CSF-1R antibody formulations

Abstract

The present invention relates to a stable CSF-1R antibody liquid pharmaceutical formulation comprising from 40 mg / mL to 200 mg / mL of an antibody against CSF-1R, from 0.01% (w / v) to 0.1% (w / v) of a surfactant, from 5 mM to 100 mM of a buffer and from 10 mM to 500 mM of at least one stabilizer; the pH value ranges from 4.5 to 7.0.

Description

[0001] This application is a divisional application of the invention application with the application date of September 12, 2019, the Chinese application number 201980067033.9, and the invention name “CSF-1R Antibody Preparation”. Technical Field

[0002] The present invention relates to a preparation of an anti-CSF-1R antibody molecule, a method for preparing the preparation and the use of the preparation. Background Art

[0003] Human CSF-1 receptor (CSF-1R; colony stimulating factor 1 receptor; alias: M-CSF receptor; macrophage colony stimulating factor 1 receptor, Fms proto-oncogene, c-fms, SEQ ID NO: 13) has been known since 1986 (Coussens, L., et al., Nature 320 (1986) 277-280). CSF-1R is a receptor for CSF-1 (colony stimulating factor 1, also known as M-CSF, macrophage colony stimulating factor) and mediates the biological effects of this cytokine (Sherr, CJ et al., Cell 41 (1985) 665-676). The cloning of colony stimulating factor-1 receptor (CSF-1R) (also known as c-fms) was first reported by Roussel, MF et al. (Nature 325 (1987) 549-552). The paper showed that CSF-1R has transforming potential, depending on changes in the C-terminal tail of the protein, including loss of inhibitory tyrosine 969 phosphorylation that binds to Cbl, thereby mediating downregulation of the receptor (Lee, PS et al., Embo J. 18 (1999) 3616-3628). The researchers also identified a second ligand for CSF-1R called interleukin-34 (IL-34) (Lin, H. et al., Science 320 (2008) 807-811).

[0004] Colony-stimulating factor 1 (CSF-1) and its receptor CSF-1R regulate the migration, differentiation, and survival of macrophages and their precursors. CSF-1R is a member of the receptor protein tyrosine kinase (rPTK) family of growth factor receptors, which includes several known proto-oncogenes. Diffuse tenosynovial giant cell tumor (TGCT) of soft tissue (also known as pigmented villonodular synovitis [PVNS]) is a rare proliferative disease affecting large joints characterized by overexpression of CSF-1. In most patients with TGCT, chromosomal translocations involving the gene encoding CSF-1 result in overexpression of this cytokine by cells of the synovial lining. This results in a massive recruitment of CSF-1R-expressing cells, which are primarily non-malignant mononuclear and multinuclear cells that form the bulk of the neoplastic mass. Marginal resection or complete synovectomy remains the treatment of choice for TGCT, but the condition sometimes requires limb resection due to local destructiveness and recurrent tumor growth. Emactuzumab is an anti-CSF-1R antibody that has been shown to successfully treat this rare disease (Cassier, P. et al., Lancet Oncol. 16 (2015) 949-956).

[0005] The main biological effect of CSF-1R signaling is the differentiation, proliferation, migration and survival of hematopoietic precursor cells to the macrophage lineage (including osteoclasts). The activation of CSF-1R is mediated by its ligands CSF-1 (M-CSF) and IL-34. The binding of CSF-1 (M-CSF) to CSF-1R induces the formation of homodimers and the activation of kinases through tyrosine phosphorylation (Li, W. et al., EMBO Journal. 10 (1991) 277-288; Stanley, ER et al., Mol. Reprod. Dev. 46 (1997) 4-10).

[0006] The biologically active homodimeric CSF-1 binds to CSF-1R within subdomains D1 to D3 of the extracellular domain of the CSF-1 receptor (CSF-1R-ECD). CSF-1R-ECD contains five immunoglobulin-like subdomains (designated D1 to D5). Subdomains D4 to D5 of the extracellular domain (CSF-1R-ECD) are not involved in CSF-1 binding (Wang, Z. et al., Molecular and Cellular Biology 13 (1993) 5348-5359). Subdomain D4 is involved in dimerization (Yeung, YG. et al., Molecular & Cellular Proteomics 2 (2003) 1143–1155; Pixley, FJ et al., Trends Cell Biol 14 (2004) 628-638). WO 2011 / 070024 A1 describes antibodies that bind to the human CSF-1R fragment delD4 of SEQ ID NO: 11 (a human CSF-1R fragment in which the D4 subdomain of the human CSF-1R-ECD is missing). These antibodies block the receptor dimerization interface via their epitopes located within D4 and D5 and are therefore unique. One of the antibodies is Emactuzumab or RG7155. The CDR and VH / VL sequences of the antibodies are as disclosed herein.

[0007] As part of the protein drug group, antibody molecules are very susceptible to physical and chemical degradation. Chemical degradation includes any process involving modification of proteins by bond formation or cleavage to obtain new chemical entities. A variety of chemical reactions are known to affect proteins. These reactions may involve hydrolysis (including cleavage of peptide bonds) and deamidation, isomerization, oxidation and decomposition. Physical degradation refers to changes in higher-order structures and includes denaturation, surface adsorption, aggregation and precipitation. Protein stability is affected by the characteristics of the protein itself (e.g., amino acid sequence, glycosylation pattern) and external influencing factors (such as temperature, solvent pH, excipients, interface or shear rate). Therefore, determining the optimal formulation conditions is important for protecting proteins from degradation reactions during production, storage and administration. (Manning, MC et al. (1989), "Stability of protein pharmaceuticals", Pharm Res 6(11), 903-918; Zheng, JY, Janis, LJ (2005), "Influence of pH, buffer species, and storage temperature on physicochemical stability of a humanized monoclonal antibody LA298", Int. J. Pharmaceutics 308, 46-51). It is particularly difficult to obtain stable liquid formulations of therapeutic antibodies when the formulation should contain high concentrations of the antibody.

[0008] Therefore, one object of the present invention is to provide a stable formulation of an anti-CSF-1R antibody, in which as few excipients as possible are required, which enables the desired dosing and convenient administration of the antibody to a patient.

[0009] The formulation of the present invention exhibits good stability after storage at the expected storage temperature of 2°C to 8°C for 24 months without the formation of visible particles, so that it can be administered intravenously without the use of an in-line filter, with greater ease of administration. Shaking and multiple freeze-thaw steps are applied to the liquid formulation to simulate physical stress conditions that may occur during drug production or transportation. After applying shaking and freeze-thaw stress, the formulation of the present invention exhibits good stability. Summary of the invention

[0010] The present invention relates to a stable high-dose pharmaceutical preparation of an antibody binding to CSF-1R, a method for preparing the preparation and the use of the preparation.

[0011] In one aspect, the present invention relates to a pharmaceutical preparation comprising:

[0012] 40 mg / mL to 200 mg / mL of anti-CSF-1R antibody;

[0013] 0.01% (w / v) to 0.1% (w / v) surfactant;

[0014] 5mM to 100mM buffer;

[0015] 10 mM to 500 mM of at least one stabilizer;

[0016] The pH value is in the range of 4.5 to 7.0.

[0017] In a specific aspect, the anti-CSF-1R antibody comprises: a heavy chain variable region comprising a heavy chain CDR1 (CDR-H1) of SEQ ID NO: 1, CDR-H2 of SEQ ID NO: 2, and CDR-H3 of SEQ ID NO: 3; and a light chain variable region comprising a light chain CDR1 (CDR-L1) of SEQ ID NO: 4, CDR-L2 of SEQ ID NO: 5, and CDR-L3 of SEQ ID NO: 6. More specifically, the anti-CSF-1R antibody comprises: a heavy chain variable region comprising an amino acid sequence of SEQ ID NO: 7; and a light chain variable region comprising an amino acid sequence of SEQ ID NO: 8.

