILTGSLGAFDI" corresponding to Kabat residues 99-1 14 of SEQ ID NO:16.

The light chain variable region (SEQ ID NO:17) of human monoclonal antibody 0233-0000-0327 contains the following CDRs: LC-CDR1 "RASQDISSWLA" corresponding to Kabat residues 24-34 of SEQ ID NO:17; LC-CDR2 "lASSLQS" corresponding to Kabat residues 50-56 of SEQ ID NO: 17 and LC-CDR3 "QQANSFPFT" corresponding to Kabat residues 89-97 of SEQ ID NO:17. The heavy chain variable region (SEQ ID NO:18) of the human monoclonal antibody 0233-0000-0327 contains the following CDRs: HC-CDR1 "DYAMH" corresponding to Kabat residues 31-35 of SEQ ID NO:18, HC-CDR2

"GVSWNSGAIGYADSVKG" corresponding to Kabat residues 50-66 of SEQ ID NO:18 and HC-CDR3 "DARWLVEEDYQYYGMDV" corresponding to Kabat residues 99-1 15 of SEQ ID NO:18.

The light chain variable region (SEQ ID NO:19) of the monoclonal human antibody 0233-0000-0296 contains the following CDRs: LC-CDR1 "RASQDISTWLA" corresponding to Kabat residues 24-34 of SEQ ID NO:19, LC-CDR2 "ITSTLHI" corresponding to Kabat residues 50-56 of SEQ ID NO:19 and LC-CDR3 "QQAYSFPFT" corresponding to Kabat residues 89-98 of SEQ ID NO:19. The heavy chain variable region (SEQ ID NO:20) of the human monoclonal antibody 0233-0000-0296 contains the following CDRs: HC-CDR1 "DYAMH" corresponding to Kabat residues 31-35 of SEQ ID NO:20, HC-CDR2

"GISWNSGGIGYADSVQG" corresponding to Kabat residues 50-66 of SEQ ID NO:20 and HC-CDR3 " D AR WLVE E D YYYYG M D V" corresponding to Kabat residues 99-1 15 of SEQ ID NO:20.

The light chain variable region (SEQ ID NO:21 ) of the monoclonal human antibody 0233-0000-0426 contains the following CDRs: LC-CDR1 "SGDILGDKYAC" corresponding to Kabat residues 23-33 of SEQ ID NO:21 , LC-CDR2 "QDIKRPS" corresponding to Kabat residues 49-55 of SEQ ID NO:21 and LC-CDR3 "QAWDSTTPVV" corresponding to Kabat residues 88-97 of SEQ ID NO:21 . The heavy chain variable region (SEQ ID NO:22) of the monoclonal antibody 0233-0000-0426 contains the following CDRs: HC-CDR1 "SYDIN" corresponding to Kabat residues 31-35 of SEQ ID NO:22, HC-CDR2

"WMNPNTDDTGYAQKFQG" corresponding to Kabat residues 50-66 of SEQ ID NO:22 and HC-CDR3 Ύ WS VTS W KW N D D H YYYYG M D V" corresponding to Kabat residues 99-1 19 of SEQ ID NO:22. The light chain variable region (SEQ ID NO:41 ) of the monoclonal human antibody

0233-0000-0418 contains the following CDRs: LC-CDR1 "RSSQSLLDSDDGNTYMD" corresponding to Kabat residues 24-40 of SEQ ID NO:41 , LC-CDR2 "MGFYRAS" corresponding to Kabat residues 56-62)of SEQ ID NO:41 and LC-CDR3 "MQRIEFPST" corresponding to Kabat residues 95-103 of SEQ ID NO:41 . The heavy chain variable region (SEQ ID NO:42) of the monoclonal antibody 0233-0000-0418 contains the following CDRs: HC-CDR1 "TSGVGVG" corresponding to Kabat residues 31-37 of SEQ ID NO:42, HC-CDR2 "LIYWDDVKRYSPSLRR" corresponding to Kabat residues 52-67 of SEQ ID NO:42 and HC- CDR3 "YNWKMRVD" corresponding to Kabat residues 100-107 of SEQ ID NO:42. The light chain variable region (SEQ ID NO:7) of the monoclonal murine antibody

0233-0000-0028 contains the following CDRs: LC-CDR1 "KASQDVGTAVG" corresponding to Kabat residues 24-34 of SEQ ID NO:7, LC-CDR2 "WASTRHT" corresponding to Kabat residues 50-56 of SEQ ID NO:7 and LC-CDR3 "QQYSSNPT" corresponding to Kabat residues 89-96 of SEQ ID NO:7. The heavy chain variable region (SEQ ID NO:8) of the monoclonal murine antibody 0233-0000-0028 contains the following CDRs: HC-CDR1 "NYGMN" corresponding to Kabat residues 31-35 of SEQ ID NO:8, HC-CDR2

"WINTYTGEPTYADDFKG" corresponding to Kabat residues 50-66 of SEQ ID NO:8 and HC- CDR3 "S LRRN PYYYAM DY" corresponding to Kabat residues 99-1 1 1 of SEQ ID NO:8. The light chain variable region (SEQ ID NO:9) of the monoclonal murine antibody

0233-0000-0128 contains the following CDRs: LC-CDR1 "KTSTDIDDDMN" corresponding to Kabat residues 24-34 of SEQ ID NO:9, LC-CDR2 "EGNTLRP" corresponding to Kabat residues 50-56 of SEQ ID NO:9 and LC-CDR3 "LQSANMPFT" corresponding to Kabat residues 89-97 of SEQ ID NO:9. The heavy chain variable region (SEQ ID NO:10) of the monoclonal murine antibody 0233-0000-0128 contains the following CDRs: HC-CDR1 "SYAMS" corresponding to Kabat residues 31 -35 of SEQ ID NO: 10, HC-CDR2 "TISSGGSYTYYPDSVKG" corresponding to Kabat residues 50-66 of SEQ ID NO: 10 and HC-CDR3 " G P Y LTTAT PS FTY" corresponding to Kabat residues 99-1 1 1 of SEQ ID NO: 10. The light chain variable region (SEQ ID NO:23) of rabbit monoclonal antibody 0233-

0000-01 10 contains the following CDRs: LC-CDR1 "QASESISSYLT" corresponding to Kabat residues 24-34 of SEQ ID NO:23, LC-CDR2 "YASTLAS" corresponding to Kabat residues 50-56 of SEQ ID NO:23 and LC-CDR3 "LGVYSYSRDDGIA" corresponding to Kabat residues 89-101 of SEQ ID NO:23. The heavy chain variable region (SEQ ID NO:24) of rabbit monoclonal antibody 0233-0000-01 10 contains the following CDRs: HC-CDR1 "SSYYMC" corresponding to Kabat residues 31-36 of SEQ ID NO:24, HC-CDR2

"CIYTAWDGASYANWAKG" corresponding to Kabat residues 51-67 of SEQ ID NO:24 and HC-CDR3 "AMGSSDGANNL" corresponding to Kabat residues 99-109 of SEQ ID NO:24. The light chain variable region (SEQ ID NO:29) of rabbit monoclonal antibody 0233-

0000-0271 contains the following CDRs: LC-CDR1 "QASQSIGGNLA" corresponding to Kabat residues 24-34 of SEQ ID NO:29, LC-CDR2 "DASKLAS" corresponding to Kabat residues 50-56 of SEQ ID NO:29 and LC-CDR "QCTYGSSGNIGNG" corresponding to Kabat residues 89-101 of SEQ ID NO:29. The heavy chain variable region (SEQ ID NO:30) of rabbit monoclonal antibody 0233-0000-0271 contains the following CDRs: HC-CDR1 "SYAMI" corresponding to Kabat residues 30-34 of SEQ ID NO:30, HC-CDR2

"Fl DTGGSAYYASWAKG" corresponding to Kabat residues 49-64 of SEQ ID NO:30 and HC- CDR3 "A L YVYS D VYTA F N I " corresponding to Kabat residues 95-108 of SEQ ID NO:30. The light chain variable region (SEQ ID NO:33) of rabbit monoclonal antibody 0233-

0000-0270 contains the following CDRs: LC-CDR1 "QASQSISSYLS" corresponding to Kabat residues 24-34 of SEQ ID NO:33, LC-CDR2 "RTSTLES" corresponding to Kabat residues 50-56 of SEQ ID NO:33 and LC-CDR3 "RTSTLES" corresponding to Kabat residues 89-102 of SEQ ID NO:33. The heavy chain variable region (SEQ ID NO:34) of rabbit monoclonal antibody 0233-0000-0270 contains the following CDRs: HC-CDR1 "SYYHIC" corresponding to Kabat residues 30-35 of SEQ ID NO:34, HC-CDR2 "C I YAAS G DTWYATWVN A" corresponding to Kabat residues 50-66 of SEQ ID NO:34 and HC-CDR3

"GPRYVSSSGAGPYCLDL" corresponding to Kabat residues 98-1 14 of SEQ ID NO:34. Thus in one embodiment of this invention, the antibody comprises the light chain variable region of SEQ ID NO:1 1 and/ or the heavy chain variable region of SEQ ID NO:12 or fragments thereof. In another embodiment of present invention, the antibody comprises light chain CDR1 with residues 24-34 of SEQ ID NO 1 1 , and/or light chain CDR2: comprising residues 50-56 of SEQ ID NO 1 1 and/or light chain CDR comprising residues (89-97) of SEQ ID NO 1 1 and/or heavy chain CDR1 comprising residues 31-35 of SEQ ID NO 12 and/or HC- CDR2 comprising residues 50-66 of SEQ ID NO: 12 and HC-CDR3 comprising residues 99- 1 15 of SEQ ID NO:12. In an further embodiment of this invention, the antibody comprises the light chain variable region of SEQ ID NO:13 and/or heavy chain variable region SEQ ID NO:14, or fragments thereof. In another embodiment of the present invention, the antibody comprises light chain CDR1 with residues 24-34 of SEQ ID NO:13 and/or LC-CDR2 with residues 50-56 of SEQ ID NO:13 and/or LC-CDR3 with residues 89-97 of SEQ ID NO:13 and/or heavy chain CDR1 comprising residues 31-35 of SEQ ID NO:14 and/or HC-CDR2 "comprising residues 50-66 of SEQ ID NO:14 and/or HC-CDR3 comprising residues 90-1 15 of SEQ ID NO:14.

In a further embodiment of the present invention, the antibody comprises the light chain variable region (SEQ ID NO:15) and/or heavy chain variable region (SEQ ID NO:16) or fragments thereof. In another embodiment the antibody comprises light chain CDR1 comprising residues 24-40 of SEQ ID NO: 15 and/or LC-CDR2 comprising residues 56-62 of SEQ ID NO:15 and/or LC-CDR3 comprising 95-103 of SEQ ID NO:15 and/or heavy chain CDR1 comprising residues 31-35 of SEQ ID NO:16 and/or HC-CDR2 comprising residues 50-66 of SEQ ID NO:16 and/or HC-CDR3 comprising residues 99-1 14 of SEQ ID NO:16.

In yet another embodiment of the current invention, the antibody comprises light chain variable region of SEQ ID NO: 17 and/or heavy chain variable region of SEQ ID NO: 18 or fragment thereof. In another embodiment the antibody comprises light chain CDR1 comprising residues 24-34 of SEQ ID NO: 17 and/or LC-CDR2 comprising residues 50-56 of SEQ ID NO:17 and/or LC-CDR3 comprising residues 89-97 of SEQ ID NO:17 and/or heavy chain CDR1 comprising residues 31-35 of SEQ ID NO:18 and/or HC-CDR2 comprising residues 50-66 of SEQ ID NO:18 and/or HC-CDR3 comprising residues 99-1 15 of SEQ ID NO:18.

In other embodiment of the current invention, the antibody comprises the light chain variable region of(SEQ ID NO: 19 and/or the heavy chain variable region of SEQ ID NO:20 or a fragment thereof. In another embodiment, the antibody comprises a light chain CDR1 comprising residues 24-34 of SEQ ID NO: 19 and/or LC-CDR2 comprising residues 50-56 of SEQ ID NO:19 and/or LC-CDR3 comprising residues 89-98 of SEQ ID NO:19 and/or heavy chain CDR1 comprising residues 31-35 of SEQ ID NO:20 and/or HC-CDR2 comprising residues 50-66 of SEQ ID NO:20 and/or HC-CDR3 comprising residues 99-1 15 of SEQ ID NO:20.

In further embodiment of the current invention, the antibody comprises the light chain variable region SEQ ID NO:21 and/or heavy chain variable region SEQ ID NO:22, or fragment thereof. In another embodiment the antibody comprises light chain CDR1 comprising residues 23-33 of SEQ ID NO:21 and/or LC-CDR2 comprising residues 49-55 of SEQ ID NO:21 and/or LC-CDR3 comprising residues 88-97 of SEQ ID NO:21 and/or HC- CDR1 comprising residues 31-35 of SEQ ID NO:22 and/or HC-CDR2 comprising residues 50-66 of SEQ ID NO:22 and/or HC-CDR3 comprising residues 99-1 19 of SEQ ID NO:22. In one embodiment of the present invention, the antibody comprises a light chain variable region of SEQ ID NO:41 and/or heavy chain variable region of SEQ ID NO:42, or a fragment thereof. In further embodiment the antibody comprises a LC-CDR1 comprising residues 24-40 of SEQ ID NO:41 and/or LC-CDR2 comprising 56-62 of SEQ ID NO:41 and/or LC-CDR3 comprising residues 95-103 of SEQ ID NO:41 and/or heavy chain CDR1 comprising residues 31 -37 of SEQ ID NO:42 and/or HC-CDR2 comprising residues 52-67 of SEQ ID NO:42 and/or HC-CDR3 comprising residues 100-107 of SEQ ID NO 42.

In another embodiment of the current invention, the antibody comprises light chain variable region SEQ ID NO:7 and/or the heavy chain variable region of SEQ ID NO:8, or fragment thereof. In further embodiment the antibody comprises light chain CDR1 comprising residues 24-34 of SEQ ID NO:7 and/or LC-CDR2 comprising residues 50-56 of SEQ ID NO:7 and/or LC-CDR3 comprising residues 89-96 of SEQ ID NO:7 and/or heavy chain CDR1 comprising residues 31 -35 of SEQ ID NO:8 and/or HC-CDR2 comprising residues 50-66 of SEQ ID NO:8 and/or HC-CDR3 comprising residues 99-1 1 1 of SEQ ID NO:8.

In further embodiment of the present invention the antibody comprises light chain variable region of SEQ ID NO:9 and/or heavy chain variable region of SEQ ID NO:10. In other embodiment the antibody comprises light chain CDR1 comprising residues 24-34 of SEQ ID NO:9 and/or LC-CDR2 comprising residues 50-56 of SEQ ID NO:9 and/or LC-CDR3 residues 89-97 of SEQ ID NO:9 and/or heavy chain CDR1.comprising residues 31-35 of SEQ ID NO:10 and/or HC-CDR2 comprising residues 50-66 of SEQ ID NO:10 and/or HC- CDR3 comprising residues 99-1 1 1 of SEQ ID NO:10.

In one embodiment of the current invention, the antibody comprises the light chain variable region of SEQ ID NO:23 and/or heavy chain variable region of SEQ ID NO:24, or fragment thereof. In other embodiment the antibody comprises a light chain CDR1 comprising residues 24-34 of SEQ ID NO:23 and/or LC-CDR2 comprising 50-56 of SEQ ID NO:23 and/or LC-CDR3 comprising residues 89-101 of SEQ ID NO:23 and/or HC-CDR1 comprising residues 31 -36 of SEQ ID NO:24 and/or HC-CDR2 comprising residues 51 -67 of SEQ ID NO:24 and/or HC-CDR3 comprising residues 99-109 of SEQ ID NO:24.

In further embodiment of the current invention, the antibody comprises the light chain variable region of SEQ ID NO:29 and/or heavy chain variable region of SEQ ID NO:30, or a fragment thereof. In another embodiment the antibody comprises a light chain CDR1 comprising residues 24-34 of SEQ ID NO:29 and/or LC-CDR2 comprising residues 50-56 of SEQ ID NO:29 and/or LC-CDR comprising residues 89-101 of SEQ ID NO:29 and/or HC- CDR1 comprising residues 30-34 of SEQ ID NO:30 and/or HC-CDR2 comprising residues 49-64 of SEQ ID NO:30 and/or HC-CDR3 comprising residues 95-108 of SEQ ID NO:30. In one embodiment of the present invention, the antibody comprises a light chain variable region of SEQ ID NO:33 and/or heavy chain variable region of SEQ ID NO:34, or a fragment thereof. In further embodiment the antibody comprises a LC-CDR1 comprising residues 24-34 of SEQ ID NO:33 and/or LC-CDR2 comprising 50-56 of SEQ ID NO:33 and/or LC-CDR3 comprising residues 89-102 of SEQ ID NO:33 and/or heavy chain CDR1 comprising residues 30-35 of SEQ ID NO:34 and/or HC-CDR2 comprising residues 50-66 of SEQ ID NO:34 and/or HC-CDR3 comprising residues 98-1 14 of SEQ ID NO:34.

In other embodiments, the VH and/or VL amino acid sequences may be 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the sequences set forth above.

From the fact that descriptions and definitions of epitopes, dependant on the epitope mapping method used, are obtained at different levels of detail, it follows that comparison of epitopes for different Abs on the same Ag can similarly be conducted at different levels of detail. Epitopes described at the amino acid level, e.g. determined from an X-ray structure, are said to be identical if they contain the same set of amino acid residues. Epitopes are said to overlap if at least one amino acid is shared by the epitopes. Epitopes are said to be separate (unique) if no amino acid residue is shared by the epitopes. In the absence of X-ray structures the binding regions of antibodies can be described using other technologies aimed at clarifying regions of an antigen that interacts with the antigen binding region. Hydrogen exchange mass spectrometry (HX-MS) identifies regions of the antigen "protected" by the binding of the antibody or antigen binding region thereof. Protected regions are thus an estimate of the epitope as the binding of the antibody or antigen binding region thereof reduces the hydrogen exchange of the bound regions.

As can be seen in example 8 herein pro-coagulant anti-FVa antibodies of the inventions have been found to bind to various regions of the activated FV molecule. In one embodiment the antibodies bind the activated human FV molecule.

An aspect of the invention relates to a monoclonal anti-FVa antibody or antigen- binding fragment thereof that displays a pro-coagulant effect, wherein the anti-FVa antibody or antigen-binding fragment thereof binds to one or more of

a) the A3 domain of the FVa light chain,

b) the C2 domain of the FVa light chain,

c) the A1 domain the FVa heavy chain and/or

d) the A2 domain of the FVa heavy chain.

In one embodiment the antibody or fragment thereof binds one or more of aa K34- I54 and aa I 164-Q182 in the A1 domain of human FVa.

In one embodiment the antibody or fragment thereof binds one or more of aa 485 to 493, 532-542, aa 553-554, aa 577-580 and 588-600 in the C2 domain of human FVa. In one embodiment the antibody or fragment thereof binds two of aa 485 to 493, 532-542, aa 553- 554, aa 577-580 and 588-600 in the C2 domain of human FVa. In one embodiment the antibody or fragment thereof binds three of aa 485 to 493, 532-542, aa 553-554, aa 577-580 and 588-600 in the C2 domain of human FVa. In one embodiment the antibody or fragment thereof binds to aa 532-542, aa 553-554, aa 577-580 and 588-600 in the C2 domain of human FVa. In one embodiment the antibody or fragment thereof binds all of aa 485 to 493, 532-542, aa 553-554, aa 577-580 and 588-600 in the C2 domain of human FVa. In one embodiment the antibody or fragment thereof binds to one or more regions including aa E330-V331 and N534-C539 in the A2 domain. In one embodiment the antibody or fragment thereof binds to at least two regions including aa E330-V331 and N534-C539 in the A2 domain.

In one embodiment the antibody or fragment thereof binds one or more of aa F325- V331 and N534-C539 in the A2 domain of human FVa.

In one embodiment the antibody or fragment thereof binds one or more of aa E330- Y335, S356-S376 and N534-F552 in the A2 domain of human FVa.

In one embodiment the antibody or fragment thereof binds two of aa E330-Y335,

S356-S376 and N534-F552 in the A2 domain of human FVa

In one embodiment the antibody or fragment thereof binds to all of aa E330-Y335, S356-S376 and N534-F552 in the A2 domain of human FVa

In one embodiment the antibody or fragment thereof binds to both the A2, and A3 domains, such as aa E330-Y335, S356-S376 and N534-F552 in the A2 domain of human FVa and Y211-M226 in the A3.

In one embodiment the antibody or fragment thereof binds to both the A2, A3 and C2 domains, such as aa E330-Y335, S356-S376 and N534-F552 in the A2 domain of human FVa and Y211-M226 in the A3 and aa F514-L530, aa D553-L554 and aa L555-S579 of the C2 domain.

In one embodiment the antibody or fragment thereof binds aa Y211-M226 in the A3 domain of human FVa. In further embodiment the pro-coagulant anti-FVa antibody or antigen-binding fragment competes with a reference antibody for binding to human FVa. Surface Plasmon resonance can be used to determine if an antibody in question can bind in the presence of another antibody. If both antibodies can bind the antibodies does not compete, whereas only one of two competing antibodies can bind at the same time.

In on embodiment the antibody or antigen binding fragment competes with an antibody disclosed herein as defined in table 27 and 28.

In one embodiment of the current invention, the antibody competes in binding to FVa with antibody 0233-0000-01 10 or a fragment thereof (such as 0233-0000-0150). In another embodiment of the invention, the antibody competes in binding to FVa with antibody 0233-0000-0028. In a further embodiment the antibody competes in binding to FVa with antibody 0233-0000-0128. In other embodiment the antibody competes in binding to FVa with antibody 0233-0000-0159. In other embodiment the antibody competes in binding to FVa with antibody 0233-0000-418.

In one embodiment the invention relates to a monoclonal anti-FVa antibody or antigen-binding fragment thereof that displays a pro-coagulant effect, wherein the anti-FVa antibody or antigen-binding fragment thereof binds to

a) the A3 domain of the FVa light chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:41 and a variable heavy chain amino acid sequence of SEQ ID NO:42, b) the A3 domain of the FVa light chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:29 and a variable heavy chain amino acid sequence of SEQ ID NO:30, c) the C2 domain of the FVa light chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:23, and a variable heavy chain amino acid sequence of SEQ ID NO:24, d) the C2 domain of the FVa light chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:1 1 , and a variable heavy chain amino acid sequence of SEQ ID NO:12, e) the C2 domain of the FVa light chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:3, and a variable heavy chain amino acid sequence of SEQ ID NO:4, f) the C2 domain of the FVa light chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:29 and a variable heavy chain amino acid sequence of SEQ ID NO:30, g) the A1 domain the FVa heavy chain competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:5, and a variable heavy chain amino acid sequence of SEQ ID NO:6, h) the A2 domain of the FVa heavy chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:29 and a variable heavy chain amino acid sequence of SEQ ID NO:30, i) the A2 domain of the FVa heavy chain competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:9, and a variable heavy chain amino acid sequence of SEQ ID NO:10 or, j) the A2 domain of the FVa heavy chain competes with a reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID

NO:29, and a variable heavy chain amino acid sequence of SEQ ID NO:30.

In one embodiment of current invention, the epitope of the antibody comprises residues of the heavy chain of FVa (SEQ ID NO:37) within the sequence from K34 to I54. In one embodiment the antibody or antibody fragment binds K34 to I54 of the heavy chain of FVa.

Thus in an embodiment of the current invention, the epitope of the antibody comprises one or more of the residues K34, K35, I36, V37, Y38, R39, E40, Y41 , E42, P43, Y44, F45, K46, K47, E48, K49, P50, Q51 , S52, T53 and I54,

In another embodiment the epitope of the antibody comprises residues on the heavy chain of FVa (SEQ ID NO:37) within the sequence from 1164 to Q182. In one embodiment the antibody or antibody fragment binds 1164 to Q182 of the heavy chain of FVa.

Thus in an embodiment of the current invention, the epitope of the antibody comprises one or more of the residues 1164, C165, K166, K167, G168, T169, L170, T171 , E172, G173, G174, T175, Q176, K177, T178, F179, D180, K181 and/or Q182 (SEQ ID NO:37).

In a further embodiment the epitope of the antibody comprises residues of the heavy chain of FVa (SEQ ID NO:37) within the sequence from K34 to I54 and residues on the heavy chain of FVa (SEQ ID NO:37) within the sequence from 1164 to Q182. In a further embodiment the antibody or antibody fragment binds K34 to I54 and 1164 to Q182 and of the heavy chain of FVa.

Thus in an embodiment of the current invention, the epitope of the antibody comprises one or more of the residues K34, K35, I36, V37, Y38, R39, E40, Y41 , E42, P43, Y44, F45, K46, K47, E48, K49, P50, Q51 , S52, T53, I54 and one or more of the residues 1164, C165, K166, K167, G168, T169, L170, T171 , E172, G173, G174, T175, Q176, K177, T178, F179, D180, K181 and/or Q182 (SEQ ID NO:37).

In one embodiment the epitope of an antibody comprises residues of the heavy chain of FVa (SEQ ID NO:37) within the sequence from F325 to V331 . In one embodiment the antibody or antibody fragment binds F325 to V331 of the heavy chain of FVa.

In one embodiment the epitope of an antibody comprises residues of the heavy chain of FVa (SEQ ID NO:37) within the sequence from N534 to C539. In one embodiment the antibody or antibody fragment binds N534 to C539 of the heavy chain of FVa.

Thus in an embodiment of the current invention, the epitope of the antibody comprises one or more of the residues F325, I326, A327, A328, E329, E330, V331 and one or more of N534, I535, N536, K537, F538, and C539. In one embodiment the epitope of an antibody comprises residues of the heavy chain of FVa (SEQ ID NO:37) within the sequence from E330 to Y335

In one embodiment the epitope of an antibody comprises residues of the heavy chain of FVa (SEQ ID NO:37) within the sequence from S356 to S376.

In one embodiment the epitope of an antibody comprises residues of the heavy chain of FVa (SEQ ID NO:37) within the sequence from N534 to F552.

In one embodiment the antibody binds FVa within one or more of the sequences from E330 to Y335, S356 to S376 and N534 to F552. In one embodiment the antibody binds FVa within two of the sequences from E330 to Y335, S356 to S376 and N534 to F552. In one embodiment the antibody binds FVa within the three sequences from E330 to Y335, S356 to S376 and N534 to F552.

The epitope of the antibody may thus comprise residues within the regions E330 to Y335, S356 to S376, and N534 to F552 of the FVa heavy-chain sequence and those residues are E330, V331 , I332, W333, D334, Y335 and S356, N357, Q358, I359, G360, K361 , H362, Y363, K364, K365, V366, M367, Y368, T369, Q370, Y371 , E372, D373, E374, S375, S376 and N534, I535, N536, K537, F538, C539, E540, N541 , P542, D543, E544, V545, K546, R547, D548, D549, P550, K551 , F552.

In one embodiment the epitope of an antibody comprises residues of the light chain of FVa (SEQ ID NO:38) within the sequence from F514 to L530, In one embodiment the epitope of an antibody comprises residues of the light chain of FVa (SEQ ID NO:38) within the sequence from D553 to L554

In one embodiment the epitope of an antibody comprises residues of the light chain of FVa (SEQ ID NO:38) within the sequence from L555 to S579

In one embodiment the epitope of an antibody comprises residues of the light chain of FVa (SEQ ID NO:38) within the sequence from F514 to L530 from D553 to L554 and from L555 to S579.

In one embodiment of the current invention, the epitope of the antibody comprises residues on the light chain of FVa (SEQ ID NO:38) within the sequence from A532 to L554 (SEQ ID NO:38).

In another embodiment the epitope of the antibody comprises residues on the light chain of FVa (SEQ ID NO:38) within the sequence from Y577 to Y580.

In further embodiment the epitope of the antibody comprises residues on the light chain of FVa (SEQ ID NO:38) within the sequence from G588 to M600.

In one embodiment the antibody or antibody fragment binds one or more of N532 to L554, A532 to L554 and/or Y577 to Y580 of the heavy chain of FVa.

Thus in an embodiment of the current invention, the epitope of the antibody comprises one or more of the residues A532, Q533, G534, R535, V536, N537, A538, W539, Q540, A541 , K542, A543, N544, N545, N546, K547, Q548, W549, L550, E551 , I552, D553, L554 and/or one or more of the residues K478, S479, YS480 and/or one or more of the residues G588, V589, E590, W591 , K592, P593, Y594, R595, L596, K597, S598, S599, and M600 (SEQ ID NO:38).

In one embodiment of the current invention, the epitope of the antibody comprises residues on the light chain of FVa (SEQ ID NO:38), within the sequence from N532 to A543. In one embodiment of the current invention, the epitope of the antibody comprises residues on the light chain of FVa (SEQ ID NO:38) within the sequence from A532 to L554 (SEQ ID NO:38). In another embodiment the epitope of the antibody comprises residues on the light chain of FVa (SEQ ID NO:38) within the sequence from D553 to L554. In a further embodiment the epitope of the antibody comprises residues on the light chain of FVa (SEQ ID NO:38) within the sequence from G588 to M600.

Thus in an embodiment of the invention, the epitope of the antibody one or more of the residues A532, Q533, G534, R535, V536, N537, A538, W539, Q540, A541 , K542 and/or A543 and one or more of the residues D553 and/or L554 and/or one or more of the residues G588, V589, E590, W591 , K592, P593, Y594, R595, L596, K597, S598, S599 and/or M600.

In one embodiment the epitope of an antibody comprises residues of the light chain of FVa (SEQ ID NO:38) within the sequence from Y21 1 to M226.

In one embodiment the antibody or antibody fragment binds Y21 1 to M226 of the FVa light chain.

Thus in an embodiment of the current invention, the epitope of the antibody comprises one or more of the residues Y21 1 , E212, K213, K214, S215, R216, S217, S218, W219, R220, L121 , T222, S223, S224, E225, and M226.

In one embodiment the epitope comprises at least 5 of the amino acid residues Y21 1 , E212, K213, K214, S215, R216, S217, S218, W219, R220, L1221 , T222, S223, S224, E225, and M226 and in further embodiments at least 8, 10, 12 or 14 amino acid residues.

