CN118786147A - Methods for once-monthly administration of FVIII mimetic bispecific antibodies - Google Patents

Abstract

The present invention relates generally to the use of bispecific FVIII mimetic antibodies in the treatment of hemophilia, such as hemophilia a with or without inhibitors, in particular methods of treating the disease, dosage regimens and compositions as used in such methods.

Description

Methods for once-monthly administration of FVIII mimetic bispecific antibodies

Technical Field

The present invention relates to methods of administering Factor VIII mimetic antibodies to hemophiliacs.

Incorporation by Reference of Sequence Listing

This application is submitted with a sequence listing in electronic form. The entire contents of the sequence listing are incorporated herein by reference.

Background Art

In patients with coagulopathy, such as in people with hemophilia A and B, various steps of the coagulation cascade become dysfunctional due to, for example, a lack or insufficient presence of functional coagulation factors. This dysfunction of a part of the coagulation cascade results in inadequate blood clotting and potentially life-threatening bleeding, or damage to internal organs such as joints.

Factor VIII (FVIII) deficiency, commonly known as hemophilia A, is a congenital bleeding disorder that affects approximately 420,000 people worldwide, of whom approximately 105,000 are currently diagnosed.

Patients with hemophilia A can receive coagulation factor replacement therapy, such as exogenous FVIII. Conventional treatment consists of replacement therapy, which is provided as prophylaxis or on-demand treatment of bleeding episodes. Until recently, prophylactic treatment for patients with severe hemophilia A was intravenous injections of plasma-derived FVIII or recombinant FVIII or its long-acting variants up to three times a week.

However, such patients are at risk of developing neutralizing antibodies (so-called inhibitors) against such exogenous factors, rendering previously effective therapies ineffective. Hemophilia A patients with inhibitors are non-limiting examples of a partially congenital, partially acquired coagulopathy. Patients who have developed FVIII inhibitors cannot be treated with conventional replacement therapy.

Exogenous clotting factors can only be administered intravenously, which is quite inconvenient and uncomfortable for the patient. For example, infants and young children may have to undergo surgery to insert an intravenous catheter into a chest vein to ensure intravenous access. This puts them at great risk of developing bacterial infections.

Drug emicizumab Also known as ACE910, it has been approved for subcutaneous prophylactic treatment of hemophilia A with or without inhibitors. Emicizumab is a humanized bispecific anti-FIX(a)/anti-FX(a) monoclonal antibody developed by Chugai Pharmaceuticals/Roche Pharmaceuticals for the treatment of hemophilia A. Emicizumab is designed to mimic FVIII cofactor function (see Sampei et al.: (2013) PLoS One, 8, e57479 and WO2012/067176).

WO2015/194233 and WO2018/047813 disclose dosage regimens allegedly useful for administering emicizumab.

For example, WO2018/021450, WO2020/025672 and WO2021/152066 also disclose FVIII-mimicking anti-FIX(a) anti-FX(a) bispecific antibodies and their use as procoagulants for the treatment of hemophilia A.

There is a need in the art for improved methods of administering specific bispecific FVIII mimetic antibodies to patients suffering from hemophilia A, such as hemophilia A with or without inhibitors.

Summary of the invention

The present invention relates to methods of administering compounds that act as a replacement for FVIII in patients suffering from coagulopathy, in particular patients lacking functional coagulation factor VIII (FVIII), such as hemophilia A patients, including hemophilia A patients with inhibitors. In particular, bispecific antibodies that can be used to treat hemophilia A with or without inhibitors, wherein the bispecific antibodies are capable of binding to FIX (SEQ ID NO: 1) or an activated form thereof and FX (SEQ ID NO: 2) or an activated form thereof.

In one aspect, the invention relates to a bispecific antibody for use in the treatment of hemophilia A with or without inhibitors, wherein the bispecific antibody comprises

An anti-FIX(a) antibody or an antigen-binding fragment thereof capable of binding to FIX (SEQ ID NO: 1) and/or its activated form (FIXa), comprising a heavy chain and a light chain, and

An anti-FX(a) antibody or an antigen-binding fragment thereof capable of binding to FX (SEQ ID NO: 2) and/or its activated form (FXa), comprising a heavy chain and a light chain,

in

The heavy chain of the anti-FIX(a) antibody or antigen-binding fragment thereof comprises

NO:3, 4 and 5 represent the CDR1-3 sequences, and

The light chain of the anti-FIX(a) antibody or antigen-binding fragment thereof comprises

NO: 8, 9 and 10 represent the CDR1-3 sequence, and

The heavy chain of the anti-FX(a) antibody or antigen-binding fragment thereof comprises the following:

NO:13, 14 and 15 represent the CDR1-3 sequence, and

The light chains of the anti-FX(a) antibody or antigen-binding fragment thereof comprise the light chains represented by SEQ ID

NO:18, 19 and 20 represent the CDR1-3 sequence, and

wherein the bispecific antibody is to be administered subcutaneously to a human patient in a composition comprising the bispecific antibody,

The loading dose includes

- about 9 mg of the bispecific antibody, administered to a patient weighing 5 kg to <15 kg, or

- about 40 mg of the bispecific antibody, administered to a patient weighing 15 kg to < 45 kg, or

- about 92 mg of the bispecific antibody, administered to a patient weighing 45 kg or more,

The maintenance dose includes

- About 9 mg of the bispecific antibody, administered once a month to a patient weighing 5 kg to <15 kg

kg patients, or

- about 20 mg of the bispecific antibody, administered once a month to a patient weighing 15 kg to

For patients <45 kg, or

- about 46 mg of the bispecific antibody, administered once a month to a patient weighing 45 kg or more,

wherein the first maintenance dose is administered one month after administration of the loading dose; and

Therein a steady state plasma concentration of the bispecific antibody is provided in the range of about 2 μg/mL to about 18 μg/mL, such as 3 to 9 μg/mL, such as 6.5 μg/mL.

In one embodiment, the bispecific antibody comprises a first heavy chain comprising SEQ ID NO: 7, a first light chain comprising SEQ ID NO: 12, a second heavy chain comprising SEQ ID NO: 17, and a second light chain comprising SEQ ID NO: 22 (mAb1).

In one aspect, the invention relates to a pharmaceutical composition comprising a bispecific antibody as described herein.

In one embodiment, the bispecific antibody is administered subcutaneously to a human patient in a pharmaceutical composition comprising the bispecific antibody as described herein.

In one aspect, the bispecific antibody as disclosed herein is administered once a month.

In a preferred embodiment, the antibody is administered as a loading dose followed by a maintenance dose, as disclosed herein.

In another aspect, the invention relates to a kit comprising a composition comprising the bispecific antibody (eg, in an injection device) and instructions for use.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 shows SEQ ID NOs: 3-22 in tabular format.

Figure 2 shows the average curve of mAb1 concentration in patient plasma. Pre-dose measurements below the lower limit of quantitation were set to 0. Due to the logarithmic axis, concentration 0 is reported as 1e-2 μg/mL. The vertical lines represent the two pharmacokinetic (PK) phases. Days 56-63 of the PK phase were used for cohorts 1-3 and 5 (once a week), while days 56-84 were used for cohort 4 (once every 4 weeks). Mean +/- SEM.

Fig. 3 shows the visual prediction check of the fit of the PK model and the data observed in the FRONTIER1 MAD part.Data points are the individual mAb1 plasma concentrations over time.The solid line represents the median of the observed data, and the dotted line represents the mAb1 plasma concentration median predicted by the model relative to time.The dotted line represents the 5th percentile (below) and the 95th percentile (above) of the model prediction obtained by 1000 test simulations with the PK model.The model fully captures the median trend line and variability of the data in all queues.

Fig. 4 shows the peak thrombin level of the patient treated with the mAb1 of increasing dosage and the emicilizumab (emi) of clinical recommended dosage.Blood plasma samples were collected from the patient treated with mAb1 (multiple ascending dose (MAD) queue) of different dosages, and these patients started treatment with emicilizumab or carried out established prevention with emicilizumab at different time points during the whole treatment period.Potential FVIII activity was neutralized by adding anti-FVIII antibody, and thrombin generation test was carried out in vitro.Solid line represents the in vitro sample of human plasma from healthy subjects, and these samples are made to be similar to hemophilia A with anti-FVIII antibody, and mAb1 or emicilizumab of different concentrations are added.The dotted line represents the average mAb1 plasma concentration (C _avg is calculated based on PK stage) of each specified queue.

Figure 5 shows the predicted typical PK profiles of mAb1 in subjects with different body weights using a single loading dose followed by a maintenance dose, as exemplified in Table 8. PK simulations confirmed that all subjects, regardless of body weight, should achieve mAb1 plasma concentrations within the therapeutic range defined by a minimum concentration (2 μg/mL) comparable to FRONTIER1 MAD Cohort 2 (C2) and a maximum concentration (18 μg/mL) defined by _Cmax in MAD Cohort 5. This was demonstrated in all QW, Q2W, and QM dosing intervals.

Sequence Description

SEQ ID NO: 1 represents the amino acid sequence of human coagulation factor IX.

SEQ ID NO: 2 represents the amino acid sequence of human coagulation factor X.

SEQ ID NOs: 3-22 represent amino acid sequences belonging to or derived from components of the bispecific antibody "mAb1" mentioned herein, as shown below:

SEQ ID NOs: 3, 4 and 5 represent complementarity determining regions (CDRs) 1-3, respectively, of the heavy chain of the anti-FIX(a) antibody component of mAbl.

SEQ ID NO: 6 represents the heavy chain variable domain ( _VH ) of the anti-FIX(a) antibody component of mAbl.

SEQ ID NO:7 represents the full-length heavy chain of the anti-FIX(a) antibody component of mAbl.

SEQ ID NOs: 8, 9 and 10 represent CDRs 1-3 of the light chain of the anti-FIX(a) antibody component of mAbl, respectively.

SEQ ID NO: 11 represents the light chain variable domain ( _VL ) of the anti-FIX(a) antibody component of mAbl.

SEQ ID NO: 12 represents the full-length light chain of the anti-FIX(a) antibody component of mAbl.

SEQ ID NOs: 13, 14 and 15 represent CDRs 1-3 of the heavy chain of the anti-FX(a) antibody component of mAb1, respectively.

SEQ ID NO: 16 represents the heavy chain variable domain ( _VH ) of the anti-FX(a) antibody component of mAbl.