[0018] The formulation according to the present invention may be provided in a liquid form, a lyophilized form or a liquid form reconstituted from a lyophilized form.

[0019] Antibodies against CSF-1R that can be used in the pharmaceutical preparation according to the invention are described in detail below.

[0020] In a preferred aspect, the concentration of the anti-CSF-1R antibody contained in the formulation according to the present invention is in the range of 40 mg / mL to 100 mg / mL, preferably in the range of 40 mg / mL to 75 mg / mL, more preferably in the range of 40 mg / mL to 60 mg / mL. A particularly preferred concentration is 50 mg / mL.

[0021] On the other hand, the pharmaceutical preparation comprises a surfactant in a concentration range of 0.01% (w / v) to 0.1% (w / v). In the preparation of the present invention, the concentration of the surfactant is described as a percentage, expressed as weight / volume (w / v). Preferably, the pharmaceutical preparation comprises a surfactant in a concentration range of 0.02% to about 0.05% (w / v) and most preferably 0.04% (w / v). The pharmaceutical preparation according to claim 1 or 2, wherein the surfactant is a polysorbate. The preferred surfactant used in the present invention is polyoxyethylene sorbitan fatty acid ester (i.e., polysorbate), preferably polysorbate 20 or polysorbate 80. In a specific aspect, the surfactant is polysorbate 20.

[0022] On the other hand, the pharmaceutical preparation according to the present invention comprises a buffer. Preferably, the buffer is a histidine buffer. The histidine buffer is a buffer with histidine (usually L-histidine) as a buffer. Most preferably, it is an L-histidine / HCl buffer, which comprises a mixture of L-histidine or L-histidine and L-histidine hydrochloride, and pH adjustment is achieved by hydrochloric acid. Unless otherwise indicated, the term "histidine" used herein to describe the buffer refers to L-histidine / HCl buffer, particularly histidine chloride buffer. In one aspect, the concentration of the buffer is in the range of 10mM to 30mM, more particularly 20mM.

[0023] Preferably, the pH of the formulation is in the range of 5.0 to 6.5, particularly about 6.0. Therefore, the pH of the formulation is preferably 6.0. Regardless of which buffer is used, the pH can be adjusted using acids or bases known in the art (e.g., hydrochloric acid, acetic acid, phosphoric acid, sulfuric acid and citric acid, sodium hydroxide and potassium hydroxide).

[0024] On the other hand, the pharmaceutical formulation according to the present invention comprises at least one stabilizer, the at least one stabilizer being selected from the group consisting of salt, sugar and amino acid. In a particular aspect, the at least one stabilizer is a sugar, in particular an oligosaccharide selected from the group consisting of sucrose, trehalose, lactose, maltose and raffinose. More particularly, the sugar is sucrose. In a preferred aspect, the stabilizer or sugar is present in a concentration in the range of 140mM to 250mM, in particular in the range of 210mM ​​to 230mM. Preferably, the sugar is present in a concentration of 220mM.

[0025] On the other hand, the pharmaceutical formulation according to the present invention comprises a first stabilizer selected from the group consisting of salts, sugars and amino acids and methionine as a second stabilizer. In a preferred aspect, the first stabilizer is present at a concentration of 120mM to 300mM, and the second stabilizer methionine is present at a concentration of 5mM to 25mM. More preferably, methionine is present at a concentration of 10mM.

[0026] The anti-CSF-1R antibody contained in the formulation of the present invention preferably binds to the human CSF-1R fragment delD4 (SEQ ID NO: 11) and the human CSF-1R extracellular domain (SEQ ID NO: 12) at a ratio of 1:50 or less.

[0027] In a particular aspect, the present invention relates to a pharmaceutical preparation comprising

[0028] 40 mg / mL to 100 mg / mL of anti-CSF-1R antibody;

[0029] 20 mM L-histidine;

[0030] 0.03% (w / v) to 0.05% (w / v) polysorbate 20;

[0031] 210mM ​​to 230mM sucrose;

[0032] Optionally 5 mM to 25 mM methionine;

[0033] The pH value is 6.0±0.5.

[0034] In a specific aspect, the pharmaceutical preparation according to the invention comprises

[0035] 50 mg / mL anti-CSF-1R antibody;

[0036] 20 mM L-histidine;

[0037] 0.04% (w / v) polysorbate 20;

[0038] 220 mM sucrose;

[0039] 10 mM methionine;

[0040] The pH value is 6.0±0.5.

[0041] In another aspect, a pharmaceutical preparation is provided herein, comprising

[0042] 50 mg / mL anti-CSF-1R antibody;

[0043] 20 mM L-histidine;

[0044] 0.04% (w / v) polysorbate 20;

[0045] 220 mM sucrose;

[0046] The pH value is 6.0±0.5.

[0047] In another aspect, a pharmaceutical preparation is provided herein, comprising

[0048] 50 mg / mL anti-CSF-1R antibody;

[0049] 20 mM L-histidine;

[0050] 0.04% (w / v) Poloxamer 188;

[0051] 220 mM sucrose;

[0052] The pH value is 6.0±0.5.

[0053] In another aspect, a pharmaceutical preparation is provided herein, comprising

[0054] 50 mg / mL anti-CSF-1R antibody;

[0055] 20 mM L-histidine;

[0056] 0.04% (w / v) polysorbate 20;

[0057] 130 mM sodium chloride;

[0058] The pH value is 6.0±0.5.

[0059] In another aspect, a pharmaceutical preparation is provided herein, comprising

[0060] 50 mg / mL anti-CSF-1R antibody;

[0061] 20 mM L-histidine;

[0062] 0.04% (w / v) polysorbate 20;

[0063] 130 mM sodium chloride;

[0064] 10 mM methionine;

[0065] The pH value is 6.0±0.5.

[0066] In another aspect, a pharmaceutical preparation is provided herein, comprising

[0067] 50 mg / mL anti-CSF-1R antibody;

[0068] 20 mM L-histidine;

[0069] 0.04% (w / v) Poloxamer 188;

[0070] 130 mM sodium chloride;

[0071] The pH value is 6.0±0.5.

[0072] In another aspect, the pharmaceutical preparation according to the present invention comprises

[0073] 50 mg / mL anti-CSF-1R antibody;

[0074] 20 mM L-histidine;

[0075] 0.04% (w / v) polysorbate 20;

[0076] 220 mM sucrose;

[0077] The pH value is 5.5±0.5.

[0078] In another aspect, the pharmaceutical preparation according to the present invention comprises

[0079] 50 mg / mL anti-CSF-1R antibody;

[0080] 20 mM L-histidine;

[0081] 0.04% (w / v) polysorbate 20;

[0082] 220 mM sucrose;

[0083] 10 mM methionine;

[0084] The pH value is 5.5±0.5.

[0085] In another aspect, the pharmaceutical preparation according to the present invention comprises

[0086] 50 mg / mL anti-CSF-1R antibody;

[0087] 20 mM L-histidine;

[0088] 0.04% (w / v) Poloxamer 188;

[0089] 220 mM sucrose;

[0090] The pH value is 5.5±0.5.

[0091] In another aspect, a pharmaceutical preparation is provided herein, comprising

[0092] 50 mg / mL anti-CSF-1R antibody;

[0093] 20 mM L-histidine;

[0094] 0.04% (w / v) polysorbate 20;

[0095] 130 mM sodium chloride;

[0096] The pH value is 5.5±0.5.

[0097] In another aspect, a pharmaceutical preparation is provided herein, comprising

[0098] 50 mg / mL anti-CSF-1R antibody;

[0099] 20 mM L-histidine;

[0100] 0.04% (w / v) polysorbate 20;

[0101] 130 mM sodium chloride;

[0102] 10 mM methionine;

[0103] The pH value is 5.5±0.5.

