formulation

Abstract

The present invention relates to the field of pharmaceutical formulations. More specifically, the present invention relates to a liquid formulation comprising an anti-TG2 antibody and a method for producing such a formulation. The liquid formulation according to the present invention is stable when stored at a temperature of about 2 to 25° C. for an appropriate period of time.

Description

[Technical field]

[0001] The present invention relates to the field of pharmaceutical formulations. More particularly, it relates to liquid formulations comprising anti-TG2 antibodies and methods for producing such formulations. The liquid formulations according to the present invention are stable when stored at a temperature of about 2-25° C. for an appropriate period of time. [Background technology]

[0002] Tissue transglutaminase (TG2) is an enzyme that forms crosslinks between proteins via epsilon (gamma-glutamyl) lysine crosslinks. High expression of TG2 leads to abnormal protein crosslinks and is associated with several pathologies, such as various types of tissue scarring, formation of neurofibrillary tangles in some brain diseases, and resistance to chemotherapy in some cancers. Various TG2 inhibitors, such as small molecules, silencing RNAs, or antibodies (e.g., US Pat. No. 5,399,411, US Pat. No. 5,499,421 or US Pat. No. 5,499,431) have been disclosed for the potential treatment of TG2-mediated disorders. Antibodies against TG2 have been described in the literature, but no stable formulations have been proposed so far.

[0003] When preparing pharmaceutical compositions containing biologically active proteins such as antibodies, the compositions must be formulated so that the protein is stable for an appropriate period of time. Loss of protein activity / stability can result from chemical or physical instability of the protein, especially due to denaturation, aggregation, oxidation. Therefore, the resulting product may not be pharma- ceutical acceptable. The use of excipients is known to increase the stability of a given protein, but the stabilizing effect of these excipients depends largely on the nature of the excipient and the biologically active protein itself. There remains a need for formulations containing anti-TG2 antibodies as active ingredients at high concentrations, which formulations are stable for a reasonable period of time and are suitable for use in injections, such as intravenous or subcutaneous injections, that would be useful for administration in the treatment of diseases or disorders mediated by TG2. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] International Publication No. 2006 / 100679 [Patent Document 2] International Publication No. 2012 / 146901 [Patent Document 3] International Publication No. 2013 / 175229 Summary of the Invention [Problem to be solved by the invention]

[0005] It is an object of the present invention to provide a novel formulation comprising an anti-TG2 antibody. More specifically, the formulation is preferably a stable liquid formulation comprising a high concentration of an anti-TG2 antibody. The present invention also provides a method for preparing the liquid formulation according to the present invention. The liquid formulations described herein are useful for administration in the treatment of diseases or disorders mediated by TG2.

[0006] In a first aspect, the present invention provides a stable liquid formulation comprising or consisting of an anti-TG2 antibody, a buffer that maintains a pH of 5.0 or about 5.0 to 6.0 or about 6.0, an amino acid stabilizer, and optionally a polysorbate surfactant. In a preferred embodiment, the buffer is a histidine buffer, and the amino acid is arginine or an arginine salt (such as arginine-HCl). The buffer maintains a pH of 5.5 or about 5.5 (e.g., 5.5±0.2). In a further preferred embodiment, the anti-TG2 antibody is in an amount of 50 or about 50 mg / mL to 300 or about 300 mg / mL. Preferably, the anti-TG2 antibody comprises a light chain variable region defined by SEQ ID NO:1 and a heavy chain variable region defined by SEQ ID NO:2.

[0007] In a second aspect, the present invention provides a method for producing a stable liquid formulation, comprising forming a mixture of an anti-TG2 antibody with a buffer, an amino acid stabilizer, and optionally a polysorbate surfactant. In a preferred embodiment, the buffer is histidine and the amino acid is arginine or an arginine salt (such as arginine-HCl). In a further preferred embodiment, the buffer maintains a pH of 5.0 or about 5.0 to 6.0 or about 6.0 (such as 5.5±0.2). In a further preferred embodiment, the anti-TG2 antibody is in an amount of 50 or about 50 mg / mL to 300 or about 300 mg / mL. Preferably, the anti-TG2 antibody comprises a light chain variable region as defined in SEQ ID NO:1 and a heavy chain variable region as defined in SEQ ID NO:2.

[0008] In a third aspect, there is provided an article of pharmaceutical or veterinary manufacture comprising a container containing a stable liquid formulation according to the invention.

[0009] In a fourth aspect, the present invention provides a stable liquid formulation according to the invention for use in therapy.

[0010] In a fifth aspect, the present invention provides a method of treating a disease or disorder by administering a stable liquid formulation according to the present invention.

[0011] Definition: - the term "about" means approximately or nearly, and in the context of numerical values ​​described herein, preferably means ±10% around the recited or claimed numerical value. - When a range of values ​​is stated or claimed, the range is intended to include the stated values.

[0012] - As used herein, the term "anti-TG2 antibody" intends an antibody molecule that binds to tissue transglutaminase (TG2) protein, an enzyme that forms crosslinks between proteins via epsilon (gamma-glutamyl) lysine bridges. Examples of such antibodies are described in WO 2013 / 175229. Without being limited thereto, an anti-TG2 antibody that can be used according to the present invention comprises, for example, a light chain variable region as defined in SEQ ID NO:1 and a heavy chain variable region as defined in SEQ ID NO:2.