SEQ ID NO: 17 represents the full-length heavy chain of the anti-FX(a) antibody component of mAbl.

SEQ ID NOs: 18, 19 and 20 represent CDRs 1-3 of the light chain of the anti-FX(a) antibody component of mAb1, respectively.

SEQ ID NO: 21 represents the light chain variable domain ( _VL ) of the anti-FX(a) antibody component of mAbl.

SEQ ID NO: 22 represents the full length light chain of the anti-FX(a) antibody component of mAb1.

In addition to the electronic sequence listing submitted herewith, Figure 1 also presents SEQ ID NOs: 3-22 in tabular form.

DETAILED DESCRIPTION

The present invention relates to methods of administering bispecific antibodies and compositions comprising such bispecific antibodies, which serve as a replacement for FVIII in patients suffering from coagulopathy, in particular patients lacking functional coagulation factor VIII (FVIII), such as hemophilia A patients, including hemophilia A patients with inhibitors and hemophilia A patients without inhibitors, in particular a once-monthly dosing regimen.

In order that the present invention may be more easily understood, some terms are first defined.

The terms “a” or “an” are intended to mean “one or more.” When the terms “comprise” and variations such as “comprising” precede recitation steps or elements, it is intended to mean that adding further steps or elements is optional and not excluded.

The term "about" is used herein to mean approximately, roughly, or around. When the term "about" is used in conjunction with a numerical range, it modifies the range by extending the boundaries above and below the numerical values. Generally, the term "about" can modify a numerical value up or down (higher or lower) to 10% above or below the stated value.

The term "annualized bleeding rate" (ABR) refers to the number of treated bleeding episodes (including spontaneous and traumatic bleeding) a patient experiences in a defined period of time, extrapolated to one year. For example, two bleeds in six months would represent an ABR of four.

The term antibody includes, but is not limited to, bivalent antibodies, such as bispecific antibodies. A full-length antibody comprises at least four polypeptide chains: two heavy chains (HC) and two light chains (LC) connected by disulfide bonds. One class of immunoglobulins of particular interest in medicine is IgG. In humans, the IgG class can be divided into four subclasses: IgG1, IgG2, IgG3, and in a preferred embodiment, IgG4, based on the sequence of the constant region of its heavy chain. Light chains can be divided into two types: kappa chains and lambda chains, based on differences in their sequence composition. An IgG molecule consists of two heavy chains interconnected by two or more disulfide bonds and two light chains each connected to a heavy chain by a disulfide bond. An IgG heavy chain can comprise one heavy chain variable domain ( _VH ) and up to three heavy chain constant ( _CH ) domains: _CH1 , _CH2 , and _CH3 . A light chain can comprise a light chain variable domain ( _VL ) and a light chain constant domain ( _CL ). The _VH and _VL regions can be further subdivided into highly variable regions called complementarity determining regions (CDRs) or hypervariable regions (HvRs), interspersed with more conserved regions called framework regions (FRs). The _VH and _VL domains are usually composed of three CDRs and four FRs, which are arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The heavy and light chain variable domains containing the hypervariable regions (CDRs) constitute a structure capable of interacting with antigens, while the constant region of the antibody can mediate the binding of immunoglobulins to host tissues or factors, including but not limited to various cells of the immune system (effector cells), Fc receptors, and the first component of the C1 complex of the classical complement system, C1q.

Antibodies or fragments thereof can be defined according to their complementary determining regions (CDRs). The term "complementarity determining region" or "CDR" as used herein refers to the region of an antibody in which the amino acid residues involved in antigen binding are located. CDRs can be identified as regions with the highest variability in the amino acid alignment of the antibody variable domains. Databases such as the Kabat database can be used for CDR identification, for example, CDRs are defined as 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) of 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). Typically, the numbering of amino acid residues in this region is performed by the method described by 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, and unless contradicted by context, numbering according to Kabat is used herein.

The term "bispecific antibody" as used herein refers to an antibody that is capable of binding to two different antigens or two different epitopes on the same antigen.

The term "fixed dose" of a bispecific antibody refers to a dose administered to patients whose body weight falls within a predetermined range (e.g., 15 kg to <45 kg). Thus, a fixed dose is not provided in mg/kg doses, but rather in absolute amounts of the bispecific antibody.

As used herein, the term "human antibody" is intended to include antibodies having variable domains in which at least a portion of the framework regions and/or at least a portion of the CDR regions are derived from human germline immunoglobulin sequences. For example, a human antibody may have variable domains in which both the framework regions and the CDR regions are derived from human germline immunoglobulin sequences. In addition, if the antibody contains a constant region, the constant region or a portion thereof is also derived from a human germline immunoglobulin sequence. Preferably, the human antibody is a monoclonal antibody.

As used herein, "mAb1" refers to a bispecific antibody comprising an anti-FIX(a) antibody capable of binding to FIX (SEQ ID NO: 1) and/or its activated form (FIXa), comprising a heavy chain and a light chain, and an anti-FX(a) antibody capable of binding to FX (SEQ ID NO: 2) and/or its activated form (FXa), comprising a heavy chain and a light chain, wherein the heavy chain of the anti-FIX(a) antibody comprises SEQ ID NO: 7, the light chain of the anti-FIX(a) antibody comprises SEQ ID NO: 12, the heavy chain of the anti-FX(a) antibody comprises SEQ ID NO: 17, and the light chain of the anti-FX(a) antibody comprises SEQ ID NO: 22. The bispecific antibody comprises CDR sequences represented by SEQ ID NOs: 3, 4, 5 and 8, 9, 10 and 13, 14, 15 and 18, 19, 20.

The term "dosage regimen" or "administration regimen" includes a treatment regimen based on a defined set of doses. For example, in one embodiment, the invention describes a dosage regimen for treating hemophilia A with or without inhibitors, wherein the bispecific antibody is first administered as a loading dose, followed by a maintenance dose comprising the same or lower amount of the bispecific antibody as the loading dose.

The term "administering" refers to administering a substance (eg, mAbl) for a therapeutic purpose (eg, treating hemophilia A with or without inhibitors).

A "dose" can be administered by a single administration or multiple consecutive administrations. For example, a 60 mg dose can be administered by a single 60 mg administration or two consecutive administrations of 30 mg each, while a 120 mg dose can be administered, for example, by three consecutive administrations of 40 mg. Continuous administration of doses within 1 hour after administration of the first dose constitutes a dose (e.g., a loading dose that cannot be easily administered by a single administration).

The term "FIX (SEQ ID NO: 1) and/or its activated form (FIXa)" may also be referred to as "FIX/FIXa", or simply as "FIX(a)".

The term "FX (SEQ ID NO: 2) and/or its activated form (FXa)" may also be referred to as "FX/FXa" or simply as "FX(a)".

The term "heavy chain" includes a full-length heavy chain. A full-length heavy chain includes a variable region domain _VH and three constant region domains _CH1 , _CH2 and _CH3 . _{The VH} domain is located at the amino terminus of the polypeptide, while the _CH domain is located at the carboxyl terminus, with _CH3 closest to the -COOH terminus.

The term "light chain" as used herein includes a full-length light chain. A full-length light chain includes a variable region domain V _L and a constant region domain _CL . The variable region domain of the light chain is located at the amino terminus of the polypeptide. As described herein, a light chain includes a kappa chain and a lambda chain.

The term "kit" refers to a packaged product that includes components and instructions for use for administering a bispecific antibody (such as mAb1) to treat a condition. The kit preferably includes a box or container containing the components of the kit. The box or container is labeled or has a protocol approved by the Food and Drug Administration (or corresponding competent authority).

The term "loading dose" as used herein refers to the first dose of a bispecific antibody administered to a patient at the beginning of a treatment regimen. Typically, a loading dose is intended to achieve therapeutically relevant plasma concentrations of the bispecific antibody in a patient within a short period of time.

The term "loading phase" refers to a period of time during which a patient is treated, including administration of a bispecific antibody to the patient to induce a clinical response. Depending on the desired dosing frequency, the "loading phase" typically lasts from one week to one month and is triggered by the administration of the first loading dose. The loading phase precedes the administration of the first maintenance dose.

The term "maintenance dose" as used herein relates to the dose of the bispecific antibody administered to the patient at a certain time point after the administration of the loading dose.

The term "maximum plasma concentration" ( _Cmax ) refers to the highest concentration of the bispecific antibody observed in the plasma of a patient following administration of the bispecific antibody to the patient.

The term "mean plasma concentration" (or " _Cavg ") refers to the average plasma concentration of the bispecific antibody at steady state over the dosing interval.

The term "steady-state _Cmax of mAb1 plasma concentration" refers to the state in which the maximum plasma concentration of mAb1 after administration does not differ from dose to dose. In one embodiment, the steady-state _Cmax of mAb1 plasma concentration is about 18 μg/mL. In another embodiment, the steady-state _Cmax of mAb1 plasma concentration is about 9 μg/mL.

The term "steady-state _Cmin of mAb1 plasma concentration" refers to a state in which the minimum plasma concentration of mAb1 after administration does not differ from dose to dose. In one embodiment, the steady-state _Cmin of mAb1 plasma concentration is about 2 μg/mL. In another embodiment, the steady-state _Cmin of mAb1 plasma concentration is about 3 μg/mL.

The term "serum or plasma half-life" refers to the time required for half the amount of a substance administered to a patient to be metabolized or eliminated from the patient's serum or plasma by normal biological processes.

The term "prophylactic treatment" refers to the administration of a therapy for the treatment of hemophilia A with or without inhibitors, wherein such treatment is intended to control, manage, prevent or reduce the occurrence and/or severity of one or more symptoms of hemophilia A with or without inhibitors, such as bleeding episodes, such as one or more spontaneous bleeding episodes, and/or joint damage.

The term "treat" or "treatment" refers to reducing one or more symptoms of hemophilia A with or without inhibitors, such as the frequency of spontaneous or uncontrolled bleeding episodes. However, "treat" is not necessarily a cure.

The term " _Tmax " refers to the time at which a substance reaches a maximum concentration in the patient's plasma after administration of the substance to the patient.