[0104] In another aspect, a pharmaceutical preparation is provided herein, comprising

[0105] 50 mg / mL anti-CSF-1R antibody;

[0106] 20 mM L-histidine;

[0107] 0.04% (w / v) Poloxamer 188;

[0108] 130 mM sodium chloride;

[0109] The pH value is 5.5±0.5.

[0110] In another aspect, the pharmaceutical preparation according to the present invention comprises

[0111] 50 mg / mL anti-CSF-1R antibody;

[0112] 20 mM L-histidine;

[0113] 0.04% (w / v) polysorbate 20;

[0114] 240 mM trehalose;

[0115] The pH value is 6.0±0.5.

[0116] In another aspect, the pharmaceutical preparation according to the present invention comprises

[0117] 50 mg / mL anti-CSF-1R antibody;

[0118] 20 mM L-histidine;

[0119] 0.04% (w / v) polysorbate 20;

[0120] 240 mM trehalose;

[0121] 10 mM methionine;

[0122] The pH value is 6.0±0.5.

[0123] In another aspect, the pharmaceutical preparation according to the present invention comprises

[0124] 50 mg / mL anti-CSF-1R antibody;

[0125] 20 mM L-histidine;

[0126] 0.04% (w / v) Poloxamer 188;

[0127] 240 mM trehalose;

[0128] The pH value is 6.0±0.5.

[0129] In another aspect, the present invention relates to a pharmaceutical formulation as described above for the treatment of cancer or metastasis. In another aspect, the pharmaceutical formulation as described above is used to treat bone loss. In another aspect, the pharmaceutical formulation as described above is used to treat inflammatory diseases such as inflammatory bowel disease. In a preferred aspect, the pharmaceutical formulation as described above is used to treat pigmented villonodular synovitis (PVNS) or tenosynovial giant cell tumor (TGCT).

[0130] On the other hand, the pharmaceutical formulation as described above is used in combination with another therapeutic agent, particularly another immunotherapy. In a specific aspect, the pharmaceutical formulation is used in combination with an agent that blocks the interaction of PD-L1 / PD-1. On the other hand, the present invention relates to the use of a pharmaceutical formulation as described above for the preparation of a medicament for treating cancer or metastasis. On the other hand, there is provided the use of a pharmaceutical formulation as described above for the preparation of a medicament for treating bone loss. On the other hand, there is provided the use of a pharmaceutical formulation as described above for the preparation of a medicament for treating inflammatory diseases such as inflammatory bowel disease. In a preferred aspect, there is provided the use of a pharmaceutical formulation as described above for the preparation of a medicament for treating pigmented villonodular synovitis (PVNS) or tenosynovial giant cell tumor (TGCT). DETAILED DESCRIPTION

[0131] The present invention relates to a stable pharmaceutical formulation comprising an anti-CSF-1R antibody.

[0132] The term "pharmaceutical formulation" or "pharmaceutical composition" refers to a preparation that is in a form that permits the biological activity of the active ingredient to be clearly effective, and contains no additional components that would be toxic to a subject to which the formulation would be administered.

[0133] The term "liquid" as used herein in connection with the formulations according to the present invention refers to a formulation that is liquid at atmospheric pressure and at a temperature of at least about 2°C to about 8°C.

[0134] The term "lyophilized" as used herein in conjunction with the formulation according to the present invention refers to a formulation produced by a freeze-drying method known per se in the art. The solvent (e.g., water) is removed by freezing, the ice is then sublimed under vacuum, and the remaining water is desorbed at high temperatures. The lyophilisate typically has a residual moisture content of about 0.1% (w / w) to 5% (w / w) and is present in the form of a powder or physically stable cake. The lyophilisate is characterized by rapid dissolution after the addition of a reconstitution medium.

[0135] The term "reconstitution medium" as used herein in conjunction with the formulation according to the present invention refers to a formulation that has been lyophilized and redissolved by adding a reconstitution medium. Suitable reconstitution media include, but are not limited to, water for injection (WFI), bacteriostatic water for injection (BWFI), sodium chloride solution (e.g., 0.9% (w / v) NaCl), glucose solution (e.g., 5% glucose), a surfactant-containing solution (e.g., 0.01% polysorbate 20), a pH buffer solution (e.g., phosphate buffer solution).

[0136] The formulations according to the invention are physiologically well tolerated, may be easily prepared, can be accurately dispensed and are stable with respect to decomposition products and aggregates during storage, during repeated freeze-thaw cycles and under mechanical stress.

[0137] A "stable" formulation is one in which a protein (eg, an antibody) substantially retains its physical and chemical stability, as well as its biological activity, upon storage.

[0138] "Stable liquid pharmaceutical antibody formulations" are liquid antibody formulations that are not observed to change significantly after being stored at refrigerated temperatures (2 to 8 ° C) for at least 12 months (particularly 2 years, more particularly 3 years). The stability criteria are as follows: measured by size exclusion chromatography (SEC-HPLC), no more than 10% (particularly 5%) of the antibody monomers are degraded. In addition, by visual analysis, the solution is colorless or clear to slightly milky white. The protein concentration of the formulation does not vary by more than + / -10%. Aggregates of no more than 10% (particularly 5%) are formed. Stability is measured by known methods in the art such as UV spectroscopy, size exclusion chromatography (SEC-HPLC), ion exchange chromatography (IE-HPLC), turbidimetry, and visual inspection.

[0139] The term "antibody" encompasses various forms of antibody structures, including but not limited to complete antibodies and antibody fragments. The antibodies according to the present invention are particularly human antibodies, humanized antibodies, chimeric antibodies, antibody fragments or further genetically engineered antibodies, as long as they retain the properties according to the present invention. More particularly, the antibodies are humanized monoclonal antibodies, especially recombinant humanized antibodies. In a specific aspect, the humanized antibody is of human IgG1 isotype.

[0140] The term "humanized antibody" refers to an antibody in which the framework or "complementarity determining region" (CDR) has been modified to contain CDRs of an immunoglobulin with different specificity compared to the parent immunoglobulin. In a preferred embodiment, the mouse CDRs are transplanted into the framework region of a human antibody to prepare a "humanized antibody". See, for example, the following documents: Riechmann, L. et al., Nature 332 (1988) 323-327; and Neuberger, MS et al., Nature 314 (1985) 268-270. Particularly preferred CDRs correspond to those sequences representing the antigens that recognize the above-mentioned chimeric antibodies. Other forms of "humanized antibodies" encompassed by the present invention are antibodies in which the constant regions have been additionally modified or altered relative to the original antibodies to produce the properties according to the present invention, particularly with respect to C1q binding and / or Fc receptor (FcR) binding.

[0141] "Anti-CSF-1R antibodies" are antibodies that specifically bind to human CSF-1R. Particularly useful anti-CSF-1R antibodies are, for example, those described in PCT Publication WO 2011 / 070024 A1, which is incorporated herein by reference in its entirety. These antibodies are unique in that they bind to human CSF-1R fragment delD4 (comprising extracellular subdomains D1-D3 and D5, SEQ ID NO: 11) and human CSF-1R extracellular domain (CSF-1R-ECD) (comprising extracellular subdomains D1-D5, SEQ ID NO: 12) at a ratio of 1:50 or less. By virtue of their epitopes located within D4 and D5, they are able to block the receptor dimerization interface.