[0013] - The term "antibody" as used herein includes, but is not limited to, monoclonal antibodies, polyclonal antibodies and recombinant antibodies produced by recombinant techniques as known in the art. "Antibody" includes antibodies of any species, in particular mammalian antibodies, such as human antibodies of any isotype, such as IgG1, IgG2a, IgG2b, IgG3, IgG4, IgE, IgD and antibodies produced as dimers of these basic structures, including pentamers such as IgGA1, IgGA2 or IgM, as well as modified variants thereof; non-human primate antibodies, such as antibodies from chimpanzees, baboons, rhesus monkeys or cynomolgus monkeys; rodent antibodies, such as antibodies from mice or rats; rabbit or goat or horse antibodies; camelid antibodies (e.g. antibodies from camels or llamas such as Nanobodies®) and derivatives thereof; antibodies of avian species, such as chicken antibodies; or antibodies of fish species, such as shark antibodies. The term "antibody" also refers to a "chimeric" antibody in which a first portion of at least one heavy and / or light chain antibody sequence is from a first species and a second portion of said heavy and / or light chain antibody sequence is from a second species. Chimeric antibodies of interest herein include "primatized" antibodies that contain variable domain antigen-binding sequences from a non-human primate (e.g., an Old World monkey such as a baboon, rhesus monkey, or cynomolgus monkey) and human constant region sequences. A "humanized" antibody is a chimeric antibody that contains sequences from a non-human antibody. In most cases, a humanized antibody is a human antibody (recipient antibody) in which residues from the recipient's hypervariable regions are replaced by residues from the hypervariable regions [or complementarity determining regions (CDRs)] of a non-human species (donor antibody) such as mouse, rat, rabbit, chicken, non-human primate, etc., to provide the desired specificity, affinity, and activity. In most cases, residues of the human (recipient) antibody outside the CDRs, i.e., in the framework regions (FR), are further replaced by corresponding non-human residues. Furthermore, humanized antibodies may comprise residues that are not found in the recipient antibody or in the donor antibody.These modifications are made to further refine the properties of the antibody. Humanization reduces the immunogenicity of non-human antibodies in humans, facilitating the application of antibodies in the treatment of human diseases. Humanized antibodies and techniques for their production are well known in the art. The term "antibody" also refers to human antibodies, which can be generated as an alternative to humanization. For example, it is possible to generate transgenic animals (e.g. mice) that are capable of producing a full repertoire of human antibodies upon immunization in the absence of endogenous mouse antibody production. Other methods for obtaining human antibodies / antibody fragments in vitro are based on display technologies, such as phage display and ribosome display technology, using recombinant DNA libraries that are at least partially artificially generated or generated from a donor immunoglobulin variable (V) domain gene repertoire. Phage and ribosome display technologies for generating human antibodies are well known in the art. Human antibodies can also be generated from isolated human B cells ex vivo immunized with an antigen of interest, which can then be fused to generate hybridomas and screened for optimal human antibodies. The term "antibody" refers to both glycosylated and aglycosylated antibodies. Furthermore, as used herein, the term "antibody" refers not only to full-length antibodies, but also to antibody fragments, more particularly antigen-binding fragments thereof. Antibody fragments contain at least one heavy or light chain immunoglobulin domain and bind to one or more antigens, as known in the art. Examples of antibody fragments according to the present invention include Fab, modified Fab, Fab', modified Fab', F(ab')2, Fv, Fab-Fv, Fab-dsFv, Fab-Fv-Fv, scFv and Bis-scFv fragments. The fragments may also be single domain antibodies (dAbs) such as diabodies, tribodies, triabodies, tetrabodies, minibodies, sdAbs, VL, VH, VHH or camelid antibodies (e.g. from camel or llama such as Nanobodies®) and VNAR fragments.An antigen-binding fragment according to the invention may also comprise a Fab linked to one or two scFvs or dsscFvs, each scFv or dsscFv binding the same or different targets (e.g. one scFv or dsscFv that binds a therapeutic target and one scFv or dsscFv that increases half-life, e.g. by binding albumin). Examples of such antibody fragments include FabdsscFv (also called BYbe®) or Fab-(dsscFv)2 (also called TrYbe®, see e.g. WO 2015 / 197772). The antibody molecules as defined above, including antigen-binding fragments thereof, are known in the art.

[0014] - The term "stability" as used herein refers to the physical, chemical and conformational stability (including maintenance of biological potency) of an anti-TG2 antibody formulation according to the present invention. Instability of said antibody formulation may be caused by chemical degradation or aggregation of the antibody to form higher order polymers, deglycosylation, modified glycosylation, oxidation or other structural modification that reduces at least one biological activity of the antibody.