Pharmaceutical formulation

In another aspect, the present invention provides compositions and formulations comprising molecules of the invention, such as the antibodies and fragments thereof, polynucleotides, vectors and cells described herein. For example, the invention provides a pharmaceutical composition that comprises one or more of the antibodies or antibody fragments of the invention, formulated together with a pharmaceutically acceptable carrier.

Accordingly, one object of the invention is to provide a pharmaceutical formulation comprising such an antibody or antibody fragment which is present in a concentration from 0.25 mg/ml to 250 mg/ml, and wherein said formulation has a pH from 2.0 to 10.0. The formulation may further comprise one or more of a buffer system, a preservative, a tonicity agent, a chelating agent, a stabilizer, or a surfactant, as well as various combinations thereof. The use of preservatives, isotonic agents, chelating agents, stabilizers and surfactants in pharmaceutical compositions is well-known to the skilled person. Reference may be made to Remington: The Science and Practice of Pharmacy, 19th edition, 1995.

In one embodiment, the pharmaceutical formulation is an aqueous formulation. Such a formulation is typically a solution or a suspension, but may also include colloids, dispersions, emulsions, and multi-phase materials. The term "aqueous formulation" is defined as a formulation comprising at least 50% w/w water. Likewise, the term "aqueous solution" is defined as a solution comprising at least 50 % w/w water, and the term "aqueous suspension" is defined as a suspension comprising at least 50 %w/w water. In another embodiment, the pharmaceutical formulation is a freeze-dried formulation, to which the physician or the patient adds solvents and/or diluents prior to use.

In a further aspect, the pharmaceutical formulation comprises an aqueous solution of such an antibody, and a buffer, wherein the antibody is present in a concentration from 1 mg/ml or above, and wherein said formulation has a pH from about 2.0 to about 10.0. Treatment

It is apparent from the above that the antibodies of the invention are useful in treatment of diseases or disorder where an increase in thrombin is desired. As mentioned above Hemophilia and in particular Hemophilia A is a key indication where stimulation of the blood coagulation cascade is useful

An aspect of the invention relates to an FVa antibody as described herein above for use in a method of treatment. An aspect of the invention relates to the use of an FVa antibody as described herein above for use in the preparation of a pharmaceutical product for use in a method of treatment. The method of treatment is in further embodiment related to the treatment of Coagulopathy, such as Haemophilia described herein.

In one embodiment the invention relates to a method of treatment of a coagulopathy comprising the steps of administering a therapeutically effect amount of an anti-FVa antibody to a subject in need and thereby treating coagulopathy.

As described above treatment may be prophylactic, palliative and/or symptomatic.

Prophylactic treatment may also be termed prevention e.g. at treatment where the medicament is administered in the absence of symptoms to reduce the occurrence and/or severity of symptoms. In a patient with Haemophilia, such as Haemophilia A, compounds of the inventions may be use both as prevention/prophylactic treatment and/or symptomatic treatment. Administration

An antibody (or fragment thereof) of the invention may be administered parenterally, such as intravenously, such as intramuscularly, such as subcutaneously. Alternatively, an antibody of the invention may be administered via a non-parenteral route, such as perorally or topically. An antibody of the invention may be administered prophylactically. An antibody of the invention may be administered therapeutically (on demand).

Depending on the treatment the antibodies or fragments thereof may be

administered in response to symptoms or at regular interval when aiming for a prophylactic treatment.

While certain features of the invention have been illustrated and described herein, many modifications, substitutions, changes, and equivalents will now occur to those of ordinary skill in the art. It is, therefore, to be understood that the appended embodiments are intended to cover all such modifications and changes as fall within the true spirit of the invention.

Embodiments 1. A monoclonal anti-activated Factor V (FVa) or an anti- Factor V (FV) antibody or an

antigen-binding fragment thereof, that when bound to FVa displays pro-coagulant effect in haemophilia

2. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that binds to FVa, for prophylactic treatment of haemophilia A.

3. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa improves FVa co-factor activity to activated Factor X. 4. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa protects FVa from inactivation by activated protein C (APC).

5. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases thrombin generation. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases thrombin generation in the presence of APC. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa increases thrombin generation in the presence of exogenously added APC in a human plasma-based thrombin generation assay. A monoclonal antibody or an antigen-binding fragment thereof, according to embodiment 7, wherein the thrombin generation is more than three times the standard deviation of the background human plasma-based thrombin generation assay with added APC without antibody. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases thrombin generation in the presence of APC, measured as thrombin generation parameter "Peak Thrombin in nM" in a haemophilia A patient plasma based thrombin generation assay. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases thrombin generation in the presence of APC, measured as thrombin generation parameter "Velocity Index in nM per minute", in haemophilia A patient plasma based thrombin generation assay. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases thrombin generation in the presence of APC, measured as thrombin generation parameter "endogenous thrombin potential (ETP) in nM x minute", in haemophilia A patient plasma based thrombin generation assay. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases thrombin generation in protein C (PC) deficient plasma. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa increases thrombin generation in protein C (PC) deficient plasma, measured in a thrombin generation assay. 14. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa increases thrombin generation in PC deficient plasma, measured as "Endogenous Thrombin Potential in nM minute" (ETP) in a thrombin generation assay. 15. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa increases thrombin generation in PC deficient plasma with added anti-FVI 11 antibody, measured as "Endogenous Thrombin Potential in nM minute (ETP)" in a thrombin generation assay. 16. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases thrombin generation in normal human plasma.

17. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa increases thrombin generation in normal human plasma, measured in a thrombin generation assay.

18. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa increases thrombin generation in normal human plasma, measured as "Endogenous Thrombin Potential in nM minute" (ETP) in a thrombin generation assay.

19. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to FVa increases thrombin generation in normal human plasma with added anti-FVIII antibody, measured as "Endogenous Thrombin Potential in nM ^χ minute" (ETP) in a thrombin generation assay.

20. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response. 21 . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in normal blood.

22. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in haemophilia A-like blood.

23. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in haemophilia A blood. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates thrombelastography, for prophylactic treatment of haemophilia. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in blood with added anti- Factor VIII antibody. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response measured as clot time (R) in seconds. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in normal blood, measured as clot time (R) in seconds. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates thrombelastography in haemophilia A-like blood measured as clot time (R) in seconds. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in haemophilia A blood measured as clot time (R) in seconds. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in blood with added anti- Factor VIII antibody, measured as clot time (R) in seconds. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in blood in the presence of APC, measured as clot time (R) in seconds. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates thromboelastographic response in blood, for prophylactic treatment of haemophilia. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response measured as "Maximum Rate of Thrombin Generation" (MTG). A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in normal blood, measured as "Maximum Rate of Thrombin Generation" (MTG). A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in haemophilia A-like blood measured as "Maximum Rate of Thrombin Generation" (MTG). A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in haemophilia A blood measured as "Maximum Rate of Thrombin Generation" (MTG). A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in blood with added anti- Factor VIII antibody, measured as "Maximum Rate of Thrombin Generation" (MTG). A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in blood in the presence of APC, measured as "Maximum Rate of Thrombin Generation" (MTG). A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in blood, for prophylactic treatment of haemophilia. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates the thromboelastographic response in blood in the presence of Thrombo-modulin (TM), measured as "Maximum Rate of Thrombin Generation" (MTG). A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation in normal blood. 43. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation in haemophilia A-like blood.

44. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation in haemophilia A blood.

45. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation, for prophylactic treatment of haemophilia. 46. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation in blood with added anti-Factor VIII antibody.

47. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation in blood in the presence of APC.

48. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation in blood, for prophylactic treatment of

haemophilia. 49. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa stimulates clot formation in blood in the presence of thrombomodulin (TM).

50. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation.

51 . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation in normal blood.

52. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation in haemophilia A-like blood.

53. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation in haemophilia A blood.

54. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation in haemophilia.

55. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation in blood with added anti-Factor VIII antibody. 56. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation in blood in the presence of APC.

57. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation in blood, for prophylactic treatment of haemophilia.

58. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases clot formation in blood in the presence of TM.

59. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reduces clotting time.

60. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reduces clotting time in normal blood.

61 . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reduces clotting time in haemophilia A-like blood.

62. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reduces clotting time in haemophilia A blood.

63. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that

reduces clotting time, for prophylactic treatment of haemophilia.

64. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reduces clotting time in blood with added anti-Factor VIII antibody.

65. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reduces clotting time in blood in the presence of APC.

66. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reduces clotting time in blood in the presence of TM. 67. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reduces clotting time in blood in the presence of APC and FVII/rFVII. 68. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa enhances thrombin generation in human platelet rich plasma (PRP) in haemophilia A-like conditions in the presence of TM.

69. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa enhances thrombin generation in human platelet rich plasma (PRP) in haemophilia A-like conditions in the presence of TM measured as thrombin generation parameter "Peak Thrombin in nM" in thrombin generation assay.

70. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa enhances thrombin generation in human platelet rich plasma (PRP) in the presence of anti-human FVIII polyclonal antibodies and TM.

71 . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa enhances thrombin generation in human platelet rich plasma (PRP) in the presence of anti-human FVIII polyclonal antibodies and TM measured as thrombin generation parameter "Peak Thrombin in nM" in thrombin generation assay.

72. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa enhances thrombin generation in haemophilia, for prophylactic treatment of haemophilia.

73. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reverses TM inhibiting effect on thrombin generation in haemophilia, for prophylactic treatment of haemophilia A.

74. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reverses TM inhibiting effect on thrombin generation in human platelet rich plasma (PRP) in haemophilia A-like conditions.

75. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reverses TM inhibiting effect on thrombin generation in human platelet rich plasma (PRP) in haemophilia A-like conditions measured as thrombin generation parameter "Peak Thrombin in nM" in thrombin generation assay.

76. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reverses TM inhibiting effect on thrombin generation in human platelet rich plasma (PRP) in the presence of anti-human FVIII polyclonal antibodies. 77. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reverses TM inhibiting effect on thrombin generation in human platelet rich plasma (PRP) in the presence of anti-human FVIII polyclonal antibodies measured as thrombin generation parameter "Peak Thrombin in nM" in thrombin generation assay.

78. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reverses TM inhibiting effect on thrombin generation in human haemophilia A.

79. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa reverses TM inhibiting effect on thrombin generation.

80. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that when bound to FVa increases thrombin generation in haemophilia, for prophylactic treatment of haemophilia.

81 . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of the preceding embodiment that binds to the C2 domain of FVa, for prophylactic treatment of haemophilia.

82. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 6, 20, 41 , 50, 59 or 73, that binds to the C2 domain of FVa.

83. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, that binds to the C2 domain of FVa. 84. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 81 , 82 or 83 that binds to a binding region located on the C2 domain of FVa.

85. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiment 84, wherein the antibody binds to a binding region comprising one or more of the amino acid residues N532 to A543, D553-L554, and G588 to M600 of SEQ ID NO:38. 86. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiment 84, wherein the binding region comprises one or more of the amino acid residues: N532, A532, Q533, G534, R535, V536, N537, A538, W539, Q540, A541 , K542, A543, D553, L554, G588, V589, E590, W591 , K592, P593, Y594, R595, L596 K597, S598, S599 or M600 of SEQ ID NO:38.

87. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiment 84, wherein the binding region comprises at least one of the amino acid residues, A532, Q533, G534, R535, V536, N537, A538, W539, Q540, A541 , K542 and A543 of SEQ ID NO:38; and at least one of the amino acid residues D553 or L554 of SEQ ID NO:38; and at least one of amino acid residues G588, V589, E590, W591 , K592, P593, Y594, R595, L596 K597, S598, S599 or M600 of SEQ ID NO:38.

88. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiment 84, wherein the binding region comprises amino acid residues, A532, Q533, G534, R535, V536, N537, A538, W539, Q540, A541 , K542, A543, D553, L554, G588, V589, E590, W591 , K592, P593, Y594, R595, L596 K597, S598, S599 and M600 of SEQ ID NO:38.

89. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 81 , 82 or 83 that binds to a binding region located on the C2 domain of FVa, for prophylactic treatment of haemophilia.

90. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, wherein the light chain of said antibody or antigen-binding fragment comprises a CDR1 sequence comprising residues 24-34 SEQ ID NO: 23 (QASESISSYLT), and/or a CDR2 sequence comprising residues 50-56 of SEQ ID NO:

23 (YASTLAS), and/or a CDR3 sequence comprising residues 89-101 of SEQ ID NO: 23 (LGVYSYSRDDGIA).

91 . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90, wherein the heavy chain of said antibody or antigen- binding fragment comprises a CDR1 sequence comprising residues 31 -36 SEQ ID NO:

24 (SSYYMC), and/or a CDR2 sequence comprising residues 51-67 of SEQ ID NO: 24 (CIYTAWDGASYANWAKG), and/or a CDR3 sequence comprising residues 99-109 of

SEQ ID NO: 24 (AMGSSDGANNL). 92. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-91 , wherein said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO: 23

(QASESISSYLT), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID

NO: 23 (YASTLAS), and/or a LC-CDR3 sequence comprising residues 89-101 of SEQ ID NO: 23 (LGVYSYSRDDGIA), and/or a HC-CDR1 sequence comprising residues 31 -36 SEQ ID NO: 24 (SSYYMC), and/or a HC-CDR2 sequence comprising residues 51-67 of SEQ ID NO: 24 (CIYTAWDGASYANWAKG), and/or a HC-CDR3 sequence comprising residues 99-109 of SEQ ID NO: 24 (AMGSSDGANNL).

93. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92, wherein the light chain variable domain (VL) of said antibody or antigen-binding fragment comprises SEQ ID NO: 23, and/or the heavy chain variable domain (VH) of said antibody or antigen-binding fragment comprises SEQ ID

NO: 24.

94. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof according to any of embodiments 1 -93, that competes with an antibody or antigen-binding fragment wherein the light chain variable domain (VL) of said antibody or antigen-binding fragment comprises SEQ ID NO: 23, and/or the heavy chain variable domain (VH) of said antibody or antigen-binding fragment comprises SEQ ID NO: 24, in binding to the C2 domain of FVa. 95. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 6, 20, 41 , 50, 59 or 73, that competes with an antibody or antigen-binding fragment wherein the light chain variable domain (VL) of said antibody or antigen-binding fragment comprises SEQ ID NO: 23, and/or the heavy chain variable domain (VH) of said antibody or antigen-binding fragment comprises SEQ ID NO: 24, in binding to the C2 domain of FVa.

96. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, that competes with an antibody or antigen-binding fragment wherein the light chain variable domain (VL) of said antibody or antigen-binding fragment comprises SEQ ID NO: 23, and/or the heavy chain variable domain (VH) of said antibody or antigen-binding fragment comprises SEQ ID NO: 24, in binding to the C2 domain of FVa.

97. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, wherein the light chain of said antibody or antigen-binding fragment comprises a CDR1 sequence comprising residues 23-33 SEQ ID NO:21 (SGDILGDKYAC), and/or a CDR2 sequence comprising residues 49-55 of SEQ ID NO:21 (QDIKRPS), and/or a CDR3 sequence comprising residues 88-97 of SEQ ID NO:21 (QAWDSTTPW).

98. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90 or 97, wherein the heavy chain of said antibody or antigen- binding fragment comprises a CDR1 sequence comprising residues 31 -35 SEQ ID NO:22 (SYDIN), and/or a CDR2 sequence comprising residues 50-66 of SEQ ID NO:22

(WMNPNTDDTGYAQKFQG), and/or a CDR3 sequence comprising residues 99-1 19 of SEQ ID NO:22 (YWS VTS WKWN D D H YYYYGM D V).

99. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, 97 or 98 wherein said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 23-33 SEQ ID NO:21

(SGDILGDKYAC), and/or a LC-CDR2 sequence comprising residues 49-55 of SEQ ID NO:21 (QDIKRPS), and/or a LC-CDR3 sequence comprising residues 88-97 of SEQ ID NO:21 (QAWDSTTPW), and/or a HC-CDR1 comprising residues 31-35 SEQ ID NO:22 (SYDIN), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:22 (WMNPNTDDTGYAQKFQG), and/or a HC-CDR3 sequence comprising residues 99-1 19 of SEQ ID NO:22 (YWSVTSWKWNDDHYYYYGMDV).

100. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92 or 97-99 wherein the light chain variable domain (VL) of said antibody or antigen-binding fragment comprises SEQ ID NO: 21 , and/or the heavy chain variable domain (VH) of said antibody or antigen-binding fragment comprises SEQ ID NO: 22.

101. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, that competes with an antibody or antigen-binding fragment wherein the light chain variable domain (VL) of said antibody or antigen-binding fragment comprises SEQ ID NO: 21 , and/or the heavy chain variable domain (VH) of said antibody or antigen-binding fragment comprises SEQ ID NO: 22, in binding to the C2 domain of FVa. 102. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 6, 20, 41 , 50, 59 or 73, that competes with an antibody or antigen-binding fragment wherein the light chain variable domain (VL) of said antibody or antigen-binding fragment comprises SEQ ID NO: 21 , and/or the heavy chain variable domain (VH) of said antibody or antigen-binding fragment comprises SEQ ID NO: 22, in binding to the C2 domain of FVa.

103. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, that competes with an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO: 21 , and/or a heavy chain variable domain (VH) comprising SEQ ID NO: 22, in binding to the C2 domain of FVa.

104. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, that competes with an antibody or antigen-binding fragment according to any of the preceding embodiments that competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 or SEQ ID NO:21 , and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24 or SEQ ID NO:22, in binding to the C2 domain of FVa. 105. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 6, 20, 41 , 50, 59 or 73, that competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO: 23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO: 24 and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, in binding to the C2 domain of FVa.

106. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, that competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO: 23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO: 24 and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, in binding to the C2 domain of FVa. 107. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89 wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-40 SEQ ID NO:15 (KSSQSVLYSSNNKNYLA), and/or a LC-CDR2 sequence comprising residues 56-62 of SEQ ID NO:15 (WASTRES), and/or a LC-CDR3 sequence comprising residues 95-103 of SEQ ID NO:15 (QQYYSTPWT).

108. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90 or 107, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO:16 (SYDIN), and/or a HC-CDR2 sequence comprising residues 50-66 of

SEQ ID NO:16 (WMNPNSGNTGYALKFQG), and/or a HC-CDR3 sequence comprising residues 99-1 14 of SEQ ID NO:16 (RTYYDILTGSLGAFDI).

109. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, 107 or 108, wherein said antibody or antigen- binding fragment comprises a LC-CDR1 sequence comprising residues 24-40 SEQ ID NO:15 (KSSQSVLYSSNNKNYLA), and/or a LC-CDR2 sequence comprising residues 56- 62 of SEQ ID NO: 15 (WASTRES), and/or a LC-CDR3 sequence comprising residues 95- 103 of SEQ ID NO:15 (QQYYSTPWT), and/or a HC-CDR1 comprising residues 31 -35 SEQ ID NO:16 (SYDIN), and/or a HC-CDR2 sequence comprising residues 50-66 of

1 10. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92 or 107-109 wherein the light chain variable domain (VL) of said antibody or antigen-binding fragment comprises SEQ ID NO: 15, and/or the heavy chain variable domain (VH) of said antibody or antigen-binding fragment comprises SEQ ID NO: 16. . A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof according to any of the previous embodiments, that competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO: 15, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 6, 20, 41 , 50, 59 or 73 or 84-1 12 that competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:15, and/or a heavy chain variable domain (VH) comprising

SEQ ID NO:16, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, that competes with an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, in binding to the C2 domain of FVa. . An monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof according to any of the previous embodiments, that competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 , and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID

NO:15, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, in binding to the C2 domain of FVa, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 84-1 14 that competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 , and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, in binding to the C2 domain of FVa, in binding to the C2 domain of FVa. 1 16. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, that competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 , and a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15, and a heavy chain variable domain (VH) comprising SEQ ID NO:16, in binding to the C2 domain of FVa, in binding to the C2 domain of FVa.

1 17. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:19 (RASQDISTWLA), and/or a LC-CDR2 sequence comprising residues 50- 56 of SEQ ID NO:19 (ITSTLHI), and/or a LC-CDR3 sequence comprising residues 89-97 of SEQ ID NO:19 (QQAYSFPFT).

1 18. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90 or 1 17 wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35

SEQ ID NO:20 (DYAMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:20 (GISWNSGGIGYADSVQG), and/or a HC-CDR3 sequence comprising residues 99-1 15 of SEQ ID NO:20 (DARWLVEEDYYYYGMDV). 1 19. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, 1 17 or 1 18 wherein said antibody or antigen- binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO: 19 (RASQDISTWLA), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:19 (ITSTLHI), and/or a LC-CDR3 sequence comprising residues 89-97 of SEQ ID NO:19 (QQAYSFPFT), and/or a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO:20 (DYAMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:20 (GISWNSGGIGYADSVQG), and/or a HC-CDR3 sequence comprising residues 99-1 15 of SEQ ID NO:20 (DARWLVEEDYYYYGMDV). 120. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92 or 1 17-1 19 with a light chain variable domain (VL) comprising SEQ ID NO:19, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20. 121. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:19, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, in binding to the C2 domain of FVa.

122. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79 or 84-121 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:19, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, in binding to the C2 domain of FVa.

123. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:19, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, in binding to the C2 domain of FVa. 124. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, that competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 , and a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15, and a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19, and a heavy chain variable domain (VH) comprising SEQ ID NO:20, in binding to the C2 domain of FVa, in binding to the C2 domain of FVa.

125. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 84-124 that competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 , and a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15, and a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19, and a heavy chain variable domain (VH) comprising SEQ ID NO:20, in binding to the C2 domain of FVa, in binding to the C2 domain of FVa. 126. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, that competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 , and a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15, and a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19, and a heavy chain variable domain (VH) comprising SEQ ID NO:20, in binding to the C2 domain of FVa, in binding to the C2 domain of FVa.

127. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:1 1 (RASQDISHWLA), and/or a LC-CDR2 sequence comprising residues 50- 56 of SEQ ID NO:1 1 (IASTLQT), and/or a LC-CDR3 sequence comprising residues 89- 97 of SEQ ID NO:1 1 (QQSNSFPLT). 128. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90 or 127, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO:12 (DYAMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:12 (GISWNSGAIGYADSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 15 of SEQ ID NO:12 (DARWLVEEDYQYYGLDV).

129. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, 127 or 128, wherein said antibody or antigen- binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:1 1 (RASQDISHWLA), and/or a LC-CDR2 sequence comprising residues 50-56 of

SEQ ID NO:1 1 (IASTLQT), and/or a LC-CDR3 sequence comprising residues 89-97 of SEQ ID NO:1 1 (QQSNSFPLT) and the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31-35 SEQ ID NO:12 (DYAMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:12 (GISWNSGAIGYADSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 15 of

SEQ ID NO: 12 (DARWLVEEDYQYYGLDV).

130. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92 or 127-129, wherein said antibody or antigen- binding fragment comprises a light chain variable domain (VL) comprising SEQ ID

NO:1 1 , and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12.

131. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, in binding to the C2 domain of FVa.

132. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79 or 84-131 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:1 1 , and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, in binding to the C2 domain of FVa. 133. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, in binding to the C2 domain of FVa.

134. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, in binding to the C2 domain of FVa.

135. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 84-134 wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, in binding to the C2 domain of FVa. 136. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising

SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, in binding to the C2 domain of FVa. 137. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:17 (RASQDISSWLA), and/or a LC-CDR2 sequence comprising residues 50- 56 of SEQ ID NO:17 (IASSLQS), and/or a LC-CDR3 sequence comprising residues 89- 97 of SEQ ID NO:17 (QQANSFPFT).

138. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90 or 137, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO:18 (DYAMH), and/or a HC-CDR2 sequence comprising residues 50-66 of

SEQ ID NO:18 (GVSWNSGAIGYADSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 15 of SEQ ID NO:18 ( D ARWLVE E D YQYYG M D V) .

139. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, 137 or 139, wherein said antibody or antigen- binding fragment comprises a light chain LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:17 (RASQDISSWLA), and/or a LC-CDR2 sequence comprising residues 50- 56 of SEQ ID NO:17 (IASSLQS), and/or a LC-CDR3 sequence comprising residues 89- 97 of SEQ ID NO:17 (QQANSFPFT), and the heavy chain of said antibody or antigen- binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35 SEQ ID

NO:18 (DYAMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:18 (GVSWNSGAIGYADSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 15 of SEQ ID NO:18 (DARWLVEEDYQYYGMDV). 140. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92 or 137-139, wherein said antibody or antigen- binding fragment comprises a light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18. 141. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, in binding to the C2 domain of FVa.

142. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79 or 84-141 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, in binding to the C2 domain of FVa.

143. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, in binding to the C2 domain of FVa. 144. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and a heavy chain variable domain

(VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and a heavy chain variable domain (VH) comprising SEQ ID NO:18, in binding to the C2 domain of FVa. 145. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 84-144 wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and a heavy chain variable domain (VH) comprising SEQ ID NO:18, in binding to the C2 domain of FVa.

146. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising

SEQ ID NO:17, and a heavy chain variable domain (VH) comprising SEQ ID NO:18, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-35 SEQ ID NO:39 (SASSSISSNYLH), and/or a LC-CDR2 sequence comprising residues 51- 57 of SEQ ID NO:39 (RTSNLAS), and/or a LC-CDR3 sequence comprising residues 90- 98 of SEQ ID NO:39 (QQGSSIPLT). . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90 or 147 wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO:40 (NYGMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:40 (MIYYDSSKMNYADTVKG), and/or a HC-CDR3 sequence comprising residues 99-107 of SEQ ID NO:40 (PTSHYVVDV). . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, 147 or 148, wherein said antibody or antigen- binding fragment wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-35 SEQ ID NO:39

(SASSSISSNYLH), and/or a LC-CDR2 sequence comprising residues 51-57 of SEQ ID NO:39 (RTSNLAS), and/or a LC-CDR3 sequence comprising residues 90-98 of SEQ ID NO:39 (QQGSSIPLT), and wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31-35 SEQ ID NO:40 (NYGMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:40 (MIYYDSSKMNYADTVKG), and/or a HC-CDR3 sequence comprising residues 99-107 of SEQ ID NO:40 (PTSHYVVDV). 150. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92 or 146-149, wherein said antibody or antigen- binding fragment comprises a light chain variable domain (VL) comprising SEQ ID NO:39, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40. 151. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, in binding to the C2 domain of FVa.

152. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79 or 84-151 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, in binding to the C2 domain of FVa.

153. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:39, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, in binding to the C2 domain of FVa. 154. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 84-154, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen- binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID

NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, in binding to the C2 domain of FVa.

157. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:13 (RASQDISNWLA), and/or a LC-CDR2 sequence comprising residues 50- 56 of SEQ ID NO:13 (ITSTLHI), and/or a LC-CDR3 sequence comprising residues 89-97 of SEQ ID NO:13 (QQANSFPFT).

158. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90 or 157, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35

SEQ ID NO:14 (DYAMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:14 (GISWNSGSTGYADSVQG), and/or a HC-CDR3 sequence comprising residues 99-1 15 of SEQ ID NO:14 (DARWLVEEDYYYYGMDV). 159. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, 157 or 158, wherein said antibody or antigen- binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:13 (RASQDISNWLA), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:13 (ITSTLHI), and/or a LC-CDR3 sequence comprising residues 89-97 of SEQ ID NO:13 (QQANSFPFT), and/or a HC-CDR1 sequence comprising residues 31-35 SEQ ID NO:14 (DYAMH), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:14 (GISWNSGSTGYADSVQG), and/or a HC-CDR3 sequence comprising residues 99-1 15 of SEQ ID NO:14 (DARWLVEEDYYYYGMDV). 160. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92 or 157-159, wherein said antibody or antigen- binding fragment comprises a light chain variable domain (VL) comprising SEQ ID NO:13, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14. 161. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:13, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, in binding to the C2 domain of FVa.

162. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79 or 84-161 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:13, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, in binding to the C2 domain of FVa.

163. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:13, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, in binding to the C2 domain of FVa. 164. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof

according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:13 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, in binding to the C2 domain of FVa. 165. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 84-164, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen- binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising

SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:13 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17 and/or a heavy chain variable domain (VH) comprising SEQ

ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:13 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 23-33 SEQ ID NO:43 (SGDKLESKYAC), and/or a LC-CDR2 sequence comprising residues 49-

55 of SEQ ID NO:43 (HDDKRPS), and/or a LC-CDR3 sequence comprising residues 88- 97 of SEQ ID NO:43 (QAWDSSTPW). . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-90 or 167, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO:44 (SYDIN), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:44 (WMNPNSGNTGYAQKFQG), and/or a HC-CDR3 sequence comprising residues 99-1 19 of SEQ ID NO:44 (YFSSTSWKWDDDYFYYYGMDV). . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-89, 167 or 169, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 23-33 SEQ ID NO:43 (SGDKLESKYAC), and/or a LC-CDR2 sequence comprising residues 49-55 of SEQ ID NO:43 (HDDKRPS), and/or a LC-CDR3 sequence comprising residues 88-97 of SEQ ID NO:43 (QAWDSSTPVV), and the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO:44 (SYDIN), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:44 (WMNPNSGNTGYAQKFQG), and/or a HC-CDR3 sequence comprising residues 99-1 19 of SEQ ID NO:44

(YFSSTSWKWDDDYFYYYGMDV). . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 84-92 or 167-169, wherein said antibody or antigen- binding fragment comprises a light chain variable domain (VL) comprising SEQ ID

NO:43, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:44. . A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:43, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:44, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79 or 84-171 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:43, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:44, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:43, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:44, in binding to the C2 domain of FVa. 174. A monoclonal anti-FVa/anti-FV antibody or antigen binding fragment thereof according to any of the previous embodiments, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:13 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:43 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:44, in binding to the C2 domain of FVa. 175. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 84-174, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen- binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:13 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:43 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:44, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:23 and a heavy chain variable domain (VH) comprising SEQ ID NO:24, and/or an antibody or an antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:21 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:22, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:15 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:16, and/or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:19 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:20, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:1 1 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:12, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:17, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:18, and/or an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:39 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:40, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:13 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:14, and/or an antibody or antigen-binding fragment with light chain variable domain (VL) comprising SEQ ID NO:43 and/or a heavy chain variable domain (VH) comprising SEQ ID NO:44, in binding to the C2 domain of FVa. 177. A binding region on the C2 domain of the activated Factor V.