[0142] "Specific binding" means that the binding is selective for the antigen and can be distinguished from unwanted or non-specific interactions. The ability of an antigen binding portion to bind to a specific antigenic determinant can be measured by enzyme-linked immunosorbent assay (ELISA) or other techniques familiar to those skilled in the art (e.g., surface plasmon resonance (SPR) technology (analyzed on, for example, a BIAcore instrument) (Liljeblad et al., Glyco J 17, 323-329 (2000)) and traditional binding assays (Heeley, Endocr Res 28, 217-229 (2002)). In one embodiment, the degree of binding of the antigen binding portion to an unrelated protein, as measured by SPR, is less than about 10% of the degree of binding of the antigen binding portion to the antigen. In certain embodiments, the dissociation constant (K) of an antigen binding portion or an antibody comprising the antigen binding portion that binds to an antigen is measured. D ) is ≤1 μM, ≤100 nM, ≤10 nM, ≤1 nM, ≤0.1 nM, ≤0.01 nM or ≤0.001 nM (e.g., 10 -8 M or lower, e.g. 10 -8 M to 10 -13 M, for example 10 -9 M to 10 -13 M).

[0143] In a specific aspect, the anti-CSF-1R antibody comprises: a heavy chain variable region comprising a heavy chain CDR1 (CDR-H1) of SEQ ID NO: 1, CDR-H2 of SEQ ID NO: 2, and CDR-H3 of SEQ ID NO: 3; and a light chain variable region comprising a light chain CDR1 (CDR-L1) of SEQ ID NO: 4, CDR-L2 of SEQ ID NO: 5, and CDR-L3 of SEQ ID NO: 6. More specifically, the anti-CSF-1R antibody comprises: a heavy chain variable region comprising an amino acid sequence of SEQ ID NO: 7; and a light chain variable region comprising an amino acid sequence of SEQ ID NO: 8. In a particularly preferred aspect, the anti-CSF-1R antibody is a human IgG1 antibody and comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence of SEQ ID NO: 9, and the light chain comprises an amino acid sequence of SEQ ID NO: 10. The antibody is called Emactuzumab or RG7155.

[0144] The term "variable region" or "variable domain" refers to the domain of an antibody heavy or light chain that is involved in binding the antibody to an antigen. The variable domains of the heavy and light chains of natural antibodies (VH and VL, respectively) generally have similar structures, with each domain comprising four conserved framework regions (FRs) and three complementarity determining regions (CDRs). A single VH or VL domain may be sufficient to confer antigen binding specificity. As used herein in conjunction with variable region sequences, "Kabat numbering" refers to the numbering system proposed by Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health, Bethesda, MD (1991).

[0145] As used herein, the term "hypervariable region" or "HVR" refers to each region of an antibody variable domain that is highly variable in sequence and determines antigen binding specificity. HVRs contain amino acid residues from "complementarity determining regions" (CDRs). Typically, an antibody contains six CDRs; three in VH (CDR-H1, CDR-H2, and CDR-H3), and three in VL (CDR-L1, CDR-L2, and CDR-L3). Exemplary CDRs herein include:

[0146] (a) At amino acid residues 26-32 (L1), 50-52 (L2), 91-96 (L3), 26-32 (H1),

[0147] The hypervariable loops occurring at 53-55 (H2) and 96-101 (H3) (Chothia and Lesk, J.

[0148] Mol.Biol.196:901-917(1987));

[0149] (b) present at amino acid residues 24-34 (L1), 50-56 (L2), 89-97 (L3), 31-35b

[0150] (H1), 50-65 (H2), and 95-102 (H3) CDRs (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Edition, Public Health Service, National Institutes of Health, Bethesda, MD

[0151] (1991)); and

[0152] (c) present at amino acid residues 27c-36 (L1), 46-55 (L2), 89-96 (L3), 30-35b

[0153] Antigenic contact points at (H1), 47-58 (H2), and 93-101 (H3) (MacCallum et al., J. Mol. Biol. 262:732-745 (1996)).

[0154] Unless otherwise indicated, CDRs are determined according to the method described in Kabat et al. (supra). Those skilled in the art will appreciate that CDR names may also be determined according to the method described in Chothia (supra), McCallum (supra), or any other scientifically accepted nomenclature system.

[0155] "Framework" or "FR" refers to the variable domain residues excluding the complementarity determining regions (CDRs). The FR of a variable domain is usually composed of the following four FR domains: FR1, FR2, FR3, and FR4. Therefore, CDR and FR sequences usually appear in the following sequence in VH (or VL): FR1-CDR-H1 (L1)-FR2-CDR-H2 (L2)-FR3-CDR-H3 (L3)-FR4.

[0156] The term "epitope" includes any polypeptide determinant that is capable of specific binding to an antibody. In certain embodiments, epitope determinants include chemically active surface groups of molecules such as amino acids, sugar side chains, phosphoryl or sulfonyl groups, and in certain embodiments, may have specific three-dimensional structural characteristics and / or specific charge characteristics. An epitope is a region of an antigen that binds to an antibody.

[0157] "Constant region" or "constant domain" does not directly participate in the binding of antibodies to antigens, but exhibits a variety of effector functions. According to the different amino acid sequences of the constant regions of their heavy chains, antibodies or immunoglobulins can be divided into the following categories: IgA, IgD, IgE, IgG and IgM, and some of them can be further divided into subclasses (isotypes), such as IgG1, IgG2, IgG3 and IgG4, IgA1 and IgA2. The antibody used in the present invention is particularly an IgG type antibody, more particularly an IgG1 or IgG4 human subtype antibody.

[0158] The term "Fc region" herein is used to define the C-terminal region of an immunoglobulin heavy chain, which contains at least a portion of a constant region. The term includes native sequence Fc regions and variant Fc regions. In one aspect, the human IgG heavy chain Fc region extends from Cys226 or from Pro230 to the carboxyl terminus of the heavy chain. However, the antibody produced by the host cell can undergo post-translational cleavage of one or more, particularly one or two amino acids from the C-terminus of the heavy chain. Therefore, the antibody produced by the host cell by expressing a specific nucleic acid molecule encoding a full-length heavy chain can include a full-length heavy chain, or the antibody can include a cleavage variant of the full-length heavy chain. This may be a situation where the last two C-terminal amino acids of the heavy chain are glycine (G446) and lysine (K447, according to the Kabat EU index). Therefore, the C-terminal lysine (Lys447) or C-terminal glycine (Gly446) and lysine (Lys447) in the Fc region may be present or may not be present. If not otherwise specified, the amino acid sequence of the heavy chain comprising the Fc region is represented herein as having no C-terminal glycine-lysine dipeptide. In one aspect, the heavy chain comprising the Fc region as specified herein is included in an antibody according to the present invention, and the heavy chain comprises additional C-terminal glycine-lysine dipeptide (G446 and K447, numbered according to the EU index of Kabat). In one aspect, the heavy chain comprising the Fc region as specified herein is included in an antibody according to the present invention, and the heavy chain comprises additional C-terminal glycine residues (G446, numbered according to the EU index of Kabat). Unless otherwise specified herein, the numbering of amino acid residues in the Fc region or constant region is according to the EU numbering system, which is also referred to as the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th edition, Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0159] The concentration of the anti-CSF-1R antibody contained in the pharmaceutical formulation is in the range of 40 to 200 mg / mL, particularly in the range of 40 to 100 mg / mL, more particularly in the range of 40 to 60 mg / mL, and most particularly 50 mg / mL.

[0160] The term "surfactant" as used herein means a pharmaceutical surface active agent. Preferably, a nonionic surfactant is used. Examples of pharmaceutical surfactants include, but are not limited to, polyoxyethylene sorbitan fatty acid esters (Tween), polyoxyethylene alkyl ethers (Brij), alkylphenyl polyoxyethylene ethers (Triton X), polyoxyethylene-polyoxypropylene copolymers (Poloxamers, Pluronic) and sodium dodecyl sulfate (SDS). Preferred polyoxyethylene sorbitan fatty acid esters are polysorbate 20 (polyoxyethylene sorbitan monolaurate, available under the trade name Tween 20 TM Sold under the trade name Tween 80 TM Preferred polyoxyethylene-polyoxypropylene copolymers are sold under the trade name F68 or Poloxamer 188 TM Preferred polyoxyethylene alkyl ethers are sold under the trade name Brij TM The preferred alkylphenyl polyoxyethylene ethers are sold under the trade name Triton X, and the most preferred is tert-octylphenoxypolyethoxyethanol (sold under the trade name Triton X-100 TM The preferred surfactant used in the present invention is polyoxyethylene sorbitan fatty acid ester, preferably polysorbate 20 or polysorbate 80, most preferably polysorbate 20. Another preferred surfactant is poloxamer 188 TM .