[0015] - The term "stable formulation" refers to a formulation in which the protein of interest (here, an anti-TG2 antibody) essentially retains its physical, chemical and biological properties upon storage. A variety of analytical methods are within the knowledge of the skilled artisan to measure the stability of an antibody in a formulation (see some examples in the Examples section). Stability is typically assessed at a selected temperature (e.g., ≦60° C., 2-8° C., 25° C., 35° C. or higher) for a selected period of time (e.g., 3 months, 6 months, 12 months or more). Since an antibody, once formulated, is typically stored in a refrigerator (typically 2-8° C.) or at room temperature (typically 15-25° C.) before being administered to a patient, it is important that the formulated antibody is stable over time at a temperature range of at least 2-25° C., e.g., 2-8° C. and 25° C., as indicated herein. To conclude about the stability over a given period of time, various values ​​can be used, for example but not limited to: 1) less than 90% of the antibody is in monomeric form, 2) the change in the monomeric form of the antibody is less than 10% (compared to the initial data), 3) the amount of high molecular weight species (HMW or HMWS; also referred to herein as aggregates) is less than 5%, or 4) the pH fluctuates less than ±0.2 units (compared to the initial data).

[0016] - The term "buffer" as used herein refers to a solution of a compound known to be safe in a pharmaceutical or veterinary formulation and effective to maintain or control the pH of the formulation in the pH range desired for the formulation. Acceptable buffers for controlling pH at moderately acidic to moderately basic pH include, but are not limited to, phosphate, acetate, citrate, arginine, histidine and TRIS (2-amino-2-hydroxymethyl-1,3,-propanediol, the term including pharma- ceutically acceptable salts thereof) buffers.

[0017] - The term "surfactant" as used herein refers to a soluble compound that can be used specifically to increase the water solubility of hydrophobic, oily substances or to increase the miscibility of two substances with different hydrophobicity. For this reason, these polymers are often used in industrial applications, cosmetics and pharmaceuticals. They are also used as model systems for drug delivery applications, especially to alter the absorption of drugs and their delivery to target tissues. Well-known surfactants include polysorbates (polyoxyethylene derivatives; Tweens) and poloxamers (i.e. copolymers based on ethylene oxide and propylene oxide, also known as Pluronics®).

[0018] - The term "stabilizing agent", "stabilizer" or "isotonicity agent" as used herein is a compound that is physiologically acceptable and confers suitable stability / tonicity to the formulation. It significantly prevents the net flow of water through cell membranes in contact with the formulation. Compounds such as glycerin are often used for such purposes. Other suitable stabilizers include, but are not limited to, amino acids or proteins (e.g., glycine or albumin), salts (e.g., sodium chloride), and sugars (e.g., dextrose, mannitol, sucrose and lactose).

[0019] - The term "vial" or "container" as used herein refers broadly to a reservoir suitable for holding an anti-TG2 antibody formulation in liquid form. Examples of vials that can be used in the present invention include ampoules, tubes, bottles, syringes (such as pre-filled syringes), cartridges, and other reservoirs suitable for delivering an anti-TG2 antibody formulation to a patient via injection, preferably intravenous or subcutaneous injection.

[0020] - The term "solvent" as used herein refers to an aqueous or non-aqueous liquid solvent. The choice of solvent depends inter alia on the solubility of the drug compound in the solvent and the mode of administration. Aqueous solvents may consist of water alone or water and one or more miscible solvents, and may also contain dissolved solutes such as sugars, buffers, salts, and other excipients. The more commonly used non-aqueous solvents are short-chain organic alcohols such as methanol, ethanol, propanol, short-chain ketones such as acetone, and polyhydric alcohols such as glycerol. According to the present invention, the preferred solvents are aqueous solvents such as water or saline.

[0021] - The term "treating" or "treatment" of a disease state includes: (i) inhibiting the disease state, i.e., preventing the onset of the disease state or its clinical symptoms, or (ii) palliating the disease state, i.e., causing the temporary or permanent regression of the disease state or its clinical symptoms.

[0022] - The terms "preventing" or "prevention" of a disease state include keeping clinical symptoms of the disease state from developing in a subject who may be exposed to or susceptible to the disease state, but who has not yet experienced or exhibited symptoms of the disease state. In all embodiments of the present invention, the "pharmaceutical composition" may also be called, without any distinction, a "stable pharmaceutical composition".

[0023] Detailed description of the invention: The present invention is based on a combination of an arginine-based stabilizer and a histidine buffer that maintains a pH of 5.0-6.0, which allows the preparation of pharmaceutical compositions suitable for human use of anti-TG2 antibodies, preferably at high concentrations, without affecting the processability of the pharmaceutical composition and the long-term stability of the antibody. The inventors have found that the pharmaceutical compositions according to the present invention are stable over time, particularly when stored at about 2-25°C, as shown in the Examples section at 2-8°C and 25°C.

[0024] A primary object of the present invention is a stable liquid formulation comprising or consisting of an anti-TG2 antibody, a buffer that maintains a pH of about 5.0 to about 6.0, and an amino acid stabilizer. In a preferred embodiment, the buffer is a histidine buffer and the amino acid stabilizer is selected from the group consisting of arginine or an arginine salt (such as arginine-HCl).