178. A binding region on the C2 domain of FVa, wherein said binding region comprises one or more of the amino acid residues A532 to A543, D553-L554, and G588 to M600 of SEQ ID NO:38.

179. A binding region on the C2 domain of FVa according to embodiment 177 or 178, wherein said binding region comprises one or more of the amino acid residues: A532, Q533, G534, R535, V536, N537, A538, W539, Q540, A541 , K542, A543, D553, L554, G588, V589, E590, W591 , K592, P593, Y594, R595, L596 K597, S598, S599 or M600 of

SEQ ID NO:38.

180. A binding region on the C2 domain of FVa according to embodiment 177, 178 or 179, wherein said binding region comprises at least one of the amino acid residues ,A532, Q533, G534, R535, V536, N537, A538, W539, Q540, A541 , K542 and A543 of

SEQ ID NO:38; and at least one of the amino acid residues D553 or L554 of SEQ ID NO:38; and at least one of amino acid residues G588, V589, E590, W591 , K592, P593, Y594, R595, L596 K597, S598, S599 or M600 of SEQ ID NO:38. 181. A binding region on the C2 domain of FVa according to embodiment 177, 178, 179 or 180, wherein said binding region comprises amino acid residues A532, Q533, G534, R535, V536, N537, A538, W539, Q540, A541 , K542, A543, D553, L554, G588, V589, E590, W591 , K592, P593, Y594, R595, L596 K597, S598, S599 and M600 of SEQ ID NO:38.

182. A monoclonal activated Factor V (FVa) or an anti- Factor V (FV) antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 177-181 displays pro-coagulant effect in haemophilia A. 183. A monoclonal activated Factor V (FVa) or an anti- Factor V (FV) antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 177-181 , displays pro-coagulant effect in haemophilia A.

184. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that binds to the binding region of embodiments 177-181 , for prophylactic treatment of haemophilia A. 185. A monoclonal anti-FVa/anti-FV antibody or an antigen binding-fragment thereof, that when bound to the binding region of embodiments 177-181 , improves FVa co-factor activity to activated Factor X.

186. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 177-181 , protects FVa from

inactivation by activated protein C (APC). 187. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 177-181 , increases thrombin generation.

188. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 177-181 , increases thrombin generation in the presence of APC.

189. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 177-181 , increases thrombin generation in the presence of exogenously added APC in a human plasma-based thrombin generation assay.

190. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 149 and/or 150.

191. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 149 and/or 150 in binding to FVa.

192. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 149 and/or 150 in binding to the binding region of any of embodiments 177-181 . 193. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 92 and/or 93. 194. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 92 and/or 93 in binding to FVa.

195. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 92 and/or 93 in binding to the binding region of any of embodiments 177-181.

196. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 109 and/or 1 10.

197. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 109 and/or 1 10 in binding to FVa.

198. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 109 and/or 1 10 in binding to the binding region of any of embodiments 177-181 . 199. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 99 and/or 100.

200. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 99 and/or 100 in binding to FVa.

201. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 99 and/or 100 in binding to the binding region of any of embodiments 177-181. 202. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 1 19 and/or 120. 203. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 1 19 and/or 120 in binding to FVa.

204. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 1 19 and/or 120 in binding to the binding region of any of embodiments 177-181 .

205. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 129 and/or 130.

206. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 129 and/or 130 in binding to FVa.

207. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 129 and/or 130 in binding to the binding region of any of embodiments 177-181 . 208. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 139 and/or 140.

209. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 139 and/or 140 in binding to FVa.

210. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 139 and/or 140 in binding to the binding region of any of embodiments 177-181 . 21 1. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 159 and/or 160. 212. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 159 and/or 160 in binding to FVa.

213. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 159 and/or 160 in binding to the binding region of any of embodiments 177-181 .

214. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody is the antibody or antigen- binding fragment of embodiments 169 and/or 170.

215. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 182-189, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 169 and/or 170 in binding to FVa.

216. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 169 and/or 170 in binding to the binding region of any of embodiments 177-181 . 217. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 81 , 82, 83 or 84 that binds to a binding region located on the C2 domain of FVa.

218. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiment 217, wherein the antibody binds to a binding region on the activated Factor F and wherein said binding region comprises one of more of the amino acid residues F325 to V331 and N534 to C539 of the FVa heavy-chain sequence (SEQ ID NO:37). 219. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiment 217, wherein the antibody binds to a binding region on activated FV and wherein the binding site comprises one or more of the amino acid residues F325, 1326, A327, A328, E329, E330, V331 , N534, I535, N536, K537, F538, and C539 of the FVa heavy-chain sequence (SEQ ID NO:37).

220. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiment 217, wherein the antibody binds to a binding region on the activated Factor F and wherein said binding region comprises one of more of the amino acid residues F325, I326, A327, A328, E329, E330 or V331 of the FVa heavy-chain sequence (SEQ ID NO:37), and wherein said binding region comprises at least one of the amino acid residues N534, I535, N536, K537, F538, or C539 of the FVa heavy-chain sequence (SEQ ID NO:37).

221. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiment 217, 218, 219 or 220, wherein the antibody binds to a binding region on activated FV and wherein the binding site comprises amino acid residues F325, I326, A327, A328, E329, E330, V331 , N534, I535, N536, K537, F538, and C539 of the

FVa heavy-chain sequence (SEQ ID NO:37).

222. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 , wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34

SEQ ID NO:9 (KTSTDIDDDMN), and/or a LC-CDR2 sequence comprising residues 50- 56 of SEQ ID NO:9 (EGNTLRP), and/or a LC-CDR3 sequence comprising residues 89- 97 of SEQ ID NO:9 (LQSANMPFT). 223. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-222, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO: 10 (SYAMS), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:10 (TISSGGSYTYYPDSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 1 1 of SEQ ID NO:10 (GPYLTTATPSFTY).

224. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-223, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprises residues 24-34 SEQ ID NO:9 (KTSTDIDDDMN), and/or a LC-CDR2 sequence comprising residues 50-

56 of SEQ ID NO:9 (EGNTLRP), and/or a LC-CDR3 sequence comprising residues 89- 97 of SEQ ID NO:9 (LQSANMPFT) and the heavy chain of said antibody comprises a HC-CDR1 sequence comprising residues 31-35 SEQ ID NO:10 (SYAMS), and/or a HC- CDR2 sequence comprising residues 50-66 of SEQ ID NO:10

(TISSGGSYTYYPDSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 1 1 of SEQ ID NO: 10 (GPYLTTATPSFTY).

225. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-224, wherein said antibody or antigen-binding fragment comprises a light chain variable domain (VL) comprising SEQ ID NO:9, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:10.

226. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of embodiments 1-84 or 217-225, wherein said antibody or antigen- binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:9, and a heavy chain variable domain

(VH) comprising SEQ ID NO:10, in binding to the C2 domain of FVa.

227. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 217, 218, 219, 220 or 221 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:9, and a heavy chain variable domain (VH) comprising SEQ ID NO:10, in binding to the C2 domain of FVa. 228. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:9, and a heavy chain variable domain (VH) comprising SEQ ID NO:10, in binding to the C2 domain of FVa.

229. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of embodiments 1-84 or 217-228, wherein said antibody or antigen- binding fragment competes with an antibody or antigen-binding fragment with a light chain comprising a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:9

(KTSTDIDDDMN), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:9 (EGNTLRP), and/or a LC-CDR3 sequence comprising residues 89-97 of SEQ ID NO:9 (LQSANMPFT) and/or a heavy chain comprising a HC-CDR1 sequence comprising residues 31 -35 SEQ ID NO:10 (SYAMS), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:10 (TISSGGSYTYYPDSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 1 1 of SEQ ID NO:10 (GPYLTTATPSFTY), in binding to the C2 domain of FVa.

230. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 , wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34

SEQ ID NO:33 (QASQSISSYLS), and/or a LC-CDR2 sequence comprising residues 50- 56 of SEQ ID NO:33 (RTSTLES), and/or a LC-CDR3 sequence comprising residues 89- 102 of SEQ ID NO:33 (QSNYYSSGSSYENA). 231. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 or 230, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 30-35 SEQ ID NO:34 (SYYHIC), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:34 (CIYAASGDTWYATWVNA), and/or a HC-CDR3 sequence comprising residues 98-1 14 of SEQ ID NO:34 (GPRYVSSSGAGPYCLDL).

232. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 , 230 or 231 , wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:33 (QASQSISSYLS), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:33 (RTSTLES), and/or a LC-CDR3 sequence comprising residues 89-102 of SEQ ID NO:33 (QSNYYSSGSSYENA) and the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 30-35 SEQ ID NO:34 (SYYHIC), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:34 (CIYAASGDTWYATWVNA), and/or a HC-

CDR3 sequence comprising residues 98-1 14 of SEQ ID NO:34

(GPRYVSSSGAGPYCLDL).

233. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 or 229-232, wherein said antibody or antigen- binding fragment comprises a light chain variable domain (VL) comprising SEQ ID NO:33, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:34.

234. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of embodiments 1-84, 217-233, wherein said antibody or antigen- binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:33, and a heavy chain variable domain (VH) comprising SEQ ID NO:34, in binding to the C2 domain of FVa. 235. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 217, 218, 219, 220 or 221 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:33, and a heavy chain variable domain (VH) comprising SEQ ID NO:34, in binding to the C2 domain of FVa.

236. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:33, and a heavy chain variable domain (VH) comprising SEQ ID NO:34, in binding to the C2 domain of FVa.

237. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of embodiments 1-84 or 217-236, wherein said antibody or antigen- binding fragment competes with an antibody or an antigen-binding fragment thereof with a light chain comprising a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:33 (QASQSISSYLS), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:33 (RTSTLES), and/or a LC-CDR3 sequence comprising residues 89-102 of SEQ ID NO:33 (QSNYYSSGSSYENA) and/or a heavy chain comprising comprises a HC-CDR1 sequence comprising residues 30-35 SEQ ID NO:34 (SYYHIC), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:34 (CIYAASGDTWYATWVNA), and/or a HC-CDR3 sequence comprising residues 98-1 14 of SEQ ID NO:34

(GPRYVSSSGAGPYCLDL), in binding to the C2 domain of FVa. 238. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof according to any of embodiments 1-84, 217-233, wherein said antibody or antigen- binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:33, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:34, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain (VL) comprising SEQ ID NO:9, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:10, in binding to the C2 domain of FVa. 239. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 217, 218, 219, 220 or 221 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:33, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:34, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain

(VL) comprising SEQ ID NO:9, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:10, in binding to the C2 domain of FVa.

240. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment thereof, with a light chain variable domain (VL) comprising SEQ ID NO:33, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:34, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain (VL) comprising SEQ ID NO:9, and/or a heavy chain variable domain (VH) comprising

SEQ ID NO:10, in binding to the C2 domain of FVa.

241. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of embodiments 1-84 or 217-236, wherein said antibody or antigen- binding fragment comprises a light chain comprising a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:33 (QASQSISSYLS), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:33 (RTSTLES), and/or a LC-CDR3 sequence comprising residues 89-102 of SEQ ID NO:33 (QSNYYSSGSSYENA) and/or a heavy chain comprising comprises a HC-CDR1 sequence comprising residues 30-35 SEQ ID NO:34 (SYYHIC), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:34 (CIYAASGDTWYATWVNA), and/or a HC-CDR3 sequence comprising residues 98-1 14 of SEQ ID NO:34 (GPRYVSSSGAGPYCLDL) or/and with an antibody or an antigen-binding fragment thereof with a light chain comprising a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:9 (KTSTDIDDDMN), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:9 (EGNTLRP), and/or a LC-CDR3 sequence comprising residues 89-97 of SEQ ID NO:9 (LQSANMPFT) and/or a heavy chain comprising a HC-CDR1 sequence comprising residues 31-35 SEQ ID NO:10 (SYAMS), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:10 (TISSGGSYTYYPDSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 1 1 of SEQ ID NO: 10 (GPYLTTATPSFTY), in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 , wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:29 (QASQSIGGNLA), and/or a LC-CDR2 sequence comprising residues 50-

56 of SEQ ID NO:29 (DASKLAS), and/or a LC-CDR3 sequence comprising residues 89- 10 of SEQ ID NO:29 (QCTYGSSGNIGNG). . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 or 242, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 30-34 SEQ ID NO:30 (SYAMI), and/or a HC-CDR2 sequence comprising residues 49-64 of SEQ ID NO:30 (FIDTGGSAYYASWAKG), and/or a HC-CDR3 sequence comprising residues 95-108of SEQ ID NO:30 (A LYVYS D VYTA F N I ) . . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 , 242 or 243, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:29 (QASQSIGGNLA), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:29 (DASKLAS), and/or a LC-CDR3 sequence comprising residues 89-10 of SEQ ID NO:29 (QCTYGSSGNIGNG) and the heavy chain of said antibody or antigen-binding fragment comprises a HC-CDR1 sequence comprising residues 30-34 SEQ ID NO:30 (SYAMI), and/or a HC-CDR2 sequence comprising residues 49-64 of SEQ ID NO:30 (FIDTGGSAYYASWAKG), and/or a HC- CDR3 sequence comprising residues 95-108 of SEQ ID NO:30 (A LYVYS D V YT A F N I ) . 245. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 217-221 or 242-244, wherein said antibody or antigen- binding fragment comprises a light chain variable domain (VL) comprising SEQ ID NO:29, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:30.

246. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of embodiments 1-84, 217-245, wherein said antibody or antigen- binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:29, and a heavy chain variable domain (VH) comprising SEQ ID NO:30, in binding to the C2 domain of FVa.

247. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 217, 218, 219, 220 or 221 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:29, and a heavy chain variable domain (VH) comprising SEQ ID NO:30, in binding to the C2 domain of FVa. 248. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or antigen- binding fragment with light chain variable domain (VL) comprising SEQ ID NO:29, and a heavy chain variable domain (VH) comprising SEQ ID NO:30, in binding to the C2 domain of FVa.

249. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof

according to any of embodiments 1-84 or 217-248, wherein said antibody competes with an antibody or an antigen-binding fragment thereof with a light chain comprising a LC- CDR1 sequence comprising residues 24-34 SEQ ID NO:29 (QASQSIGGNLA), and/or a

LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:29 (DASKLAS), and/or a LC-CDR3 sequence comprising residues 89-10 of SEQ ID NO:29 (QCTYGSSGNIGNG) and a heavy chain comprising a HC-CDR1 sequence comprising residues 30-34 SEQ ID NO:30 (SYAMI), and/or a HC-CDR2 sequence comprising residues 49-64 of SEQ ID NO:30 (FIDTGGSAYYASWAKG), and/or a HC-CDR3 sequence comprising residues 95-

108 of SEQ ID NO:30 (ALYVYSDVYTAFNI), in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof according to any of embodiments 1-84, 217-249, wherein said antibody or antigen- binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:33, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:34, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain (VL) comprising SEQ ID NO:9, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:10, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain (VL) comprising SEQ ID NO:29, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:30, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 1 , 2, 3, 4, 5, 20, 41 , 50, 59, 73, 79, 217, 218, 219, 220 or 221 , wherein said antibody or antigen-binding fragment competes with an antibody or antigen-binding fragment with a light chain variable domain (VL) comprising SEQ ID NO:33, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:34, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain (VL) comprising SEQ ID NO:9, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:10, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain (VL) comprising SEQ ID NO:29, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:30, in binding to the C2 domain of FVa. . A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 7, 8, 12, 16, 24, 32, 39, 45, 48, 54, 57, 63, 72, 78 or 80, wherein said antibody or antigen-binding fragment competes with an antibody or an antigen-binding fragment thereof, with a light chain variable domain (VL) comprising SEQ ID NO:33, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:34, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain (VL) comprising SEQ ID NO:9, and/or a heavy chain variable domain (VH) comprising

SEQ ID NO:10, and/or with an antibody or an antigen-binding fragment comprising a light chain variable domain (VL) comprising SEQ ID NO:29, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:30, in binding to the C2 domain of FVa. 253. A monoclonal anti-FVa/anti-FV antibody an antigen-binding fragment thereof according to any of embodiments 1-84 or 217-236, wherein said antibody or antigen- binding fragment comprises a light chain comprising a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:33 (QASQSISSYLS), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:33 (RTSTLES), and/or a LC-CDR3 sequence comprising residues 89-102 of SEQ ID NO:33 (QSNYYSSGSSYENA) and/or a heavy chain comprising comprises a HC-CDR1 sequence comprising residues 30-35 SEQ ID NO:34 (SYYHIC), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:34 (CIYAASGDTWYATWVNA), and/or a HC-CDR3 sequence comprising residues 98-1 14 of SEQ ID NO:34 (GPRYVSSSGAGPYCLDL) or/and with an antibody or an antigen-binding fragment thereof with a light chain comprising a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:9 (KTSTDIDDDMN), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:9 (EGNTLRP), and/or a LC-CDR3 sequence comprising residues 89-97 of SEQ ID NO:9 (LQSANMPFT) and/or a heavy chain comprising a HC-CDR1 sequence comprising residues 31-35 SEQ ID NO:10

(SYAMS), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:10 (TISSGGSYTYYPDSVKG), and/or a HC-CDR3 sequence comprising residues 99-1 1 1 of SEQ ID NO: 10 (GPYLTTATPSFTY), and/or with an antibody or an antigen-binding fragment thereof with a light chain comprising a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:29 (QASQSIGGNLA), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:29 (DASKLAS), and/or a LC-CDR3 sequence comprising residues 89-10 of SEQ ID NO:29 (QCTYGSSGNIGNG) and a heavy chain comprising a HC-CDR1 sequence comprising residues 30-34 SEQ ID NO:30 (SYAMI), and/or a HC- CDR2 sequence comprising residues 49-64 of SEQ ID NO:30 (FIDTGGSAYYASWAKG), and/or a HC-CDR3 sequence comprising residues 95-108 of SEQ ID NO:30

(ALYVYS D VYTAF N I ), in binding to the C2 domain of FVa.

254. A binding region on the C2 domain of the activated Factor V. 255. A binding region on the C2 domain of FVa, wherein said binding region comprises one or more of the amino acid residues F325-V331 or N534- C539 (SEQ ID NO:37).

256. A binding region on the C2 domain of FVa according to embodiment 254 or 255, wherein said binding region comprises one or more of the amino acid residues: F325, I326, A327, A328, E329, E330, V331 , N534, I535, N536, K537, F538 or C539 (SEQ ID

NO:37). 257. A binding region on the C2 domain of FVa according to embodiment 254, 255 or 256, wherein said binding region comprises at least one of the amino acid residues F325, 1326, A327, A328, E329, E330 or V331 (SEQ ID NO:37) and at least one of the amino acid residues N534, I535, N536, K537, F538 or C539 (SEQ ID NO:37).

258. A binding region on the C2 domain of FVa according to embodiment 254, 255, 256 or 257, wherein said binding region comprises the amino acid residues F325, I326, A327, A328, E329, E330, V331 , N534, I535, N536, K537, F538 and C539 (SEQ ID NO:37).

259. A monoclonal activated Factor V (FVa) or an anti- Factor V (FV) antibody or an

antigen-binding fragment thereof, that when bound to the binding region of embodiments 254-258 displays pro-coagulant effect in haemophilia A. 260. A monoclonal activated Factor V (FVa) or an anti- Factor V (FV) antibody or an

antigen-binding fragment thereof, that when bound to the binding region of embodiments 254-258, displays pro-coagulant effect in haemophilia A.

261. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that binds to the binding region of embodiments 254-258, for prophylactic treatment of haemophilia A.

262. A monoclonal anti-FVa/anti-FV antibody or an antigen binding-fragment thereof, that when bound to the binding region of embodiments 254-258, improves FVa co-factor activity to activated Factor X.

263. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 254-258, protects FVa from

inactivation by activated protein C (APC).

264. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 254-258, increases thrombin generation. 265. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 254-258, increases thrombin generation in the presence of APC. 266. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that when bound to the binding region of embodiments 254-258, increases thrombin generation in the presence of exogenously added APC in a human plasma-based thrombin generation assay. 267. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 259-266, wherein said antibody is the antibody or antigen- binding fragment of embodiments 224 and/or 225.

268. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 259-266, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 224 and/or 225 in binding to FVa.

269. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 224 and/or 225 in binding to the binding region of any of embodiments 254-258.

270. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 259-266, wherein said antibody is the antibody or antigen- binding fragment of embodiments 232 and/or 233.

271. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 259-266, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 232 and/or 233 in binding to FVa. 272. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 232 and/or 233 in binding to the binding region of any of embodiments 254-258.

273. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 259-266, wherein said antibody is the antibody or antigen- binding fragment of embodiments 244 and/or 245. 274. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to embodiments 259-266, wherein said antibody competes with the antibody or antigen-binding fragments of embodiments 244 and/or 245 in binding to FVa. 275. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment of embodiments 244 and/or 245 in binding to the binding region of any of embodiments 254-258.

276. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-

CDR1 sequence comprising residues 24-34 SEQ ID NO:7 (KASQDVGTAVG), and/or a LC-CDR2 sequence comprising residues 50-56 of SEQ ID NO:7 (WASTRHT), and/or a LC-CDR3 sequence comprising residues 89-96 of SEQ ID NO:7 (QQYSSNPT). 277. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC- CDR1 sequence comprising residues 31 -35 SEQ ID NO:8 (NYGMN), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:8 (WINTYTGEPTYADDFKG), and/or a HC-CDR3 sequence comprising residues 99-1 1 1 of SEQ ID NO:10

(SLRRNPYYYAMDY).

278. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 276-277, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-34 SEQ ID NO:7 (KASQDVGTAVG), and/or a LC-CDR2 sequence comprising residues 50-

56 of SEQ ID NO:7 (WASTRHT), and/or a LC-CDR3 sequence comprising residues 89- 96 of SEQ ID NO:7 (QQYSSNPT) and the heavy chain of said antibody comprises a HC- CDR1 sequence comprising residues 31 -35 SEQ ID NO:8 (NYGMN), and/or a HC-CDR2 sequence comprising residues 50-66 of SEQ ID NO:8 (WINTYTGEPTYADDFKG), and/or a HC-CDR3 sequence comprising residues 99-1 1 1 of SEQ ID NO:10

(SLRRNPYYYAMDY).

279. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 276-278, wherein said antibody or antigen-binding fragment comprises a light chain variable domain (VL) comprising SEQ ID NO:7, and/or a heavy chain variable domain (VH) comprising SEQ ID NO:8.

280. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that competes with any of the antibodies or antigen-binding fragments of embodiments 276-

279.

281. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, wherein the light chain of said antibody or antigen-binding fragment comprises a LC- CDR1 sequence comprising residues 24-40 SEQ ID NO:41 (RSSQSLLDSDDGNTYMD), and/or a LC-CDR2 sequence comprising residues 56-62 of SEQ ID NO:41 (MGFYRAS), and/or a LC-CDR3 sequence comprising residues 95-103 of SEQ ID NO:41

(MQRIEFPST). 282. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, wherein the heavy chain of said antibody or antigen-binding fragment comprises a HC- CDR1 sequence comprising residues 31-37 SEQ ID NO:42 (TSGVGVG), and/or a HC- CDR2 sequence comprising residues 52-67 of SEQ ID NO:42 (LIYWDDVKRYSPSLRR), and/or a HC-CDR3 sequence comprising residues 100-107 of SEQ ID NO:42

(YNWKMRVD).

283. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 or 282, wherein the light chain of said antibody or antigen-binding fragment comprises a LC-CDR1 sequence comprising residues 24-40 SEQ ID NO:41 (RSSQSLLDSDDGNTYMD), and/or a LC-CDR2 sequence comprising residues 56-62 of SEQ ID NO:41 (MGFYRAS), and/or a LC-CDR3 sequence comprising residues 95-103 of SEQ ID NO:41 (MQRIEFPST) and the heavy chain of said antibody comprises a HC-CDR1 sequence comprising residues 31-37 SEQ ID NO:42

(TSGVGVG), and/or a HC-CDR2 sequence comprising residues 52-67 of SEQ ID NO:42 (LIYWDDVKRYSPSLRR), and/or a HC-CDR3 sequence comprising residues 100-107 of

SEQ ID NO:42 (YNWKMRVD).

284. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281-283, wherein said antibody or antigen-binding fragment comprises a light chain variable domain (VL) comprising SEQ ID NO:41 , and/or a heavy chain variable domain (VH) comprising SEQ ID NO:42. 285. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof, that competes with any of the antibodies or antigen-binding fragments of embodiments 281 - 284.

286. A monoclonal activated Factor V (FVa) or an anti-Factor V (FV) antibody or an

antigen-binding fragment thereof according to any of embodiments 276, 277, 278, 279 or 280, that when bound to the C2 domain of FVa displays pro-coagulant effect in haemophilia A.

287. A monoclonal activated Factor V (FVa) or an anti- Factor V (FV) antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa displays pro-coagulant effect in haemophilia A.

288. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, for prophylactic treatment of haemophilia A. 289. A monoclonal anti-FVa/anti-FV antibody or an antigen binding-fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa improves FVa co-factor activity to activated Factor X.

290. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa protects FVa from inactivation by activated protein C (APC).

291. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa increases thrombin generation.

292. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa increases thrombin generation in the presence of APC. 293. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa increases thrombin generation in the presence of exogenously added APC in a human plasma-based thrombin generation assay.

294. A monoclonal activated Factor V (FVa) or an anti- Factor V (FV) antibody or an antigen-binding fragment thereof according to any of embodiments 276, 277, 278, 279 or 280, that when bound to the C2 domain of FVa displays pro-coagulant effect in haemophilia A.

295. A monoclonal activated Factor V (FVa) or an anti- Factor V (FV) antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa displays pro-coagulant effect in haemophilia A.

296. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, for prophylactic treatment of haemophilia A. 297. A monoclonal anti-FVa/anti-FV antibody or an antigen binding-fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa improves FVa co-factor activity to activated Factor X.

298. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa protects FVa from inactivation by activated protein C (APC).

299. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa increases thrombin generation.

300. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa increases thrombin generation in the presence of APC. 301. A monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment thereof according to any of embodiments 281 , 282, 283, 284 or 285, that when bound to the C2 domain of FVa increases thrombin generation in the presence of exogenously added APC in a human plasma-based thrombin generation assay.

302. Use of a monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment

thereof that binds to activated Factor V (FVa) for prophylactic treatment of haemophilia A.

303. Use of a monoclonal anti-FVa/anti-FV antibody or an antigen-binding fragment

thereof that binds to FVa, as a pro-coagulant in prophylactic treatment of haemophilia A.

304. Use according to embodiments 302 or 303, wherein the monoclonal anti-FVa/anti- FV antibody or antigen-binding fragment thereof, is the antibody of embodiments 92 and/or 93.

305. Use according to embodiments 302 or 303, wherein the monoclonal anti-FVa/anti- FV antibody or antigen-binding fragment thereof, is the antibody of any of embodiments

92, 93, 149, 150, 99, 100, 109, 1 10, 1 19, 120, 129, 130, 139, 140, 159, 160, 169 and/or 170.

306. Use according to embodiments 302 or 303, wherein the monoclonal anti-FVa/anti- FV antibody or antigen-binding fragment thereof, is the antibody of any of embodiments 224, 225, 232, 233, 244 and/or 245. 307. Use according to embodiments 302 or 303, wherein the monoclonal anti-FVa/anti- FV antibody or antigen-binding fragment thereof, is the antibody of embodiments 278 and/or 279.

308. Use according to embodiments 302 or 303, wherein the monoclonal anti-FVa/anti- FV antibody or antigen-binding fragment thereof, is the antibody of embodiments 283 and/or 284.

309. A pharmaceutical composition comprising the antibody of embodiments 92 and/or 93.

310. A pharmaceutical composition comprising the antibody of any of embodiments 92,

93, 149, 150, 99, 100, 109, 1 10, 1 19, 120, 129, 130, 139, 140, 159, 160, 169 and/or 170. . A pharmaceutical composition comprising the antibody of any of embodiments 224, 225, 232, 233, 244 and/or 245. . A pharmaceutical composition comprising the antibody of embodiments 278 and/or 279. . A pharmaceutical composition comprising the antibody of embodiments 283 and/or 284. . An antigen-binding fragment of the antibody of any of embodiments 1 -176 or 182- 301 , which is a Fab, Fab', F(ab)2, F(ab')2, Fv, single-chain Fv, dsFv, Fd or a dAb fragment, a VH, VL, VhH, or V-NAR domains, a monovalent molecule, minibody, diabody, triabody, tetrabody or kappa body, or an IgNAR. . A variant of the antibody of any of embodiments 1 -176 or 182-301 , which is a deletion variant or an insertion variant. . A nucleic acid construct that encodes the antibody of any of embodiments 1 -176 or 182-301 .

A recombinant vector that comprises the nucleic acid construct of embodiment 316. . An isolated cell that expresses the antibody of any of embodiments 1-176 or182- 301. . An isolated cell that comprises the nucleic acid construct of embodiment 316 or the vector of embodiment 318.

Examples

Example 1 - Antibody generation.