The present invention relates to a method of administering a bispecific antibody that acts as a replacement for FVIII in patients suffering from coagulopathy, in particular patients lacking functional coagulation factor VIII (FVIII), such as hemophilia A patients, including hemophilia A patients with inhibitors and hemophilia A patients without inhibitors. In particular, a bispecific antibody or antigen-binding fragment thereof that can be used to treat hemophilia A with or without inhibitors, wherein the antibody is capable of binding to FIX (SEQ ID NO: 1) or an activated form thereof and FX (SEQ ID NO: 2) or an activated form thereof.

In one such embodiment, the heavy chain of the anti-FIX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 3, 4 and 5, respectively, and the light chain of the anti-FIX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 8, 9 and 10, respectively, and the heavy chain of the anti-FX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 13, 14 and 15, respectively, and the light chain of the anti-FX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 18, 19 and 20, respectively.

In one such embodiment, the anti-FIX(a) antibody or antigen-binding fragment thereof comprises a heavy chain variable domain represented by SEQ ID NO:6 and a light chain variable domain represented by SEQ ID NO:11, and the anti-FX(a) antibody or antigen-binding fragment thereof comprises a heavy chain variable domain represented by SEQ ID NO:16 and a light chain variable domain represented by SEQ ID NO:21.

In a preferred embodiment, the bispecific antibody is of the IgG4 isotype.

In preferred embodiments, the bispecific antibodies are human antibodies.

In one embodiment, the heavy chain of the anti-FIX(a) antibody comprises SEQ ID NO:7 and the light chain of the anti-FIX(a) antibody comprises SEQ ID NO:12, and the heavy chain of the anti-FX(a) antibody comprises SEQ ID NO:17 and the light chain of the anti-FX(a) antibody comprises SEQ ID NO:22 (also referred to herein as mAb1).

The properties of bispecific antibodies have been described in WO2020/025672, which is incorporated herein by reference.

As is apparent from the above, a bispecific antibody may therefore be characterized according to its CDR sequences, its variable domain sequences or its complete heavy and light chain sequences. For the avoidance of doubt, reference to mAbl requires the presence of a heavy chain and a light chain defined by SEQ ID NOs: 7 and 12 and 17 and 22 herein. The term "mAbl" is used interchangeably with the term "bimAbl".

The methods disclosed herein comprise administering a subcutaneous injection of a bispecific antibody to a patient once a month. Preferably, administration of one or more loading doses precedes the monthly dosing regimen.

Therefore, in a first aspect of the present invention, there is provided a method for treating hemophilia A with or without inhibitors, comprising administering to a patient in need thereof an effective amount of a bispecific antibody, the method comprising

a) administering at least one loading dose of the bispecific antibody to the patient;

as well as

b) administering to the patient at least one maintenance dose of the bispecific antibody after administration of the final loading dose.

In one aspect, such a bispecific antibody is mAbl.

In a preferred embodiment, the loading dose and the maintenance dose are selected based on the patient's weight, and in particular, the patient's weight range (also referred to herein as a "weight band").

In such an embodiment, patients weighing 5 kg to <15 kg are grouped together.

In another such embodiment, patients weighing 15 kg to <45 kg are grouped together.

In yet another such embodiment, patients weighing 45 kg or more are grouped together.

In one embodiment, the loading dose and maintenance dose are each a fixed dose suitable for patients weighing 5 kg to <15 kg.

In one embodiment, the loading dose and maintenance dose are each a fixed dose suitable for patients weighing 15 kg to <45 kg.

In one embodiment, the loading dose and maintenance dose are each a fixed dose suitable for patients weighing 45 kg or more.

In one embodiment, the loading dose and the maintenance dose are identical in terms of the amount of bispecific antibody to be delivered, measured in mg.

In one embodiment, the loading dose and the maintenance dose measured in mg are not identical with respect to the amount of bispecific antibody to be delivered.

The method of administration as disclosed herein includes a once-monthly dosing regimen, such regimen including specific patient weight bands and dosages carefully designed by the inventors to allow for safe and effective treatment.

Due to the long half-life of mAb1, the limited inter-subject variability of plasma concentration, the large therapeutic window, and the near-maximal effect of administration (determined by the inventor), it is considered feasible to use a simplified weight belt to replace continuous weight-based administration. In addition to providing safe and effective hemostatic coverage, fixed-volume injection administration based on a weight belt is also more convenient than administration by kg body weight, does not require dose calculation, and reduces the risk of medication errors. It is intended to simplify administration while still considering differences in plasma concentrations associated with body weight and drug product strength (see Example 1, Table 6). This mode of administration is suitable for injection devices, such as, but not limited to, pre-filled pen syringes for subcutaneous administration of mAb1.

Once-monthly dosing regimen range

In a preferred embodiment, a loading dose comprising about 5 mg to about 15 mg, such as 9 mg, of the bispecific antibody, such as mAb1, is administered to a patient weighing 5 kg to <15 kg, and a loading dose comprising about 35 mg to about 45 mg, such as 40 mg, of the bispecific antibody, such as mAb1, is administered to a patient weighing 15 kg to <45 kg, and a loading dose comprising about 85 mg to about 95 mg, such as 92 mg, of the bispecific antibody, such as mAb1, is administered to a patient weighing 45 kg or more.

In a preferred embodiment, a maintenance dose comprising about 7 mg to about 11 mg, such as 9 mg, of the bispecific antibody, such as mAb1, is administered to a patient weighing 5 kg to <15 kg, and a maintenance dose comprising about 18 mg to about 22 mg, such as 20 mg, of the bispecific antibody, such as mAb1, is administered to a patient weighing 15 kg to <45 kg, and a maintenance dose comprising about 44 mg to about 48 mg, such as 46 mg, of the bispecific antibody, such as mAb1, is administered to a patient weighing 45 kg or more.

In a preferred embodiment, the first maintenance dose is administered one month after the loading dose.

In a preferred embodiment, administration of the monthly maintenance dose is continued as long as treatment is necessary.

In one embodiment, a method of treating hemophilia, such as hemophilia A with or without inhibitors, is provided, wherein the method comprises administering to said patient a composition comprising a bispecific antibody capable of binding to FIX(a) and FX(a), wherein said administration provides a steady-state plasma concentration of said bispecific antibody in the range of 2 μg/mL to about 18 μg/mL, preferably in the range of about 3 μg/mL to about 9 μg/mL, such as 6 to 7 μg/mL, such as 6.5 to 7 μg/mL.

In a preferred embodiment, the bispecific antibody comprises an anti-FIX(a) antibody or an antigen-binding fragment thereof capable of binding to FIX (SEQ ID NO: 1) and/or its activated form (FIXa), comprising a heavy chain and a light chain, and an anti-FX(a) antibody or an antigen-binding fragment thereof capable of binding to FX (SEQ ID NO: 2) and/or its activated form (FXa), comprising a heavy chain and a light chain, wherein the heavy chain of the anti-FIX(a) antibody or its antigen-binding fragment comprises the CDR1-3 sequences represented by SEQ ID NOs: 3, 4 and 5, respectively, and the light chain of the anti-FIX(a) antibody or its antigen-binding fragment comprises the CDR1-3 sequences represented by SEQ ID NOs: 8, 9 and 10, respectively, and the heavy chain of the anti-FX(a) antibody or its antigen-binding fragment comprises the CDR1-3 sequences represented by SEQ ID NOs: 13, 14 and 15, respectively, and the light chain of the anti-FX(a) antibody or its antigen-binding fragment comprises the CDR1-3 sequences represented by SEQ ID NOs: 18, 19 and 20, respectively.

In a more preferred embodiment, the anti-FIX(a) antibody or antigen-binding fragment thereof comprises a heavy chain variable domain represented by SEQ ID NO:6 and a light chain variable domain represented by SEQ ID NO:11, and the anti-FX(a) antibody or antigen-binding fragment thereof comprises a heavy chain variable domain represented by SEQ ID NO:16 and a light chain variable domain represented by SEQ ID NO:21.

In a most preferred embodiment, the heavy chain of the anti-FIX(a) antibody comprises SEQ ID NO:7 and the light chain of the anti-FIX(a) antibody comprises SEQ ID NO:12, and the heavy chain of the anti-FX(a) antibody comprises SEQ ID NO:17 and the light chain of the anti-FX(a) antibody comprises SEQ ID NO:22.

For mAbl characterized by its heavy and light chain CDR sequences or heavy and light chain variable domain sequences or full-length heavy and light chain sequences, the loading dose in a once-monthly dosing regimen

In one embodiment, the composition comprising the bispecific antibody is administered subcutaneously as a loading dose in a once-monthly dosing regimen, which can be a dose of 8.5, 8.6, 8.7, 8.8, 8.9, 9, 9.1, 9.2, 9.3, 9.4 or 9.5 mg, preferably 9 mg in patients weighing 5 kg to <15 kg.

In one embodiment, the composition comprising the bispecific antibody is administered subcutaneously as a loading dose in a once-monthly dosing regimen, which can be a dose of 39.5, 39.6, 39.7, 39.8, 39.9, 40, 40.1, 40.2, 40.3, 40.4 or 40.5 mg, preferably 40 mg in patients weighing 15 kg to <45 kg.

In one embodiment, the composition comprising the bispecific antibody is administered subcutaneously as a loading dose in a once-monthly dosing regimen, which in patients weighing 45 kg or more may be a dose of 91.5, 91.6, 91.7, 91.8, 91.9, 92, 92.1, 92.2, 92.3, 92.4 or 92.5 mg, preferably 92 mg.

For mAbl characterized by its heavy and light chain CDR sequences or heavy and light chain variable domain sequences or full-length heavy and light chain sequences, the maintenance dose in a once-monthly dosing regimen

In one embodiment, the composition comprising the bispecific antibody is administered subcutaneously at a maintenance dose in a once-monthly dosing regimen, which can be 8.5, 8.6, 8.7, 8.8, 8.9, 9, 9.1, 9.2, 9.3, 9.4 or 9.5 mg, preferably a dose of 9 mg in patients weighing 5 kg to <15 kg.

In one embodiment, the composition comprising the bispecific antibody is administered subcutaneously at a maintenance dose in a once-monthly dosing regimen, which in patients weighing 15 kg to <45 kg can be a dose of 19.5, 19.6, 19.7, 19.8, 19.9, 20, 20.1, 20.2, 20.3, 20.4 or 20.5 mg, preferably 20 mg.