[0025] The invention further provides a method for producing any of the stable liquid formulations of anti-TG2 antibody described herein, comprising combining the anti-TG2 antibody with a buffer, an amino acid stabilizer and optionally a surfactant, such as a polysorbate surfactant. Said step is typically carried out by buffer exchange according to conventional procedures. As an example, to prepare a suitable stable formulation, a predetermined amount of anti-TG2 antibody is buffer exchanged with a histidine buffer that maintains a pH of 5.0 or about 5.0-6.0 or about 6.0, and an amino acid stabilizer (preferably arginine or an arginine salt, such as arginine-HCl). After buffer exchange, the formulation is filtered (final filtration). Depending on the target concentration of the antibody, the formulation can be concentrated between the buffer exchange step and the final filtration. If the formulation contains a surfactant, it is preferably added after the concentration step (if any). Each of these compounds (i.e., anti-TG2 antibody, buffer, amino acid stabilizer and optionally, surfactant) can be used according to the concentrations, pH and / or ratios described herein. The resulting mixture is dispensed into containers. Variations on this process will be apparent to those skilled in the art.

[0026] The invention also provides a pharmaceutical or veterinary article of manufacture comprising a container containing any of the stable liquid formulations described herein, the formulation comprising or consisting of an anti-TG2 antibody, a buffer, an amino acid stabilizer, and optionally a surfactant. Each of these components (i.e., the anti-TG2 antibody, the buffer, the amino acid stabilizer, and optionally a surfactant) may be used according to the concentrations, pH, and / or ratios described herein.

[0027] Also described is packaging that provides instructions for use. Preferably, the anti-TG2 antibody used according to the present invention comprises in its entirety (see also Table A): 1) a light chain variable domain having the sequence defined in SEQ ID NO:1 and a heavy chain variable domain having the sequence defined in SEQ ID NO:2; 2) a light chain variable domain having at least 80% identity or similarity, preferably 90% identity or similarity, to the sequence defined in SEQ ID NO:1 and a heavy chain variable domain having at least 80% identity or similarity, preferably 90% identity or similarity, to the sequence defined in SEQ ID NO:2; 3) a light chain having the sequence defined in SEQ ID NO:3 and a heavy chain having the sequence defined in SEQ ID NO:4, or 4) A light chain having at least 80% identity or similarity, preferably 90% identity or similarity, to the sequence defined in SEQ ID NO:3, and a heavy chain having at least 80% identity or similarity, preferably 90% identity or similarity, to the sequence defined in SEQ ID NO:4.

[0028] [Table A]

[0029] In the context of the present invention as a whole, the amount of anti-TG2 antibody in the formulation is preferably 50 or about 50 mg / mL to 300 or about 300 mg / mL, preferably 100 or about 100 mg / mL to 250 or about 250 mg / mL or more preferably 100 or about 100 mg / mL to 220 or about 220 mg / mL, such as 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, 200, 210 or 220 mg / mL. Alternatively, the anti-TG2 antibody is preferably present in the protein formulation in an amount expressed as weight per 100 mL (% w / v). In such a case, the anti-TG2 antibody is contained in the formulation of the present invention in an amount of about 5 to 30 or about 30% w / v, preferably about 10 to 25 or about 25% w / v, or more preferably about 10 to about 22% w / v, for example, 10.0, 10.5, 11.0, 11.5, 12.0, 12.5, 13.0, 13.5, 14.0, 14.5, 15.0, 15.5, 16.0, 16.5, 17.0, 17.5, 18.0, 18.5, 19.0, 19.5, 20.0, 20.5, 21.0, 21.5, or 22.0% w / v. The anti-TG2 antibody may, for example, comprise a light chain variable region defined by SEQ ID NO:1 and a heavy chain variable region defined by SEQ ID NO:2.

[0030] Preferred buffers according to the present invention are generally histidine buffers (such as L-histidine) that maintain a pH in the range of about 5.0 to about 6.0, preferably in the range of about 5.2 to about 5.8, such as 5.2, 5.3, 5.4, 5.5, 5.6, 5.7 and 5.8. More preferably, the pH is 5.5 or about 5.5. In all embodiments of the present invention, unless otherwise indicated, pH values ​​are measured at room temperature and are preferably within ±0.1 or ±0.2 of the target pH unit (e.g., 5.5±0.1 or 5.5±0.2).

[0031] In the context of the present invention as a whole, the buffer concentration is preferably 10 or about 10 to 100 or about 100 mM. In a preferred embodiment, the buffer concentration is 20 or about 20 to 80 or about 80, more preferably about 40 to about 60 mM, for example 40, 45, 50, 55 or 60 mM. Preferably, the buffer concentration is 50 or about 50 mM.