Mouse generated antibodies

Conventional mice (RBF, NMRCF1 ) and HK or HL -Kymice (Kymab, Cambridge,

UK) were immunized biweekly with 20 μg human FVa (SEQ ID NO: 37 and 38) using a standard protocol emulsified in Sigma Adjuvant System®. The Kymice were used to generate antibodies with human variable regions. Ten days after the last immunization mice were chin bled and the sera were tested in a direct ELISA. Nunc immunoplates were coated with 1 μg/ml of human FVa (HTI) and incubated overnight at 4°C. Plates were blocked with blocking buffer (PBS with 0.05% Tween20) for 15 min and were washed with PBS/0.05%Tween20. Serum was added with a starting dilution of 1 :50 and followed by threefold serial dilution and the plates were incubated for 1 hour at room temperature. After another wash, HRP labelled gt anti-mouse Fc-HRP was added at a concentration of 1 μg/ml, and incubated for 1 hour. After washing, plates were developed with TMB-substrate (Kem- EN-Tec) as described by the manufacturer. Absorbance at 450 nm was measured on an ELISA-reader. Only mice, which showed a specific response towards human FVa were considered for fusions and mice with the highest titer were selected. Mice with positive titer were boosted i.v. with human FVa and sacrificed three days later. The spleen was removed aseptically and dispersed to a single cell suspension. Fusion of spleenocytes with myeloma cells (FOX-Ny or Ag8-X63) was done by standard electrofusion. Cells were seeded in microtiter plates and cultured for 13 days under selection with HAT/HT. Supernatants were screened in a direct ELISA on human FVa. Nunc immunoplates were coated with 1 μg/ml of peptide of and incubated overnight at 4°C. Plates were blocked with blocking buffer (PBS with 0.05% Tween20) for 15 min and were washed with PBS/0.05%Tween20. Culture supernatants from the hybridoma cells were added and the plates were incubated for 1 hour at room temperature. After another wash, HRP labelled gt anti-mouse Fc-HRP was added at a concentration of 1 μg/ml, and incubated for 1 hour. After washing, plates were developed with TMB-substate (Kem-EN-Tec) as described by the manufacturer. Absorbance at 450 nm was measured on an ELISA-reader. After the confirmation screen by ELISA human FVa specific hybridoma supernatants were further tested in a Bio-Layer Interferometry (BLI) assay on an Octett system from Fortebio. Using anti-murine Fey specific tip human FVa specific antibodies were captured from the supernatant. Afterwards human FVa was bound to the surface by dipping the tips into a 50 nM hFVa solution. After the association phase the tips were transferred to buffer in order to determine the dissociation of hFVa Rabbit generated antibodies

Rabbits (New Zealand White) were immunized biweekly four times with 50 μg human FVa (SEQ ID NO: 37 and 38) using emulsified in Sigma Adjuvant System® a standard protocol.

The hFVa immunized rabbits were sacrificed, the spleen removed and passed through a nylon mesh. After thorough wash, the spleen cells were frozen in aliquots in liquid N₂.

Six of 384 wells were seeded with 12.500ο/75μΙΛ/νβΙΙ of irradiated (5000 Rads) EL4.B5 mouse thymoma cells expressing the CD40-ligand crucial for B-cell proliferation, differentiation and Ig-production. The cells were cultured at 37^°C and 5% C0₂ for 24 to 48 hours in medium supplemented with 100ng/ml mBAFF, 600ng/ml RblL-2 and 5% of a PMA- activated splenic supernatant.

Frozen spleen cells were thawed, washed in FACS buffer (PBS + 1 % BSA) and 2 vials of 15x10⁶ rabbit splenocytes were resuspended in 200 μΙ FACS buffer. To block unspecific binding, 500 nM FVIII was added to the cells. The suspension was incubated for 10 min, hFVa biotin (10 μg/ml) was added and the cells incubated for further 30 min on ice.

Cells were washed three times in Facs buffer and resuspended in 200 μΙ Facs buffer with 10 μΙ Donkey anti Rabbit Fitc and 5 μΙ Streptavidin-Pe. The cells were incubated for 30 min on ice.

Facs buffer was added up to 1 ml with 1 μΙ SYTOX Red dead cell stain and the cells incubated further 20 min on ice. The cells were then washed twice in Facs Buffer, resuspended in a desired volume and filtered through a 30 μηη filcon.

Cells were acquired on a FACSAria Sorter. Gates were initially set on lymphocytes, then live lymphocytes and finally on single live lymphocytes. This final gate was used when sorting gate was set on the antigen specific B-cells identified as those simultaneously binding to both anti rabbit IgG and biotinylated hFVa. Cells were sorted as 1 cell/well into 384-well plates already seeded with the EL4B5 cells. The cells were then kept in the incubator at 37^°C and 5% C0_2.

Seven days post sorting, all supernatants were tested in a binding assays to investigate presence of antigen specific IgG. Cells from all positive wells were picked and further processed. Example 2 - Identification of pro-coagulant anti-Factor Va antibodies in human haemophilic plasma by thrombin generation assay

For identifying anti-FVa antibodies with pro-coagulant effect, anti-FVa antibodies were identified, that were capable of increasing thrombin generation in the presence of exogenously added APC in a human plasma-based thrombin generation assay. The purified test antibodies were tested in the final assay at 0 nM - 500 nM, and the assay was run at room temperature. Briefly, human haemophilia A (HA) (factor VIII deficient) plasma (Georg King Medical, #0800) stored at -80C was thawed in water at 37C for 5 min, and then stored at room temperature until use. 18 μΙ plasma was added to a 384-well microtiter plate (Perkin Elmer, #6007659), and then 2 μΙ antibody solution (in 20 nM Tris, pH 7.4) was added, and the antigen binding was allowed to proceed for 20 min. Subsequently, 5 μΙ of a solution, containing APC (Haematologic Technologies Inc, #HCAPC-0080) spiked (100-fold dilution) into a prepared PPP-Reagent LOW reagent (Thrombinoscope, #TS31 .00), was added to the assay, resulting in APC at 2 nM, tissue factor at 1 pM and phospholipids at 4 uM in the final assay. Without incubation, 5 μΙ of a prepared FluCa reagent (Thrombinoscope, #TS50.00) was added, and a continuous reading of fluorescence was done every 30 sec for 2hrs. The thrombogram was calculated as the first derivative of the integral fluorescence curve, and the ETP and peak thrombin parameters were calculated from the thrombogram, and used in the evaluation of thrombin generation.

Antibodies that were capable of increasing thrombin generation in the presence of exogenously added APC in a human plasma-based thrombin generation assay were categorized as hits. During the screening of 500 antibodies, 35 antibodies were categorized as hits. Table 1 lists 21 antibodies out of the 35 identified as hits, including three low and non-functioning antibodies determined by the plateau from the concentration-response curve.

Table 1 . Platelet-poor plasma (PPP) TGT results.

Compound

(Origin of variable region) PPP ECso ( nM) Plateau

0233-0000-0005 (murine) 130 Low

0233-0000-0006 (murine) >100 Low

0233-0000-0008 (murine) 71 High

0233-0000-0028 (murine) 86 High

0233-0000-0056 (murine) 50 High Compound

(Origin of variable region) PPP ECso ( nM) Plateau

0233-0000-0110 (rabbit) 32 High

0233-0000-0150 (Fab of 0110) 52 High

0233-0000-0128 (murine) 148 High

0233-0000-0143 (rabbit) 58 High

0233-0000-0159 (murine) 55 High

0233-0000-0161 (murine) n/a High

0233-0000-0222 (rabbit) 134 High

0233-0000-0263 (rabbit) 5 High

0233-0000-0270 (rabbit) 80 High

0233-0000-0271 (rabbit) 5 High

0233-0000-0293 (human) n/a High

0233-0000-0296 (human) 83 High

0233-0000-0302 (human) 91 High

0233-0000-0327 (human) 101 High

0233-0000-0334 (human) 1 14 High

0233-0000-0337 (human) 342 High

0233-0000-0368 (murine) 100 High

0233-0000-0376 (human) n/a Non-functional

0233-0000-0418 (human) 38 High

0233-0000-0422 (rabbit) 54 High

0233-0000-0426 (human) 93 High

Example 3 - Stimulation of thrombin generation in normal and severe haemophilia A patient plasma Materials and Methods

The amount of thrombin generated in plasma was measured by Calibrated Automated Thrombography (Hemker et al., "Calibrated Automated Thrombin Generation Measurement in Clotting Plasma," Pathophysiol Haemost Thromb. 33:4-15 (2003); Hemker et al., "Thrombin Generation in Plasma: Its Assessment via the Endogenous Thrombin Potential," Thromb Haemost. 74:134-138 (1995)). In a 96-well plate, 72 μΙ_ of factor VIII deficient plasma pool (<1 % residual activity, platelet-poor) from severe haemophilia A patients lacking factor VIII inhibitor (George King Bio-Medical, Overland Park, Kans.) was incubated with 8 μΙ_ of antibody (or antibody fragment, HEPES-BSA buffer or recombinant Factor FVIIa) for 10 minutes at 37° C. Reactions were started by adding 20 μΙ_ of either Thrombinoscope PPP (5 pM tissue-factor and 4 μΜ phospholipid) or Thrombinoscope PPP LOW (1 pM tissue-factor and 4 μΜ phospholipid) and mixing with 20 μί fluorogenic substrate (Z-Gly-Gly-Arg-AMC) in HEPES-BSA buffer including 0.1 M CaCI₂. All reagents were pre- warmed to 37° C. The development of a fluorescent signal at 37° C was monitored at 20 second intervals using a Fluoroskan Ascent reader (Thermo Labsystems OY, Helsinki, Finland). Fluorescent signals were corrected by the reference signal from the thrombin calibrator samples (Hemker et al., "Calibrated Automated Thrombin Generation

Measurement in Clotting Plasma," Pathophysiol Haemost Thromb. 33:4-15 (2003)) and actual thrombin generation in nM was calculated as previously described (Hemker et al., "Thrombin Generation in Plasma: Its Assessment via the Endogenous Thrombin Potential," Thromb Haemost. 74:134-138 (1995)). Thrombin generation parameters peak thrombin, velocity index and endogenous thrombin potential (etp) were calculated as previously described (Hemker et al., "Data management in thrombin generation," Thromb Res 131 :3-1 1 (2013)).

Thrombin generation

Tables 2, 3 and 4 below set forth the thrombin generation parameters peak thrombin, velocity index and endogenous thrombin potential, respectively, determined in haemophilia A plasma in the presence of either Buffer, Factor VI 11 (1 % or 10%), or antibody 0233-0000-0005 (90 nM). Both in the absence and presence of APC or thrombo-modulin (TM), antibody 0233-0000-0005 yields a stronger stimulation of thrombin generation than 10% Factor VIII.

Table 2 lists Thrombin generation as a thrombin generation parameter Peak

Thrombin (in nM) from thrombin generation in haemophilia A patient plasma in the presence of various concentrations of Activated Protein C (APC) or thrombo-modulin (TM) and either Buffer, 1 % Factor VIII (0.01 lll/mL), 10% Factor VIII (0.1 IU/ml_) or 90 nM NNC-0233-0000- 0005. Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP LOW (1 pM tissue-factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal.

Table 2. Thrombin generation in haemophilia A patient plasma with different concentration APC, TM and either buffer, 1 % FVIII, 10% FVIII or 90 nM 0233-0000-0005

Table 3 lists Thrombin generation as thrombin generation parameter Velocity Index (in nM per minute) from thrombin generation in haemophilia A patient plasma in the presence of various concentrations of Activated Protein C (APC) or thrombo-modulin (TM) and either Buffer, 1 % Factor VIII (0.01 lll/mL), 10% Factor VIII (0.1 lll/mL) or 90 nM NNC-0233-0000- 0005. Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP LOW (1 pM tissue-factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal. Table 3. Thrombin generation in haemophilia A patient plasma with different concentration of

APC, TM and either buffer, 1 % FVIII, 10% FVIII or 90 nM 0233-0000-0005

In table 4, Thrombin generation is show as thrombin generation parameter endogenous thrombin potential (ETP, in nM minute) from thrombin generation in haemophilia A patient plasma in the presence of various concentrations of Activated Protein C (APC) or thrombo-modulin (TM) and either Buffer, 1 % Factor VIII (0.01 lU/mL), 10% Factor VIII (0.1 lU/mL) or 90 nM NNC-0233-0000-0005. Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP LOW (1 pM tissue-factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal.

Table 4. Thrombin generation in haemophilia A patient plasma with different concentration of APC, TM and either buffer, 1 % FVIII, 10% FVIII or 90 nM 0233-0000-0005

Dose-response

Tables 5, 6 and 7 below set forth the thrombin generation parameters peak thrombin, velocity index and endogenous thrombin potential, respectively, determined haemophilia A plasma in the presence of activated protein C (APC, 5 nM) and increasing concentrations (5 nM to 500 nM) of three antibodies (0233-0000-0005, 0233-0000-0028, 0233-0000-01 10). All three antibodies stimulated thrombin generation (as judged by either of the parameters) but to different extent, with 0233-0000-01 10 providing the most pronounced stimulation.

Table 5, shows the thrombin generation parameter Peak Thrombin (in nM) from thrombin generation in haemophilia A patient plasma in the presence of Activated Protein C (5 nM) and various antibody concentrations (5 nM to 500 nM). Last column indicates the number of independent experiments (n). Thrombin generation was triggered by

Thrombinoscope PPP (5 pM tissue-factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal. In the absence of antibody, no thrombin generation was observed under these conditions. Table 5. Thrombin generation in haemophilia A patient plasma in the presence of APC (5 nM) and various antibody concentrations (5 nM to 500 nM)

Table 6 shows thrombin generation parameter Velocity Index (in nM per minute) from thrombin generation in haemophilia A patient plasma in the presence of Activated Protein C (5 nM) and various antibody concentrations (5 nM to 500 nM). Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP (5 pM tissue-factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal. In the absence of antibody, no thrombin generation was observed under these conditions. Table 6. Thrombin generation in haemophilia A patient plasma in the presence of APC (5 nM) and various antibody concentrations (5 nM to 500 nM).

Table 7 lists the thrombin generation parameter endogenous thrombin potential (ETP, in nM minute) from thrombin generation in haemophilia A patient plasma in the presence of Activated Protein C (5 nM) and various antibody concentrations (5 nM to 500 nM) is demonstrated in table 7. Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP (5 pM tissue- factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal. In the absence of antibody, no thrombin generation was observed under these conditions.

Table 7. Thrombin generation in haemophilia A patient plasma in the presence of APC (5 nM) and various antibody concentrations (5 nM to 500 nM)

Table 8 shows the thrombin generation parameters peak thrombin, velocity index and endogenous thrombin potential determined in haemophilia A plasma in the absence or presence of activated protein C (APC, 5 nM) and various concentrations (250 nM or 500 nM) of six antibodies or antibody fragments. Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP (5 pM tissue- factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal.

In the absence of APC, some antibodies appeared to have a negative effect on all three thrombin generation parameters, whereas others were either without effect or stimulated thrombin generation. However, in the presence of APC (5 nM) all antibodies stimulated thrombin generation as judged by all three parameters.

Table 8. Thrombin generation in haemophilia A patient plasma in the absence or presence of APC (5 nM) and various antibody concentrations (250 or 500 nM)

Tables 9, 10 and 1 1 below set forth the thrombin generation parameters peak thrombin, velocity index and endogenous thrombin potential, respectively, determined in normal human plasma or protein C deficient plasma in the absence or presence of a neutralising anti-Factor VIII polyclonal antibody (0.1 mg/mL) and increasing concentrations of 0233-0000-0005 (0 nM to 500 nM). In both normal and protein C-deficient plasma 0233- 0000-0005 dose-dependently stimulated thrombin generation as judged by peak thrombin and velocity index both in the absence and presence of a neutralising anti-Factor VIII antibody.

Table 9 shows thrombin generation parameter Peak Thrombin (in nM) from thrombin generation in human normal plasma or protein C deficient plasma (PC-def.) in the presence various concentrations of 0233-0000-0005 (0 nM to 500 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (0.1 mg/mL). Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP (5 pM tissue-factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal. Table 9. Thrombin generation in normal or PC-def. plasma with 0233-0000-0005 (0 nM to 500 nM) with or without anti-Factor VIII antibody (0.1 mg/mL)

Table 10 shows the thrombin generation parameter Velocity Index (in nM per minute) from thrombin generation in human normal plasma or protein C deficient plasma (PC-def.) in the presence various concentrations of 0233-0000-0005 (0 nM to 500 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (0.1 mg/mL). Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP (5 pM tissue-factor and 4 μΜ phospholipid). NS indicates no thrombin generation signal.

Table 10. Thrombin generation in normal or PC-def. plasma with 0233-0000-0005 (0 nM to 500 nM) with or without anti-Factor VIII antibody (0.1 mg/mL)

0233-0000-0005 Velocity Index (nM per minute) a-FVIII Concentration (nM) PC-def plasma Normal plasma n no 0 49.6 55.0 2 no 20 45.8 59.2 1 no 50 51.6 63.7 1 no 100 60.9 79.9 1 no 200 62.1 77.7 1 no 500 65.6 89.3 1 yes 0 0.8 1.6 yes 20 1.7 3.3 1 yes 50 1.7 3.5 1 yes 100 1.1 4.1 1 yes 200 2.4 3.7 1 0233-0000-0005 Velocity Index (nM per minute) α-FVIII Concentration (nM) PC-def plasma Normal plasma n yes 500 2.0 4.7 1

Thrombin generation parameter Endogenous Thrombin Potential (ETP, in nMxminute) from thrombin generation in human normal plasma or protein C deficient plasma (PC-def.) in the presence various concentrations of 0233-0000-0005 (0 nM to 500 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (0.1 mg/mL) is demonstrated in table 1 1 . Last column indicates the number of independent experiments (n). Thrombin generation was triggered by Thrombinoscope PPP (5 pM tissue-factor and 4 μΜ

phospholipid). NS indicates no thrombin generation signal.

Table 1 1. Thrombin generation in normal or PC-def. plasma with 0233-0000-0005 (0 nM to 500 nM) with or without anti-Factor VIII antibody (0.1 mg/mL)

Example 4 - Stimulation of thrombelastography in haemophilia A-like blood

The elastic properties of blood during thrombus formation were measured by thrombelastography using a TEG® hemostasis analyzer (U.S. Pat. No. 5,223,227, and Luddington, RJ, "Thrombelastography/thromboelastometry.," Clin Lab Haematol. 27:81 -90 (2005)). The TEG® hemostasis analyzer monitors the elastic properties of blood as it is induced to clot under a low shear environment resembling sluggish venous blood flow. The patterns of changes in shear elasticity of the developing clot enable the determination of the kinetics of clot formation, as well as the strength and stability of the formed clot; in short, the mechanical properties of the developing clot. In a TEG® a total volume of 340 μΙ_ of preheated (37°C) human whole blood (that had been incubated with combinations of compound, neutralising polyclonal anti-Factor VIII antibody, activated protein C or thrombo-modulin) was recalcified using 20 μΙ_ calcium chloride (0.2 M) and the TEG analysis was initiated. The clotting time (R) was defined as the time from initiation to amplitude had reached 2 mm, and Maximum Rate of Thrombus Generation (MTG) was defined as the global maximum of the first derivative of amplitude in time.

Results

Tables 12A and 12B, demonstrate the thrombelastography parameters R (clot time) and Maximum Rate of Thrombin Generation (MTG), respectively, determined in normal and haemophilia A-like human blood (i.e. in the absence or presence of a neutralising anti-Factor VIII polyclonal antibody, 0.1 mg/mL) and increasing concentrations of antibodies 0233-0000- 01 10 and 0233-0000-0150 (0 nM to 400 nM). In haemophilia A-like blood both 0233-0000- 01 10 and 0233-0000-0150 stimulated thrombelastography by dose-dependently reducing the clotting time, and 0233-0000-0150 furthermore dose-dependently increased the MTG.

Table 12A shows the thrombelastography parameter clot time (R, in seconds) from thrombelastography in normal or haemophilia A-like whole blood (i.e. in the absence or presence of a neutralising anti-Factor VIII polyclonal antibody, 0.1 mg/mL) in the presence of various concentrations of antibody or antibody fragment (0 nM to 400 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (a-FVIII, 0.1 mg/mL). Last column indicates the number of independent experiments (n). Thrombelastography was initiated using 40,000- fold diluted Innovin. ND indicates not determined.

Table 12A. Clot time in normal or haemophilia A-like whole blood, with 0 nM to 400 nM antibody or antibody fragments

Table 12B shows the thrombelastography parameter Maximum Rate of Thrombus Generation (MTG, in 100xmm per second) from thrombelastography in normal or

haemophilia A-like whole blood (i.e. in the absence or presence of a neutralising anti-Factor VIII polyclonal antibody, 0.1 mg/mL) in the presence of various concentrations of antibody or antibody fragment (0 nM to 400 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (a-FVIII, 0.1 mg/mL). Last column indicates the number of independent experiments (n). Thrombelastography was initiated using 40,000-fold diluted Innovin. ND indicates not determined.

Table 12B. TMG in normal or haemophilia A-like whole blood, with 0 nM to 400 nM antibody or antibody fragments

Tables 13A and 13B below set forth the thrombelastography parameters R (clot time) and Maximum Rate of Thrombin Generation (MTG), respectively, determined in normal and haemophilia A-like human blood (i.e. in the absence or presence of a neutralising anti- Factor VIII polyclonal antibody, 0.1 mg/mL) in the presence of activated protein C (APC, 5 nM) and increasing concentrations of 0233-0000-01 10 and 0233-0000-0150 (0 nM to 400 nM). In normal and haemophilia A-like blood both 0233-0000-01 10 and 0233-0000-0150 stimulated thrombelastography by dose-dependently reducing the clotting time in the presence of APC. In normal blood both antibodies also increased the MTG in the presence of APC. Table 13A shows thrombelastography parameter clot time (R, in seconds) from thrombelastography in normal or haemophilia A-like whole blood in the presence of Activated Protein C (5 nM) and various concentrations of antibody or antibody fragment (0 nM to 400 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (a-FVIII, 0.1 mg/mL). Last column indicates the number of independent experiments (n). Thrombelastography was initiated using 40,000-fold diluted Innovin. ND indicates not determined.

Table 13A. Clot time in normal or haemophilia A-like whole blood with ACP (5 nM) and various concentrations of antibody or antibody fragment (0 nM to 400 nM)

Table 13B shows thrombelastography parameter Maximum Rate of Thrombus Generation (MTG, in 100xmm per second) from thrombelastography in normal or haemophilia A-like whole blood (i.e. in the absence or presence of a neutralising anti-Factor VIII polyclonal antibody, 0.1 mg/mL) in the presence of Activated Protein C (5 nM) and various concentrations of antibody or antibody fragment (0 nM to 400 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (a-FVIII, 0.1 mg/mL). Last column indicates the number of independent experiments (n). Thrombelastography was initiated using 40,000- fold diluted Innovin. ND indicates not determined. Table 13B. TMG in normal or haemophilia A-like whole blood with ACP (5 nM) and various concentrations of antibody or antibody fragment (0 nM to 400 nM)

Tables 14A and 14B, below set forth the thrombelastography parameters R (clot time) and Maximum Rate of Thrombin Generation (MTG), respectively, determined in normal and haemophilia A-like human blood (i.e. in the absence or presence of a neutralising anti- Factor VIII polyclonal antibody, 0.1 mg/mL) in the presence of Thrombo-modulin (TM, 5 nM) and increasing concentrations of 0233-0000-01 10 and 0233-0000-0150 (0 nM to 400 nM). In normal and haemophilia A-like blood both 0233-0000-01 10 and 0233-0000-0150 stimulated thrombelastography by dose-dependently reducing the clotting time in the presence of TM. In normal blood both antibodies also increased the MTG in the presence of TM.

Table 14A shows thrombelastography parameter clot time (R, in seconds) from thrombelastography in normal or haemophilia A-like whole blood in the presence of

Thrombo-modulin (5 nM) and various concentrations of antibody or antibody fragment (0 nM to 400 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (a-FVIII, 0.1 mg/mL) is set forth in table 16. Last column indicates the number of independent

experiments (n). Thrombelastography was initiated using 40,000-fold diluted Innovin. ND indicates not determined.

Table 14A. Clot time in normal or haemophilia A-like whole blood with Thrombo-modulin (5 nM) and various concentrations of antibody or antibody fragment (0 nM to 400 nM)

Table 14B shows hrombelastography parameter Maximum Rate of Thrombus Generation (MTG, in 100xmm per second) from thrombelastography in normal or haemophilia A-like whole blood in the presence of Thrombo-modulin (5 nM) and various concentrations of antibody or antibody fragment (0 nM to 400 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (a-FVIII, 0.1 mg/mL). Last column indicates the number of independent experiments (n). Thrombelastography was initiated using 40,000- fold diluted Innovin. ND indicates not determined. Table 14B. MTG in normal or haemophilia A-like whole blood with Thrombo-modulin (5 nM) and various concentrations of antibody or antibody fragment (0 nM to 400 nM)

Table 15A and 15B, below, show the thrombelastography parameters R (clot time) and Maximum Rate of Thrombin Generation (MTG), respectively, determined in normal and haemophilia A-like human blood (i.e. in the absence or presence of a neutralising anti-Factor VIII polyclonal antibody, 0.1 mg/mL) in the presence of activated protein C (APC, 5 nM) and increasing concentrations of recombinant activated Factor VII (rFVIIa), 0233-0000-01 10 or 0233-0000-0150 (0 nM to 400 nM). In normal and haemophilia A-like blood both rFVIIa, 0233-0000-01 10 and 0233-0000-0150 stimulated thrombelastography by dose-dependently reducing the clotting time in the presence of APC. In normal blood rFVIIa and both antibodies also increased the MTG in the presence of APC. Table 15A shows thrombelastography parameter clot time (R, in seconds) from thrombelastography in normal or haemophilia A-like whole blood in the absence and presence of Activated Protein C (APC, 5 nM) and various concentrations of recombinant activated Factor VII (rFVIIa) or antibody or antibody fragment (0 nM to 400 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (a-FVIII, 0.1 mg/mL). Last column indicates the number of independent experiments (n). Thrombelastography was initiated using kaolin. ND indicates not determined.

Table 15A. Clot time in normal or haemophilia A-like whole blood with or without 5 nM APC and various concentrations of rFVIIa or antibody or antibody fragment (0 nM to 400 nM)

Table 15B shows thrombelastography Maximum Rate of Thrombus Generation (MTG, in 100xmm per second) from thrombelastography in normal or haemophilia A-like whole blood in the absence and presence of Activated Protein C (APC, 5 nM) and various concentrations of recombinant activated Factor VII (rFVIIa) or antibody or antibody fragment (0 nM to 400 nM) with or without a neutralising polyclonal anti-Factor VIII antibody (a-FVIII, 0.1 mg/mL). Last column indicates the number of independent experiments (n).

Thrombelastography was initiated using kaolin. ND indicates not determined. Table 15B. MTG in normal or haemophilia A-like whole blood with or without 5 nM APC and various concentrations of rFVIIa or antibody or antibody fragment (0 nM to 400 nM)

Table 16 shows thrombelastography parameters clot time (R, in seconds) and MTG from thrombelastography in normal blood and blood made haemophilia A-like by incubation with a neutralising polyclonal anti-FVIII antibody (0.1 mg/mL). Various concentrations of antibody or antibody fragment (0 nM to 1000 nM) was tested in the absence of APC.

Thrombelastography was initiated with 40000-fold diluted Innovin®. All compounds dose- dependently shortened the clotting time and increased the MTG under haemophilia A-like conditions in human whole blood.

Table 16 Clot time (R) and MTG in normal (no anti-FVIII pAb) and haemophilia A-like whole blood in the presence of various concentrations antibody or antibody fragment (0 nM to 1000 nM) in the absence of APC. Values are mean ± standard deviation of the number of independent experiments (donors) indicated in parentheses.

Anti-FVIII pAb Cone Compound TEG Clot time (R) TEG MTG

(nM) (s) (100*mm/min) no 0 Buffer 398 ± 81 (8) 17.0 ± 4.4 (8) yes 0 Buffer 1945 ± 916 (8) 4.3 ± 1.7 (8) yes 10 0233-0000-01 10 3185 ± 1800 (3) 1 .7 ± 1.0 (3) yes 30 0233-0000-01 10 2305 ± 1461 (3) 2.9 ± 2.1 (3) yes 100 0233-0000-01 10 1572 ± 790 (3) 3.9 ± 3.6 (3) Anti-FVIII pAb Cone Compound TEG Clot time (R) TEG MTG

(nM) (s) (100*mm/min) yes 300 0233-0000-01 10 1723 ± 557 (3) 3.3 ± 2.6 (3) yes 1000 0233-0000-01 10 1840 ± 991 (3) 2.9 ± 3.0 (3) yes 10 0233-0000-0270 1813 ± 440 (3) 3.6 ± 1.2 (3) yes 30 0233-0000-0270 1450 ± 321 (3) 4.3 ± 1.4 (3) yes 100 0233-0000-0270 1 175 ± 465 (3) 5.3 ± 2.1 (3) yes 300 0233-0000-0270 837 ± 192 (3) 6.6 ± 2.7 (3) yes 1000 0233-0000-0270 740 ± 1 18 (3) 7.3 ± 2.7 (3) yes 10 0233-0000-0271 1465 ± 399 (3) 3.7 ± 1.1 (3) yes 30 0233-0000-0271 842 ± 145 (3) 4.9 ± 1.4 (3) yes 100 0233-0000-0271 718 ± 105 (3) 7.2 ± 2.1 (3) yes 300 0233-0000-0271 673 ± 63 (3) 8.3 ± 2.0 (3) yes 10 0233-0000-0418 4910 ± 3347 (3) 1 .2 ± 0.9 (3) yes 30 0233-0000-0418 3390 ± 2342 (3) 1 .5 ± 1.0 (3) yes 100 0233-0000-0418 2525 ± 1716 (3) 2.8 ± 2.3 (3) yes 300 0233-0000-0418 1783 ± 1 141 (3) 4.1 ± 3.4 (3) yes 1000 0233-0000-0418 1955 ± 1399 (3) 4.3 ± 2.9 (3)

Table 17 shows relative thrombelastography parameters clot time , R (in seconds) and MTG from thrombelastography in normal blood and blood made haemophilia A-like by incubation with a neutralising polyclonal anti-FVI 11 antibody (0.1 mg/mL). Various concentrations of antibody or antibody fragment (0 nM to 1000 nM) was tested in the absence of APC.

Thrombelastography was initiated with 40000-fold diluted Innovin®.

To render values comparable across individual donors (which show significant variation in absolute clotting times), the measured clotting time was expressed relative to a signal window ranging from clot time determined under normal conditions (0%) to clot time determined under haemophilia A-like conditions (100%) in the same donors in the absence of drug (Buffer).

Likewise, MTG was expressed relative to a signal window ranging from MTG determined under under haemophilia A-like conditions (0%) to MTG determined under normal conditions (100%). All compounds dose-dependently shortened the relative clotting time under haemophilia A-like conditions in human whole blood. Compounds 0233-0000- 0150, 0233-0000-0270 and 0233-0000-0271 dose-dependently stimulated the relative MTG.

Table 17 Relative clot time and MTG in normal (no anti-FVI 11 pAb) and haemophilia A-like whole blood in the presence of various concentrations antibody or antibody fragment (0 nM to 1000 nM) in the absence of APC. Values are mean ± standard deviation of the number of independent experiments (donors) indicated in parentheses.