[0161] The term "buffer" as used herein means a pharmaceutical excipient that is used to stabilize the pH of a pharmaceutical formulation. Suitable buffers are well known in the art and can be found in the literature. Preferred pharmaceutical buffers include, but are not limited to, histidine buffer, citrate buffer, succinate buffer, acetate buffer, arginine buffer, phosphate buffer, or mixtures thereof. Therefore, the buffer is a histidine salt, citrate, succinate, acetate, malate, phosphate, and lactate. Buffers of particular interest include L-histidine or a mixture of L-histidine and L-histidine hydrochloride or L-histidine acetate, the pH of which is adjusted by an acid or base as known in the art. The above-mentioned buffer is usually used in an amount of about 5mM to about 100mM, particularly 10mM to about 30mM and more particularly about 20mM. Independently of the buffer used, the pH can be adjusted to a value in the range of 4.5 to 7.0, in particular to a value in the range of 5.0 to 6.0, and most particularly to pH 6.0 ± 0.03 with acids or bases known in the art, such as hydrochloric, acetic, phosphoric, sulfuric and citric acids, sodium hydroxide and potassium hydroxide.

[0162] The term "stabilizer" refers to a pharmaceutical excipient that is used to protect active pharmaceutical ingredients and / or preparations from chemical and / or physical degradation during production, storage and application. Stabilizers include, but are not limited to, sugars, amino acids, polyols (e.g., mannitol, sorbitol, xylitol, dextran, glycerol, arabitol, propylene glycol, polyethylene glycol), cyclodextrins (e.g., hydroxypropyl-β-cyclodextrin, sulfobutylethyl-β-cyclodextrin, β-cyclodextrin), polyethylene glycols (e.g., PEG 3000, PEG 3350, PEG 4000, PEG 6000), albumins (human serum albumin (HSA), bovine serum albumin (BSA)), salts (e.g., sodium chloride (saline), magnesium chloride, calcium chloride), chelating agents (e.g., EDTA) as defined below. The stabilizer used in particular in the present invention is selected from the group consisting of sugars, polyols, and amino acids. The stabilizer may be present in the formulation in an amount of about 10 mM to about 500 mM, particularly in an amount of about 140 to about 250 mM, and more particularly in an amount of about 210 mM to about 240 mM. More particularly, sucrose or trehalose is used as a stabilizer in an amount of about 220 mM to about 240 mM.

[0163] The term "sugar" as used herein includes monosaccharides and oligosaccharides. Monosaccharides are monomeric carbohydrates that cannot be hydrolyzed by acid, including simple sugars and their derivatives (such as amino sugars). Sugars are usually used in their D configuration. Examples of monosaccharides include glucose, fructose, galactose, mannose, sorbose, ribose, deoxyribose, neuraminic acid. Oligosaccharides are carbohydrates composed of more than one monomeric sugar unit, and these monomeric sugar units are connected by branched or linear glycosidic bonds. The monomeric sugar units in oligosaccharides can be the same or different. Depending on the number of monomeric sugar units, oligosaccharides are disaccharides, trisaccharides, tetrasaccharides, pentasaccharides, etc. Compared with polysaccharides, monosaccharides and oligosaccharides are soluble in water. Examples of oligosaccharides include sucrose, trehalose, lactose, maltose and raffinose. The preferred sugar used in the present invention is sucrose and trehalose (i.e., α, α-D-trehalose), most preferably sucrose. Available trehalose is trehalose dihydrate. The sugar may be present in the formulation in an amount of about 10 mM to about 500 mM, preferably in an amount of about 200 mM to about 300 mM, more preferably in an amount of about 220 mM to about 250 mM, especially in an amount of about 220 mM or about 240 mM, most preferably in an amount of about 220 mM.

[0164] As used herein, the term "amino acid" refers to a pharmaceutical organic molecule having an amino moiety located at the α-position of a carboxyl group. Examples of amino acids include, but are not limited to, arginine, glycine, ornithine, lysine, histidine, glutamic acid, aspartic acid, isoleucine, leucine, alanine, phenylalanine, tyrosine, tryptophan, methionine, serine, proline. The amino acid employed in each case is preferably an L-type amino acid. Basic amino acids such as arginine, histidine or lysine are preferably used in the form of their inorganic salts (advantageously in the form of hydrochloride, i.e. as amino acid hydrochloride). The preferred amino acid used in the present invention is methionine. The concentration of methionine used is preferably from about 5mM to about 25mM, and most preferably about 10mM.

[0165] A subgroup in stabilizers is lyoprotectants. The term "lyoprotectant" means a pharmaceutical excipient that is used to protect unstable active ingredients (e.g., proteins) from unstable conditions during freeze drying and subsequent storage and reconstitution. Lyoprotectants include, but are not limited to, groups consisting of sugars, polyols (such as sugar alcohols) and amino acids. Preferred lyoprotectants may be selected from the group consisting of: sugars, such as sucrose, trehalose, lactose, glucose, mannose, maltose, galactose, fructose, sorbose, raffinose, neuraminic acid; amino sugars, such as glucosamine, galactosamine, N-methylglucosamine ("methylglucamine"); polyols, such as mannitol and sorbitol; and amino acids, such as arginine and glycine; or mixtures thereof. The lyoprotectant is generally present in an amount of about 10 mM to 500 mM, preferably in an amount of about 10 mM to about 300 mM, and more preferably in an amount of about 100 mM to about 300 mM.

[0166] Another subgroup of stabilizers is antioxidants. The term "antioxidant" refers to a pharmaceutical excipient that is used to prevent oxidation of the active pharmaceutical ingredient. Antioxidants include, but are not limited to, ascorbic acid, glutathione, cysteine, methionine, citric acid, EDTA. Antioxidants can be used in an amount of 0.01 mM to about 100 mM, preferably in an amount of about 5 mM to about 50 mM, and more preferably in an amount of about 5 mM to about 25 mM.

[0167] The preparation according to the present invention may also include one or more tonicity agents. The term "tonicity agent" refers to a pharmaceutical excipient for adjusting the tonicity of the preparation. The preparation may be a hypotonic, isotonic or hypertonic preparation. Isotonicity is generally related to the osmotic pressure of the solution, which is generally relative to the osmotic pressure of human serum (about 250-350mOsmol / kg). The preparation according to the present invention may be a hypotonic, isotonic or hypertonic preparation, but is preferably an isotonic preparation. An isotonic preparation is a liquid or a liquid reconstituted from a solid form (e.g., from a lyophilized form), and represents a solution having the same tonicity as a certain other solution such as a physiological saline solution and serum compared thereto. Suitable tonicity agents include, but are not limited to, sodium chloride, potassium chloride, glycerol, and any component (particularly glucose) selected from the group of amino acids or sugars. Tonicity agents are generally used in an amount of about 5mM to about 500mM. In stabilizers and tonicity agents, a group of compounds can play two roles simultaneously, i.e., they are used as stabilizers and tonicity agents simultaneously. Examples of such compounds may be found in the group consisting of sugars, amino acids, polyols, cyclodextrins, polyethylene glycols and salts. An example of a sugar that can be used as both a stabilizer and a tonicity agent is trehalose.