[0032] In the context of the present invention as a whole, the amino acid stabilizer is selected from the group consisting of arginine or an arginine salt. Preferably, the arginine or arginine salt is L-arginine or an L-arginine salt. Its concentration is preferably 100 or about 100 mM to 300 or about 300 mM, preferably 110 or about 110 to 250 or about 250 mM, more preferably 110 or about 110 to 200 or about 200 mM, for example, 110 or about 110, 115 or about 115, 120 or about 120, 125 or about 125, 130 or about 130, 140 or about 140, 150 or about 150, 160 or about 160, 170 or about 170, 180 or about 180, 190 or about 190, 200 or about 200, 210 or about 210, 220 or about 220, 230 or about 230, 240 or about 240, 250 or about 250, 260 or about 260, 270 or about 270, 280 or about 280, 290 or about 290, 300 or about 300, 310 or about 310, 320 or about 320, 330 or about 330, 340 or about 340, 350 or about 350, 360 or about 360, 370 or about 370, 380 or about 380, 390 or about 390, 400 or about 400, 410 or about 410, 420 or about 35, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, or 200 mM. Arginine or an arginine salt also acts as a viscosity-lowering agent in the formulations of the present invention.

[0033] In the context of the overall invention, a surfactant may optionally be present. If present, it is preferably a polysorbate surfactant, such as polysorbate 20 (PS20, also known as Tween® 20) or polysorbate 80 (PS80, also known as Tween® 80). Preferably, said surfactant is present in the formulation in an amount of 0.01 or about 0.01 to 5 or about 5 mg / mL, more preferably 0.1 or about 0.1 to 1 or about 1 mg / mL, more particularly 0.1 or about 0.1 to 0.5 or about 0.5 mg / mL, such as 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45 or 0.5 mg / mL. Alternatively, said polysorbate surfactant is preferably present in the protein formulation in an amount expressed as % by weight (% w / v) per 100 mL. In such cases, the polysorbate surfactant contained in the formulation of the present invention as a whole is 0.001 to 0.5% w / v, preferably 0.01 to 0.1% w / v, more preferably 0.01 to 0.05% w / v, for example, 0.01, 0.015, 0.02, 0.025, 0.03, 0.035, 0.04, 0.045, 0.045 or 0.05% w / v.

[0034] In a preferred embodiment, the stable liquid formulation of the present invention comprises or consists, in total, of 50 or about 50-300 or about 300 mg / mL of an anti-TG2 antibody, about 10 to about 100 mM histidine at a pH of about 5.5, about 100 to about 300 mM arginine or an arginine salt, and optionally, about 0.01 to 5 mg / mL of a surfactant (such as a polysorbate surfactant).

[0035] Alternatively, the stable liquid formulation of the present invention comprises or consists of about 5 to about 30% w / v anti-TG2 antibody, about 10 to about 100 mM histidine at a pH of about 5.5, about 100 to about 300 mM arginine or an arginine salt, and optionally, 0.001 to 0.5% w / v surfactant (such as a polysorbate surfactant).

[0036] By way of illustrative (but not limiting) example, provided herein is a stable liquid formulation comprising or consisting of: i) about 125 mg / mL of an anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 125 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; ii) about 125 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 150 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; iii) about 150 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 125 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; iv) about 150 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 150 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; v) about 175 mg / mL of an anti-TG2 antibody, about 50 mM histidine buffer at or about pH 5.5, about 125 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; vi) about 175 mg / mL of an anti-TG2 antibody, about 50 mM histidine buffer at or about pH 5.5, about 150 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; vii) about 200 mg / mL anti-TG2 antibody, about 50 mM histidine buffer at pH 5.5 or about 5.5, about 125 mM arginine-HCl, and optionally about 0.02-0.05% w / v polysorbate, or viii) about 200 mg / mL anti-TG2 antibody, about 50 mM histidine buffer at pH 5.5 or about 5.5, about 150 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; Here, the anti-TG2 antibody may, for example, comprise a light chain variable region defined in SEQ ID NO:1 and a heavy chain variable region defined in SEQ ID NO:2.

[0037] Preferably, the formulations of the invention retain at least 80% of the biological activity of the anti-TG2 antibody at the time of formulation and / or packaging for a period of at least 12 months (prior to first use). Anti-TG2 antibody activity can be measured according to routine methods such as Elisa or cell-based assays.

[0038] Additional excipients for use within pharmaceutical compositions of the present invention include, but are not limited to, a stabilizer, a bulking agent, a solubilizing agent, or combinations thereof.

[0039] The present invention also provides a container comprising a pharmaceutical composition according to the present invention. In particular, the container may be, without limitation, a vial, ampoule, tube, bottle, cartridge or syringe (such as a pre-filled syringe) containing the pharmaceutical composition.

[0040] The container may be part of a kit-of-parts comprising one or more containers containing a pharmaceutical composition of the invention, a delivery device such as a syringe, prefilled syringe, auto-injector, needle-free device, implant or patch, or other parental administration device, and instructions for use.

[0041] The liquid formulation of the present invention can be stored for at least about 12 months to about 36 months. Under preferred storage conditions, prior to first use, the formulation is stored away from bright light (preferably in the dark) at a temperature of about 2-25°C, e.g., room temperature (25 or about 25°C) or 2-8°C (see examples below). The formulation minimizes loss of the active principle, i.e., anti-TG2 antibody. The formulation has also been found to be resistant to acidification and formation of protein aggregates, while having an acceptable viscosity (preferably about 30 cP or less).