Anti-FVIII pAb Cone Compound Relative TEG Clot Relative TEG MTG

(nM) time , R (%)

(%)

no 0 Buffer 0 100

yes 0 Buffer 100 0

yes 10 0233-0000-01 10 151 ± 25 (3) -14 ± 5 (3) yes 30 0233-0000-01 10 93 ± 12 (3) -8 ± 2 (3) yes 100 0233-0000-01 10 63 ± 8 (3) -3 ± 8 (3) yes 300 0233-0000-01 10 88 ± 47 (3) -6 ± 4 (3) yes 1000 0233-0000-01 10 77 ± 17 (3) -8 ± 5 (3) yes 10 0233-0000-0270 84 ± 48 (3) -0 ± 8 (3) yes 30 0233-0000-0270 64 ± 39 (3) 6 ± 5 (3) yes 100 0233-0000-0270 49 ± 34 (3) 12 ± 13 (3) yes 300 0233-0000-0270 29 ± 21 (3) 21 ± 15 (3) yes 1000 0233-0000-0270 22 ± 14 (3) 26 ± 14 (3) yes 10 0233-0000-0271 67 ± 47 (3) 1 ± 8 (3) yes 30 0233-0000-0271 29 ± 22 (3) 9 ± 1 1 (3) yes 100 0233-0000-0271 22 ± 16 (3) 26 ± 10 (3) yes 300 0233-0000-0271 18 + 1 1 (3) 34 ± 6 (3) yes 10 0233-0000-0418 351 ± 137 (3) -39 ± 1 1 (3) yes 30 0233-0000-0418 224 ± 1 14 (3) -36 ± 10 (3) yes 100 0233-0000-0418 151 ± 94 (3) -20 ± 7 (3) yes 300 0233-0000-0418 98 ± 64 (3) -6 ± 22 (3) yes 1000 0233-0000-0418 1 1 1 ± 82 (3) -4 ± 17 (3) Table 18 shows thrombelastography parameters clot time (R) and MTG from thrombelastography in normal blood and blood made haemophilia A-like by incubation with a neutralising polyclonal anti-FVIII antibody (0.1 mg/mL). Various concentrations of antibody or antibody fragment (0 nM to 1000 nM) were tested in the presence of APC (5 nM).

Thrombelastography was initiated with 40000-fold diluted Innovin®. All compounds dose- dependently shortened the clotting time and increased the MTG under haemophilia A-like conditions in human whole blood in the presence of APC (5 nM).

Table 18 Clot time (R) and MTG in normal (no anti-FVIII pAb) and haemophilia A-like whole blood in the presence of various concentrations antibody or antibody fragment (0 nM to 1000 nM) in the presence of APC ( 5 nM). Values are mean ± standard deviation of the number of independent experiments (donors) indicated in parentheses.

Anti-FVIII pAb Cone Compound TEG Clot time (R) TEG MTG

(nM) (s) (100*mm/min) no 0 Buffer 838 ± 218 (8) 15.0 ± 4.1 (8) yes 0 Buffer 10593 ± 392 (8) 0.5 ± 0.2 (8) yes 10 0233-0000-01 10 10800 ± 0 (3) 0.3 ± 0.1 (3) yes 30 0233-0000-01 10 7842 ± 5124 (3) 0.7 ± 0.5 (3) yes 100 0233-0000-01 10 7608 ± 5528 (3) 1.1 ± 1.5 (3) yes 300 0233-0000-01 10 7585 ± 5569 (3) 1.4 ± 2.0 (3) yes 1000 0233-0000-01 10 6075 ± 6682 (2) 1.5 ± 1.7 (2) yes 10 0233-0000-0270 10623 ± 306 (3) 0.3 ± 0.2 (3) yes 30 0233-0000-0270 9622 ± 1239 (3) 1.2 ± 0.8 (3) yes 100 0233-0000-0270 6777 ± 2885 (3) 1.7 ± 1.2 (3) yes 300 0233-0000-0270 2817 ± 2203 (3) 4.2 ± 4.7 (3) yes 1000 0233-0000-0270 1538 ± 820 (3) 4.1 ± 4.1 (3) yes 10 0233-0000-0271 4775 ± 3721 (3) 1.1 ± 0.2 (3) yes 30 0233-0000-0271 1512 ± 423 (3) 2.6 ± 2.3 (3) yes 100 0233-0000-0271 1092 ± 371 (3) 4.4 ± 3.6 (3) yes 300 0233-0000-0271 925 ± 212 (3) 5.4 ± 2.9 (3) yes 10 0233-0000-0418 7583 ± 5220 (3) 0.4 ± 0.4 (3) yes 30 0233-0000-0418 4642 ± 5383 (3) 2.1 ± 1.6 (3) yes 100 0233-0000-0418 4013 ± 4489 (3) 3.1 ± 2.7 (3) Anti-FVIII pAb Cone Compound TEG Clot time (R) TEG MTG

(nM) (s) (100*mm/min) yes 300 0233-0000-0418 3863 ± 4288 (3) 3.0 ± 2.5 (3) yes 1000 0233-0000-0418 3555 ± 4057 (3) 4.0 ± 2.8 (3)

Table 19 shows relative thrombelastography parameters clot time, R (in seconds) and MTG from thrombelastography in normal blood and blood made haemophilia A-like by incubation with a neutralising polyclonal anti-FVIII antibody (0.1 mg/mL). Various

concentrations of antibody or antibody fragment (0 nM to 1000 nM) was tested in the presence of APC (5 nM). Thrombelastography was initiated with 40000-fold diluted Innovin®. To render values comparable across individual donors (which show significant variation in absolute clotting times), the measured clotting time was expressed relative to a signal window ranging from clot time determined under normal conditions (0%) to clot time determined under haemophilia A-like conditions (100%) in the same donors in the absence of drug (Buffer). Likewise, MTG was expressed relative to a signal window ranging from MTG determined under under haemophilia A-like conditions (0%) to MTG determined under normal conditions (100%). All compounds dose-dependently shortened the relative clotting time and relative MTG under haemophilia A-like conditions in human whole blood in the presence of APC (5 nM).

Table 19 Relative clot time and MTG in normal (no anti-FVIII pAb) and haemophilia A-like whole blood in the presence of various concentrations antibody or antibody fragment (0 nM to 1000 nM) in the presence of APC (5 nM). Values are mean ± standard deviation of the number of independent experiments (donors) indicated in parentheses.

Example 5 - Enhancement of thrombin generation by anti-FVa antibodies in platelet rich plasma under haemophilia A like conditions in the presence of thrombomodulin

Anti-FVa antibodies were tested for their capacity to reverse the inhibitory effect of thrombomodulin in a thrombin generation assay including platelets. Generated thrombin was measured using a fluorogenic method from Thrombinoscope®. In brief, human platelet rich plasma (PRP) was obtained by centrifugation of citrated human whole blood at 220g for 20 min. The upper phase containing platelets was collected and the remaining sample was centrifuged at 2500g for 10 min to obtain platelet poor plasma (PPP) used to adjust the platelet concentration to a final concentration of 150000 plts/μΙ. The PRP was made haemophilic by 30 min incubation with a sheep anti-human FVIII polyclonal antibody (0.1 mg/ml) (HTI #Z0429). The haemophilic-like PRP were let to incubate with anti-FVa antibodies together with thrombomodulin (25 nM) (HTI#4202) and Innovin (1 pM) (Siemens #B4212-50) for 30 min (RT) before the reaction was started by addition of a combination of protease activated recertor-1 (PAR-1 ) activation peptide (30 μΜ) (SFLLRN; Bachem #H- 2936) and the GPVI activating Convulxin (100 ng/ml) (Pentapharm #1 19-02) together with the FluCa reagent to the samples in 96-well Nunc Microwell round bottom well plates (Nunc #268152). The fluorescent signal from the substrate was detected in a ThermoFisher Fluoroskan plate reader (Fisher Scientific). The thrombin concentration was calculated using a Thrombin calibrator provided by Thrombinoscope® according to their instructions.

The results showed a dose dependent reversal of the thrombomodulin (TM) inhibiting effect on thrombin generation by antibody 0233-0000-01 10, as shown in table 20. Thrombin generation was measured under induced haemophilia A conditions (sheep anti- human FVIII polyclonal antibodies) with a fixed platelet concentration of 150000 plts/μΙ and with added thrombomodulin (25 nM) and innovin (1 pM). The reaction was started with platelet activation through the addition of PAR-1 activating peptide SFLLRN (30 μΜ) and the GPIV agonist Convulxin (100 ng/ml). Peak thrombin generation (nM) is shown as mean values. NA indicates not applicable.

Table 20. Dose dependent reversal of the thrombomodulin (TM) inhibiting effect on thrombin generation by antibody 0233-0000-01 10

Data from other anti-FVa antibodies at two concentrations (50nM and 500nM) are presented in table 21 as percent of peak thrombin generated by 0233-0000-01 10 (500nM) under the conditions described above.

Table 21. Peak thrombin generation by various anti-FVa antibodies in human PRP with induced haemophilia A containing thrombomodulin (25nM)

Example 6 - Evaluating prior-art anti-FVa monoclonal antibodies in human haemophilic plasma by thrombin generation assay

Commercially available antibodies were procured to be tested in the same assay, that we identified the antibodies represented in this patent (see example 2). The commercial antibodies usually represented the same antibody clone across multiple products from different vendors in different buffer conditions.

The commercial antibodies were tested at the highest possible concentration to maximize the chance of identifying a pro-coagulant antibody, and the assay was run at room temperature. Briefly, human haemophilia A (HA) (factor VIII deficient) plasma (Georg King Medical, #0800) stored at -80C was thawed in water at 37C for 5 min, and then stored at room temperature until use. 18 μΙ_ plasma was added to a 384-well microtiter plate (Perkin Elmer), and then 2 μΙ_ antibody solution (in 20 nM Tris, pH 7.4) was added, and the antigen binding was allowed to proceed for 20 min. Subsequently, 5 μΙ_ of a solution, containing APC (Haematologic Technologies Inc, #HCAPC-0080) spiked (100-fold dilution) into a prepared PPP-Reagent LOW reagent (Thrombinoscope, #TS31 .00), was added to the assay, resulting in APC at 2 nM, tissue factor at 1 pM and phospholipids at 4 μΜ in the final assay. Without incubation, 5 μΙ_ of a prepared FluCa reagent (Thrombinoscope, #TS50.00) was added, and a continuous reading of fluorescence was done every 30 sec for 2hrs. The thrombogram was calculated as the first derivative of the integral fluorescence curve, and the ETP and peak thrombin parameters were calculated from the thrombogram, and used in the evaluation of thrombin generation. Table 22 lists the results of all the antibodies tested, along with the background (APC only) and the 0233-0000-01 10 antibody disclosed in this invention. The result value is the peak height relative to the 0233-0000-01 10 antibody.

As can be seen, one of the commercially available antibodies showed similar effect in the assay, as the 0233-0000-01 10 antibody, that is the AHV-5102 antibody, from Pierce Biotechnology, with product id MA1 -43001 . Another antibody, the 3H367 antibody, from US Biological, with id F001 1 -23 had a peak height of 0,8 compared with the reference 0233- 0000-01 10 antibody. As can be seen in example 7, that shows the binning of different antibodies, including the AHV-5102 antibody, from Pierce Biotechnology, with product id MA1 -43001 and the 3H367 antibody, from US Biological, with id F001 1 -23, neither of those antibodies actually bind to activated Factor V. Table 22. Platelet-poor plasma TGT results of commercially available antibodies

Clone Vendor Product Id Peak Height

0233-0000-0110 1 ,0

Background 0,4

701 GenWay Biotech, GWB-B41 D1 F 0,3

Inc.

701 abeam ab20951 0,4

802 Creative Biomart CABT-27898MH 0,4

1 B9 Creative Biomart CABT-18075MH 0,0

1C3 Creative Biomart CABT-18076MH 0,5 Clone Vendor Product Id Peak Height

2E3-C2 Creative Biomart CABT-18080MH 0,3

2F11 Creative Biomart CABT-18077MH 0,3

3F4-2C6 Creative Biomart CABT-27895MH 0,3

3H365 US Biological F001 1-21 0,4

3H367 US Biological F001 1-23 0,8

3H369 US Biological F001 1-25 0,4

4D7-2D4 Creative Biomart CABT-27896MH 0,4

4H5 Creative Biomart CABT-27892MH 0,4

6A5 Santa Cruz sc-13512 0,3

Biotechnology, inc.

7G1 Creative Biomart CABT-18078MH 0,5

8E6 Creative Biomart CABT-18079MH 0,3

A812 Creative Biomart CABT-20174MH 0,4

A813 Creative Biomart CABT-20175MH 0,4

AHV-5102 LSBio LS-C23351 0,4

AHV-5102 Novus Biologicals NBP1 -9501 1 0,6

AHV-5102 Molecular MA-HFV-2 0,6

Innovations

AHV-5102 HTI AHV-5102 0,7

AHV-5102 Pierce MA1-43001 1 ,0

Biotechnology

AHV-5108 Gentauer/Molecular MA-HFV-3 0,0

Innovations

AHV-5108 Novus Biologicals NBP1 -95012 0,2

AHV-5108 abeam ab61413 0,3

AHV-5108 LSBio LS-C23352 0,3

AHV-5108 Pierce MA1 -43002 0,4

Biotechnology

AHV-5110 Molecular MA-HFV-1 0,0

Innovations

AHV-5110 Novus Biologicals NBP1 -95010 0,0

AHV-5110 Santa Cruz sc-73458 0, 1 Clone Vendor Product Id Peak Height

Biotechnology, inc.

AHV-5110 HTI AHV-51 10 0,1

AHV-5110 Gentauer/Molecular MA-HFV-4 0,2

Innovations

AHV-5110 Novus Biologicals NBP1 -95013 0,3

AHV-5112 Molecular MA-HFV-5 0,2

Innovations

AHV-5112 LSBio LS-C36300 0,3

AHV-5112 LSBio LS-C23354 0,4

AHV-5146 abeam ab61348 0,4

AHV-5146 Novus Biologicals NBP1 -95015 0,5

AHV-5146 HTI AHV-5146 0,5

AHV-5146 Molecular MA-HFV-6 0,5

Innovations

B056M Creative Biomart CABT-27897MH 0,4

B10 Acris Antibodies 19702 0,3

B10 LSBio LS-C83328 0,3

B10 Pierce MA1-10827 0,3

Biotechnology

B10 abeam ab1015 0,4

B38 Acris Antibodies 19701 0,4

B38 LSBio LS-C83327 0,4

CBC-MOR101 abeam ab1009 0,5

DCD-NPS202 Creative Biomart CABT-27899MH 0,4

EPR5191 abeam ab108614 0, 1

EPR5191 LSBio LS-C138518 0,2

FQS6202 Creative Biomart CABT-27894RH 0,2

GMA-044 GMA Antibodies GMA-044 0,0

GMA-044 Santa Cruz sc-65945 0, 1

Biotechnology, inc.

M5020721 Antibodies Online ABIN283599 0,2

M5020721 LSBio LS-C84453 0,4 Clone Vendor Product Id Peak Height n/a GenWay Biotech, GWB-4ED1 12 0,3

Inc.

n/a abeam ab61347 0,4

n/a American 235 0,5

Diagnostica, Inc.

Example 7 - Binning of anti-FVa monoclonal antibodies

Surface plasmon resonance (Biacore T200) was used to address competition among the identified antibodies (see table 1 in example 1 ).

Initially, the antibodies were divided into two groups those competing with 0233-

0000-01 10 for binding to FVa and those not competing. In brief, antibodies were captured on an anti-mouse IgG or an anti-human IgG coated CM5 chip depending on the origin of the individual antibodies. The binding of human FVa either alone or in complex with antibody was monitored. All antibodies assigned to bin 1 compete with 0233-0000-01 10 for binding to FVa.

The antibodies that were found not to compete with 0233-0000-01 10 for binding to FVa was further subdivided by direct coating of the antibodies on a CM5 chip and measuring binding of FVa alone or incubated with the respective antibodies. Based on these results the antibodies were divided into additional four bins.

The antibodies assigned to bin 5 (commercially available antibodies, see example 6, above) bind the activation peptide of FV and thus do not bind activated FVa and does thus not fall in any of the binds of the antibodies identified herein. The result of this binning can be seen in table 23. Table 23 - Binning of anti-FVa monoclonal antibodies

Binning of antibodies against 0233-0000-0150 (fab of -01 10) that binds to bin 1 of table 23, were done in a Bio-Layer Interferometry (BLI) assay on an Octett system from Fortebio. Using anti-murine Fey specific tip human FVa specific antibodies were captured from the supernatant. Afterwards human FVa was bound to the surface by dipping the tips into a 50nM hFVa solution. The tips were directly transferred after the association to wells containing 50nM fab and the binding was measured for ~10s. Antibodies which did not bind in the same bin as -0150 showed significant binding in these first 10s, whereas antibodies in the same bin as 0150 didn't bind in the presence of 0150.

Table 24. Binning of anti-FVa monoclonal antibodies against 0233-0000-0150

Example 8 - Epitope mapping

This example identifies the interaction surface on human plasma FVa when binding the antibodies 0233-0000-0005, 0233-0000-0006, 0233-0000-01 10, 0233-0000-0128, 0233- 0000-0271 , 0233-0000-0453 (Fab of 0233-0000-0418), and 0233-0000-0302, respectively. Residues in the sequence for FVa light chain (SEQ ID NO 38) used in this example are numbered as the first amino acid residue being no. 1546. The numbering can be converted to the sequence numbering for SEQ ID NO 38 by subtracting 1545 from the residue number used in this example. E.g. the residue S1546 used in this example corresponds to residue S1 in SEQ ID NO 38.

Hydrogen exchange mass spectrometry (HX-MS) exploits that the change in mass when hydrogen is exchanged with deuterium can be followed over time using mass spectrometry. When an antibody binds to FVa, amino acid residues located in the binding interface are protected from hydrogen exchange with the solvent. Pepsin digestion allows identification of peptides that are part of the epitope that show slower rates of hydrogen exchange when the antibody is bound. The borders of an epitope can be further defined by using information from overlapping peptides. If a peptide that is not protected from hydrogen- deuterium exchange is partially overlapping with a protected peptide, the common sequence between the two peptides will not be part of the epitope. The N-terminal amino group of the peptide backbone back-exchanges rapidly with the solvent during the chromatography and any deuterium label on that position will be lost. Therefore, it is not possible to conclude whether the N-terminal residue of a peptide is part of an epitope or not. HX-MS can also reveal other features such as structural flexibility/stabilization and changes in these features upon binding of an antibody. These experiments were performed using human FVa purified from human blood plasma. Solutions of FVa alone or in the presence of one of the antibodies or Fabs 0233-

0000-0005, 0233-0000-0006, 0233-0000-01 10, 0233-0000-0128, 0233-0000-0271 , 0233- 0000-0453, or 0233-0000-0302 were diluted 25-fold in 95% deuterated imidazole buffer (20 mM imidazole, 150 mM sodium chloride, 10 mM calcium chloride, pH 7.4). Non-deuterated controls were prepared by diluting into protiated imidazole buffer. The hydrogen exchange experiments were performed on a nanoAcquity UPLC system with HDX technology (Waters Corporation, Milford, MA, USA) which includes the HD-x PAL auto sampler (LEAP

Technologies Inc., Carrboro, NC, USA) for automated sample preparation and an ultra-high performance liquid chromatography (UPLC) system. The UPLC tubing, pre- and analytical columns and switching valves were located in a chamber cooled to 0.3°C. The trypsin digestion column was stored at 15°C. Hydrogen exchange reactions were performed at 20°C. Mass analysis was performed online using a Waters SYNAPT G2 HDMS mass spectrometer.

A volume containing 100 pmol of FVa with or without 120 pmol of one of the antibodies or antibody fragments was diluted into deuterated imidazole buffer. At the time intervals from 15 seconds to 4 hours 50 μΙ of the sample was quenched in 50 μΙ 1 .35 mM Tris (2-carboxyethyl) phosphine adjusted to pH 2.7 and held at 3°C. 99 μΙ of the quenched solution was immediately injected and passed over either a Porozyme immobilized pepsin column (2.1 mm x 30 mm) (Applied Biosystems, Life Technologies Corporation, Carlsbad, CA, USA) (0233-0000-0005, 0233-0000-0006, 0233-0000-01 10, 0233-0000-0128, and 0233-0000- 0302) or a Waters Enzymate column (2.1 mm x 30 mm) (0233-0000-0271 or 0233-0000- 0453) and trapped on a Waters VanGuard BEH C18 1.7 μηη (2.1 mm 5 mm) column using a 5% methanol, 0.1 % formic acid mobile phase and either a 100 μΙ/min flow rate (0233-0000- 0005, 0233-0000-0006, 0233-0000-01 10, 0233-0000-0128, and 0233-0000-0302) or a flow rate sequence of 400 μΙ/min for 15 seconds, 10 μΙ/min for 2 minutes, and 400 μΙ/min for 1 minute and 45 seconds (0233-0000-0271 or 0233-0000-0453). The peptides were separated on a Waters UPLC BEH C18 1.7 μηι (1 .0 mm x 100 mm) column using a 9 min 10 - 40 % acetonitrile gradient containing 0.1 % formic acid at a 40 μΙ/min flow-rate. The mass spectrometer was run in positive ion mode with ion mobility separation enabled. The instrument parameters used were 3.2 kV capillary, 30 V sample cone, and 4 V extraction cone offsets, 850 ml/min flow of desolvation gas and 25 ml/min cone gas flow. The source block was heated to 120°C and the desolvation gas to 350°C. Lock-mass correction data using acquired using the 1 + ion of Leucine-enkephalin (m/z 556.2771 ) as reference compound and applied during data analysis. The ions were separated by ion mobility in 3 mbar pressure of nitrogen maintained by a 90 ml/min gas flow. The trap and transfer regions were flushed with a 2 ml/min gas flow of argon gas to trap the ions before and after ion mobility separation and for collision induced dissociation in MSe-type experiments. For peptide identification MSe-type experiments using trap collision offsets of 6 V (low-energy scan) and either a low activation (10-30 V), medium activation (15-35 V) or high activation (20-45 V) energy ramps (elevated energy/fragmentation scan) were performed. Deuterated samples were analysed using the 6 V low energy trap collision offset only. For further details see Andersen et al, Int. J. Mass Spectrom. 302:139-148 (201 1 ). The MSE-data was analysed using Waters ProteinLynx Global Server 2.5 and peptides of FVa were identified that covered 82 % (0233-0000-0005 and 0233-0000-0006, see table 25), 87 % (0233-0000-01 10, 0233-0000-0128, and 0233-0000-0302, see table 26), and 83 % (0233-0000-0271 or 0233-0000-0453, see table 27) of the protein sequence, respectively. The HX-MS data files were analysed using Waters DynamX 2.0 that automatically applies lock-mass correction and determines the degree of deuterium incorporation in each peptide. In addition, all data was manually inspected to ensure correct peak assignment and calculation of deuterium incorporation.

Table 25: Epitope mapping of mAbs 0233-0000-0005 and 0233-0000-0006 on human plasma FVa