In one embodiment, the composition comprising the bispecific antibody is administered subcutaneously at a maintenance dose in a once-monthly dosing regimen, which in patients weighing 45 kg or more may be a dose of 45.5, 45.6, 45.7, 45.8, 45.9, 46, 46.1, 46.2, 46.3, 46.4 or 46.5 mg, preferably 46 mg.

In one embodiment, the administration method as disclosed herein provides an ABR of 1, 2, 3, 4 or 5, or in the range of 0-3, such as 1-3 or 2-3, or in the range of 1-5, such as 1-2, 1-3, 1-4, 2-3, 2-4, 2-5, 3-4, 3-5 or 4-5.

In one embodiment, the method of administration includes administering one, two or three further loading doses - referred to as "extended loading doses" to distinguish them from the initial loading dose - if the patient does not achieve an adequate clinical response at the end of the initial loading period. The dose and dosing interval during the extended loading period are generally the same as those during the initial loading period, but can be changed if the competent healthcare professional has reason to believe that the patient may benefit from a change such as increasing the dose of the bispecific antibody or more frequent dosing.

In one embodiment, the first maintenance dose is administered one month after the patient is administered the loading dose.

In one embodiment, _{T 1/2} is about 30.4 days.

In one embodiment, T _max is about 9.1 days.

In preferred embodiments, the treatment as disclosed herein is a prophylactic treatment.

In one embodiment, the methods of administration as disclosed herein reduce spontaneous bleeding or bleeding episodes in a patient susceptible to such spontaneous bleeding or bleeding episodes.

Pharmaceutical preparations

In one aspect of the present invention, pharmaceutical compositions suitable for use in the methods of administration disclosed herein are provided.

Such a pharmaceutical composition comprises the bispecific antibody, preferably present in a concentration of 1 mg/mL to 100 mg/mL, such as 2 mg/mL to 100 mg/mL, such as 2 mg/mL to 60 mg/mL, and a pH in the range of 5.5 to 7.5, preferably in the range of 6.0-6.5, such as about 6.3, such as 6.3.

In more preferred embodiments, the concentration of the bispecific antibody is 2 mg/mL, 5 mg/mL, 11.25 mg/mL, 25 mg/mL or 57.5 mg/mL.

Such pharmaceutical compositions are suitable for use with, but not limited to, fixed dose injection devices, such as injection devices configured to administer 0.8 ml per injection.

In a preferred embodiment, the pharmaceutical composition is an aqueous formulation. In one embodiment, the bispecific antibody is mAbl.

The pharmaceutical composition may further comprise one or more of a buffer system, a preservative, a tonicity agent, a chelating agent, a stabilizer or a surfactant and 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 composition comprises 1 mg/mL to 100 mg/mL of a bispecific antibody such as mAbl, L-arginine or L-arginine hydrochloride, L-histidine, and a surfactant, and the pH is in the range of 5.5 to 7.0.

In one embodiment, the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of a bispecific antibody such as mAbl, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, polysorbate 20 or polysorbate 80, and a pH in the range of 5.5-7.0.

In one embodiment, the pharmaceutical composition comprises 2 mg/mL, 5 mg/mL, 11.25 mg/mL, 25 mg/mL or 57.5 mg/mL of the bispecific antibody mAb1, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, about 0.02% polysorbate 20, and a pH of about 6.3.

In a preferred embodiment, the pharmaceutical composition comprises about 2 mg/mL, about 5 mg/mL, about 11.25 mg/mL, about 25 mg/mL or about 57.5 mg/mL of the bispecific antibody mAb1, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, about 0.02% polysorbate 20, and a pH of about 6.3.

In a preferred embodiment, the pharmaceutical composition comprises 2 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02% polysorbate 20, and a pH of 6.3.

In another preferred embodiment, the pharmaceutical composition comprises 5 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02% polysorbate 20, and a pH of 6.3.

In another preferred embodiment, the pharmaceutical composition comprises 11.25 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02% polysorbate 20, and a pH of 6.3.

In another preferred embodiment, the pharmaceutical composition comprises 25 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02% polysorbate 20, and a pH of 6.3.

In yet another preferred embodiment, the pharmaceutical composition comprises 57.5 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02% polysorbate 20, and a pH of 6.3.

In one embodiment, the pharmaceutical composition comprises 2 mg/mL, 5 mg/mL, 11.25 mg/mL, 25 mg/mL or 57.5 mg/mL of the bispecific antibody mAb1, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, about 0.02 w/v % polysorbate 20, and a pH of about 6.3.

In a preferred embodiment, the pharmaceutical composition comprises about 2 mg/mL, about 5 mg/mL, about 11.25 mg/mL, about 25 mg/mL or about 57.5 mg/mL of the bispecific antibody mAb1, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, about 0.02 w/v % polysorbate 20, and a pH of about 6.3.

In a preferred embodiment, the pharmaceutical composition comprises 2 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02 w/v % polysorbate 20, and a pH of 6.3.

In another preferred embodiment, the pharmaceutical composition comprises 5 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02 w/v % polysorbate 20, and a pH of 6.3.

In another preferred embodiment, the pharmaceutical composition comprises 11.25 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02 w/v % polysorbate 20, and a pH of 6.3.

In another preferred embodiment, the pharmaceutical composition comprises 25 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02 w/v % polysorbate 20, and a pH of 6.3.

In yet another preferred embodiment, the pharmaceutical composition comprises 57.5 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02 w/v % polysorbate 20, and a pH of 6.3.

In one embodiment, a pharmaceutical composition comprising a mAbl concentration of 2 mg/mL is used to administer a 1.6 mg dose of mAbl.

In one embodiment, a pharmaceutical composition comprising a mAbl concentration of 5 mg/mL is used to administer a 4 mg dose of mAbl.

In one embodiment, a pharmaceutical composition comprising a mAbl concentration of 11.25 mg/mL is used to administer a 9 mg dose of mAbl.

In one embodiment, a pharmaceutical composition comprising a mAbl concentration of 25 mg/mL is used to administer a 20 mg dose of mAbl.

In one embodiment, a pharmaceutical composition comprising a mAbl concentration of 57.5 mg/mL is used to administer a 46 mg dose of mAbl.

In some embodiments, a pharmaceutical composition as disclosed herein is intended for use in and/or contained in an injection device.

In preferred embodiments, the injection device is a fixed dose device, such as a device configured to deliver a single predetermined dose of a pharmaceutical composition or a device that delivers multiple predetermined doses of a pharmaceutical composition, the latter sometimes referred to as a multiple fixed dose device or a fixed dose multi-shot device. In some embodiments, the injection device is a disposable, pre-filled, multi-dose device. In some embodiments, the injection device is a disposable, pre-filled, single-shot device.

In one embodiment, the pharmaceutical composition of the invention is administered using an injection device comprising a cannula having a needle gauge in the range of 26 to 36.

In one embodiment, the loading dose and/or the maintenance dose may each be administered as a single injection, wherein the entire loading dose and/or the maintenance dose is administered as a single administration, ie, wherein the entire dose is administered all at once.

In some embodiments, the loading dose and/or maintenance dose is administered as multiple smaller doses, for example, as a total of 2, 3, or 4 smaller doses that constitute a complete loading dose or maintenance dose. As a non-limiting example, a loading dose of 80 mg of a bispecific antibody can be administered as three smaller doses of 60 mg each. Alternatively, for example, two 50 mg doses and one 80 mg dose may also be administered consecutively.

The pharmaceutical composition may be administered as a subcutaneous injection of at least 0.05 mL of injection solution to achieve the desired dose measured in mg.

For example, for a pharmaceutical composition comprising 100 mg/mL of mAb1 compound, a volume of 100 μL is required to provide a dose of 10 mg.

The necessary volume will depend on the concentration of the bispecific antibody in the pharmaceutical composition to be administered, as lower volumes will generally result in the need for additional injections, while larger volumes will generally result in discomfort at the injection site for the patient. Typically, a volume of 0.08 to 1.5 mL, preferably 0.2 to 1 mL, more preferably 0.6 to 0.9 mL, more preferably 0.8 mL of injection solution is administered per injection.

The pharmaceutical compositions described above can be combined to allow a specific dose to be administered using a (fixed) injection solution volume of 0.8 mL per injection - as disclosed herein. For example, two injections of 0.8 mL of a pharmaceutical composition containing 57.5 mg/mL mAb1 will allow a cumulative dose of 92 mg mAb1. In another example, one injection of 0.8 mL of a pharmaceutical composition containing 11.25 mg, followed by one injection of 0.8 mL of a pharmaceutical composition containing 57.5 mL, will allow a cumulative dose of 55 mg mAb1.

The pharmaceutical compositions may be administered at the same or different injection sites.

In another general aspect, the present invention relates to a kit comprising an injection device containing a pharmaceutical composition comprising mAb 1 and one or more pharmaceutically acceptable carriers. The kit comprises instructions for subcutaneous administration of the pharmaceutical composition for the treatment of hemophilia A with or without inhibitors.

Further implementation plans

1. A dosage regimen for treating hemophilia A with or without inhibitors comprising subcutaneous administration of a bispecific antibody comprising

An anti-FIX(a) antibody or an antigen-binding fragment thereof capable of binding to FIX (SEQ ID NO: 1) and/or its activated form (FIXa), comprising a heavy chain and a light chain, and an anti-FX(a) antibody or an antigen-binding fragment thereof capable of binding to FX (SEQ ID NO: 2) and/or its activated form (FXa), comprising a heavy chain and a light chain,

wherein the heavy chain of the anti-FIX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 3, 4 and 5, respectively, and

The light chain of the anti-FIX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 8, 9 and 10, respectively, and

The heavy chain of the anti-FX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 13, 14 and 15, respectively, and

The light chain of the anti-FX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 18, 19 and 20, respectively;

This is followed by a monthly maintenance dose.

wherein the loading dose comprises the same or higher amount of the bispecific antibody as the maintenance dose.

2. The dosage regimen according to embodiment 1, wherein the anti-FIX(a) antibody or antigen-binding fragment thereof comprises

a heavy chain variable domain represented by SEQ ID NO: 6 and a light chain variable domain represented by SEQ ID NO: 11, and

The anti-FX(a) antibody or antigen-binding fragment thereof comprises a heavy chain variable domain represented by SEQ ID NO:16 and a light chain variable domain represented by SEQ ID NO:21.