[0042] The present invention provides stable liquid formulations of anti-TG2 antibodies for use in therapy. For example, the stable liquid formulations of anti-TG2 antibodies described herein are suitable for pharmaceutical or veterinary use. The present invention also provides methods of treating a disease or disorder by administering the stable liquid formulations of anti-TG2 antibodies.

[0043] The stable liquid formulation containing the anti-TG2 antibody of the present invention can be administered to improve or treat a disease or disorder mediated by TG2. Such diseases or disorders mediated by TG2 include, for example, celiac disease, wound healing disorders, scar formation, keloids and hypertrophic scars, ocular scar formation, inflammatory bowel disease, macular degeneration, Graebe's ophthalmopathy, drug-induced ergotism, psoriasis, fibrotic diseases or fibrosis-related diseases, atherosclerosis, restenosis, inflammatory diseases, autoimmune diseases, neurodegenerative / neurological diseases (e.g., Huntington's disease, Alzheimer's disease, Parkinson's disease, polyglutamine deficiency syndrome, etc.). , spinobulbar muscular atrophy, dentatorubral-pallidoluysian atrophy, spinocerebellar ataxia 1, 2, 3, 6, 7 and 12, rubro-pallidal atrophy, spinocerebellar palsy) and / or cancer (e.g. glioblastoma such as glioblastoma of Li-Fraumeni syndrome and sporadic glioblastoma, malignant melanoma, pancreatic ductal adenocarcinoma, myeloid leukemia, acute myeloid leukemia, myelodysplastic syndrome, myeloproliferative syndrome, gynecological cancer, Kaposi's sarcoma, leprosy, collagen colitis).

[0044] The pharmaceutical compositions of the present invention can be administered in a therapeutically effective amount. As used herein, the term "therapeutically effective amount" refers to the amount of therapeutic agent (i.e., antibody) required to treat, ameliorate, or prevent a disease or disorder mediated by TG2, or to exhibit a detectable therapeutic, pharmacological, or prophylactic effect. For any antibody, the therapeutically effective amount can be initially estimated either in cell culture assays or in animal models (usually rodents, rabbits, dogs, pigs, primates). The animal models can also be used to determine appropriate concentration ranges and routes of administration. Such information can then be used to determine useful doses and routes of administration for administration in humans.

[0045] In treating the above diseases and / or disorders, the appropriate dosage will vary depending, for example, on the particular antibody employed, the subject being treated, the mode of administration, and the nature and severity of the condition being treated. In certain embodiments, the pharmaceutical compositions of the present invention are administered intravenously or subcutaneously. When administered intravenously, the administration can be by bolus injection or continuous infusion. Depending on the mode of administration, the formulations described herein can be diluted with a solvent (such as NaCl) prior to use.

[0046] The pharmaceutical compositions according to any embodiment of the present invention may also be administered by intramuscular injection. The pharmaceutical compositions may be injected using injection devices such as syringes, auto-injectors, needle-free devices, hand-held devices, implants and patches.

[0047] The liquid pharmaceutical formulations of the invention are suitably administered to a patient at one time or over a series of treatments and can be administered to a patient at any time following diagnosis; they can be administered as the sole treatment or in combination with other drugs or treatments useful in the treatment of conditions such as those previously described herein.

[0048] The antibody may be the only active ingredient in the liquid pharmaceutical formulation. Alternatively, the antibody may be administered in combination with one or more other therapeutically active ingredients, e.g., simultaneously, sequentially, or separately. As used herein, an active ingredient refers to an ingredient that has a pharmacological effect, such as a therapeutic effect, at an appropriate dose. In some embodiments, the antibody in the pharmaceutical composition may be accompanied by other active ingredients, including other antibodies or non-antibody components, administered by the same or different administration route, to treat other inflammatory or autoimmune diseases. In one embodiment, the subject is administered simultaneously or sequentially (before and / or after) other antibody components, such as anti-TNF antibodies, or non-antibody components, such as small drug molecules.

[0049] The following examples are provided to further illustrate the preparation of the formulations and compositions of the present invention. The scope of the present invention should not be construed as consisting solely of the following examples. EXAMPLES

[0050] material: Anti-TG2 antibody: The anti-TG2 monoclonal antibody (mAb) used in the following examples comprises a light chain variable region defined in SEQ ID NO:1 and a heavy chain variable region defined in SEQ ID NO:2, and is designated mAb1 in the following examples.

[0051] method: Protein concentration: The protein concentration was calculated using ultraviolet-visible spectroscopy according to the following formula: Concentration (mg / mL)=[(A280) / a×b] where A280 = absorbance at 280 nm (AU), a = mass extinction coefficient (1.34 mL mg -1 cm -1 ), b=path length (1 cm).

[0052] Aggregation and fragmentation: Size exclusion chromatography (SEC), e.g., size exclusion ultra-performance liquid chromatography (SE-UPLC), was used according to standard methods. The percentage values ​​of the peak areas of the main species and species eluting before and after the main species, designated high molecular weight (HMW) and low molecular weight (LMW), respectively, were evaluated.