[0168] As used herein, the term "polyol" refers to a pharmaceutical alcohol having more than one hydroxyl group. Suitable polyols include, but are not limited to, mannitol, sorbitol, glycerol (glycerol), dextran, arabitol, propylene glycol, polyethylene glycol and combinations thereof. Polyols can be used in an amount of 10 mM to about 500 mM, particularly in an amount of about 10 mM to about 250 mM, and more particularly in an amount of about 200 mM to about 250 mM.

[0169] The preparation may also include adjuvants, such as preservatives, wetting agents, emulsifiers and dispersants. Can be sterilized and added with various antibacterial and antifungal agents (e.g. parabens, chlorobutanol, phenol, sorbic acid, etc.) to ensure that there is no microorganism. Preservatives are usually used in an amount of about 0.001% (w / v) to about 2% (w / v). Preservatives include, but are not limited to, ethanol, benzyl alcohol, phenol, metacresol, chlorometacresol, methylparaben or propylparaben, benzalkonium chloride.

[0170] Pharmaceutical preparations may also include adjuvants, such as preservatives, wetting agents, emulsifiers and dispersants. Can be sterilized and by adding various antibacterial and antifungal agents (e.g. parabens, chlorobutanol, phenol, sorbic acid, etc.) to ensure that there is no microorganism. Preservatives are usually used in an amount of about 0.001% (w / v) to about 2% (w / v). Preservatives include but are not limited to ethanol, benzyl alcohol, phenol, metacresol, chlorometacresol, methylparaben or propylparaben, benzalkonium chloride.

[0171] Preparations of the present invention can be used by a variety of methods known in the art. As will be appreciated by those skilled in the art, routes of administration and / or modes will vary according to desired results. For using preparations of the present invention by certain routes of administration, it may be necessary to dilute the preparation with a diluent. Pharmaceutical diluents include saline, glucose, Ringer's solution, and aqueous buffered solutions.

[0172] Preferably, the preparation according to the present invention is administered intravenously (IV), subcutaneously (SC) or any other parenteral administration mode (such as those known in the pharmaceutical field). In a preferred aspect, the pharmaceutical preparation is administered by IV infusion. When administered by intravenous injection, it can be administered as a push or continuous infusion. For example, the pharmaceutical preparation of the present invention can be diluted with a sterile saline solution and administered with an infusion pump commonly used in clinical settings.

[0173] The phrases "parenteral administration" and "administered parenterally" as used herein mean modes of administration other than enteral and topical administration, which are usually administered by injection, and include, but are not limited to, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, intratracheal, subcutaneous, subcutaneous, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injection and infusion.

[0174] The pharmaceutical formulation according to the present invention is suitable for administration to the patient at one time or over a series of treatments and may be administered to the patient at any time from diagnosis; it may be administered as a sole therapy or in combination with other drugs or therapies for the treatment of the conditions mentioned above.

[0175] The anti-CSF-1R antibody may be the only active ingredient in the liquid pharmaceutical composition. Alternatively, the anti-CSF-1R antibody may be administered in combination with one or more other therapeutic active ingredients, for example, simultaneously, sequentially or separately. As used herein, the term "active ingredient" refers to an ingredient that has a pharmacological effect, such as a therapeutic effect, at a relevant dose. Therefore, the anti-CSF-1R antibody in the liquid pharmaceutical composition may be accompanied by other active ingredients, including other antibody components, such as CD40 antibodies, VEGF antibodies, or agents that block PD-L1 / PD-1 interactions. In particular, the agent that blocks PD-L1 / PD-1 interactions is an anti-PD-L1 antibody or an anti-PD1 antibody. More particularly, the agent that blocks PD-L1 / PD-1 interactions is selected from the group consisting of atezolizumab, durvalumab, pembrolizumab and nivolumab. In a specific aspect, the agent that blocks PD-L1 / PD-1 interactions is atezolizumab. In a specific aspect, the CD40 antibody is selicrelumab. In another preferred aspect, the VEGF antibody is bevacizumab (Avastin).

[0176] Pharmaceutical compositions are suitable for comprising therapeutically effective amounts of antibodies. As used herein, the term "therapeutically effective amount" refers to the amount of therapeutic agent required to treat, improve or prevent a target disease or condition or to exhibit a detectable therapeutic, pharmacological or preventive effect. For any antibody, the therapeutically effective amount can initially be estimated by cell culture assays or animal models (usually in rodents, rabbits, dogs, pigs or primates). Animal models can also be used to determine appropriate concentration ranges and routes of administration. This information can then be used to determine useful doses and routes of administration in humans.

[0177] For human subjects, the exact therapeutically effective amount will depend on the severity of the disease state, the general health of the subject, the age, weight and sex of the subject, diet, time and frequency of administration, drug combination, reaction sensitivity and tolerance / response to therapy. The effective amount can be determined by routine experimentation and is at the discretion of the clinician. In a specific aspect, the anti-CSF-1R antibody is administered at a (fixed) dose of 600-1200 mg, particularly at a dose of 750-1100 mg, more particularly at a dose of 750-1000 mg, and even more particularly at a dose of 900-1000 mg. In a preferred aspect, the dose is 1000 mg.

[0178] In a preferred aspect of the invention, the pharmaceutical preparation is used in a treatment cycle. In particular, the length of the treatment cycle is between 2 weeks and 4 weeks, preferably between 18 days and 24 days. More preferably, the length of the treatment cycle is (approximately) 3 weeks.

[0179] The pharmaceutical preparation can be conveniently provided in a unit dosage form containing a predetermined amount of an anti-CSF-1R antibody. In a specific aspect, the pharmaceutical preparation can be provided in a vial and stored at 2°C to 8°C. In a preferred aspect, the pharmaceutical preparation will be provided in a vial with a size of 20 mL. In another preferred aspect, the pharmaceutical preparation will be provided in a vial with a size of 50 mL.

[0180] Stable formulations to be used for in vivo administration must be sterile. This is easily achieved by filtering with a sterile filtration membrane. The formulation must have a certain degree of fluidity so that it can be delivered by a syringe or an infusion system. In addition to water, the carrier can also be an isotonic buffered saline solution, ethanol, a polyol (such as glycerol, propylene glycol and liquid polyethylene glycol, etc.) and a suitable mixture thereof. In view of their high stability, the pharmaceutical formulation according to the present invention can be administered by iv without the need for an inline filter, and is therefore much more convenient to handle than conventional formulations that need to be administered by an inline filter. An inline filter such as a 2-way filter must be installed in the infusion line of the iv drug. To prevent any particles, air or microorganisms from being present in the iv infusion solution or tubing. Particles of 5 to 20 microns or larger have the ability to block blood flow through the lung capillaries, which can lead to complications such as pulmonary embolism. Foreign particles can also cause phlebitis at the injection site, and filters can help reduce the incidence of phlebitis.

[0181] The stable pharmaceutical formulation according to the present invention can be prepared by methods known in the art (e.g., ultrafiltration-diafiltration, dialysis, addition and mixing, lyophilization, reconstitution, and combinations thereof). Examples of the preparation of the formulation according to the present invention can be found below.

[0182] Thus, the present invention includes a method for preparing a formulation according to the present invention. The method comprises buffer exchanging the antibody with a diafiltration buffer comprising a desired buffer composition and, if desired, concentrating the antibody by diafiltration, then adding an excipient (e.g., sucrose, sodium chloride, methionine) as a stock solution to the antibody solution, then adding a surfactant as a stock solution to the antibody / excipient solution, and finally adjusting the antibody concentration to the desired final concentration using a buffer solution to achieve the final excipient and surfactant concentrations.