Sequence Start End 0005 0006

YVAAQGISW 7 15 None None

KKIVYREYEPYF 34 45 Protection None

KKIVYREYEPYFKKEKPQSTISGLLGPTL 34 62 Protection None

REYEPYFKKEKPQSTISGL 39 57 Protection None

REYEPYFKKEKPQSTISGLLGPTL 39 62 Protection None

YEPYFKKEKPQSTISGLLGPTL 41 62 Protection None

KKEKPQSTISGLLGPTL 46 62 Protection None

ISGLLGPTL 54 62 None None

YAEVGDIIKVHFKNKADKPLSIHPQGIRY 63 91 None None

YAEVGDIIKVHFKNKADKPLSIHPQGIRYS

63 94 None None KL

YAEVGDIIKVHFKNKADKPLSIHPQGIRYS

63 99 None None KLSEGAS

IIKVHFKNKADKPLSIHPQGIRY 69 91 None None

IIKVHFKNKADKPLSIHPQGIRYSKL 69 94 None None

IKVHFKNKADKPLSIHPQGIRY 70 91 None None

IKVHFKNKADKPLSIHPQGIRYSK 70 93 None None

KVHFKNKADKPLSIHPQGIRY 71 91 None None

VHFKNKADKPLSIHPQGIRY 72 91 None None

PLSIHPQGIRY 81 91 None None

YLDHTFPAE 100 108 None None

YLDHTFPAEKMDD 100 1 12 None None

YLDHTFPAEKMDDAVAPGRE 100 1 19 None None

LDHTFPAEKMDD 101 1 12 None None

LDHTFPAEKMDDAVAPGRE 101 1 19 None None

DHTFPAEKMDDAVAPGRE 102 1 19 None None

YEWSISEDSGPTHDDPPCL 122 140 None None

WSISEDSGPTHDDPPCL 124 140 None None

WSISEDSGPTHDDPPCLTHIYYSHE 124 148 None None

WSISEDSGPTHDDPPCLTHIYYSHENL 124 150 None None

ISEDSGPTHDDPPCL 126 140 None None

Low Low

DSGPTHDDPPCL 129 140

exchange exchange

DSGPTHDDPPCLTHIYYSHENL 129 150 None None

THIYYSHENL 141 150 None None

Low Low

IEDFNSGL 151 158

exchange exchange

Low Low

FNSGLIGPLL 154 163

exchange exchange

IGPLLICKKGTLTEGGTQKTFDKQ 159 182 Protection None

GPLLICKKGTLTEGGTQKTFDKQ 160 182 Protection None

ICKKGTLTEGGTQKTFDKQ 164 182 Protection None Sequence Start End 0005 0006

DESKSWSQSSSL 191 202 None None

Low Low

CAHDHISWHL 220 229

exchange exchange

Low Low

CAHDHISWHLLG 220 231

exchange exchange

Low Low

CAHDHISWHLLGM 220 232

exchange exchange

Low Low

AHDHISWHLLGM 221 232

exchange exchange

MSSGPEL 232 238 None None

FSIHFNG 239 245 None None

FSIHFNGQVL 239 248 None None

FSIHFNGQVLEQNHHKVSA 239 257 None None

IISSLTPKHLQAGMQ 279 293 None None

IISSLTPKHLQAGMQA 279 294 None None

IISSLTPKHLQAGMQAY 279 295 None None

AYIDIKNCPKKTRNLKKITREQRRHMKR

294 325 None None WEYF

YIDIKNCPKKTRNLKKITREQRRHMKRW

295 325 None None EYF

YAP VI PAN M 335 343 None None

PVIPANM 337 343 None None

DKKYRSQHLDNF 344 355 None None

DKKYRSQHLDNFSNQIGKHYKKVMYTQ

344 371 None None Y

FSNQIGKHYKKVMY 355 368 None None

FSNQIGKHYKKVMYTQY 355 371 None None

FSNQIGKHYKKVMYTQYEDES 355 375 None None

FTKHTVNPNM 376 385 None None

FTKHTVNPNMKEDGILGPIIRA 376 397 None None

TKHTVNPNMKEDGILGPIIRA 377 397 None None

TKHTVNPNMKEDGILGPIIRAQVRDTL 377 403 None None

KEDGILGPIIRAQVRDTL 386 403 None None

Low Low

GILGPIIRA 389 397

exchange exchange

Low Low

LGPIIRA 391 397

exchange exchange

PIIRAQVRDTL 393 403 None None

IRAQVRDTLKIVFKNMASR 395 413 None None

QVRDTLKIVF 398 407 None None

KNMASRPYSIYPHGVTF 408 424 None None

KNMASRPYSIYPHGVTFSPYE 408 428 None None

IRAVQPGETYT 444 454 None None

YTYKWNIL 453 460 None None

YKWNILE 455 461 None None

LTRPYYSDVD 473 482 None None

VDIMRDIASGL 481 491 None None

LLICKSRSLDRRGIQ 495 509 None None

LLICKSRSLDRRGIQRAAD 495 513 None None

ICKSRSLDRRGIQRAAD 497 513 None None Sequence Start End 0005 0006

Low Low

IEQQAVF 514 520

exchange exchange

AVFDENKSWYLED 521 533 None None

AVFDENKSWYLEDNINKF 521 538 None None

FDENKSWYLEDNINKFCENPDEVKRDDP

523 552 None None KF

NINKFCENPDEVKRDDPKF 534 552 None None

CENPDEVKRDDPKF 539 552 None None

PDEVKRDDPKF 542 552 None None

Low Low

IMSTING 557 563

exchange exchange

Low Low

CFDDTVQWHF 575 584

exchange exchange

Low Low

FDDTVQWHF 576 584

exchange exchange

Low Low

DDTVQWHF 577 584

exchange exchange

CSVGTQNEIL 585 594 None None

Low Low

TIHFTGHS 595 602

exchange exchange

Low Low

TIHFTGHSFIYGKRHEDTL 595 613

exchange exchange

Low Low

FIYGKRHEDTL 603 613

exchange exchange

Low Low

FIYGKRHEDTLTL 603 615

exchange exchange

Low Low

IYGKRHEDTL 604 613

exchange exchange

LTLFPMRGESVT 613 624 None None

TLFPMRGESVT 614 624 None None

FPMRGESVT 616 624 None None

VTMDNVGTW 625 633 None None

MDNVGTW 627 633 None None

MLTSMNSSPRSKKLRL 634 649 None None

MLTSMNSSPRSKKLRLKF 634 651 None None

ATRKMHDRLEPEDEESDADYDYQNRL 677 702 None None

ISWDYSE 1561 1567 None None

DSDDIPEDTTYKKVVF 1577 1592 None None

SDDIPEDTTYKKVVF 1578 1592 None None

PEDTTYKKWFRKYLDST 1582 1599 None None

FTKRDPRGEYEEHL 1600 1613 None None

FTKRDPRGEYEEHLGILG 1600 1617 None None

FTKRDPRGEYEEHLGILGPIIRA 1600 1622 None None

TKRDPRGEYEEHL 1601 1613 None None

TKRDPRGEYEEHLGILG 1601 1617 None None

TKRDPRGEYEEHLGILGPIIRA 1601 1622 None None

PRGEYEEHLGILGPIIRA 1605 1622 None None

Low Low

EEHLGILGPIIRA 1610 1622

exchange exchange

PIIRAEVDD 1618 1626 Low Low Sequence Start End 0005 0006

exchange exchange

DVIQVRFKNLASRPYSL 1626 1642 None None

Low Low

VIQVRFKNL 1627 1635

exchange exchange

VIQVRFKNLASRPYSLHAHGLS 1627 1648 None None

IQVRFKNLASRPYSL 1628 1642 None None

QVRFKNLASRPYSL 1629 1642 None None

VRFKNLASRPYSLHAHGLS 1630 1648 None None

YEKSSEGKTYEDDSPEW 1649 1665 None None

YVWHATERSGPESPGSA 1680 1696 None None

YVWHATERSGPESPGSACRA 1680 1699 None None

WAYYSAVNPEKDIHSGL 1700 1716 None None

YYSAVNPEKDIHSGL 1702 1716 None None

YSAVNPEKDIHSGL 1703 1716 None None

YSAVNPEKDIHSGLIGPLL 1703 1721 None None

SAVNPEKDIHSGLIGPLL 1704 1721 None None

ICQKGILHKDSNMPMD 1722 1737 None None

MTFDEKKSWYYEKKSRSSWRLTSSE 1746 1770 None None

DEKKSWYYEKKSRSSWRLTSSE 1749 1770 None None

Low Low

MKKSHEFHAING 1771 1782

exchange exchange

Low Low

MKKSHEFHAINGMI 1771 1784

exchange exchange

IYSLPGLKM 1784 1792 None None

YSLPGLKM 1785 1792 None None

HLLNIGGSQDIHVVHFHGQTL 1801 1821 None None

LNIGGSQDIHWHF 1803 1816 None None

LNIGGSQDIHWHFHGQTLL 1803 1822 None None

LENGNKQHQLGVW 1822 1834 None None

LENGNKQHQLGVWPLL 1822 1837 None None

LENGNKQHQLGVWPLLPGSF 1822 1841 None None

ENGNKQHQLGVWPLLPGSF 1823 1841 None None

PLLPGSF 1835 1841 None None

Low Low

MKASKPGW 1846 1853

exchange exchange

Low Low

MKASKPGWWLL 1846 1856

exchange exchange

Low Low

KASKPGWWLLNT 1847 1858

exchange exchange

NTEVGENQRAGMQTPFL 1857 1873 None None

TEVGENQRAGMQTPFL 1858 1873 None None

EVGENQRAGMQTPFL 1859 1873 None None

Low Low

QRAGMQTPFL 1864 1873

exchange exchange

Low Low

QRAGMQTPFLIM 1864 1875

exchange exchange

IMDRDCRMPMGL 1874 1885 None None

STGIISD 1886 1892 None None

STGIISDSQIKASE 1886 1899 None None Sequence Start End 0005 0006

GIISDSQIKASEF 1888 1900 None None

IISDSQIKASEF 1889 1900 None None

ARLNNGGSYNAW 1909 1920 None None

ARLNNGGSYNAWSVEKL 1909 1925 None None

NAWSVEKL 1918 1925 None None

FASKPWIQV 1929 1937 None None

FASKPWIQVD 1929 1938 None None

Low Low

MQKEVIITG 1939 1947

exchange exchange

VIITGIQTQGAKHYLKS 1943 1959 None None

ITGIQTQGAKHYLKSCY 1945 1961 None None

TGIQTQGAKHYLKSCY 1946 1961 None None

IQTQGAKHYLKSCY 1948 1961 None None

IQTQGAKHYLKSCYTTE 1948 1964 None None

QINWQIF 1973 1979 None None

IKENQFDPPIVA 1999 2010 None None

IKENQFDPPIVARY 1999 2012 None None

YIRISPTRA 2012 2020 None None

IRISPTRA 2013 2020 None None

IRISPTRAYNRPTL 2013 2026 None None

IRISPTRAYNRPTLRL 2013 2028 None None

ELQGCEVNGCSTPLGMENGKIENKQITA

2029 2058 None None SS

EVNGCSTPLGMENGKIENKQITASS 2034 2058 None None

EVNGCSTPLGMENGKIENKQITASSF 2034 2059 None None

CSTPLGMENGKIENKQITASSF 2038 2059 None None

SSFKKSWWGDYWEPFRARL 2057 2075 None None

FKKSWWGDY 2059 2067 None None

FKKSWWGDYWEPFRARL 2059 2075 None None

KKSWWGDYWEPFRARL 2060 2075 None None

Low Low

YWEPFRARL 2067 2075

exchange exchange

Low Low

WEPFRARL 2068 2075

exchange exchange

Low Low

EPFRARL 2069 2075

exchange exchange

NAQG RVN AWQAKAN N N KQWL 2076 2095 None Protection

NAQG RVN AWQAKAN N N KQWLE 2076 2096 None Protection

NAQGRVNAWQAKANNNKQWLEIDL 2076 2099 None Protection

WQAKAN N N KQWL 2084 2095 None Protection

WQAKANNNKQWLEIDL 2084 2099 None Protection

LKIKKITA 2100 2107 None None

IITQGCKSLSSE 2108 21 19 None None

TQGCKSLSSEM 21 10 2120 None None

YVKSYTI HYSEQGVEWKPYRLKSSM 2121 2145 None Protection

YTIHYSEQ 2125 2132 None None

YTIHYSEQGVEWKPYRL 2125 2141 None Protection

YTIHYSEQGVEWKPYRLKSSM 2125 2145 None Protection

SEQGVEWKPYRL 2130 2141 None Protection Sequence Start End 0005 0006

SEQGVEWKPYRLKSSM 2130 2145 None Protection

GVEWKPYRL 2133 2141 None Protection

VDKIFEGNTNTKGHVKNFFNPPIISR 2146 2171 None None

VDKIFEGNTNTKGHVKNFFNPPIISRF 2146 2172 None None

KIFEGNTNTKGHVKNFFNPPIISRF 2148 2172 None None

Low Low

FFNPPIISRF 2163 2172

exchange exchange

Low Low

PPIISRF 2166 2172

exchange exchange

IRVIPKTWNQS 2173 2183 None None

IRVIPKTWNQSIA 2173 2185 None None

Table 26: Epitope mapping of mAbs 0233-0000-01 10, 0233-0000-0128, and 0233-0000- 0302 on human plasma FVa

Protection Protection Protection

Sequence Start End

0110 0128 0302

RQFYVAAQGISW 4 15 None None None

SVTSFKKIVYREYEPYFK

29 62 None None None KEKPQSTISGLLGPTL

KKIVYREYEPYF 34 45 None None None

KKIVYREYEPYFKKEKPQ

34 53 None None None ST

KKEKPQSTISGLLGPTL 46 62 None None None

LYAEVGDIIKVHFKNKAD

62 93 None None None KPLSIHPQGIRYSK

YAEVG DIIKVHFKNKADK

63 91 None None None PLSIHPQGIRY

GDIIKVHFKNKADKPLSIH

67 94 None None None PQGIRYSKL

IIKVHFKNKADKPLSIHPQ

69 91 None None None GIRY

SEGASYL 95 101 None None None

SEGASYLDHTFPAEKMD

95 1 12 None None None D

YLDHTFPAEKMDD 100 1 12 None None None

YLDHTFPAEKMDDAVAP

100 1 19 None None None GRE

YLDHTFPAEKMDDAVAP

100 121 None None None GREYT

AVAPGREYTYE 1 13 123 None None None

WSISEDSGPTHDDPPCL

124 148 None None None THIYYSHE

DSGPTHDDPPCL 129 140 None None None

DSGPTHDDPPCLTHIYYS

129 150 None None None HENL

IEDFNSGL 151 158 None None None

FNSGLIGPLL 154 163 Low Low Low Protection Protection Protection

Sequence Start End

0110 0128 0302 exchange exchange exchange

LICKKGTLTEGGTQKT 163 178 None None None

AVFDESKSWSQSSSL 188 202 None None None

AVFDESKSWSQSSSLM 188 203 None None None

DESKSWSQSSSL 191 202 None None None

VCAHDHISWHLLGMSSG

219 238 None None None PEL

CAHDHISWHLLGMSSGP

220 238 None None None EL

Low Low Low

AHDHISWHL 221 229

exchange exchange exchange

Low Low Low

AHDHISWHLLGM 221 232

exchange exchange exchange

Low Low Low

HDHISWHLLGM 222 232

exchange exchange exchange

LGMSSGPEL 230 238 None None None

FSIHFNG 239 245 None None None

ITLVSATST 258 266 None None None

KWIISSLTPKHLQ 277 289 None None None

IISSLTPKHLQAGMQ 279 293 None None None

IISSLTPKHLQAGMQA 279 294 None None None

IISSLTPKHLQAGMQAY 279 295 None None None

AYIDIKNCPKKTRNLKKIT

294 324 None None None REQRRHMKRWEY

YIDIKNCPKKTRNLKKITR

295 324 None None None EQRRHMKRWEY

YIDIKNCPKKTRNLKKITR

EQRRHMKRWEYFIAAEE 295 343 None Protection None

VIWDYAPVIPANM

IDIKNCPKKTRNLKKITRE

296 324 None None None QRRHMKRWEY

IDIKNCPKKTRNLKKITRE

QRRHMKRWEYFIAAEEV 296 343 None Protection None

IWDYAPVIPANM

EEVIWDYAPVIPANM 329 343 None Protection None

EVIWDYAPVIPANM 330 343 None Protection None

VIWDYAPVIPANM 331 343 None None None

VIWDYAPVI PANMDKKYR

331 355 None None None SQHLDNF

Low Low Low

WDYAPVIPANMDK 333 345

exchange exchange exchange

DKKYRSQHLDNF 344 355 None None None

FSNQIGKHYKKVMY 355 368 None None None

FSNQIGKHYKKVMYTQ 355 370 None None None

FSNQIGKHYKKVMYTQY

355 374 None None None EDE

FSNQIGKHYKKVMYTQY

355 375 None None None EDES

SNQIGKHYKKVMYTQYE 356 375 None None None Protection Protection Protection

Sequence Start End

0110 0128 0302

DES

FTKHTVNPNM 376 385 None None None

FTKHTVNPNMKEDGILG

376 397 None None None PIIRA

FTKHTVNPNMKEDGILG

376 403 None None None PIIRAQVRDTL

KEDGILGPIIRAQVRDTLK

386 407 None None None IVF

KIVFKNMASRPYSIYPHG

404 424 None None None VTF

KIVFKNMASRPYSIYPHG

404 428 None None None VTFSPYE

KNMASRPYSIYPHGVTF 408 424 None None None

KNMASRPYSIYPHGVTF

408 428 None None None SPYE

IRAVQPGET 444 452 None None None

IRAVQPGETYT 444 454 None None None

I RAVQPGETYTYKWN I LE 444 461 None None None

YTYKWNIL 453 460 None None None

YKWNILE 455 461 None None None

FDEPTENDAQC 462 472 None None None

LTRPYYSD 473 480 None None None

LTRPYYSDV 473 481 None None None

LTRPYYSDVD 473 482 None None None

VDIMRDIASGL 481 491 None None None

LLICKSRSLDRRGIQRAA

495 513 None None None D

IEQQAVF 514 520 None None None

Low Low Low

AVFDENKSWY 521 530

exchange exchange exchange

Low Low Low

AVFDENKSWYL 521 531

exchange exchange exchange

AVFDENKSWYLED 521 533 None None None

AVFDENKSWYLEDNINK

521 538 None Protection None F

AVFDENKSWYLEDNINK

521 552 None Protection None FCENPDEVKRDDPKF

FDENKSWYLEDNINKFC

523 552 None Protection None ENPDEVKRDDPKF

DENKSWYLED 524 533 None None None

DENKSWYLEDNINKF 524 538 None Protection None

DENKSWYLEDNINKFCE

524 552 None Protection None NPDEVKRDDPKF

WYLEDNINKF 529 538 None Protection None

LEDNINKF 531 538 None Protection None

NINKFCENPDE 534 544 None Protection None

NINKFCENPDEVKRDDP

534 552 None Protection None KF

NKFCENPDEVKRDDPKF 536 552 None Protection None Protection Protection Protection

Sequence Start End

0110 0128 0302

CENPDEVKRDDPKF 539 552 None None None

PDEVKRDDPKF 542 552 None None None

VKRDDPKF 545 552 None None None

IMSTINGYVPESITT 557 571 None None None

IMSTINGYVPESITTL 557 572 None None None

IMSTINGYVPESITTLG 557 573 None None None

STINGYVPESITT 559 571 None None None

STINGYVPESITTL 559 572 None None None

STINGYVPESITTLGF 559 574 None None None

Low Low Low

CFDDTVQWHF 575 584

exchange exchange exchange

Low Low Low

DDTVQWHF 577 584

exchange exchange exchange

Low Low Low

DTVQWHF 578 584

exchange exchange exchange

CSVGTQNEIL 585 594 None None None

TIHFTGHS 595 602 None None None

TIHFTGHSFIYGKRHEDT

595 613 None None None L

FIYGKRHEDTLTLF 603 616 None None None

LTLFPMRGESVT 613 624 None None None

TLFPMRGESVT 614 624 None None None

FPMRGES 616 622 None None None

VTMDNVGTW 625 633 None None None

VTMDNVGTWM 625 634 None None None

MLTSMNSSPRSKKLRL 634 649 None None None

TSMNSSPRSKKLRL 636 649 None None None

SSPRSKKLRLKF 640 651 None None None

SPRSKKLRLKF 641 651 None None None

PRSKKLRLKF 642 651 None None None

IFEPPESTVM 667 676 None None None

KMHDRLEPEDEESDADY

680 702 None None None DYQNRL

AAALGIR 703 709 None None None

SNNGNRRNYYIA 1546 1557 None None None

SNNGNRRNYYIAAEE 1546 1560 None None None

ISWDYSEF 1561 1568 None None None

FVQRETDIE 1568 1576 None None None

VQRETDIEDSDDIPEDTT

1569 1592 None None None YKKWF

IEDSDDIPEDTTYKKWF

1575 1599 None None None RKYLDST

IEDSDDIPEDTTYKKWF

1575 1600 None None None RKYLDSTF

SDDIPEDTTYKKVVF 1578 1592 None None None

YKKWFRKYLDST 1587 1599 None None None

YKKWFRKYLDSTF 1587 1600 None None None

FTKRDPRGEYEEHL 1600 1613 None None None

FTKRDPRGEYEEHLGILG 1600 1622 None None None Protection Protection Protection

Sequence Start End

0110 0128 0302

PIIRA

TKRDPRGEYEEHL 1601 1613 None None None

TKRDPRGEYEEHLGILG 1601 1617 None None None

TKRDPRGEYEEHLGILGP

1601 1622 None None None IIRA

TKRDPRGEYEEHLGILGP

1601 1626 None None None IIRAEVDD

TKRDPRGEYEEHLGILGP

IIRAEVDDVIQVRFKNLAS 1601 1642 None None None

RPYSL

PRGEYEEHLGILGPIIRA 1605 1622 None None None

Low Low Low

EEHLGILGPIIRA 1610 1622

exchange exchange exchange

EEHLGILGPIIRAEVDDVI

QVRFKNLASRPYSLHAH 1610 1646 None None None G

HLGILGPIIRA 1612 1622 None None None

PIIRAEVDD 1618 1626 None None None

VDDVIQVRFKNLASRPYS 1624 1641 None None None

DVIQVRFKNLASRPYSL 1626 1642 None None None

DVIQVRFKNLASRPYSLH

1626 1648 None None None AHGLS

Low Low Low

VIQVRFKNL 1627 1635

exchange exchange exchange

VIQVRFKNLASRPYSL 1627 1642 None None None

VIQVRFKNLASRPYSLHA

1627 1648 None None None HGLS

Low Low Low

IQVRFKNL 1628 1635

exchange exchange exchange

IQVRFKNLASRPYSL 1628 1642 None None None

Low Low Low

QVRFKNL 1629 1635

exchange exchange exchange

VRFKNLASRPYSLHAHG

1630 1648 None None None LS

VRFKNLASRPYSLHAHG

1630 1650 None None None LSYE

KNLASRPYSLHAHGLS 1633 1648 None None None

ASRPYSLHAHGLS 1636 1648 None None None

SRPYSLHAHGLS 1637 1648 None None None

YEKSSEGKTYEDDSPEW 1649 1665 None None None

YVWHATERSGPESPGSA 1680 1696 None None None

YVWHATERSGPESPGSA

1680 1697 None None None C

YVWHATERSGPESPGSA

1680 1699 None None None CRA

PGSACRA 1693 1699 None None None

WAYYSAVNPEKDIHSGL 1700 1716 None None None

WAYYSAVNPEKDIHSGLI

1700 1721 None None None GPLL Protection Protection Protection

Sequence Start End

0110 0128 0302

YSAVNPEKDIHSGL 1703 1716 None None None

YSAVNPEKDIHSGLIGPL

1703 1721 None None None L

SAVNPEKDIHSGL 1704 1716 None None None

SAVNPEKDIHSGLIGPLL 1704 1721 None None None

ICQKGIL 1722 1728 None None None

ICQKGILHKDSNMPMD 1722 1737 None None None

ICQKGILHKDSNMPMDM

1722 1740 None None None RE

HKDSNMPMD 1729 1737 None None None

HKDSNMPMDM 1729 1738 None None None

Low Low Low

FMTFDEKKSWY 1745 1755

exchange exchange exchange

Low Low Low

MTFDEKKSWY 1746 1755

exchange exchange exchange

DEKKSWYYEKKSRSSW

1749 1770 None None None RLTSSE

YEKKSRSSWRLTSSE 1756 1770 None None None

MKKSHEFHAINGM 1771 1783 None None None

MKKSHEFHAINGMI 1771 1784 None None None

M KKS H EF H Al N GM I YS LP

1771 1792 None None None GLKM

SHEFHAINGMIY 1774 1785 None None None

MIYSLPGLKM 1783 1792 None None None

IYSLPGLKM 1784 1792 None None None

YSLPGLKM 1785 1792 None None None

VRLHLLNIGGSQDIHWH

1798 1821 None None None FHGQTL

HLLNIGGSQDIHVVHFHG

1801 1821 None None None QTL

LNIGGSQ 1803 1809 None None None

DIHWHFHGQTL 1810 1821 None None None

IHWHFHGQTL 181 1 1821 None None None

VHFHGQTLLENGNKQHQ 1814 1830 None None None

VHFHGQTLLENGNKQHQ

1814 1833 None None None LGV

QTLLENGNKQHQLGVW

1819 1841 None None None PLLPGSF

LENGNKQHQLGVW 1822 1834 None None None

LENGNKQHQLGVWPLLP

1822 1841 None None None GSF

ENGNKQHQLGVWPLLP

1823 1841 None None None GSF

PLLPGSF 1835 1841 None None None

Low Low Low

KTLEMKASKPGWWLL 1842 1856

exchange exchange exchange

Low Low Low

MKASKPGWWLL 1846 1856

exchange exchange exchange

KASKPGW 1847 1853 Low Low Low Protection Protection Protection

Sequence Start End

0110 0128 0302 exchange exchange exchange

Low Low Low

KASKPGWWLLNT 1847 1858

exchange exchange exchange

NTEVGENQRAGMQTPFL 1857 1873 None None

VGENQRAGMQTPFL 1860 1873 None None

VGENQRAGMQTPFLIM 1860 1875 None None

Low Low Low

QRAGMQTPFLIM 1864 1875

exchange exchange exchange

IMDRDCRM 1874 1881 None None None

IMDRDCRMPMG 1874 1884 None None None

IMDRDCRMPMGL 1874 1885 None None None

IMDRDCRMPMGLSTGIIS

1874 1900 None None None DSQIKASEF

MDRDCRM 1875 1881 None None None

DRDCRMPMGLSTGIISD

1876 1900 None None None SQIKASEF

CRMPMGLSTGIISD 1879 1892 None None None

CRMPMGLSTGIISDSQIK

1879 1900 None None None ASEF

LSTGIISDSQIKASEF 1885 1900 None None None

STGIISDSQIKASEF 1886 1900 None None None

GIISDSQIKASEF 1888 1900 None None None

IISDSQIKASE 1889 1899 None None None

IISDSQIKASEF 1889 1900 None None None

Weak

FLGYWEPRL 1900 1908 None None stabilization

ARLNNGGSY 1909 1917 None None None

Weak

ARLNNGGSYNAW 1909 1920 None None stabilization

ARLNNGGSYNAWSVE 1909 1923 Stabilization None None

ARLNNGGSYNAWSVEKL 1909 1925 Stabilization None None

NAWSVEKL 1918 1925 Stabilization None None

FASKPWIQVDM 1929 1939 None None None

FASKPWIQVDMQ 1929 1940 None None None

FASKPWIQVDMQKEVIIT

1929 1961 None None None GIQTQGAKHYLKSCY

ASKPWIQVD 1930 1938 None None None

ASKPWIQVDM 1930 1939 None None None

IQVDMQKEVIITGIQTQGA 1935 1953 None None None

QKEVIITGIQTQGAKHYLK

1940 1961 None None None SCY

VIITGIQTQGAKHYLKS 1943 1959 None None None

VIITGIQTQGAKHYLKSC 1943 1960 None None None

VIITGIQTQGAKHYLKSCY

1943 1964 None None None TTE

VIITGIQTQGAKHYLKSCY

1943 1965 None None None TTEF

ITGIQTQGAKHYLKS 1945 1959 None None None

ITGIQTQGAKHYLKSCY 1945 1961 None None None Protection Protection Protection

Sequence Start End

0110 0128 0302

IQTQGAKHYLKSCY 1948 1961 None None None

IKENQFDPPIVA 1999 2010 None None None

IKENQFDPPIVAR 1999 201 1 None None None

IKENQFDPPIVARY 1999 2012 None None None

PPIVARY 2006 2012 None None None

RYIRISPTRA 201 1 2020 None None None

YIRISPTRA 2012 2020 None None None

YIRISPTRAYNRPTLRL 2012 2028 None None None

IRISPTRA 2013 2020 None None None

IRISPTRAYNRPTL 2013 2026 None None None

IRISPTRAYNRPTLRL 2013 2028 None None None

RISPTRA 2014 2020 None None None

RISPTRAYNRPTLRL 2014 2028 None None None

PTRAYNRPTLRL 2017 2028 None None None

Low Low Low

YNRPTLRL 2021 2028

exchange exchange exchange

Low Low Low

YNRPTLRLEL 2021 2030

exchange exchange exchange

Low Low Low

NRPTLRL 2022 2028

exchange exchange exchange

ELQGCEVNGCSTPLGME

2029 2059 None None Protection NGKIENKQITASSF

CSTPLGMENGKIENKQIT

2038 2058 None None None ASS

SSFKKSWWGDYWEPFR

2057 2075 None None None ARL

FKKSWWGDYWEPFRAR

2059 2075 None None None L

KKSWWGDYWEPFRARL 2060 2075 None None None

Low Low Low

YWEPFRARL 2067 2075

exchange exchange exchange

Low Low Low

WEPFRARL 2068 2075

exchange exchange exchange

Low Low Low

EPFRARL 2069 2075

exchange exchange exchange

N AQG RVN AWQAKAN N N

2076 2095 Protection None Protection KQWL

N AQG RVN AWQAKAN N N

2076 2099 Protection None Protection KQWLEIDL

WQAKAN N N KQWL 2084 2095 Protection None Protection

ANNNKQWLEI 2088 2097 None None None

LKIKKITAIITQGCKSLSSE

2100 2120 None None None M

IITQGCKSLSSE 2108 21 19 None None None

IITQGCKSLSSEM 2108 2120 None None None

Low Low Low

YVKSYTIHY 2121 2129

exchange exchange exchange Protection Protection Protection

Sequence Start End

0110 0128 0302

YVKSYTIHYSEQGVEWK

2121 2141 Protection None Protection PYRL

YTIHYSEQ 2125 2132 None None None

YTIHYSEQGVEWKPYRL 2125 2141 Protection None Protection

YTI HYSEQGVEWKPYRL

2125 2145 Protection None Protection KSSM

SEQGVEWKPYRLKSSM 2130 2145 Protection None Protection

VDKIFEGNTNTKGHVKNF

2146 2171 Stabilization None Stabilization FNPPIISR

VDKIFEGNTNTKGHVKNF

2146 2172 Stabilization None Stabilization FNPPIISRF

KIFEGNTNTKGHVKNFFN Weak Weak

2148 2172 None

PPIISRF stabilization stabilization

FFNPPIISRF 2163 2172 Stabilization None Stabilization

PPIISRF 2166 2172 Stabilization None Stabilization

IRVIPKTWNQS 2173 2183 None None None

IRVIPKTWNQSIA 2173 2185 None None None

Table 27: Epitope mapping of mAb 0233-0000-0271 and the antibody fragment 0233-0000- 0453 derived from 0233-0000-0418.

Protection Protecti o

Sequence start end 0271 n 0453

AQLRQFYVAA 1 10 None None

AQLRQFYVAAQGISW 1 15 None None

FYVAAQGISW 6 15 None None

SVTSFKKIVYREYEPYFKKEKPQSTISG None None LLGPTL 29 62

KKIVYREYEPYF 34 45 None None

KKIVYREYEPYFKKEKPQSTISGL 34 57 None None

KKEKPQSTISGLLGPTL 46 62 None None

ISGLLGPTL 54 62 None None

GLLGPTLYAE 56 65 Destabilization None

YAEVGDIIKVHFKNKADKPLSIHPQGIR Destabilization None Y 63 91

IIKVHFKNKADKPLSIHPQGIRY 69 91 Destabilization None

IIKVHFKNKADKPLSIHPQGIRYSKL 69 94 Destabilization None

SEGASYLDHTFPAEKMDD 95 1 12 Destabilization None YLDHTFPAE 100 108 Destabilization None