[0053] Acidic and basic species: The presence of acidic and basic species was assessed using isoelectric focusing capillary electrophoresis (iCE) according to standard iCE techniques (which separate proteins based on differences in charge). Typically, peaks eluting before the main peak are labeled as acidic species and those eluting after the main peak are labeled as basic species.

[0054] pH: The pH was assessed according to standard methods using a pH meter equipped with a temperature compensated electrode.

[0055] viscosity: Viscosity measurements were performed using a Rheosense MicroVisc® according to standard methods.

[0056] Example 1 - Preliminary Screening: 1.1. Selection of buffers and major excipients: A suitable formulation needed to be identified that would deliver the highest possible concentration of mAb1 (>10%, and in some cases at least >15%, i.e. at least 150 mg / mL). One of the main concerns with highly concentrated antibody formulations is viscosity, so this was the focus of the preliminary screen. The effect of four different buffers and various excipients on the viscosity of mAb1 was evaluated. mAb1 samples were buffer exchanged into the buffers listed in Table 1 according to standard methods.

[0057] [Table 1]

[0058] The different formulations were analyzed for viscosity and pH at the highest mAb1 concentration reached in each formulation (see Table 2).

[0059] [Table 2]

[0060] Based on the preliminary results, formulations F3, F7, F10, F14 and F16 were further evaluated at different mAb1 concentrations. Indeed, most of these selected formulations reached mAb1 concentrations of 220 mg / ml or more while keeping the viscosity below 20 cP. F16 was the exception, reaching mAb1 concentrations of 270 mg / ml or more, despite a viscosity of 35 cP. Most of the preselected formulations were based on histidine buffer at pH 5.5. The behavior of the selected formulations (in terms of viscosity and pH) was evaluated at decreasing concentrations, as reported in Table 3.

[0061] [Table 3]

[0062] Based on preliminary evaluation, the most promising formulations appeared to be those containing histidine and arginine as buffers (F14 and F16).

[0063] Example 2 - Comparison of Arginine Salts as Viscosity Reducers: The preformulation work of Example 1 highlighted that a formulation consisting of histidine and arginine as a buffer was most promising for formulating high concentrations of mAb1 while maintaining acceptable viscosity (<20 cP at or about 20% mAb1 in the formulation). New formulations were prepared with histidine as the buffer, arginine salts as the viscosity-lowering agent, and optional other excipients, as shown in Table 4.

[0064] [Table 4]

[0065] The different formulations were analyzed for viscosity and pH at the highest mAb1 concentration reached in each formulation (see Table 5).

[0066] [Table 5]

[0067] F1 and F14 had higher viscosities than the other formulations containing arginine and were not considered further. F12 had a much lower viscosity but contained rare excipients and was also not considered. The viscosities of the other formulations were comparable. However, the viscosity of the simpler formulation F15 was comparable to the more sophisticated formulations and was therefore selected for further stability studies.

[0068] Example 3 - Long-term (12 month) stability study of selected formulations: Based on the above examples, seven formulations were selected for long-term testing: - F1: 100 mg / mL mAb1, 50 mM histidine, 125 mM arginine-HCl, pH 5.5, 0.03% PS80; - F2: 125 mg / mL mAb1, 50 mM histidine, 125 mM arginine-HCl, pH 5.5, 0.03% PS80; - F3: 150 mg / mL mAb1, 50 mM histidine, 125 mM arginine-HCl, pH 5.5, 0.03% PS80; - F4: 175 mg / mL mAb1, 50 mM histidine, 125 mM arginine-HCl, pH 5.5, 0.03% PS80; - F5: 200 mg / mL mAb1, 50 mM histidine, 125 mM arginine-HCl, pH 5.5, 0.03% PS80; - F6: 150 mg / mL mAb1, 50 mM histidine, 125 mM arginine-HCl, pH 5.5; - F7: 100 mg / ml mAb1, 50 mM histidine, 250 mM glycine, pH 5.5 (control formulation).

[0069] The long term stability of the different formulations (tested at 5°C, 25°C and 40°C) was analyzed with respect to pH, protein concentration, charge variants (by iCE3), aggregation and fragmentation (by SE UPLC).

[0070] HMWS (see Tables 6-8): HMWS% increases faster with increasing mAb1 concentration under accelerated conditions (25°C and 40°C). No effect of surfactant (PS80) was observed (see F3 vs. F6). Overall, the newly identified formulation (F1) was more stable versus the control formulation (F7) at similar mAb1 concentrations. Stability at 5°C and 25°C was comparable between the 150 mg / mL formulation (i.e., 15%; F3) and the control formulation (F7). There was no difference at 40°C.

[0071] LMWS (see Tables 6-8): The LMWS% decreases with mAb1 concentration. The LMWS increases slightly at 5° C. and 25° C. No effect of the surfactant (PS80) was observed for the HMWS species (see F3 vs. F6).

[0072] Monomers (see Tables 6-8): Stability was comparable for F1 and F7 formulations (i.e., at comparable concentrations) at 5° C. and 25° C. Comparable stability was also observed between F3 formulation (15% mAb1) and the control formulation (F7, 10% mAb) at 5° C. and 25° C.