[0183] Alternatively, excipients can also be added as solids to the starting solution containing the antibody. If the antibody is in solid form, such as a lyophilisate, the preparation according to the present invention can be prepared by the following method: first, the antibody is dissolved in water or a buffer solution (optionally containing one or more of the excipients), and then other excipients are added as a stock solution or solid. The antibody can also be advantageously directly dissolved in a solution containing all other excipients. One or more of the excipients present in the preparation according to the present invention can be added during or at the end of the preparation of the antibody, for example, by the following means: in the final purification step implemented after the preparation of the antibody, the antibody is directly dissolved in one, more than one or preferably all of the excipients containing the preparation. If the solution containing the antibody and the excipient has not yet reached the desired pH, it is adjusted by adding an acid or a base, preferably using an acid or a base already present in the buffer system to adjust. Then sterile filtration is performed.

[0184] The stable liquid pharmaceutical preparation according to the present invention can also be provided as a lyophilized form or a liquid form reconstructed by a lyophilized form." lyophilized form "is made by a lyophilization method as known in the art. Lyophilized material generally has a residual moisture content of about 0.1% (w / w) to 5% (w / w), and exists in the form of a powder or physically stable cake. "Reconstructed form" can be obtained by rapidly dissolving the lyophilized material after adding a reconstructed medium. Suitable reconstructed media include, but are not limited to, water for injection (WFI), bacteriostatic water for injection (BWFI), sodium chloride solution (e.g., 0.9% (w / v) NaCl), glucose solution (e.g., 5% (w / v) glucose), a solution containing a surfactant (e.g., 0.01% (w / v) polysorbate 20) and a pH buffer solution (e.g., phosphate buffered solution).

[0185] The present invention further includes the use of the preparation according to the present invention for treating a disease, or the use of the preparation according to the present invention for preparing a medicament for treating a disease (especially treating cancer), and for treating pigmented villonodular synovitis (PVNS) or tenosynovial giant cell tumor (TGCT).

[0186] The term "cancer" as used herein can be, for example, lung cancer, non-small cell lung (NSCL) cancer, bronchoalveolar cell lung cancer, bone cancer, pancreatic cancer, skin cancer, head or neck cancer, cutaneous or intraocular melanoma, uterine cancer, ovarian cancer, rectal cancer, cancer of the anal region, stomach cancer, gastric cancer, In one preferred aspect, the cancer is breast cancer, colorectal cancer, melanoma, head and neck cancer, lung cancer, prostate cancer, bladder cancer, kidney cancer or ureteral cancer, renal cell carcinoma, renal pelvic cancer, mesothelioma, hepatocellular carcinoma, bile duct cancer, central nervous system (CNS) tumors, spinal axis tumors, brain stem gliomas, glioblastoma multiforme, astrocytoma, neurilemmoma, ependymoma, medulloblastoma, meningioma, squamous cell carcinoma, pituitary adenoma, lymphoma, lymphocytic leukemia, including refractory forms of any of the above cancers, or a combination of one or more of the above cancers. In a preferred aspect, the cancer is breast cancer, colorectal cancer, melanoma, head and neck cancer, lung cancer, or prostate cancer. In another preferred aspect, such cancers are breast cancer, lung cancer, colon cancer, ovarian cancer, melanoma, bladder cancer, renal cancer, kidney cancer, liver cancer, head and neck cancer, colorectal cancer, pancreatic cancer, gastric cancer, esophageal cancer, mesothelioma, prostate cancer, leukemia, lymphoma, myeloma. In a preferred aspect, such cancers are further characterized by CSF-1 or CSF-IR expression or overexpression. On the other hand, the pharmaceutical preparations of the present invention are used to treat primary tumors and new metastases simultaneously. On the other hand, the pharmaceutical preparations of the present invention are used to treat periodontitis, histiocytosis X, osteoporosis, Paget's disease of bone (PDB), bone loss caused by cancer treatment, periprosthetic osteolysis, glucocorticoid-induced osteoporosis, rheumatoid arthritis, psoriatic arthritis, osteoarthritis, inflammatory arthritis and inflammation. In another preferred aspect, the pharmaceutical preparations are used for melanoma, bladder cancer (UCB) or lung cancer (e.g., non-small cell lung (NSCL) cancer). In another preferred aspect, the pharmaceutical preparation is used for renal cell carcinoma (RCC) or head and neck squamous cell carcinoma (HNSCC). In a particularly preferred aspect, the pharmaceutical preparation of the present invention is used to treat pigmented villonodular synovitis (PVNS) or tenosynovial giant cell tumor (TGCT).

[0187] Sequence Listing:

[0188]

[0189]

[0190] Examples

[0191] The development of a liquid pharmaceutical formulation for intravenous (iv) administration according to the present invention is as follows.

[0192] Example 1: Preparation of liquid formulations for preliminary formulation screening

[0193] The huMAb CSF-1R liquid formulations F1 to F16 listed in Table 1 were prepared at a protein concentration of 50 mg / mL.

[0194] The anti-CSF-1R antibody (emactuzumab) is produced by techniques generally known in the production of recombinant proteins and the methods described in WO 2011 / 070024. To prepare the pharmaceutical formulation according to the example, the antibody is provided at a concentration of about 20 mg / mL in a 20 mM histidine buffer (L-histidine / HCl buffer) at a pH of about 5.5. The CSF-1R antibody used in the example is a humanized antibody comprising a heavy chain and a light chain, wherein the heavy chain comprises the amino acid sequence shown in SEQ ID NO: 9, and the light chain comprises the amino acid sequence shown in SEQ ID NO: 10.

[0195] The excipients of the formulation according to the present invention are widely used in practice and are known to those skilled in the art. Therefore, there is no need to explain these excipients in detail herein.

[0196] To prepare a liquid formulation, the anti-CSF-1R antibody is buffer exchanged with a diafiltration buffer containing a desired buffer composition, and the antibody concentration is concentrated to about 80 mg / mL by ultrafiltration. After the ultrafiltration operation is completed, an excipient (e.g., sucrose, methionine) as a stock solution is added to the antibody solution. Then, a surfactant is added as a 125-fold stock solution. Finally, the protein concentration is adjusted with a buffer so that the final concentration of the anti-CSF-1R antibody reaches about 50 mg / mL.

[0197] All formulations were sterile filtered through a 0.22 μm low protein binding filter and aseptically filled into 6 mL sterile glass bottles, which were sealed with rubber stoppers and aluminum crimp caps with ETFE (copolymer of ethylene and tetrafluoroethylene) coatings. The filling volume was approximately 2.4 mL. These formulations were stored for different time intervals under different ICH climate conditions (5°C, 25°C, and 40°C) and shaken (200 min at 5°C and 25°C). -1 The cells were stressed using the shaking method (shaking at a shaking frequency of 1 week) and the freeze-thaw stress method (freeze-thaw cycles five times at -80°C / +5°C).

[0198] Table 1 - Overview of different formulations

[0199]

[0200]

[0201] Before and after the stress test, the samples were analyzed using the following analytical methods

[0202] UV spectrophotometry

[0203] Size Exclusion Chromatography (SEC)

[0204] Ion Exchange Chromatography (IEC)

[0205] Solution clarity and turbidity

[0206] Analytical Protein A Chromatography

[0207] Visual inspection

[0208] UV spectrometry for protein content determination was performed using a Perkin Elmer λ35 UV spectrophotometer with a wavelength range of 240 nm to 400 nm. Pure protein samples were diluted to approximately 0.5 mg / mL with the corresponding formulation buffer. Protein concentration was calculated according to Formula 1.

[0209] Formula 1: Protein content

[0210] The UV absorption at 280 nm is corrected for light scattering at 320 nm and multiplied by a dilution factor determined from the weight and density of the neat sample and dilution buffer. The molecule is divided by the product of the optical path length d of the cuvette and the extinction coefficient ε.