YLDHTFPAEKMDD 100 1 12 Destabilization None

YLDHTFPAEKMDDAVAPGREYTYE 100 123 Destabilization None

LDHTFPAEKMDD 101 1 12 Destabilization None

DHTFPAEKMDD 102 1 12 Destabilization None

AVAPGREYTYE 1 13 123 None None

EWSISEDSGPTHDDPPCLTHIYYSH 123 147 Destabilization None

WSISEDSGPTHDDPPCL 124 140 Destabilization None

WSISEDSGPTHDDPPCLTHIYYSHENL 124 150 Destabilization None

DSGPTHDDPPCL 129 140 Destabilization None

DSGPTHDDPPCLTHIYYSHENL 129 150 Destabilization None

THIYYSHENL 141 150 None None

Low exchange Low

FNSGLIGPLL 154 163 exchange

IGPLLICKKGTLTEGGTQKTFDKQ 159 182 None None

GPLLICKKGTLTEGGTQKTFDKQI 160 183 None None

ICKKGTLTEGGTQKTFDKQ 164 182 None None

FAVFDESKSWSQSSSL 187 202 None None

FAVFDESKSWSQSSSLM 187 203 None None

AVFDESKSWSQSSSLM 188 203 None None

DESKSWSQSSSLM 191 203 None None

ESKSWSQSSSLM 192 203 None None

Low exchange Low

CAHDHISWHL 220 229 exchange

Low exchange Low

CAHDHISWHLLG 220 231 exchange

Low exchange Low

CAHDHISWHLLGM 220 232 exchange

Weak None

CAHDHISWHLLGMSSGPEL 220 238 destabilization

Low exchange Low

AHDHISWHLLGM 221 232 exchange

Low exchange Low

HDHISWHLLG 222 231 exchange Low exchange Low

HDHISWHLLGM 222 232 exchange

ISWHLLGMSSGPELFSIHFN 225 244 Destabilization None

HLLGMSSGPEL 228 238 Destabilization None

LGMSSGPEL 230 238 Destabilization None

MSSGPEL 232 238 Destabilization None

FSIHFNG 239 245 Destabilization None

FSIHFNGQ 239 246 Destabilization None

FSIHFNGQVL 239 248 Destabilization None

IISSLTPKHLQA 279 290 Destabilization None

IISSLTPKHLQAGMQ 279 293 Destabilization None

IISSLTPKHLQAGMQA 279 294 Destabilization None

IISSLTPKHLQAGMQAY 279 295 Destabilization None

EEVIWDYAPVIPANM 329 343 Protection None

EVIWDYA 330 336 Protection None

EVIWDYAPVIPANM 330 343 Protection None

YAPVIPANM 335 343 None None

DKKYRSQHLDNF 344 355 None None

DKKYRSQHLDNFSNQIGKHYKKVMYT Protection None QY 344 371

FSNQIGKHYKKVMY 355 368 Protection None

FSNQIGKHYKKVMYTQY 355 371 Protection None

FSNQIGKHYKKVMYTQYEDES 355 375 Protection None

SNQIGKHYKKVMYTQY 356 371 Protection None

SNQIGKHYKKVMYTQYEDES 356 375 Protection None

EDESFTKHTVNPNM 372 385 Protection None

FTKHTVNPNMKEDGILGPIIRA 376 397 None None

FTKHTVNPNMKEDGILGPIIRAQVRDTL 376 403 None None

KEDGILGPIIRAQVRDTL 386 403 None None

KEDGILGPIIRAQVRDTLKIVF 386 407 None None

Low exchange Low

LGPIIRA 391 397 exchane

Weak None

PIIRAQVRDTL 393 403 protection QVRDTLKIVF 398 407 None None

KIVFKNMASRPYSIYPHGVTF 404 424 Destabilization None

KIVFKNMASRPYSIYPHGVTFSPYE 404 428 Destabilization None

KNMASRPYSIYPHGVTF 408 424 Destabilization None

KNMASRPYSIYPHGVTFSPYED 408 429 Destabilization None

IRAVQPGETYT 444 454 None None

YKWNILEF 455 462 Destabilization None

FDEPTENDAQC 462 472 None None

LTRPYYSDV 473 481 None None

LTRPYYSDVD 473 482 None None

Low exchange Low

MRDIASGL 484 491 exchange

Low exchange Low

RDIASGL 485 491 exchange

LLICKSRSLDRRGIQRAAD 495 513 None None

LICKSRSLDR 496 505 None None

LICKSRSLDRRGIQRAADI 496 514 None None

IEQQAVF 514 520 None None

Low exchange Low

AVFDENKS 521 528 exchange

Low exchange Low

AVFDENKSW 521 529 exchange

Low exchange Low

AVFDENKSWY 521 530 exchange

AVFDENKSWYLED 521 533 None None

AVFDENKSWYLEDNINKF 521 538 Protection None

AVFDENKSWYLEDNINKFCENPDE 521 544 Protection None

DENKSWYLED 524 533 None None

WYLEDNINKF 529 538 Protection None

NINKFCENPDE 534 544 Protection None

NINKFCENPDEVKRDDPKF 534 552 Protection None

NKFCENPDEVKRDDPKF 536 552 Protection None

Weak None

CENPDEVKRDDPKF 539 552 protection Weak None

NPDEVKRDDPKF 541 552 protection

IMSTING 557 563 None None

IMSTINGYVPESITT 557 571 None None

STINGYVPESITTL 559 572 None None

CFDDTVQWHF 575 584 None None

Low exchange Low

DTVQWHF 578 584 exchange

CSVGTQNEIL 585 594 None None

ILTIHFTGHS 593 602 None None

IHFTGHS 596 602 None None

FIYGKRHEDTL 603 613 Destabilization None

FIYGKRHEDTLTL 603 615 Destabilization None

IYGKRHEDTLTL 604 615 Destabilization None

YGKRHEDTLT 605 614 Destabilization None

TLFPMRG 614 620 None None

TLFPMRGESVT 614 624 None None

FPMRGES 616 622 None None

PMRGESVT 617 624 None None

VTMDNVGTWM 625 634 Destabilization None

DNVGTWM 628 634 Destabilization None

MLTSMNSSPRSKKLRLKF 634 651 None None

NSSPRSKKLRLKF 639 651 None None

DADYDYQNRL 693 702 None None

AAALGIR 703 709 None None

SNNGNRRNYYIA 1546 1557 None None

SNNGNRRNYYIAA 1546 1558 None None

SNNGNRRNYYIAAE 1546 1559 None None

SNNGNRRNYYIAAEE 1546 1560 None None

ISWDYSE 1561 1567 None None

VQRETDIEDSDDIPEDTTYKKVVF 1569 1592 None None

IEDSDDIPEDTT 1575 1586 None None

DSDDIPEDTTYKKVVF 1577 1592 None None

YKKVVFRKYLDST 1587 1599 None None FTKRDPRGEYEEHL 1600 1613 None None

FTKRDPRGEYEEHLGILG 1600 1617 None None

FTKRDPRGEYEEHLGILGPIIRA 1600 1622 None None

TKRDPRGEYEEHL 1601 1613 None None

TKRDPRGEYEEHLGILG 1601 1617 None None

TKRDPRGEYEEHLGILGPIIRA 1601 1622 None None

TKRDPRGEYEEHLGILGPIIRAEVDD 1601 1626 None None

Low exchange Low

EEHLGILGPIIRA 1610 1622 exchange

Low exchange Low

HLGILGPIIRA 1612 1622 exchange

EVDDVIQVRFKNLASRPYSLHAHGLS 1623 1648 None None

DVIQVRFKNLASRPYSL 1626 1642 None None

Low exchange Low

VIQVRFKNL 1627 1635 exchange

VIQVRFKNLASRPYSLHAHGLS 1627 1648 None None

VRFKNLASRPYSL 1630 1642 None None

VRFKNLASRPYSLHAHGLS 1630 1648 None None

KNLASRPYSL 1633 1642 None None

KNLASRPYSLHAHGLS 1633 1648 None None

YEKSSEGKTYEDDSPEW 1649 1665 None None

EKSSEGKTYEDDSPEW 1650 1665 None None

YVWHATERSGPESPGSA 1680 1696 None None

YVWHATERSGPESPGSACRA 1680 1699 None None

WAYYSAVNPEKDIHSGL 1700 1716 None None

WAYYSAVNPEKDIHSGLIGPLL 1700 1721 None None

YYSAVNPEKDIHSGL 1702 1716 None None

YYSAVNPEKDIHSGLIGPLL 1702 1721 None None

YSAVNPEKDIHSGLIGPL 1703 1720 None None

YSAVNPEKDIHSGLIGPLL 1703 1721 None None

SAVNPEKDIHSGLIGPLL 1704 1721 None None

ICQKGIL 1722 1728 None None

ICQKGILHKDSNMPMD 1722 1737 None None

ICQKGILHKDSNMPMDMRE 1722 1740 None None Low exchange Low

FMTFDEKKSWY 1745 1755 exchange

Low exchange Low

MTFDEKKSWY 1746 1755 exchange

Low exchange Low

DEKKSWY 1749 1755 exchange

DEKKSWYYEKKSRSSWRLTSSE 1749 1770 Protection Protection

YYEKKSRSSWRLTSSE 1755 1770 Protection Protection

YEKKSRSSWRL 1756 1766 Protection Protection

YEKKSRSSWRLTSSE 1756 1770 Protection Protection

YEKKSRSSWRLTSSEM 1756 1771 Protection Protection

Low exchange Low

MKKSHEFHAINGM 1771 1783 exchange

Low exchange Low

MKKSHEFHAINGMI 1771 1784 exchange

Low exchange Low

KKSHEFHAINGMI 1772 1784 exchange

MIYSLPGLKM 1783 1792 None None

WVRLHLLNIGGSQ 1797 1809 Destabilization None

VRLHLLNIGGSQ 1798 1809 Destabilization None

VRLHLLNIGGSQDIHVVHF 1798 1816 Destabilization None

VRLHLLNIGGSQDIHVVHFHGQTL 1798 1821 Destabilization None

LNIGGSQ 1803 1809 Destabilization None

Low exchange Low

DIHWHF 1810 1816 exchange

DIHWHFHGQTL 1810 1821 Destabilization None

DIHWHFHGQTLL 1810 1822 Destabilization None

LENGNKQHQLGVW 1822 1834 Destabilization None

LENGNKQHQLGVWPLL 1822 1837 Destabilization None

LENGNKQHQLGVWPLLPGSF 1822 1841 Destabilization None

ENGNKQHQLGVWPLLPGSF 1823 1841 Destabilization None

PLLPGSF 1835 1841 Destabilization None

Destabilization Low

KTLEMKASKPGWWLL 1842 1856 exchange Destabilization Low

MKASKPGW 1846 1853 exchange

MKASKPGWWLLNT 1846 1858 Destabilization None

KASKPGW 1847 1853 Destabilization None

Destabilization Low

KASKPGWWLL 1847 1856 exchange

Destabilization Low

KASKPGWWLLNT 1847 1858 exchange

WLLNTEVGENQRAGMQTPFL 1854 1873 Destabilization None

NTEVGENQRAGMQTPFL 1857 1873 Destabilization None

EVGENQRAGMQTPFL 1859 1873 Destabilization None

EVGENQRAGMQTPFLIM 1859 1875 Destabilization None

NQRAGMQTPFL 1863 1873 Destabilization None

QRAGMQTPFL 1864 1873 Destabilization None

Destabilization Low

QRAGMQTPFLIM 1864 1875 exchange

IMDRDCRMPMG 1874 1884 Destabilization None

IMDRDCRMPMGL 1874 1885 Destabilization None

LSTGIISDSQIKASEF 1885 1900 Destabilization None

STGIISD 1886 1892 Destabilization None

STGIISDSQIKASEF 1886 1900 Destabilization None

GIISDSQIKASEF 1888 1900 Destabilization None

IISDSQIKASEF 1889 1900 Destabilization None

FLGYWEPRL 1900 1908 Destabilization None

LGYWEPRL 1901 1908 Destabilization None

ARLNNGGSYNAW 1909 1920 Destabilization None

EFASKPWIQVDM 1928 1939 Destabilization None

FASKPWIQ 1929 1936 Destabilization None

FASKPWIQVD 1929 1938 Destabilization None

FASKPWIQVDM 1929 1939 Destabilization None

ASKPWIQVD 1930 1938 Destabilization None

MQKEVIITG 1939 1947 Destabilization None

QKEVIITGIQTQGAKHYLKSCY 1940 1961 Destabilization None

VIITGIQTQGAKHYLKSCY 1943 1961 Destabilization None GIQTQGAKHYLKSCY 1947 1961 Destabilization None

IQTQGAKHYLKSCY 1948 1961 Destabilization None

YFNGNSDAST 1989 1998 Destabilization None

IKENQFDPPIVA 1999 2010 Destabilization None

IKENQFDPPIVAR 1999 201 1 Destabilization None

IKENQFDPPIVARY 1999 2012 Destabilization None

KENQFDPPIVARYI 2000 2013 Destabilization None

PPIVARY 2006 2012 Destabilization None

RYIRISPTRAYNRPTLRL 201 1 2028 Destabilization None

YIRISPTRAYNRPTLRL 2012 2028 Destabilization None

IRISPTRAYNRPTL 2013 2026 Destabilization None

IRISPTRAYNRPTLRL 2013 2028 Destabilization None

IRISPTRAYNRPTLRLEL 2013 2030 Destabilization None

PTRAYNRPTLRL 2017 2028 Destabilization None

YNRPTLRLEL 2021 2030 Destabilization None

NRPTLRL 2022 2028 Destabilization None

NGKIENKQITASSF 2046 2059 None None

Low exchange Low

IENKQITASSF 2049 2059 exchange

Weak None

FKKSWWGDYWEPFRARL 2059 2075 protection

Weak None

KKSWWGD 2060 2066 protection

Weak None

KKSWWGDY 2060 2067 protection

Weak None

KKSWWGDYWEPFRARL 2060 2075 protection

Low exchange Low

YWEPFRARL 2067 2075 exchange

N AQG RVN AWQAKAN N N KQWL 2076 2095 None None

NAQGRVNAWQAKANNNKQWLEIDL 2076 2099 Protection None

WQAKAN N N KQWL 2084 2095 None None

WQAKANNNKQWLE 2084 2096 None None

WQAKAN NNKQWLEIDL 2084 2099 Protection None ANNNKQWLEI 2088 2097 None None

LKIKKITAIITQGCKSLSSE 2100 21 19 Protection None

LKI KKITAI ITQGCKSLSSEM 2100 2120 Protection None

YVKSYTIHYSEQGVEWKPYRLKSSM 2121 2145 Protection None

YTIHYSEQ 2125 2132 None None

YTI HYSEQGVEWKPYRL 2125 2141 None None

YTI HYSEQGVEWKPYRLKSSM 2125 2145 None None

SEQGVEWKPYRL 2130 2141 None None

SEQGVEWKPYRLKSSM 2130 2145 None None

GVEWKPYRLKSSM 2133 2145 None None

VDKIFEGNTNTKGHVKNFFNPPIISR 2146 2171 None None

KIFEGNTNTKGHVKNFFNPPIISRF 2148 2172 None None

FIRVIPKTWNQSIA 2172 2185 None None

IRVIPKTWNQS 2173 2183 None None

IRVIPKTWNQSIA 2173 2185 None None

IRVIPKTWNQSIAL 2173 2186 None None

Low exchange Low

IALRLEL 2184 2190 exchange

Epitope mapping of 0233-0000-0271 and 0233-0000-0418 (Fab 0233-0000-0453),

Amide hydrogen exchange of 262 peptides covering 83 % of the FVa protein sequence were followed in the presence or absence of one of the antibodies 0233-0000- 0271 or the Fab fragment 0233-0000-0453. The peptides could be divided into groups of 1 ) peptides that displayed an exchange pattern that was largely unaffected by binding of the antibody, 2) Peptides whose exchange pattern was markedly protected from hydrogen- deuterium exchange by antibody binding indicating that the peptide comprises part of the epitope or that the structure in this region is stabilized by antibody binding, 3) peptides whose exchange pattern was affected in a small, but significant, degree by antibody binding indicating that only a few residues in the peptides are part of the epitope, and 4) Peptides that displayed an increase in hydrogen-deuterium exchange, and 5) Peptides where the changes in exchange pattern upon antibody binding could not be determined due to low intrinsic hydrogen exchange in the sequence covered by the peptide. An exchange protection was interpreted as being due to antibody binding epitope if there was significant protection of more than 0.5 Da in the earliest time points of less than 2 minutes. An exchange protection was interpreted as being due to structural change or structural stabilization if the exchange protection was weaker than 0.5 Da. An increase in hydrogen- deuterium exchange was interpreted as a structural change.

Epitope mapping of mAbs 0233-0000-0005. 0233-0000-0006. 0233-0000-01 10. 0233-0000- 0128 and 0233-0000-0302

Amide hydrogen exchange of 210 peptides covering 82 % of the FVa protein sequence (0233-0000-0005 and 0233-0000-0006) and 275 peptides covering 87 % of the FVa protein sequence (0233-0000-01 10, 0233-0000-0128, and 0233-0000-0302) were followed in the presence or absence of one of the antibodies 0233-0000-0005, 0233-0000- 0006, 0233-0000-01 10, 0233-0000-0128, or 0233-0000-0302. The peptides could be divided into groups of 1 ) peptides that displayed an exchange pattern that was largely unaffected by binding of the antibody, 2) Peptides whose exchange pattern was markedly protected from hydrogen-deuterium exchange by antibody binding indicating that the peptide comprises part of the epitope or that the structure in this region is stabilized by antibody binding, 3) peptides whose exchange pattern was protected from hydrogen-deuterium exchange in a small, but significant, degree by antibody binding indicating that only a few residues in the peptides are part of the epitope, 4) Peptides where an increase in hydrogen-deuterium exchange was observed upon antibody or Fab binding indicating a structural change or destabilization, and 5) Peptides where the changes in exchange pattern upon antibody binding could not be determined due to low intrinsic hydrogen exchange in the sequence covered by the peptide. An exchange protection was interpreted as being due to antibody binding epitope if there was significant protection of more than 0.2 Da in the earliest time points of less than 2 minutes. An exchange protection was interpreted as being due to structural stabilization if the exchange protection was weak at the short incubation times and increasing at longer incubation times.

Epitope of 0233-0000-0005 on human FVa

Two segments of the FVa heavy-chain sequence were significantly affected by exchange protection upon binding of 0233-0000-0005. Peptides covering the sequence from K34 to L62 showed significant protection from hydrogen exchange upon binding of the 0233-0000- 0005 antibody. However, as peptides covering the sequence from I54 to L62 showed no protection upon antibody binding, this part of the sequence could be dismissed as being part of the epitope. Thus, part of the epitope could be confirmed to be within the sequence from K34 to I54. A second part of the sequence was also protected from hydrogen exchange upon binding of the 0233-0000-0005 antibody. This region includes peptides from 1159 to Q182. However, as peptides covering the sequence from F154 to L163 showed low intrinsic exchange this part of the sequence could not be confirmed as being part of the epitope.

Thus, part of the epitope could be confirmed to be within the sequence from 1164 to

Q182. The epitope of 0233-0000-0005 comprises residues within the regions K34 to I54 and 1164 to Q182 and those residues are K34, K35, I36, V37, Y38, R39, E40, Y41 , E42, P43, Y44, F45, K46, K47, E48, K49, P50, Q51 , S52, T53, I54, 1164, C165, K166, K167, G168, T169, L170, T171 , E172, G173, G174, T175, Q176, K177, T178, F179, D180, K181 , and Q182.

Epitope of 0233-0000-0006 on human FVa

Two segments of the FVa light-chain sequence were significantly affected by exchange protection upon binding of 0233-0000-0006. One segment of the sequence was covered by peptides from N2076 to L2099 and showed significant protection from hydrogen exchange upon binding of the 0233-0000-0006 antibody.

Thus, part of the epitope could be confirmed to be within the sequence from A2077 to L2099.

A second part of the sequence was also protected from hydrogen exchange upon binding of the 0233-0000-0006 antibody. This region included peptides from Y2121 to M2145.

However, as peptides covering the sequence from Y2125 to Q2132 showed no protection upon antibody binding, this part of the sequence could be dismissed as being part of the epitope. Thus, part of the epitope could be confirmed to be within the sequence from V2122 to Y2125 and the sequence from G2133 to M2145.

The epitope of 0233-0000-0006 comprises residues within the regions A2077 to L2099, Y2121 to Y2125, and G2133 to M2145 of the FVa light-chain sequence and those residues are A2077, Q2078, G2079, R2080, V2081 , N2082, A2083, W2084, Q2085, A2086, K2087, A2088, N2089, N2090, N2091 , K2092, Q2093, W2094, L2095, E2096, I2097, D2098, L2099, V2122, K2123, S2124, Y2125, G2133, V2134, E2135, W2136, K2137, P2138, Y2139, R2140, L2141 , K2142, S2143, S2144, and M2145. Epitope of 0233-0000-01 10 and 0233-0000-0302 on human FVa

Two segments of the FVa light-chain sequence is significantly affected by exchange protection upon binding of either of the antibodies 0233-0000-01 10 and 0233-0000-0302. One segment of the sequence was covered by peptides from N2076 to L2099 and showed significant protection from hydrogen exchange upon binding of either of the antibodies 0233- 0000-01 10 and 0233-0000-0302. However, as peptides covering the sequence from A2088 to I2097 showed no protection upon antibody binding, this part of the sequence could be dismissed as being part of the epitope.

Thus, part of the epitope could be confirmed to be within the sequence from A2077 to A2088 and D2098-L2099.

A second part of the sequence was also protected from hydrogen exchange upon binding of the 0233-0000-01 10 antibody. This region included peptides from Y2121 to M2145.

However, as a peptide covering the sequence from Y2125 to Q2132 showed no protection upon antibody binding and since a peptide covering the sequence from Y2121 to Y2129 had low intrinsic exchange, it could not be confirmed if the residues from Y2121 to Y2129 comprise part of the epitope and the sequence from S2130 to Q2132 could be dismissed as being part of the epitope, Thus, part of the epitope could be confirmed to be within the sequence from G2133 to M2145.

The epitope of 0233-0000-01 10 and 0233-0000-0302 comprises residues within the regions A2077 to A2088, D2098-L2099, and G2133 to M2145 of the FVa light-chain sequence and those residues are A2077, Q2078, G2079, R2080, V2081 , N2082, A2083, W2084, Q2085, A2086, K2087, A2088, D2098, L2099, G2133, V2134, E2135, W2136, K2137, P2138, Y2139, R2140, L2141 , K2142, S2143, S2144, and M2145.

Binding of 0233-0000-0302 to human FVa protects a further peptide covering the sequence from E2029 to F2059. However, as a peptide covering the sequence from C2038 to S2058 showed no protection upon antibody binding, this part of the sequence could be dismissed as being part of the epitope. Thus, part of the epitope could be confirmed to be within the sequence from L2030 to C2038.

Binding of either of the antibodies 0233-0000-01 10 and 0233-0000-0302 to human FVa also causes structural stabilization to parts of the C1 and C2 domains. In particular, peptides in the region from F1900 to L1908 and F1918 to L1925 are stabilised when 0233- 0000-01 10 is bound and peptides in the region V2146 to F2172 show stabilization when either of the antibodies 0233-0000-01 10 or 0233-0000-0302 is bound. Stabilization is determined by little or no protection at short incubation times of less than one minute, and a higher degree of protection at longer incubation times of several minutes.

Epitope of 0233-0000-0128 on human FVa

Two segments of the FVa heavy-chain sequence were significantly affected by exchange protection upon binding of 0233-0000-0128. One segment of the sequence was covered by peptides from Y295 to M343 and showed significant protection from hydrogen exchange upon binding of the 0233-0000-0128 antibody. However, as peptides covering the sequence from Y295 to Y324 and V331-M343 showed no protection upon antibody binding, these parts of the sequence could be dismissed as being part of the epitope. Thus, part of the epitope could be confirmed to be within the sequence from F325 to

V331 .

A second part of the sequence was also protected from hydrogen exchange upon binding of the 0233-0000-0128 antibody. This region includes peptides from A521 to F552. However, as peptides covering the sequence from A521 to D533 and the sequence from C539 to F552 showed no protection upon antibody binding, these parts of the sequence could be dismissed as being part of the epitope.

Thus, part of the epitope could be confirmed to be within the sequence from N534 to C539.

The epitope of 0233-0000-0128 comprises residues within the regions F325 to V331 and N534 to C539 of the FVa heavy-chain sequence and those residues are F325, I326, A327, A328, E329, E330, V331 , N534, I535, N536, K537, F538, and C539. Epitope of 0233-0000-0453 (and 0233-0000-0418) on human FVa

One segment of the FVa light-chain sequence was significantly affected by exchange protection upon binding of 0233-0000-0453. This segment of the sequence is covered by peptides from D1749 to M1771 and showed significant protection from hydrogen- deuterium exchange upon binding of the 0233-0000-0453 Fab. However, as peptides covering the sequence from F1745 to Y1755 showed low intrinsic exchange this part of the sequence could not be confirmed to be part of the epitope.

Thus, residues in the sequence from Y1756 to M1771 could be confirmed to contribute to the epitopes of 0233-0000-0453 and 0233-0000-0528 on human FVa and these residues are Y1756, E1757, K1758, K1759, S1760, R1761 , S1762, S1763, W1764, R1765, L1766, T1767, S1768, S1769, E1770, and M1771.

Epitope mapping of 0233-0000-0271 on human FVa

Three segments of the FVa heavy-chain sequence were significantly affected by exchange protection upon binding of 0233-0000-0271. One segment of the sequence is covered by peptides from E329 to M343 and showed significant protection from hydrogen exchange upon binding of the 0233-0000-0271 antibody. However, as peptides covering the sequence from Y335 to M343 show no protection upon antibody binding, this part of the sequence can be dismissed as being part of the epitope.

Thus, part of the epitope could be confirmed to be within the sequence from E330 to

Y335.

A second part of the sequence was also protected from hydrogen exchange upon binding of the 0233-0000-0271 antibody. This region includes peptides from D344 to M385. However, as peptides covering the sequence from D344 to F355 and the sequence from S376 to A397 showed no protection upon antibody binding, these parts of the sequence could be dismissed as being part of the epitope.

Thus, part of the epitope could be confirmed to be within the sequence from S356 to

S376.

A third part of the sequence was also protected from hydrogen exchange upon binding of the 0233-0000-0271 antibody. This region includes peptides from A521 to F552. However, as peptides covering the sequence from A521 to Y530 showed low intrinsic exchange, this segment could not be confirmed to contribute to the epitope, and as peptides covering the sequence from A521 to D533 showed no protection upon antibody binding, the sequence from L531 to D533 could be dismissed as being part of the epitope.

Thus, part of the epitope could be confirmed to be within the sequence from N534 to

F552.

The epitope of 0233-0000-0271 comprises residues within the regions E330 to Y334, S356 to S376, and N534 to F552 of the FVa heavy-chain sequence and those residues are E330, V331 , I332, W333, D334, Y335, S356, N357, Q358, I359, G360, K361 , H362, Y363, K364, K365, V366, M367, Y368, T369, Q370, Y371 , E372, D373, E374, S375, S376, N534, I535, N536, K537, F538, and C539, E540, N541 , P542, D543, E544, V545, K546, R547, D548, D549, P550, K551 , F552.

Further parts of the FVa sequence was also protected from hydrogen exchange upon binding of the 0233-0000-0271 antibody as protection was also observed in the regions from Y1756 to M1771 , F2059 to L2075, D2098 to L2099, L2100 to S2124.

The Y1756 to M1771 region is shared with the epitope of 0418 and D2098 to L2099 with several antibodies binding C2, while F2059 to L2075 and 2100 to 2124 are specific for 0271 .

Upon binding of 0233-0000-0271 to human FVa changes in hydrogen-deuterium exchange were observed that could not be attributed to an epitope. These include sequences where hydrogen-deuterium exchange increased. This observation suggests that 0233-0000-0271 cause structural changes or destabilization when binding to human FVa. Protection was observed in the regions from Y1756 to M1771 , F2059 to L2075, 2098 to 2099, 2100 to 2124. This suggests that binding of 0233-0000-0271 to human FVa causes structural changes or that 0233-0000-0271 has an alternative binding site in this region. Example 9 - Cloning and sequencing of anti-FVa monoclonal antibodies variable light chain (VLC) and variable heavy chains (VHC) cDNA from isolated hybridoma cells

This example describes the cloning and sequencing of the variable regions of murine, human and rabbit heavy chain and light chain sequences of anti-FVa/FV antibodies listed in table 31 in example 10 here below.

Murine and human antibodies:

Total RNA was extracted from hybridoma cells using the RNeasy-Mini Kit (Qiagen) and used as template for cDNA synthesis. cDNA was synthesized in a 5'-RACE reaction using the SMART™ RACE cDNA amplification kit (Clontech). Subsequent target amplification of HC and LC sequences was performed by PCR using Phusion Hot Start polymerase (Finnzymes) and the universal primer mix (UPM) included in the SMART™ RACE kit as forward primer. A reverse primer with the following sequence was used for VHC amplification:

5'-CCCTTGACCAGGCATCCCAG-3'

A reverse primer with the following sequence was used for VLC amplification: 5'-GCTCTAGACTAACACTCATTCCTGTTGAAGCTCTTG-3'

The PCR products were separated by gel electrophoresis, extracted using the GFX PCR DNA & Gel Band Purification Kit (GE Healthcare Bio-Sciences) and cloned for sequencing using a Zero Blunt TOPO PCR Cloning Kit and chemically competent TOP10 E.coli (Invitrogen). DNA plasmid material, for sequencing was obtained from plasmid preparations generated by a standard alkaline lysis protocol using a DNA miniprep kit (Qiagen). Sequencing was performed at MWG Biotech (Martinsried/Germany) using M13uni(-21 )/M13rev(-29) sequencing primers. Sequences were analyzed and annotated using the VectorNTI program. All kits and reagents were used according to the

manufacturer's instructions.

A single unique murine kappa type LC and a single unique murine HC, subclass antibody was identified for each of the hybridomas. Amino acid sequences for the variable heavy chain and variable light chain sequences are specified as SEQ IDs NO: 1 -22 (the leader peptide sequences are not included). The in-house software AbAnalyzer was used for CDR identification of the antibodies. The software generate alignments at regions of homology in the variable regions of the sequence of the antibody with a "standard" Kabat numbered sequence. The numbering of amino acid residues is performed by the method described in Kabat et al. (1991 ), Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242.

See also the section "Brief description of sequences" in this application, the sequence listing and the CDR sequences that are annotated in table 28, 29 and 30.

Rabbit antibodies:

B-cells positive in the ELISA were harvested. Cells were washed with 100 μΙ_ cold phosphate buffered saline (PBS: 137mM NaCI; 2,7mM KCI; 10 mM Na₂HP0₄; 1 .8 mM KH₂P0₄) followed by centrifugation at 200G for 5 minutes. 1 10μΙ PBS was aspirated from each well followed by addition of 100 μΙ_ PBS and centrifugation performed as above. 1 10μΙ PBS was aspirated after centrifugation and 100μΙ H₂0 supplemented to 1 1Ι/μΙ_ with RNasin® Plus RNase Inhibitor (Promega, Madison, Wl, USA) were added. Mix was placed at -80^°C until use. In the following this mix is designated "template". Heavy chain (VH) amplification was performed as the following using the QIAGEN

OneStep RT-PCR kit (QIAGEN, Valencia, CA, USA): 30 μΙ_ template were mixed with the following components: 10 μΙ RNAse free water, 5 x QIAGEN OneStep RT-PCR Buffer 1x, 10 μΙ_ dNTP mix (containing 10 mM 2,0 μΙ_ 400 μΜ of each dNTP), 2 μΙ_ QIAGEN OneStep RT- PCR enzyme mix, 3 μΙ_ HC primer A (to a final concentration of 0,6 μΜ) and 3 μΙ_ HC primer B (to a final concentration of 0,6 μΜ).

Light chain (VL) amplification was performed as described above except that HC primer A was replaced with LC primer A and HC primer B was replaced with LC primer B. Samples were moved to a thermo cycler and subjected to the following: 1 cycle: 50 °C/30 min, 1 cycle: 95 °C/15 min, 40 cycles: 94 °C/30 sec, 54 °C/30 sec, 72 °C/1 min, 1 cycle: 72 °C/10 min, 1 cycle: 8 °C/∞

Following amplification aliquots of samples were analysed by agarose gel electrophoresis using 0,7% gels. Fragments of 450 bp were indicative of successful amplification. 1.8μΙ AMPure XP reagent (Agencourt Bioscience Corporation, Beverly,

Massachusetts, USA) was added per 1 .0μΙ_ of PCR product and incubated for 5 minutes at RT. The sample was placed on an Agencourt SPRI Plate 96 Super Magnet Plate (Agencourt Bioscience Corporation, Beverly, Massachusetts, USA) for 2 minutes to separate beads from the solution. The cleared supernatant was aspirated. 200 μΙ_ 70% ethanol was added to the sample followed by incubation for 30 seconds at RT. Sample was removed from the

Agencourt SPRIPIate 96 Super Magnet Plate and 40 μΙ_ reagent grade water was added to the sample and mixed by pipetting 10 times. Beads were separated from solution by moving the sample to the Agencourt SPRIPIate 96 Super Magnet Plate for 1 minute. The eluant was transferred to a new container.

The VH and VL fragments were transferred to CMV promoter-based expression vectors (pTT vectors) suitable for transient expression in the HEK293-EBNA (HEK293-6E) expression system as described by Durocher et al. Nucleic Acids Res. 2002 Jan 15;30(2):E9. The VH of the antibody in question was cloned in the CMV-based vector containing the constant region for human lgG4(S241 P) (containing a S241 P mutation in the hinge region). VL fragments was cloned in the CMV-based vector containing the constant region of the human kappa light chain.

Cloning vectors were prepared by inverse PCR on hyper-methylated vector templates. 0,5 μg vector template was methylated with the CpG Methyltransferase kit from Fermentas (Thermo Fisher Scientific, Inc, West Palm Beach, FL, USA). DNA was incubated in 50 μΙ_ 10x M. SssI Buffer, 10 μΙ_ 50x SAM and 25 μΙ M. SssI methyl transferase at 37°C for 25 min and inactivated by incubation at 65°C for 30 min. Following, DNA concentration was adjusted to 10 ng/μΙ by addition of reagent grade water.