3. The dosage regimen according to embodiment 1 or 2, wherein

The heavy chain of the anti-FIX(a) antibody comprises SEQ ID NO: 7, and the light chain of the anti-FIX(a) antibody comprises SEQ ID NO: 12, and

The heavy chain of the anti-FX(a) antibody comprises SEQ ID NO:17, and the light chain of the anti-FX(a) antibody comprises SEQ ID NO:22.

4. The dosage regimen according to any one of the preceding embodiments, wherein the bispecific antibody is administered as a pharmaceutical composition comprising the bispecific antibody.

5. The dosage regimen according to any of the preceding embodiments, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, L-arginine or L-arginine hydrochloride, L-histidine, and a surfactant at about pH 5.5 to about pH 7.

6. The dosage regimen according to any one of the preceding embodiments, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, L-arginine or L-arginine hydrochloride, L-histidine, and a surfactant, and the pH is about 6.3.

7. The dosage regimen according to any of the preceding embodiments, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, about 150 mM L-arginine, about 20 mM L-histidine, and about 0.02% polysorbate 20 or 80, at a pH of about 6.3.

8. The dosage regimen according to any of the preceding embodiments, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, and about 0.02% polysorbate 20, at a pH of about 6.3.

9. The dosage regimen according to any of the preceding embodiments, wherein the pharmaceutical composition comprises 2 mg/mL to 60 mg/mL of mAb1, such as 2 mg/mL, 5 mg/mL, 11.25 mg/mL, 25 mg/mL or 57.5 mg/mL of mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, and 0.02% polysorbate 20, at a pH of 6.3.

10. The dosage regimen according to any one of the preceding embodiments, wherein the bispecific antibody is administered to a human patient.

11. The dosage regimen according to any one of embodiments 1-10, wherein the loading dose comprises

about 5 mg to about 15 mg of the bispecific antibody, administered to a patient weighing 5 kg to <15 kg, or

about 35 mg to about 45 mg of the bispecific antibody, administered to a patient weighing 15 kg to <45 kg, or

about 85 mg to about 95 mg of the bispecific antibody, administered to a patient weighing 45 kg or more,

The maintenance dose includes

About 7 mg to about 11 mg of the bispecific antibody, administered once a month to a patient weighing 5 kg to <15 kg, or

About 18 mg to about 22 mg of the bispecific antibody, administered once a month to a patient weighing 15 kg to <45 kg, or

about 44 mg to about 48 mg of the bispecific antibody, administered once a month to a patient weighing 45 kg or more,

The first maintenance dose is administered one month after the loading dose.

12. The dosage regimen of embodiment 11, wherein a loading dose of 8.5, 8.6, 8.7, 8.8, 8.9, 9, 9.1, 9.2, 9.3, 9.4 or 9.5 mg of the bispecific antibody is administered to a patient weighing 5 kg to <15 kg, or

administering a loading dose of 39.5, 39.6, 39.7, 39.8, 39.9, 40, 40.1, 40.2, 40.3, 40.4 or 40.5 mg of the bispecific antibody to a patient weighing 15 kg to <45 kg, or

A loading dose of 91.5, 91.6, 91.7, 91.8, 91.9, 92, 92.1, 92.2, 92.3, 92.4, 92.5 mg of the bispecific antibody is administered to a patient weighing 45 kg or more, and wherein

A maintenance dose of 8.5, 8.6, 8.7, 8.8, 8.9, 9, 9.1, 9.2, 9.3, 9.4 or 9.5 mg of the bispecific antibody is administered once a month to patients weighing 5 kg to <15 kg, or

administering a maintenance dose of 19.5, 19.6, 19.7, 19.8, 19.9, 20, 20.1, 20.2, 20.3, 20.4 or 20.5 mg of the bispecific antibody once a month to a patient weighing 15 kg to <45 kg, or

Patients weighing 45 kg or more are administered a maintenance dose of 45.5, 45.6, 45.7, 45.8, 45.9, 46, 46.1, 46.2, 46.3, 46.4 or 46.5 mg of the bispecific antibody once a month.

13. The dosage regimen according to embodiment 12, wherein

administering a loading dose of about 9 mg of the bispecific antibody to a patient weighing 5 kg to <15 kg, or

administering a loading dose of about 40 mg of the bispecific antibody to a patient weighing 15 kg to <45 kg, or

A loading dose of about 92 mg of the bispecific antibody is administered to a patient weighing 45 kg or more, and wherein

administering a maintenance dose of about 9 mg of the bispecific antibody once a month to a patient weighing 5 kg to <15 kg, or

A maintenance dose of about 20 mg of the bispecific antibody is administered once a month to a patient weighing 15 kg to <45 kg, or

A maintenance dose of about 46 mg of the bispecific antibody is administered once a month to patients weighing 45 kg or more.

14. The dosage regimen according to any one of the preceding embodiments, wherein the treatment is a prophylactic treatment.

15. The dosage regimen according to any of the preceding embodiments, wherein a steady-state plasma concentration of the bispecific antibody is provided in the range of about 2 μg/mL to about 18 μg/mL.

16. The dosage regimen according to any of the preceding embodiments, wherein a steady-state plasma concentration of the bispecific antibody is provided in the range of about 3 μg/mL to about 9 μg/mL, such as 6.5 μg/mL to 7 μg/mL.

17. The dosage regimen according to any of the preceding embodiments, wherein a steady-state plasma concentration of the bispecific antibody of about 6.5 μg/mL is provided.

18. The dosage regimen of any one of the preceding embodiments, wherein the regimen provides an ABR of 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9 or 3 or 4, or an ABR in the range of 1-5.

19. The dosage regimen according to any one of the preceding embodiments, wherein the bispecific antibody is mAb1.

20. A kit comprising a) a pharmaceutical composition comprising the bispecific antibody according to any one of embodiments 1-4; and b) instructions for once-monthly subcutaneous administration of the pharmaceutical composition for the treatment of hemophilia A with or without inhibitors according to any of the preceding embodiments.

21. A method for treating hemophilia A with or without inhibitors comprising subcutaneously administering a bispecific antibody in one or more loading doses, the bispecific antibody comprising

an anti-FIX(a) antibody or an antigen-binding fragment thereof capable of binding to FIX (SEQ ID NO: 1) and/or its activated form (FIXa), comprising a heavy chain and a light chain, and an anti-FX(a) antibody or an antigen-binding fragment thereof capable of binding to FX (SEQ ID NO: 2) and/or its activated form (FXa), comprising a heavy chain and a light chain,

wherein the heavy chain of the anti-FIX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 3, 4 and 5, respectively, and

The light chain of the anti-FIX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 8, 9 and 10, respectively, and

The heavy chain of the anti-FX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 13, 14 and 15, respectively, and

The light chain of the anti-FX(a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOs: 18, 19 and 20, respectively;

This is followed by a monthly maintenance dose.

wherein the loading dose comprises the same or higher amount of the bispecific antibody as the maintenance dose, and

Therein a plasma concentration of the bispecific antibody is provided in the range of about 2 μg/mL to about 18 μg/mL.

22. The method of claim 21, wherein the anti-FIX(a) antibody or antigen-binding fragment thereof comprises

a heavy chain variable domain represented by SEQ ID NO: 6 and a light chain variable domain represented by SEQ ID NO: 11, and

The anti-FX(a) antibody or antigen-binding fragment thereof comprises a heavy chain variable domain represented by SEQ ID NO:16 and a light chain variable domain represented by SEQ ID NO:21.

23. The method of treatment according to embodiment 21 or 22, wherein

The heavy chain of the anti-FIX(a) antibody comprises SEQ ID NO: 7, and the light chain of the anti-FIX(a) antibody comprises SEQ ID NO: 12, and

The heavy chain of the anti-FX(a) antibody comprises SEQ ID NO:17, and the light chain of the anti-FX(a) antibody comprises SEQ ID NO:22.

24. The method of treatment according to any one of embodiments 21-23, wherein the bispecific antibody is administered as a pharmaceutical composition comprising the bispecific antibody.

25. The method of any one of embodiments 21-24, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, L-arginine or L-arginine hydrochloride, L-histidine, and a surfactant at about pH 5.5 to about pH 7.

26. The method of treatment according to any one of embodiments 21-25, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, L-arginine or L-arginine hydrochloride, L-histidine, and a surfactant, and the pH is about 6.3.

27. The method of any one of embodiments 21-26, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, about 150 mM L-arginine, about 20 mM L-histidine, and about 0.02% polysorbate 20 or 80, at a pH of about 6.3.

28. The method of any one of embodiments 21-27, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, and about 0.02% polysorbate 20, at a pH of about 6.3.

29. The method of any one of embodiments 21-28, wherein the pharmaceutical composition comprises 2 mg/mL to 60 mg/mL of mAb1, such as 2 mg/mL, 5 mg/mL, 11.25 mg/mL, 25 mg/mL, or 57.5 mg/mL of mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, and 0.02% polysorbate 20, at a pH of 6.3.

30. The method of any one of embodiments 21-29, wherein the bispecific antibody is administered to a human patient.

31. The method of treatment according to any one of embodiments 21-30, wherein the loading dose comprises

about 5 mg to about 15 mg of the bispecific antibody, administered to a patient weighing 5 kg to <15 kg, or

about 35 mg to about 45 mg of the bispecific antibody, administered to a patient weighing 15 kg to <45 kg, or

about 85 mg to about 95 mg of the bispecific antibody, administered to a patient weighing 45 kg or more,

The maintenance dose includes

About 7 mg to about 11 mg of the bispecific antibody, administered once a month to a patient weighing 5 kg to <15 kg, or

About 18 mg to about 22 mg of the bispecific antibody, administered once a month to a patient weighing 15 kg to <45 kg, or

about 44 mg to about 48 mg of the bispecific antibody, administered once a month to a patient weighing 45 kg or more,

The first maintenance dose is administered one month after the loading dose.