[0073] Main peak, APG and BPG (see Tables 6-8): No effect of surfactant (PS80) was observed with respect to the main peak, APG and BPG (see F3 vs. F6). The APG% of the control formulation was higher and the BPG% was lower at 25°C and 40°C than the F1 formulation (both at the same mAb1 concentration). Overall, the main peak levels were higher in F1 compared to F7 at 5°C and 25°C (no difference was observed at 40°C).

[0074] Viscosity (see table 9): Viscosity increased as expected with increasing concentration of mAb1 in the formulation, but could be kept below about 20 cP in the most concentrated formulations. The preferred formulation was 15%, with a viscosity of about 6 cP.

[0075] [Table 6-1] [Table 6-2]

[0076] [Table 7-1] [Table 7-2]

[0077] [Table 8-1] [Table 8-2]

[0078] [Table 9]

[0079] Overall conclusion: Overall, formulations F1-F5 were more stable after storage than formulations F6 and F7, especially at the same concentrations, under 5°C (T52w) and 25°C (T26w) conditions. Comparable stability was observed between formulations F3 and F7 at 5°C and 25°C. A general observation is that the higher the antibody concentration, the higher the aggregation levels. However, even at 20%, these aggregation levels were acceptable. iCE data showed no effect of the presence or absence of PS80 (formulations F3 and F6), but there was a slight increase in turbidity (visual assessment) without PS80 (formulation F6). The preferred formulation was F3 (15% of mAb1), but any of F2 (12.5% ​​of mAb1) to F5 (20% of mAb1) were acceptable and could be back-up options.

[0080] References: 1) International Publication No. 2006100679 2) International Publication No. 2012146901 3) International Publication No. 2013175229

Claims

1. A stable liquid formulation comprising an anti-TG2 antibody, a buffer that maintains a pH of about 5.0 to 6.0, and an amino acid stabilizer.

2. 2. The stable liquid formulation of claim 1, wherein the buffer is a histidine buffer.

3. 3. The stable liquid formulation of claim 2, wherein the histidine buffer maintains a pH of 5.5 or about 5.5±0.

2.

4. 2. The stable liquid formulation of claim 1, wherein the concentration of the buffer is at or about 10-100 mM, preferably 20-80 mM, or more preferably 40-60 mM.

5. 2. The stable liquid formulation of claim 1, wherein the amino acid stabilizer is arginine or an arginine salt in an amount of about 100-300 mM, preferably 110-250 mM, or more preferably 120-200 mM.

6. 6. The stable liquid formulation of claim 5, wherein the arginine salt is arginine-HCl.

7. 10. The stable liquid formulation of claim 1, further optionally comprising a polysorbate surfactant.

8. 8. The stable liquid formulation of claim 7, wherein the concentration of the polysorbate surfactant is at or about 0.005-0.1% w / v, preferably at or about 0.01-0.05% w / v.

9. 2. The stable liquid formulation of claim 1, wherein the concentration of the anti-TG2 antibody is from about 50 mg / mL to about 300 mg / mL, preferably from 110 mg / mL to 250 mg / mL, or more preferably from 115 mg / mL to 220 mg / mL.

10. 2. The stable liquid formulation of claim 1, wherein the liquid formulation comprises: i) about 125 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 125 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; ii) about 125 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 150 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; iii) about 150 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 125 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; iv) about 150 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 150 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; v) about 175 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 125 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; vi) about 175 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 150 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; vii) about 200 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 125 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate; or viii) about 200 mg / mL anti-TG2 antibody, about 50 mM histidine buffer maintained at pH 5.5 or about 5.5, about 150 mM arginine-HCl, and, optionally, about 0.02-0.05% w / v polysorbate.

11. 2. The stable liquid formulation of claim 1, wherein the anti-TG2 antibody comprises: 1) a light chain variable domain having the sequence defined in SEQ ID NO:1 and a heavy chain variable domain having the sequence defined in SEQ ID NO:2; 2) a light chain variable domain having at least 80% identity or similarity, preferably 90% identity or similarity, to the sequence defined in SEQ ID NO: 1 and a heavy chain variable domain having at least 80% identity or similarity, preferably 90% identity or similarity, to the sequence defined in SEQ ID NO: 2; 3) a light chain having the sequence defined in SEQ ID NO: 3 and a heavy chain having the sequence defined in SEQ ID NO: 4, or 4) A light chain having at least 80% identity or similarity, preferably 90% identity or similarity, to the sequence defined in SEQ ID NO:3 and a heavy chain having at least 80% identity or similarity, preferably 90% identity or similarity, to the sequence defined in SEQ ID NO:

4.

12. 12. A method for producing the stable liquid formulation of any one of claims 1 to 11, comprising forming a mixture of an anti-TG2 antibody, a histidine buffer, arginine-HCl, and optionally, a polysorbate surfactant.

13. An article of manufacture comprising a container containing the stable liquid formulation of any one of claims 1 to 11.

14. A stable liquid formulation according to any one of claims 1 to 11 for use in therapy.

15. A method for treating a disease or disorder by administering the stable liquid formulation of any one of claims 1 to 11.