Inverse amplification of the vectors was performed with Phusion High-Fidelity DNA

Polymerase (Thermo Fisher Scientific, Inc, West Palm Beach, FL, USA). 2,5 ng template was mixed with 25 μΙ_ 2xphusion mastermix; 0, 15 μί 100 μΜ Primer mskj1627; 0, 15 μί 100 μΜ Primer mskjl 628 and reagent grade water to 50 μΙ. PCR conditions were the following: 1 cycle: 98°C/ 30 sec, 30 cycles: 98°C/30sec, 66°C/30sec, 72°C/3min,

1 cycle: 72°C, 10 min, 1 cycle: 10°C/ 24h. Resulting fragments were purified with the AMpure XP kit as described above.

Purified HC and LC fragments were cloned by In-Fusion® HD Cloning (Clontech Laboratories, Inc., Mountain View, CA, USA). 2 μΙ_ 5x In-Fusion HD Enzyme Premix was incubated with 50-100 ng vector and 5-25 ng fragment in a total volume of 10 μΙ_. Reaction was incubated for 15min at 50^°C and placed on ice.

E. coli Library Efficiency® DH5aTM Competent Cells (Life technologies, Thermo Fisher Scientific, Inc, West Palm Beach, FL, USA) were used for transformation. Cells were thawed for 15 minutes and 20μΙ cells per reaction were used. 1 μί In-Fusion® HD Cloning reaction from above was added the cells. Mix was incubated on ice for 30 minutes followed by a heat-chock for 30seconds at 42°C and incubation on ice for 2 minutes. 100 μΙ S.O.C. medium was added and the reaction incubated in a shaking incubator at 450 rpm for 1 hour at 37°C. Aliquots of the mix were spread on solid LB Bouillon supplemented with ampicillin to 100 μg/mL and the plates incubated o/n at 37 °C. Resulting clones were grown o/n in liquid LB with 100 μg/mL ampicillin and DNA isolated. DNA from E. coli cultures were extracted using the Qiagen DirectPrep® 96 kit as described by the manufacturer (QIAGEN, Valencia, CA, USA). DNA sequencing was performed at MWG Biotech (Martinsried/Germany.

HEK293-6E cells were grown in suspension in FreeStyle™ 293 expression medium (Gibco) supplemented with 25 μg/ml Geneticin (Gibco, Life technologies, Thermo Fisher Scientific, Inc, West Palm Beach, FL, USA), 1 % v/v of the surfactant Pluronic F-68 (Gibco Life technologies, Thermo Fisher Scientific, Inc, West Palm Beach, FL, USA) & 1 % v/v Penicillin-Streptomycin (Gibco Life technologies, Thermo Fisher Scientific, Inc, West Palm Beach, FL, USA). Cells were cultured in Erlenmeyer shaker flasks in shaker incubators at 37°C, 8 % C02 and 125 rpm and maintained at cell densities between 0.1-1.5 x 106 cells/ml.

The cell density of cultures used for transfection was 0.9-1 .1 x 106 cells/ml. One μg DNA was used per ml cell culture. The DNA was diluted in Opti-MEM media (Gibco, Life technologies, Thermo Fisher Scientific, Inc, West Palm Beach, FL, USA) 3,0μΙ media^g DNA, mixed and incubated at room temperature (23-25 °C) for 5 min. 293Fectin™

(Invitrogen) was used as transfection reagent at a concentration of

1 μΙ per μg DNA. The 293Fectin™ was diluted 30X in Opti-MEM media (Gibco, Life technologies, Thermo Fisher Scientific, Inc, West Palm Beach, FL, USA), mixed and incubated at room temperature (23-25 °C) for 5 min. The DNA and 293Fectin solutions were mixed and left to incubate at room temperature (23-25 °C) for 25 min. The DNA-293Fectin mix was then added directly to the cell culture. The transfected cell culture was transferred to a shaker incubator at 37°C, 8 % C02 and 125 rpm. 5-6 days post transfection, cell culture supernatants were harvested by centrifugation, followed by filtration through a 0.22 μηη PES filter (Corning Life Sciences - Axygen Inc., Union City, CA, USA). When possible quantitative analysis of protein production was performed by Biolayer Interferometry using the ForteBio Octet system (Menlo Park, CA, USA) and protein A biosensors.

See also the section "Brief description of sequences" in this application, the sequence listing and the CDR sequences that are annotated in table 27, 28 and 30.

Table 27. Light chain CDR annotation

Table 28. Heavy chain CDR annotation

SEQ

HC CDR1 CDR2 CDR3

ID #

NINPANGKTNYDEKFK

0233-0000-0161 TYWMH NYV 2

N

RDYVW

0233-0000-0006 YITYSGSTNYNPSLKS WDYGGRGGFAY 4

N 0233-0000-0005 TYSVH MIWSGGSTDYNSALKS NYGNNFDF 6

Wl NTYTG E PTYAD D F K SLRRNPYYYAMD

0233-0000-0028 NYGMN 8

G Y

0233-0000-0128 SYAMS TISSGGSYTYYPDSVKG GPYLTTATPSFTY 10

GISWNSGAIGYADSVK DARWLVEEDYQY

0233-0000-0302 DYAMH 12

G YGLDV

GISWNSGSTGYADSVQ DARWLVEEDYYY

0233-0000-0337 DYAMH 14

G YGMDV

WMNPNSGNTGYALKF RTYYDILTGSLGA

0233-0000-0293 SYDIN 16

QG FDI

GVSWNSGAIGYADSVK DARWLVEEDYQY

0233-0000-0327 DYAMH 18

G YGMDV

GISWNSGGIGYADSVQ DARWLVEEDYYY

0233-0000-0296 DYAMH 20

G YGMDV

WMNPNTDDTGYAQKF YWSVTSWKWND

0233-0000-0426 SYDIN 22

QG DHYYYYGMDV

SSYYM CIYTAWDGASYANWAK

0233-0000-01 10 AMGSSDGANNL 24

C G

CIVGGSSGSTYYANWA DESERDYGDFFK

0233-0000-0222 SSYWIS 26

EG L

CIVGGGSGTTYWSRW DENDRDWGDFFK

0233-0000-0143 SYWIC 28

AEG L

ALYVYSDVYTAFN

0233-0000-0271 SYAMI FIDTGGSAYYASWAKG 30

I

0233-0000-0263 SYAMT TISTDGDTYVASWAKG DITTAYAFFSI 32

CIYAASGDTWYATWVN GPRYVSSSGAGP

0233-0000-0270 SYYHIC 34

A YCLDL

MIYYDSSKMNYADTVK

0233-000-0008 NYGMH PTSHYVVDV 40

G

TSGVGV

0233-0000-0418 LIYWDDVKRYSPSLRR YNWKMRVD 42

G

WMNPNSGNTGYAQKF YFSSTSWKWDDD

0233-0000-0422 SYDIN 44

QG YFYYYGMDV

Table 29. The SEQ ID NR location of identified CDR's

CDR Sequence SEQ ID

NNC 0233-0000-0161

LC-CDR1 RANKSVNKSGYDYMH SEQ ID NO 1 (24-38)

LC-CDR2 LASHLES SEQ ID NO 1 (54-60)

LC-CDR3 QHSRELPLT SEQ ID NO 1 (93-101 )

HC-CDR1 TYWMH SEQ ID NO 2 (31 -35)

HC-CDR2 NINPANGKTNYDEKFKN SEQ ID NO 2 (50-66)

HC-CDR3 NYV SEQ ID NO 2 (99-101 )

0233-0000-0006 LC-CDR1 RSSENIHTYLV SEQ ID NO 3 (24-34)

LC-CDR2 NAETLAE SEQ ID NO 3 (50-56)

LC-CDR3 QHHYGSPLT SEQ ID NO 3 (89-97)

HC-CDR1 RDYVWN SEQ ID NO 4 (31-36)

HC-CDR2 YITYSGSTNYNPSLKS SEQ ID NO 4 (51 -66)

HC-CDR3 WDYGGRGGFAY SEQ ID NO 4 (99-109)

0233-0000-0005

LC-CDR1 RSSQTVVHENGNTYLE SEQ ID NO 5 (24-39)

LC-CDR2 KVSNRFS SEQ ID NO 5 (55-61 )

LC-CDR3 FQGSHVPYM SEQ ID NO 5 (99-102)

HC-CDR1 TYSVH SEQ ID NO 6 (31 -35)

HC-CDR2 MIWSGGSTDYNSALKS SEQ ID NO 6 (50-65)

HC-CDR3 NYGNNFDF SEQ ID NO 6 (98-105)

0233-0000-0028

LC-CDR1 KASQDVGTAVG SEQ ID NO 7 (24-34)

LC-CDR2 WASTRHT SEQ ID NO 7 (50-56)

LC-CDR3 QQYSSNPT SEQ ID NO 7 (89-96)

HC-CDR1 NYGMN SEQ ID NO 8 (31-35)

HC-CDR2 WINTYTGEPTYADDFKG SEQ ID NO 8 (50-66)

HC-CDR3 SLRRNPYYYAMDY SEQ ID NO 8 (99-1 1 1 )

0233-0000-0128

LC-CDR1 KTSTDIDDDMN SEQ ID NO 9 (24-34)

LC-CDR2 EGNTLRP SEQ ID NO 9 (50-56)

LC-CDR3 LQSANMPFT SEQ ID NO 9 (89-97)

HC-CDR1 SYAMS SEQ ID NO 10 (31-35)

HC-CDR2 TISSGGSYTYYPDSVKG SEQ ID NO 10 (50-66)

HC-CDR3 GPYLTTATPSFTY SEQ ID NO 10 (99-1 1 1 )

0233-0000-0302

LC-CDR1 RASQDISHWLA SEQ ID NO 1 1 (24-34)

LC-CDR2 IASTLQT SEQ ID NO 1 1 (50-56)

LC-CDR3 QQSNSFPLT SEQ ID NO 1 1 (89-97)

HC-CDR1 DYAMH SEQ ID NO 12 (31 -35)

HC-CDR2 GISWNSGAIGYADSVKG SEQ ID NO 12 (50-66) HC-CDR3 DARWLVEEDYQYYGLDV SEQ ID NO 12 (99-1 15)

0233-0000-0337

LC-CDR1 RASQDISNWLA SEQ ID NO 13 (24-34)

LC-CDR2 ITSTLHI SEQ ID NO 13 (50-56)

LC-CDR3 QQANSFPFT SEQ ID NO 13 (89-97)

HC-CDR1 DYAMH SEQ ID NO 14 (31 -35)

HC-CDR2 GISWNSGSTGYADSVQG SEQ ID NO 14 (50-66)

HC-CDR3 DARWLVE E DYYYYGM DV SEQ ID NO 14 (99-1 15)

0233-0000-0293

LC-CDR1 KSSQSVLYSS N N KNYLA SEQ ID NO 15 (24-40)

LC-CDR2 WASTRES SEQ ID NO 15 (56-62)

LC-CDR3 QQYYSTPWT SEQ ID NO 15 (95-103)

HC-CDR1 SYDIN SEQ ID NO 16 (31-35)

HC-CDR2 WMNPNSGNTGYALKFQG SEQ ID NO 16 (50-66)

HC-CDR3 RTYYDILTGSLGAFDI SEQ ID NO 16 (99-1 14)

0233-0000-0327

LC-CDR1 RASQDISSWLA SEQ ID NO 17 (24-34)

LC-CDR2 IASSLQS SEQ ID NO 17 (50-56)

LC-CDR3 QQANSFPFT SEQ ID NO 17 (89-97)

HC-CDR1 DYAMH SEQ ID NO 18 (31 -35)

HC-CDR2 GVSWNSGAIGYADSVKG SEQ ID NO 18 (50-66)

HC-CDR3 DARWLVEEDYQYYGMDV SEQ ID NO 18 (99-1 15)

0233-0000-0296

LC-CDR1 RASQDISTWLA SEQ ID NO 19 (24-34)

LC-CDR2 ITSTLHI SEQ ID NO 19 (50-56)

LC-CDR3 QQAYSFPFT SEQ ID NO 19 (89-97)

HC-CDR1 DYAMH SEQ ID NO 20 (31 -35)

HC-CDR2 GISWNSGGIGYADSVQG SEQ ID NO 20 (50-66)

HC-CDR3 D ARWLVE E DYYYYGM DV SEQ ID NO 20 (99-1 15)

0233-0000-0426

LC-CDR1 SGDILGDKYAC SEQ ID NO 21 (23-33)

LC-CDR2 QDIKRPS SEQ ID NO 21 (49-55)

LC-CDR3 QAWDSTTPVV SEQ ID NO 21 (88-97) HC-CDR1 SYDIN SEQ ID NO 22 (31-35)

HC-CDR2 WMNPNTDDTGYAQKFQG SEQ ID NO 22 (50-66)

HC-CDR3 YWS VTS WKWN D D H YYYYGM D V SEQ ID NO 22 (99-1 19)

0233-0000-0110

LC-CDR1 QASESISSYLT SEQ ID NO 23 (24-34)

LC-CDR2 YASTLAS SEQ ID NO 23 (50-56)

LC-CDR3 LGVYSYSRDDGIA SEQ ID NO 23 (89-101 )

HC-CDR1 SSYYMC SEQ ID NO 24 (31-36)

HC-CDR2 C I YTAW D G AS YA N WA KG SEQ ID NO 24 (51-67)

HC-CDR3 AMGSSDGANNL SEQ ID NO 24 (99-109)

0233-0000-0222

LC-CDR1 QASENIGSALA SEQ ID NO 25 (24-34)

LC-CDR2 RASILES SEQ ID NO 25 (50-56)

LC-CDR3 QSYYGDYSVASYA SEQ ID NO 25 (89-101 )

HC-CDR1 SSYWIS SEQ ID NO 26 (31-36)

HC-CDR2 CIVGGSSGSTYYANWAEG SEQ ID NO 26 (51-68)

HC-CDR3 DESERDYGDFFKL SEQ ID NO 26 (100-1 12)

0233-0000-0143

LC-CDR1 QASENIGGALA SEQ ID NO 27 (24-34)

LC-CDR2 RASTLES SEQ ID NO 27 (50-56)

LC-CDR3 QSYYGDYSTASYA SEQ ID NO 27 (89-101 )

HC-CDR1 SYWIC SEQ ID NO 28 (32-36)

HC-CDR2 CIVGGGSGTTYWSRWAEG SEQ ID NO 28 (51-68)

HC-CDR3 DENDRDWGDFFKL SEQ ID NO 28 (100-1 12)

0233-0000-0271

LC-CDR1 QASQSIGGNLA SEQ ID NO 29 (24-34)

LC-CDR2 DASKLAS SEQ ID NO 29 (50-56)

LC-CDR3 QCTYGSSGNIGNG SEQ ID NO 29 (89-101 )

HC-CDR1 SYAMI SEQ ID NO 30 (30-34)

HC-CDR2 Fl DTGGSAYYASWAKG SEQ ID NO 30 (49-64)

HC-CDR3 ALYVYSDVYTAFNI SEQ ID NO 30 (95-108)

0233-0000-0263

LC-CDR1 QASQSISNWLA SEQ ID NO 31 (34-34) LC-CDR2 KASTLAS SEQ ID NO 31 (50-56)

LC-CDR3 QQGYTITNVENA SEQ ID NO 31 (89-100)

HC-CDR1 SYAMT SEQ ID NO 32 (30-34)

HC-CDR2 TISTDGDTYVASWAKG SEQ ID NO 32 (49-64)

HC-CDR3 DITTAYAFFSI SEQ ID NO 32 (95-105)

0233-0000-0270

LC-CDR1 QASQSISSYLS SEQ ID NO 33 (24-34)

LC-CDR2 RTSTLES SEQ ID NO 33 (50-56)

LC-CDR3 RTSTLES SEQ ID NO 33 (89-102)

HC-CDR1 SYYHIC SEQ ID NO 34 (30-35)

HC-CDR2 CIYAASGDTWYATWVNA SEQ ID NO 34 (50-66)

HC-CDR3 GPRYVSSSGAGPYCLDL SEQ ID NO 34 (98-1 14)

0233-000-0008

LC-CDR1 SASSSISSNYLH SEQ ID NO 39 (24-35)

LC-CDR2 RTSNLAS SEQ ID NO 39 (51 -57)

LC-CDR3 QQGSSIPLT SEQ ID NO 39 (90-98)

HC-CDR1 NYGMH SEQ ID NO 40 (31 -35)

HC-CDR2 M I YYDSS KM N YADTVKG SEQ ID NO 40 (50-66)

HC-CDR3 PTSHYVVDV SEQ ID NO 40 (99-107)

0233-0000-0418

LC-CDR1 RSSQSLLDSDDGNTYMD SEQ ID NO 41 (24-40)

LC-CDR2 MGFYRAS SEQ ID NO 41 (56-62)

LC-CDR3 MQRIEFPST SEQ ID NO 41 (95-103)

HC-CDR1 TSGVGVG SEQ ID NO 42 (31 -37)

HC-CDR2 LIYWDDVKRYSPSLRR SEQ ID NO 42 (52-67)

HC-CDR3 YNWKMRVD SEQ ID NO 42 (100-107)

0233-0000-0422

LC-CDR1 SGDKLESKYAC SEQ ID NO 43 (23-33)

LC-CDR2 HDDKRPS SEQ ID NO 43 (49-55)

LC-CDR3 QAWDSSTPW SEQ ID NO 43 (88-97)

HC-CDR1 SYDIN SEQ ID NO 44 (31 -35)

HC-CDR2 WMNPNSGNTGYAQKFQG SEQ ID NO 44 (50-66)

HC-CDR3 YFSSTS WKWD D DYFYYYGM DV SEQ ID NO 44 (99-1 19) Example 10 - Recombinant expression of anti-FVa antibodies

Generation of vectors for recombinant expression of anti-FVa antibodies:

A series of CMV promoter-based expression vectors (pTT vectors) were generated for transient expression of anti-FVa antibodies in EXPI293F cells (Life Technologies). The pTT vectors are developed for transient protein expression by Yves Durocher (Durocher et al. Nucleic Acid Research, 2002). In addition to the CMV promoter, the pTT-based vectors contain a pMB1 origin, an EBV origin and the Amp resistance gene. Light Chain (LC) expression vectors:

pTT-based LC vectors were generated for transient expression of mouse or human anti-FVa antibodies. Initially for each anti-FVa antibody (see table 31 ), the region

corresponding to the variable light chain (VLC) domain of the antibody was PCR amplified from an original TOPO sequencing clone, using primers containing sequences specific for the 3' and 5' region of the identified variable domain sequences. In addition, the sense primer contained a sequence complementary to the DNA sequence of the 3' end of the human CD33 signal peptide sequence. Correspondingly, the anti-sense primer contained a sequence complementary to the DNA sequence of the 5' end of the light chain constant region. The generated PCR fragment was purified using the GFX PCR Purification Kit (GE 25 Healthcare) and cloned into a PCR amplified fragment of a pTT-based vector containing the CD33 signal peptide sequence and the sequence for a human kappa or lambda constant region. The vector fragment was obtained by PCR amplification of the pTT vector using an anti-sense primer specific for the 3' end of the human CD33 signal peptide sequence and a sense primer specific for the 5' end of the light chain constant region. The vector fragment was treated with Dpnl restriction nuclease to remove template DNA and purified using the GFX PCR Purification Kit (GE Healthcare). The amplified VLC fragment was cloned in to the vector in-frame between the CD33 signal peptide and the light chain constant region using the In-Fusion® HD Cloning Kit (Clontech) according to manufacturer's instructions. The cloning reaction was subsequently transformed into E. coli for selection. The sequences of the final constructs were verified by DNA sequencing.

Heavy Chain (HC) expression vectors:

5 pTT-based HC vectors were generated for transient expression of anti-FVa antibodies. Initially for each anti-FVa antibody (see table 31 ), the region corresponding to the variable heavy chain (VHC) domain of the antibody was PCR amplified from an original TOPO sequencing clone, using primers containing sequences specific for the 3' and 5' region of the identified variable domain sequences. In addition, the sense primer contained a sequence complementary to the DNA sequence of the 3' end of the human CD33 signal peptide sequence. Correspondingly, the anti-sense primer contained a sequence

complementary to the DNA sequence of the 5' end of the heavy chain constant region. The generated PCR fragment was purified using the GFX PCR Purification Kit (GE Healthcare) and cloned into a PCR amplified fragment of a pTT-based vector containing the CD33 signal peptide sequence and sequence for a human lgG1.1 (lgG1 with the following amino acid substitutions L234A, L235E, G237A, A330S and P331 S) or lgG4 constant region. Fab molecules includes the the variable region, the human lgG4 CH1 domain and a hinge sequence of 5 amino acids (YGPPS). The vector fragment was obtained by PCR

amplification of the vector sequence using an anti-sense primer specific for the 3' end of the human CD33 signal peptide sequence and a sense primer specific for the 5' end of the heavy chain constant region. The vector fragment was treated with Dpnl restriction nuclease to remove template DNA and purified using the GFX PCR Purification Kit (GE Healthcare). The amplified VHC fragment was cloned in to the vector in-frame between the CD33 signal peptide and the heavy chain constant region using the In-Fusion® HD Cloning Kit (Clontech) according to manufacturer's instructions. The cloning reactions were subsequently transformed into E. coli for selection. The sequence of the final constructs was verified by DNA sequencing.

Recombinant expression of monoclonal antibodies:

The anti-FVa antibodies and fragments were expressed transiently in EXPI293F cells (Life 30 Technologies) by co-transfection of the pTT-based LC/HC expression vectors according to manufacturer's instructions. The following procedure describes the generic EXPI293F expression protocol.

Cell maintenance:

EXPI293Fcells were grown in suspension in Expi293™ expression medium (Life

Technologies). Cells were cultured in Erienmeyer shaker flasks in an orbital shaker incubator at 36.5°C, 8% C02 and 85-125 rpm and maintained at cell densities between 0.4-4x10E6 cells/mL. DNA Transfection: Typical, 30-300ml_ culture volumes were transfected. Separate dilutions of DNA and transfection reagent were initially prepared. Following components were mixed per 1 -mL cell culture:

1. A total of 1 μg vector DNA (0.5ug LC vector and 0.5ug HC vector) was diluted in 50μΙ_ Opti-MEM media (Gibco) and incubated at room temperature (23-25°C) for 5 min.

2. A total of 2.7μΙ_ Expifectamin™ 293 (Life Technologies) was diluted in 50μί Opti- MEM media (Gibco) and incubated at room temperature (23-25°C) for 5 min.

The two separate dilutions were mixed and incubated at room temperature (23-25°C) for 10min. The DNA-Expifectamin™ 293 mix was added directly to 1 mL EXPI293F cell culture. At the time of transfection the cell density of the EXPI293F culture should be 2.8-3.2x10E6 cells/mL. The transfected cell cultures were incubated in an orbital shaker incubator at 36.5°C, 8% C02 and 85-125rpm. 18hrs post transfection, 5μΙ Expifectamin™ 293

Transfection Enhancer 1 and 50μΙ Expifectamin™ 293 Transfection Enhancer 2 were added per 1-mL culture. 5 days post transfection, the cell culture supernatants were harvested by centrifugation, followed by filtration through a 0.22 μηη PES filter unit (Corning).

Purification of antibodies Antibodies were purified by affinity purification with MabSelect SuRe (GE Healthcare) on different Akta chromatography systems (GE Healthcare) according to the manufacturer's guidelines. E.g. cell culture supernatant was loaded onto column with affinity resin, equilibrated in PBS buffer. The column was washed with PBS buffer, pH 7.4 and bound proteins eluted with 10 mM sodium formate, pH 3.5, followed by buffer exchange to 10 mM histidine, 150 mM NaCI, pH 7.5.

Purification of Fab

Purification of recombinant Fab was performed by a combination of affinity

chromatography and size exclusion chromatography on an Akta Avant chromatography system (GE Healthcare). Cell culture supernatant was loaded onto a column with

KappaSelect resin (GE Healthcare), which had been equilibrated with buffer A (10mM Phosphate , 140 mM NaCI, 2.68 mM KCI, pH 7.4 ). The column was washed with buffer A. Bound proteins were eluted with buffer B (20 mM sodium formate, pH 3.0) and pH was neutralised with 1 M Tris, pH 8.0. Fractions containing Fab fragment were concentrated by ultrafiltration and applicated to a Superdex75 HiLoad size exclusion column (GE Healthcare) and separated with 10mM HEPES, 150 mM NaCI, pH 7.5. Fractions containing the Fab fragment were pooled. While certain features of the invention have been illustrated and described herein, many modifications, substitutions, changes, and equivalents will now occur to those of ordinary skill in the art. It is, therefore, to be understood that the appended claims are intended to cover all such modifications and changes as fall within the true spirit of the invention.

Claims

1. A monoclonal anti-activated Factor V (FVa) antibody or an antigen-binding fragment thereof that displays a pro-coagulant effect.

2. The monoclonal anti-FVa antibody or an antigen-binding fragment thereof according to claim 1 , wherein the anti-FVa antibody or antigen-binding fragment thereof increases thrombin generation.

3. The monoclonal anti-FVa antibody or an antigen-binding fragment thereof according to any of claims 1-2, wherein the anti-FVa antibody or antigen-binding fragment thereof increases thrombin generation in the presence of APC, such as in the presence of exogenously added APC in a human plasma-based thrombin generation assay.

4. The monoclonal anti-FVa antibody or an antigen-binding fragment thereof according to any of claims 1-2, wherein the anti-FVa antibody or antigen-binding fragment thereof increases thrombin generation measured by one or more of peak thrombin, the velocity index and/or endogenous thrombin potentialin om the presence of APC, in a human plasma-based thrombin generation assay, wherein the plasma is haemophilia A patient plasma.

5. The monoclonal anti-FVa antibody or an antigen-binding fragment thereof according to claim 1 -2, that stimulates the thromboelastographic response, such as by increasing the elastic properties of blood during thrombus formation measured by thrombelastography using a TEG® hemostasis analyzer.

6. The monoclonal anti-FVa antibody or an antigen-binding fragment thereof according to any of claim 1-5, that when bound to FVa improves FVa co-factor activity to activated Factor X.

7. The monoclonal anti-FVa antibody or an antigen-binding fragment thereof according to any of claim 1 -5, that when bound to FVa protects FVa from inactivation by activated protein C (APC).

8. A monoclonal anti-FVa antibody or an antigen-binding fragment thereof that binds to a) the A3 domain of the FVa light chain, b) the C2 domain of the FVa light chain, c) the A1 domain the FVa heavy chain or d) the A2 domain of the FVa heavy chain.

9. A monoclonal anti-FVa antibody or antigen-binding fragment thereof that displays a pro- coagulant effect, wherein the anti-FVa antibody or antigen-binding fragment thereof binds to a) the A3 domain of the FVa light chain, b) the C2 domain of the FVa light chain, c) the A1 domain the FVa heavy chain or d) the A2 domain of the FVa heavy chain.

10. A monoclonal anti-FVa antibody or antigen-binding fragment thereof that displays a pro- coagulant effect, wherein the anti-FVa antibody or antigen-binding fragment thereof binds to

a) the A3 domain of the FVa light chain and competes with a reference antibody,

wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:41 and a variable heavy chain amino acid sequence of SEQ ID NO:42, b) the A3 domain of the FVa light chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:29 and a variable heavy chain amino acid sequence of SEQ ID NO:30, c) the C2 domain of the FVa light chain and competes with an reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:29 and a variable heavy chain amino acid sequence of SEQ ID NO:30, d) the C2 domain of the FVa light chain and competes with a reference antibody,

wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:23, and a variable heavy chain amino acid sequence of SEQ ID NO:24, e) the C2 domain of the FVa light chain and competes with a reference antibody,

wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:1 1 , and a variable heavy chain amino acid sequence of SEQ ID NO:12, f) the C2 domain of the FVa light chain and competes with a reference antibody,

wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:3, and a variable heavy chain amino acid sequence of SEQ ID NO:4, g) the A1 domain the FVa heavy chain competes with a reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:5, and a variable heavy chain amino acid sequence of SEQ ID NO:6, h) the A2 domain of the FVa heavy chain competes with a reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID NO:9, and a variable heavy chain amino acid sequence of SEQ ID NO:10 or.

i) the A2 domain of the FVa heavy chain competes with a reference antibody, wherein said reference antibody has a variable light chain amino acid sequence of SEQ ID

NO:29, and a variable heavy chain amino acid sequence of SEQ ID NO:30.

1 1 . A monoclonal anti-FVa antibody or antigen-binding fragment thereof, wherein the CDRs of the light and heavy chain variable regions are as in the variable regions defined by SEQ ID NO 1 and SEQ ID NO 2, SEQ ID NO 3 and SEQ ID NO 4, SEQ ID NO 5 and

SEQ ID NO 6, SEQ ID NO 7 and SEQ ID NO 8, SEQ ID NO 9 and SEQ ID NO 10, SEQ ID NO 1 1 and SEQ ID NO 12, SEQ ID NO 13 and SEQ ID NO 14, SEQ ID NO 15 and SEQ ID NO 16, SEQ ID NO 17 and SEQ ID NO 18, SEQ ID NO 19 and SEQ ID NO 20, SEQ ID NO 21 and SEQ ID NO 22, SEQ ID NO 23 and SEQ ID NO 24,SEQ ID NO 25 and SEQ ID NO 26, SEQ ID NO 27 and SEQ ID NO 28, SEQ ID NO 29 and SEQ ID NO 30, SEQ ID NO 31 and SEQ ID NO 32, SEQ ID NO 33 and SEQ ID NO 34, SEQ ID NO 39 and SEQ ID NO 40, SEQ ID NO 41 and SEQ ID NO 42 or SEQ ID NO 43 and SEQ ID NO 44.

12. A antigen-binding fragment of the antibody of any of the preceding claims, which is a Fab, Fab', F(ab)2, F(ab')2, Fv, single-chain Fv, dsFv, Fd or a dAb fragment, a VH, VL, VhH, or V-NAR domains, a monovalent molecule, minibody, diabody, triabody, tetrabody or kappa body, or an IgNAR.

13. A pharmaceutical composition comprising the antibody or antigen-binding fragment of any of the preceding claims optionally comprising one or more pharmaceutical excipients..

14. A pharmaceutical composition of claim 13 for use in prophylactic, palliative and/or

symptomatic treatment of a coagulation deficiency such as haemophilia A.

15. A method for prophylactic, palliative and/or symptomatic treatment of haemophilia A

comprising administering an effective dose of an antibody or pharmaceutical composition according to any of the claims 1 -14 to a subject in need thereof and thereby treat a coagulation deficiency such as haemophilia A.