32. The method of treatment according to embodiment 31, wherein a loading dose of 8.5, 8.6, 8.7, 8.8, 8.9, 9, 9.1, 9.2, 9.3, 9.4 or 9.5 mg of the bispecific antibody is administered to a patient weighing 5 kg to <15 kg, or

administering a loading dose of 39.5, 39.6, 39.7, 39.8, 39.9, 40, 40.1, 40.2, 40.3, 40.4 or 40.5 mg of the bispecific antibody to a patient weighing 15 kg to <45 kg, or

A loading dose of 91.5, 91.6, 91.7, 91.8, 91.9, 92, 92.1, 92.2, 92.3, 92.4, 92.5 mg of the bispecific antibody is administered to a patient weighing 45 kg or more, and

in

A maintenance dose of 8.5, 8.6, 8.7, 8.8, 8.9, 9, 9.1, 9.2, 9.3, 9.4 or 9.5 mg of the bispecific antibody is administered once a month to patients weighing 5 kg to <15 kg, or

administering a maintenance dose of 19.5, 19.6, 19.7, 19.8, 19.9, 20, 20.1, 20.2, 20.3, 20.4 or 20.5 mg of the bispecific antibody once a month to a patient weighing 15 kg to <45 kg, or

Patients weighing 45 kg or more are administered a maintenance dose of 45.5, 45.6, 45.7, 45.8, 45.9, 46, 46.1, 46.2, 46.3, 46.4 or 46.5 mg of the bispecific antibody once a month.

33. The method of treatment according to embodiment 32, wherein

administering a loading dose of about 9 mg of the bispecific antibody to a patient weighing 5 kg to <15 kg, or

administering a loading dose of about 40 mg of the bispecific antibody to a patient weighing 15 kg to <45 kg, or

A loading dose of about 92 mg of the bispecific antibody is administered to a patient weighing 45 kg or more, and wherein

administering a maintenance dose of about 9 mg of the bispecific antibody once a month to a patient weighing 5 kg to <15 kg, or

administering a maintenance dose of about 20 mg of the bispecific antibody once a month to a patient weighing 15 kg to <45 kg, or

A maintenance dose of about 46 mg of the bispecific antibody is administered once a month to patients weighing 45 kg or more.

34. The method of any one of embodiments 21-33, wherein the treatment is a prophylactic treatment.

35. The method of treatment according to any one of Embodiments 21-34, wherein a plasma concentration of mAbl is provided in the range of about 3 μg/mL to about 9 μg/mL, such as 6.5 μg/mL to 7 μg/mL.

36. The method of treatment according to any one of Embodiments 21-35, wherein a plasma concentration of mAbl of about 6.5 μg/mL is provided.

37. A treatment method according to any one of embodiments 21-36, wherein the plasma concentration is a steady-state plasma concentration.

38. The method of any one of embodiments 21-37, wherein the treatment provides an ABR of 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3, or 4, or an ABR in the range of 1-5.

39. The method of any one of embodiments 21-27, wherein the pharmaceutical composition comprises 1 mg/mL to 100 mg/mL of the bispecific antibody, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, and about 0.02% polysorbate 20, at a pH of about 6.3.

40. The method of any one of embodiments 21-28, wherein the pharmaceutical composition comprises 2 mg/mL to 60 mg/mL of mAb1, such as 2 mg/mL, 5 mg/mL, 11.25 mg/mL, 25 mg/mL, or 57.5 mg/mL of mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, and 0.02% polysorbate 20, at a pH of 6.3.

41. The method of treatment according to any one of embodiments 21-37, wherein the bispecific antibody is mAb1.

42. A pharmaceutical composition comprising the bispecific antibody mAb1, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, about 0.02 w/v % polysorbate 20, and a pH of about 6.3.

43. A pharmaceutical composition according to embodiment 42, wherein the composition comprises about 2 mg/mL to about 57.5 mg/mL of the bispecific antibody mAb1, about 150 mM L-arginine hydrochloride, about 20 mM L-histidine, about 0.02 w/v % polysorbate 20, and a pH of about 6.3.

44. A kit comprising a) a pharmaceutical composition comprising the bispecific antibody according to any one of embodiments 1-19; and b) instructions for once-monthly subcutaneous administration of the pharmaceutical composition for the treatment of hemophilia A with or without inhibitors according to any one of embodiments 21-41.

45. The kit according to embodiment 20 or 44, wherein the bispecific antibody is mAb1.

In a preferred embodiment, the inventors provide a once-monthly dosing regimen for treating hemophilia A with or without inhibitors, comprising subcutaneous administration of a bispecific antibody comprising

an anti-FIX(a) antibody or an antigen-binding fragment thereof capable of binding to FIX (SEQ ID NO: 1) and/or its activated form (FIXa), comprising a heavy chain and a light chain, and an anti-FX(a) antibody or an antigen-binding fragment thereof capable of binding to FX (SEQ ID NO: 2) and/or its activated form (FXa), comprising a heavy chain and a light chain,

wherein the heavy chain of the anti-FIX(a) antibody comprises SEQ ID NO: 7, and the light chain of the anti-FIX(a) antibody comprises SEQ ID NO: 12, and

The heavy chain of the anti-FX(a) antibody comprises SEQ ID NO: 17, and the light chain of the anti-FX(a) antibody comprises SEQ ID NO: 22;

Monthly maintenance doses are then administered;

wherein the bispecific antibody is administered as a pharmaceutical composition comprising the bispecific antibody, wherein the pharmaceutical composition comprises 11.25 mg/mL, 25 mg/mL or 57.5 mg/mL of the bispecific antibody, 150 mM L-arginine hydrochloride, 20 mM L-histidine and 0.02% polysorbate 20, and the pH is about 6.3;

in

administering a loading dose of 9 mg of the bispecific antibody to a patient weighing 5 kg to <15 kg, or

administering a loading dose of 40 mg of the bispecific antibody to a patient weighing 15 kg to <45 kg, or

A loading dose of 92 mg of the bispecific antibody is administered to a patient weighing 45 kg or more, and wherein

A maintenance dose of 9 mg of the bispecific antibody is administered once a month to patients weighing 5 kg to <15 kg, or

A maintenance dose of 20 mg of the bispecific antibody is administered once a month to patients weighing 15 kg to <45 kg, or

A maintenance dose of 46 mg of the bispecific antibody is administered once a month to patients weighing 45 kg or more,

wherein a steady-state plasma concentration of the bispecific antibody is provided in the range of about 2 μg/mL to about 18 μg/mL.

In a preferred embodiment, the inventors provide a once-monthly dosing regimen for treating hemophilia A with or without inhibitors, comprising subcutaneous administration of a bispecific antibody comprising

an anti-FIX(a) antibody or an antigen-binding fragment thereof capable of binding to FIX (SEQ ID NO: 1) and/or its activated form (FIXa), comprising a heavy chain and a light chain, and an anti-FX(a) antibody or an antigen-binding fragment thereof capable of binding to FX (SEQ ID NO: 2) and/or its activated form (FXa), comprising a heavy chain and a light chain,

wherein the heavy chain of the anti-FIX(a) antibody comprises SEQ ID NO: 7, and the light chain of the anti-FIX(a) antibody comprises SEQ ID NO: 12, and

The heavy chain of the anti-FX(a) antibody comprises SEQ ID NO: 17, and the light chain of the anti-FX(a) antibody comprises SEQ ID NO: 22;

Monthly maintenance doses are then administered;

wherein the bispecific antibody is administered as a pharmaceutical composition comprising the bispecific antibody, wherein the pharmaceutical composition comprises 11.25 mg/mL, 25 mg/mL or 57.5 mg/mL of the bispecific antibody, 150 mM L-arginine hydrochloride, 20 mM L-histidine and 0.02 w/v% polysorbate 20, and the pH is about 6.3;

in

administering a loading dose of 9 mg of the bispecific antibody to a patient weighing 5 kg to <15 kg, or

administering a loading dose of 40 mg of the bispecific antibody to a patient weighing 15 kg to <45 kg, or

A loading dose of 92 mg of the bispecific antibody is administered to a patient weighing 45 kg or more, and wherein

A maintenance dose of 9 mg of the bispecific antibody is administered once a month to patients weighing 5 kg to <15 kg, or

A maintenance dose of 20 mg of the bispecific antibody is administered once a month to patients weighing 15 kg to <45 kg, or

A maintenance dose of 46 mg of the bispecific antibody is administered once a month to patients weighing 45 kg or more,

wherein a steady-state plasma concentration of the bispecific antibody is provided in the range of about 2 μg/mL to about 18 μg/mL.

Those skilled in the art will be aware of various methods for measuring and calculating the pharmacokinetic (e.g., but not limited to, _Cmax , _Tmax , serum half-life) and pharmacodynamic parameters described herein. In addition, the skilled artisan will be aware of various methods for making statistical comparisons (e.g., but not limited to, comparisons of changes from baseline to post-treatment and/or comparisons between treatment groups) and/or analyzing the pharmacokinetic and pharmacodynamic parameters described herein.

Example

List of abbreviations

CL: systemic clearance

IIV: Inter-individual variability

_Ka : absorption rate constant

MAD: multiple ascending dose

OFV: Objective Function Value

PwHA: People with hemophilia A

PK: Pharmacokinetics

SAD: Single ascending dose

SIA: sequence identical analogs

Q: Interventricular clearance

QW: Once a week

Q2W: Every two weeks

Q4W: every four weeks

QM: Once a month

RSE: relative standard error

V ₂ : Central distribution volume

V ₃ : Peripheral distribution volume

F: Relative bioavailability

Example 1: Dose escalation study

The following anti-FIX(a)/FX(a) bispecific antibody is being developed for patients with hemophilia A with or without inhibitors (PwHA), which comprises a first heavy chain and a first light chain and a second heavy chain and a second light chain, the first heavy chain comprising SEQ ID NO:7, the first light chain comprising SEQ ID NO:12, the second heavy chain comprising SEQ ID NO:17, and the second light chain comprising SEQ ID NO:22 (mAb1).

FRONTIER1 (EudraCT: 2019-000465-20; NCT04204408) aims to study the safety, tolerability, pharmacokinetics, pharmacodynamics and efficacy of a single ascending subcutaneous dose of mAb1 (bispecific antibody) in healthy participants and multiple ascending doses of mAb1 in PwHA with or without inhibitors. In addition, the study aims to provide data for dose setting in the subsequent FRONTIER study.

Methods: In the single-ascending dose (SAD) phase, healthy subjects received single ascending doses of mAb1 (targeting plasma concentrations of 0.05 to 3 μg/mL) or placebo. In the multiple-ascending dose (MAD) phase, PwHA received multiple ascending doses of mAb1, targeting mean plasma concentrations of 1 μg/mL (cohort 1, QW dosing), 3 μg/mL (cohort 2, QW dosing), 9 μg/mL (cohort 3, QW dosing, and cohort 4, Q4W dosing), or 16 μg/mL (cohort 5, QW dosing).

Nonlinear mixed effects modeling was used to analyze the mAb1 plasma concentration-time data from FRONTIER1. The development of the structural basis model of mAb1 includes a one-compartment model and a two-compartment model. The mAb1 plasma concentration-time curve is best described by a two-compartment model with a first-order elimination and absorption rate, which is parameterized as: absorption rate constant ( _ka ), systemic clearance (CL), inter-compartment clearance (Q), central distribution volume ( _V2 ), peripheral distribution volume ( _V3 ) and relative bioavailability (F, fixed to 1). The random effects were explored using the inter-individual variability (IIV, or inter-subject variability) on the main parameters of the model and the cumulative and/or proportional residual errors. In addition to model stability and parameter accuracy, the minimum decline of the objective function value (OFV) of 3.84 was considered to incorporate random effects. The final model includes IIV on CL, k _a and F, and the residual error is described by the proportional error.

Tables 1-5 list the dosing regimens used for MAD Cohorts 1-5.

Table 1: Loading and maintenance doses in MAD cohort 1 (once weekly dosing)

The loading doses administered at Week 0 and Week 1 were numbered 1 and 2.

*Based on body weight measured at week 0.

Subsequent maintenance doses will be administered starting in Week 2.

*Based on most recent body weight measured per protocol flow chart.

Table 2: MAD Cohort 2 (weekly dosing) loading and maintenance doses Loading dose numbers 1 and 2 to be administered at Week 0 and Week 1:

*Based on body weight measured at week 0.

Subsequent maintenance doses to be administered starting in Week 2:

Table 3: Loading and maintenance doses in MAD cohort 3 (once weekly dosing)

Loading doses numbered 1 and 2 to be administered at Week 0 and Week 1:

Subsequent maintenance doses to be administered starting in Week 2:

Table 4: MAD Cohort 4 (Q4W Dosing)

One loading dose to be administered at Week 0:

Subsequent doses to be administered starting in Week 4:

Table 5: MAD Cohort 5 (QW Dosing)

One loading dose to be administered at Week 0:

Subsequent maintenance doses to be administered starting in Week 2:

Results: mAb1 was well tolerated after single and multiple doses, and no related thromboembolic events or serious adverse events were reported. No anti-mAb1 antibodies were reported. Data from the SAD portion provided an estimated T _1/2 of 30.4 days and a T _max of 9.1 days. The highest mAb1 concentration observed in MAD cohort 5 was 18 μg/mL, which was evaluated as safe and effective (Figure 2).

Population PK model analysis of FRONTIER1 data suggested that the PK of mAb1 was best described by a two-compartment PK model with first-order absorption and elimination. The main covariate factors affecting mAb1 concentrations were baseline body weight and drug product strength. Parameter estimates from the population PK model analysis are collected in Table 6, and the fit of the model to the FRONTIER1 MAD data is presented in Figure 3.

Table 6: Parameter estimates for the population PK model

RSE: relative standard error

During the 12-week observation period, 15 treated bleeds were reported in 8 patients, of which 13 were observed in 6 patients from the lowest dose cohort (9 were traumatic). Both bleeds in patients from cohorts 2 and 3 were traumatic, so neither treated joint bleeds nor spontaneous bleeds were observed outside of cohort 1 (Table 7).

Conclusions: Pharmacokinetic properties support a once-monthly dosing approach. The population PK model adequately described the median trend and variability of the data, with the main covariates being baseline body weight and drug product strength. mAb1 was well tolerated, and no anti-mAb1 antibodies were reported. FRONTIER1 provides encouraging data supporting further clinical development using the dosing approach/dose regimen described herein, which was derived by accounting for the effects of body weight and drug product strength.

Table 7: Treated bleeds during 12 weeks of treatment in PwHA with multiple ascending doses of mAb1

Example 2: Thrombin generation after administration of mAbl

Factor VIII (FVI11) replacement therapy is the standard of care for patients with hemophilia A (HA). mAb1 is a bispecific antibody that binds to factors IX(a) and FX(a) and has enhanced hemostatic properties in vitro and in a mouse model of HA compared to emicizumab. FRONTIER1 (NCT04204408) is a Phase 1/2 study designed to investigate the safety, tolerability, pharmacokinetics, and pharmacodynamics of subcutaneously administered mAb1 in healthy volunteers and patients with severe HA (independent of FVI11 inhibitor status).

Peak thrombin generation and laboratory markers in response to mAbl or emicizumab were analyzed.

Methods: The Phase II portion of FRONTIER1 was an open-label study with subcutaneous administration of mAb1 over 12 weeks in 5 multiple ascending dose cohorts, targeting mean plasma concentrations of 1-18 μg/mL via weekly dosing (cohorts 1-3 and 5) or every four weeks (cohort 4); cohorts 3/4 targeted the same plasma concentrations. An additional exploratory cohort of subjects treated with emicizumab was also included for comparison.

Results: 42 subjects using mAb1 (cohorts 1 [n=7], 2 [n=9], 3 [n=8], 4 [n=8], and 5 [n=10]) and 10 subjects using emicizumab were included. Peak thrombin levels increased with increasing doses of mAb1. In in vitro experiments in which plasma samples neutralized by FVIIl were spiked with mAb1 or emicizumab before dosing, the maximum peak height was achieved at lower plasma concentrations compared with emicizumab, indicating that mAb1 had higher efficacy (see Figure 4). In cohort 2, mean peak thrombin levels were comparable between patients using emicizumab and mAb1. No dose-dependent changes in D-dimer, fibrinogen, platelet, or FIXa/FX antigen levels were observed; most values remained within the normal range. For patients taking mAb1 and emicizumab, dose-dependent increases in prothrombin fragments 1 and 2 were observed (relative change [%]: 26.08, 93.99, 323.65, 606.41, and 315.62 in cohorts 1-5, respectively, and 85.04 for emicizumab), stabilizing at steady-state.

Conclusions: A dose-dependent increase in thrombin generation was observed in mAb1 patients, reaching higher peak thrombin levels than in emicizumab patients. Laboratory parameters showed no safety signals.

Example 3: Extrapolation of the results of Examples 1 and 2 to new dosage regimens for different body weight bands

Based on the specific circumstances newly learned from the FRONTIER1 clinical trial, such as those included in Examples 1 and 2, the inventors designed the administration method disclosed herein, in particular the specific dosage regimen (including the selection of weight bands/groups), by carefully analyzing and using the new data obtained in the trial, including patient responses and observed properties of the bispecific antibody mAb1 (such as dose, T _1/2 , T _max and plasma concentration). For example, the inventors have achieved a therapeutic steady-state plasma concentration of about 2 μg/mL to about 18 μg/mL, preferably in the range of 3 to 9 μg/mL, such as 5, 5.5, 6, 6.5 or 7 μg/mL, and developed the necessary dosage regimen for achieving this steady-state plasma concentration for a specific weight band by expanding the number of weight bands from two to three to cover a body weight of more than 5 kg, thus also including pediatric patients.

Using the population PK model described in Example 1, the inventors have derived a dosing regimen that rapidly establishes steady-state mAbl plasma concentrations within the therapeutic range (2 to 18 μg/mL) with a loading dose followed by maintenance doses at dosing frequencies of QW, Q2W, and QM for typical body weights within the hemophilia population.

Examples of the dosing regimen and weight belts are shown in Table 8.

Table 8: Examples of maintenance and loading doses and body weight bands for mAb1

QM=once a month.

Examples of the mAbl PK profiles for typical subjects within each weight band and dosing frequency are shown in FIG5 (C2 = Cohort 2, _Cavg = mean plasma concentration, _Cmax = maximum plasma concentration).

Example 4: Stability of mAb1 Composition

The pharmaceutical composition analyzed in this example comprises 1-100 mg/mL of the bispecific antibody mAb1, 150 mM L-arginine hydrochloride, 20 mM L-histidine, 0.02 w/v % polysorbate 20, and a pH of about 6.3.

Stability was evaluated based on the following key parameters: appearance, high molecular weight protein (HMWP), monomer and purity. According to the stability results given in Tables 9 to 22, no trend changes or only minor trend changes were observed for the key parameters related to chemical stability (HMWP, monomer and purity) and physical stability (appearance) during storage under long-term storage conditions (5°C ± 3°C) and accelerated storage conditions (25°C ± 2°C).

In summary, the results show that these compositions are chemically and physically stable over time at 5°C and 25°C.

Analytical procedures

1. Pharmacopoeia Analytical Procedure

The following pharmacopoeial analytical procedures are used to control drug products.

Appearance

The appearance of the drug product was determined by visual inspection according to the European and Japanese Pharmacopoeias.

pH

The pH was measured by potentiometric determination according to the European, US and Japanese Pharmacopoeias.

2. Non-pharmacopoeial analytical procedures

The following non-pharmacopoeial analytical procedures are used to control drug products.

2.1 Content determined by SE-HPLC

The content was determined by SE-HPLC using isocratic elution on a size exclusion column and subsequent UV detection. The content was calculated using the area of the high molecular weight protein (HMWP) peak and the monomer peak relative to the area of the HMWP peak and the monomer peak of a reference substance with known content concentration and expressed in mg/ml.

2.2 Monomer determination by SE-HPLC

The monomer content was determined by SE-HPLC using isocratic elution on a size exclusion column and subsequent UV detection. The monomer peak area was calculated relative to the total area and expressed as a percentage.

2.3 High molecular weight protein (HMWP) determined by SE-HPLC

The HMWP content was determined by SE-HPLC using isocratic elution on a size exclusion column and subsequent UV detection. The HMWP peak area was calculated relative to the total area and expressed as a percentage.

2.4 Purity determined by CE-SDS

Purity is defined as the main peak area measured by capillary electrophoresis (CE-SDS) in the presence of sodium dodecyl sulfate. Purity is determined by CE-SDS under non-reducing conditions with UV detection. Purity is calculated as the main peak area relative to the total area, expressed as a percentage.

Stability data for compositions containing mAb1 in the concentration range of 1-100 mg/ml

The stability of compositions containing 1, 2, 5, 11.25, 57.5 and 100 mg/mL mAb1 was followed under long-term storage conditions (5°C ± 3°C) and accelerated storage conditions (25°C ± 2°C).

Results for compositions comprising mAb 1 in the range of 1 mg/ml to 100 mg/ml showed comparable stability. The compositions were chemically and physically stable.

Stability data of 1mg/ml mAb1

The stability data of the compositions comprising 1 mg/ml mAb1 under long-term storage conditions are reported in Table 9, while the stability data under accelerated storage conditions are reported in Table 10.

Table 9: Stability data of 1 mg/ml mAb1 at 5°C ± 3°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Table 10: Stability data of 1 mg/ml mAb1 at 25°C ± 2°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Stability data of 2.0mg/ml mAb1

The stability data of 2.0 mg/ml mAb1 under long-term storage conditions are reported in Table 11, while the stability data under accelerated storage conditions are reported in Table 12.

Table 11: Stability data of 2.0 mg/ml mAb1 at 5°C ± 3°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Table 12: Stability data of 2.0 mg/ml mAb1 at 25°C ± 2°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Stability data of 5.0mg/ml mAb1

The stability data of 5.0 mg/ml mAb1 under long-term storage conditions are reported in Table 13, while the stability data under accelerated storage conditions are reported in Table 14.

Table 13: Stability data of 5.0 mg/ml mAb1 at 5°C ± 3°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Table 14: Stability data of 5.0 mg/ml mAb1 at 25°C ± 2°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Stability data of 11.3mg/ml mAb1

The stability data of 11.3 mg/ml mAb1 under long-term storage conditions are reported in Table 15, while the stability data under accelerated storage conditions are reported in Table 16.

Table 15: Stability data of 11.3 mg/ml mAb1 at 5°C ± 3°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Table 16: Stability data of 11.3 mg/ml mAb1 at 25°C ± 2°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Stability data of mAb1 at 25.0 mg/ml

The stability data of 25.0 mg/ml mAb1 under long-term storage conditions are reported in Table 17, while the stability data under accelerated storage conditions are reported in Table 18.

Table 17: Stability data of 25.0 mg/ml mAb1 at 5°C ± 3°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Table 18: Stability data of 25.0 mg/ml mAb1 at 25°C ± 2°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Stability data of 57.5mg/ml mAb1

The stability data for 57.5 mg/ml mAb1 under long-term storage conditions are reported in Table 19, while the stability data under accelerated storage conditions are reported in Table 20.

Table 19: Stability data of 57.5 mg/ml mAb1 at 5°C ± 3°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Table 20: Stability data of 57.5 mg/ml mAb1 at 25°C ± 2°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Stability data of mAb1 at 100mg/ml

The stability data of 100 mg/ml mAb1 under long-term storage conditions are reported in Table 21, while the stability data under accelerated storage conditions are reported in Table 22.

Table 21: Stability data of 100 mg/ml mAb1 at 5°C ± 3°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Table 22: Stability data of 100 mg/ml mAb1 at 25°C ± 2°C

¹ Comply with: clear or almost clear, colorless or almost colorless liquid, basically free of particles

Although certain features of the present invention have been illustrated and described herein, many modifications, substitutions, changes and equivalents will now occur to those skilled in the art. Therefore, it should be understood that the appended claims are intended to cover all such modifications and changes that fall within the true scope of the invention.

Claims (15)

1. Bispecific antibodies for use in the treatment of hemophilia a with or without inhibitors, wherein the bispecific antibodies comprise

An anti-FIX (a) antibody or antigen-binding fragment thereof capable of binding to FIX (SEQ ID NO: 1) and/or an activated form thereof (FIXa), comprising a heavy chain and a light chain, and

Anti-FX (SEQ ID NO: 2) and/or activated form thereof (FXa)

FX (a) antibodies or antigen-binding fragments thereof, comprising a heavy chain and a light chain,

Wherein the method comprises the steps of

The heavy chain of the anti-FIX (a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOS 3, 4 and 5, respectively, and

The light chain of the anti-FIX (a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOS 8, 9 and 10, respectively, and

The heavy chain of said anti-FX (a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOS 13, 14 and 15, respectively, and

The light chain of the anti-FX (a) antibody or antigen-binding fragment thereof comprises the CDR1-3 sequences represented by SEQ ID NOS: 18, 19 and 20, respectively, and

Wherein the bispecific antibody is to be subcutaneously administered to a human patient in a composition comprising the bispecific antibody,

Wherein the loading dose comprises

About 9mg of said bispecific antibody, to a patient weighing 5kg to <15kg, or

About 40mg of said bispecific antibody, to a patient weighing 15kg to <45kg, or

About 92mg of said bispecific antibody, to a patient weighing 45kg or more,

Wherein the maintenance dose comprises

About 9mg of said bispecific antibody, once a month to a weight of 5kg to 5kg

Patient administration of <15kg, or

About 20mg of said bispecific antibody, administered once a month to a patient weighing 15kg to <45kg, or

About 46mg of said bispecific antibody, administered once a month to a patient weighing 45kg or more,

Wherein a first maintenance dose is administered one month after administration of the loading dose; and

Wherein a steady state plasma concentration of the bispecific antibody in the range of about 2 μg/mL to about 18 μg/mL, such as 3 to 9 μg/mL, such as 6.5 μg/mL is provided.

2. Bispecific antibody for the treatment of hemophilia a with or without inhibitor according to claim 1

Wherein the anti-FIX (a) antibody or antigen-binding fragment thereof comprises a heavy chain variable domain represented by SEQ ID NO. 6 and a light chain variable domain represented by SEQ ID NO. 11, and

Wherein the anti-FX (a) antibody or antigen-binding fragment thereof comprises a heavy chain variable domain represented by SEQ ID NO. 16 and a light chain variable domain represented by SEQ ID NO. 21.

3. The bispecific antibody for use in the treatment of hemophilia a with or without inhibitor according to claim 2, wherein

The heavy chain of the anti-FIX (a) antibody comprises SEQ ID NO. 7 and the light chain of the anti-FIX (a) antibody comprises SEQ ID NO. 12, and

The heavy chain of the anti-FX (a) antibody comprises SEQ ID NO. 17 and the light chain of the anti-FX (a) antibody comprises SEQ ID NO. 22.

4. The bispecific antibody for use in the treatment of hemophilia a with or without inhibitor according to any one of the preceding claims, wherein the bispecific antibody is mAb1.

5. The bispecific antibody for use in the treatment of hemophilia a with or without inhibitor according to any one of the preceding claims, wherein the loading dose comprises

-9Mg of said bispecific antibody, to a patient weighing 5kg to <15kg, or

-40Mg of said bispecific antibody, to a patient weighing 15kg to <45kg, or

92Mg of said bispecific antibody to a patient weighing 45kg or more,

Wherein the maintenance dose comprises

-9Mg of the bispecific antibody, administered once a month to a patient weighing 5kg to <15kg, or

-20Mg of said bispecific antibody, administered once a month to a patient weighing 15kg to <45kg, or

46Mg of said bispecific antibody, administered once a month to a patient weighing 45kg or more,

Wherein the method comprises the steps of

Providing a steady state plasma concentration of the bispecific antibody in the range of 3 to 9 μg/mL, such as 6.5 μg/mL.

6. Bispecific antibody for use in the treatment of hemophilia a with or without inhibitor, wherein the bispecific antibody is mAb1, and

Wherein mAb1 is to be subcutaneously administered to a human patient in a composition comprising mAb1,

Wherein the loading dose comprises

-9Mg of mAb1, administered to a patient weighing 5kg to <15kg, or

40Mg of mAb1, administered to a patient weighing 15kg to <45kg, or

92Mg of mAb1, administered to a patient weighing 45kg or more,

Wherein the maintenance dose comprises

9Mg of mAb1, administered once a month to a patient weighing 5kg to <15kg, or

20Mg of mAb1, administered once a month to a patient weighing 15kg to <45kg, or

46Mg of mAb1, administered once a month to patients weighing 45kg or more,

Wherein a first maintenance dose is administered one month after administration of the loading dose; and

Wherein a steady state plasma concentration of mAb1 in the range of 3 to 9 μg/mL, such as 6.5 μg/mL, is provided.

7. The bispecific antibody for use in treating hemophilia a with or without inhibitor according to any one of the preceding claims, wherein the composition comprises about 150mM L-arginine hydrochloride or L-arginine, about 20mM L-histidine, and a surfactant, such as polysorbate 20 or polysorbate 80, at a ph of about 6.3.

8. The bispecific antibody for use in treating hemophilia a with or without inhibitor according to any one of the preceding claims, wherein the composition comprises about 150mM L-arginine hydrochloride or L-arginine, about 20mM L-histidine, and about 0.02w/v% polysorbate 20 or polysorbate 80, ph about 6.3.

9. The bispecific antibody for use in treating hemophilia a with or without inhibitor according to any one of the preceding claims, wherein the composition comprises about 150mM L-arginine hydrochloride, about 20mM L-histidine, and about 0.02w/v% polysorbate 20, ph about 6.3.

10. The bispecific antibody for use in the treatment of hemophilia a with or without inhibitor according to any one of the preceding claims, wherein the composition comprises 150mM L-arginine hydrochloride, 20mM L-histidine and 0.02w/v% polysorbate 20, ph 6.3.

11. The bispecific antibody for use in treating hemophilia a with or without inhibitor according to any one of the preceding claims, wherein the composition comprises about 2mg/ml, about 5mg/ml, about 11.25mg/ml, about 25mg/ml or about 57.5mg/ml of the bispecific antibody.

12. The bispecific antibody for use in the treatment of hemophilia a with or without inhibitor according to any one of the preceding claims, wherein the treatment provides an annual bleeding rate of 0, 1,2, 3 or 4.

13. A pharmaceutical composition comprising bispecific antibody mAb1, about 150mM L-arginine hydrochloride, about 20mM L-histidine, about 0.02w/v% polysorbate 20, and a ph of about 6.3.

14. The pharmaceutical composition of claim 13, wherein the composition comprises about 2mg/mL to about 57.5mg/mL of bispecific antibody mAb1, about 150mM L-arginine hydrochloride, about 20mM L-histidine, about 0.02w/v% polysorbate 20, ph about 6.3.

15. A kit comprising a) a pharmaceutical composition comprising a bispecific antibody according to any one of claims 1-4; and b) instructions for once monthly subcutaneous administration of the pharmaceutical composition for use in the treatment of hemophilia a with or without inhibitor according to any of the preceding claims.