[0211] Size exclusion chromatography (SEC) was used to detect soluble high molecular weight substances (aggregates) and low molecular weight (LMW) hydrolysis products in the formulation. The method used a Waters Alliance 2695HPLC instrument equipped with a Waters W2487 dual absorbance detector and a Waters BioSuite 250 column. 0.2M K2HPO4 / 0.25M KCl (pH 7.0) was used as the mobile phase, and intact monomers, aggregates, and hydrolysis products were separated by an isocratic elution curve and detected at a wavelength of 280nm.

[0212] Ion exchange chromatography (IEC) is used to detect chemical degradation products that change the net charge of the antibody in the formulation. The method uses a Waters Alliance 2695HPLC instrument equipped with a Waters W2487 dual absorbance detector (detection wavelength 280nm) and a MabPac SCX-10 chromatographic column (4mm×150mm). 5mM MES and 15mM NaCl (pH 6.5); 5mMBICINE and 30mM NaCl (pH 8.5) and 5mM BICINE and 1M NaCl (pH 8.5) were used as mobile phases A, B and C, respectively, with a flow rate of 0.5mL / min.

[0213] Gradient Program:

[0214] time ％A ％B ％C 0.0 100 0 0 5.0 100 0 0 50.0 0 100 0 54.0 0 100 0 55.0 100 0 0 65.0 100 0 0 80.0 0 0 100 85.0 0 0 100 86.0 100 0 0 96.0 100 0 0

[0215] Clarity and opacity were measured by nephelometric method in Formazine Turbidity Units (FTU).The neat sample was transferred into a clear glass tube of 11 mm diameter and placed into a HACH 2100AN turbidimeter.

[0216] Analytical protein A chromatography was used to monitor the oxidation state of the four conserved methionine side chains of the Fc portion of the anti-CSF-1R antibody. The method used a Waters Alliance 2695 HPLC instrument equipped with a Waters W2487 dual absorbance detector (detection wavelength 280 nm) and a Poros A720 4.6 mm × 50 mm chromatographic column (from Applied Biosystems, USA). PBS (from Gibco, Invitrogen) and 0.1 M acetic acid, 0.15 M sodium chloride (pH 2.8) were used as mobile phases A and B, respectively, with a flow rate of 2.0 mL / min:

[0217] Gradient Program:

[0218]

[0219]

[0220] The samples were checked for the presence of visible particles using the Simplex ampoule testing device OPTIMA I.

[0221] The stability test results of formulations F1 to F16 are listed in the table below. It was determined that formulation F2 was most conducive to obtaining the highest antibody stability and an antibody formulation free of particles.

[0222]

[0223]

[0224]

[0225]

[0226]

[0227]

[0228]

[0229]

[0230]

[0231]

[0232]

[0233]

[0234]

[0235]

[0236]

[0237]

[0238]

[0239]

[0240]

[0241]

[0242] The present disclosure relates to the following embodiments:

[0243] 1. A pharmaceutical preparation comprising:

[0244] 40 mg / mL to 200 mg / mL of anti-CSF-1R antibody;

[0245] 0.01% (w / v) to 0.1% (w / v) surfactant;

[0246] 5mM to 100mM buffer;

[0247] 10 mM to 500 mM of at least one stabilizer;

[0248] The pH value is in the range of 4.5 to 7.0.

[0249] 2. The pharmaceutical preparation according to item 1, wherein the anti-CSF-1R antibody comprises: a heavy chain variable region, wherein the heavy chain variable region comprises a heavy chain CDR1 (CDR-

[0250] H1), CDR-H2 of SEQ ID NO: 2, and CDR-H3 of SEQ ID NO: 3; and

[0251] A light chain variable region comprising a light chain CDR1 of SEQ ID NO: 4

[0252] (CDR-L1), CDR-L2 of SEQ ID NO:5 and CDR-L3 of SEQ ID NO:6.

[0253] 3. The pharmaceutical preparation according to item 1 or 2, wherein the anti-CSF-1R antibody comprises:

[0254] a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:7; and a light chain variable region comprising the amino acid sequence of SEQ ID NO:8.

[0255] 4. A pharmaceutical preparation according to any one of items 1 to 3, wherein the anti-CSF-1R antibody

[0256] The concentration of the agonist is in the range of 40 mg / mL to 100 mg / mL, particularly 50 mg / mL.

[0257] 5. The pharmaceutical formulation according to any one of items 1 to 4, wherein the surfactant is polysorbate.

[0258] 6. Pharmaceutical formulation according to any one of items 1 to 5, wherein the polysorbate is present in a concentration ranging from 0.01% (w / v) to 0.1% (w / v), in particular 0.04% (w / v).

[0259] 7. The pharmaceutical formulation according to any one of items 1 to 6, wherein the buffer is a histidine buffer, in particular a histidine chloride buffer.

[0260] 8. A pharmaceutical formulation according to any one of items 1 to 7, wherein the concentration of the buffer is

[0261] In the range of 10 mM to 30 mM, in particular 20 mM.

[0262] 9. A pharmaceutical formulation according to any one of items 1 to 8, wherein the pH of the formulation is 5.0

[0263] to 6.5, particularly 6.0.

[0264] 10. The pharmaceutical formulation according to any one of items 1 to 9, wherein the at least one stabilizer is selected from the group consisting of salts, sugars and amino acids.

[0265] 11. Pharmaceutical formulation according to any one of items 1 to 10, wherein the at least one stabilizer is a sugar, in particular sucrose.

[0266] 12. Pharmaceutical formulation according to any one of items 1 to 11, wherein the at least one stabilizer is present in a concentration in the range of 140 mM to 250 mM, in particular in the range of 210 mM to 230 mM.

[0267] 13. The pharmaceutical preparation according to item 10, comprising a first stabilizer selected from the group consisting of salts, sugars and amino acids, and methionine as a second stabilizer.

[0268] 14. The pharmaceutical formulation according to claim 13, wherein the first stabilizer is present in an amount of 120 mM to 300 mM.

[0269] mM, and the second stabilizer methionine is present at a concentration of 5 mM to 25 mM.

[0270] 15. The pharmaceutical formulation according to any one of items 1 to 14, wherein the anti-CSF-1R antibody binds to the human CSF-1R fragment delD4 (SEQ ID NO: 11) and to the human CSF-1R extracellular domain (SEQ ID NO: 12) at a ratio of 1:50 or less.

[0271] 16. A pharmaceutical preparation according to any one of items 1 to 12, comprising

[0272] 40 mg / mL to 100 mg / mL of anti-CSF-1R antibody;

[0273] 20 mM L-histidine;

[0274] 0.03% (w / v) to 0.05% (w / v) polysorbate 20;

[0275] 210mM ​​to 230mM sucrose;

[0276] Optionally 5 mM to 25 mM methionine;

[0277] The pH value is 6.0±0.5.

[0278] 17. A pharmaceutical preparation according to any one of items 1 to 16, comprising

[0279] 50 mg / mL anti-CSF-1R antibody;

[0280] 20 mM L-histidine;

[0281] 0.04% (w / v) polysorbate 20;

[0282] 220 mM sucrose;

[0283] 10 mM methionine;

[0284] The pH value is 6.0±0.5.

[0285] 18. The pharmaceutical formulation according to any one of items 1 to 17, which is in liquid form, lyophilized form or liquid form reconstituted from a lyophilized form.

[0286] 19. A pharmaceutical formulation according to any one of items 1 to 18 for use in the treatment of cancer.

[0287] 20. A pharmaceutical formulation according to any one of items 1 to 19, for use in combination with another therapeutic agent, in particular an agent that blocks the PD-L1 / PD-1 interaction.

[0288] 21. The pharmaceutical formulation according to any one of items 1 to 18, for use in the treatment of pigmented villonodular synovitis (PVNS) or tenosynovial giant cell tumor (TGCT).

[0289] 22. Use of a formulation according to any one of items 1 to 18 for the preparation of a medicament useful for treating cancer or treating pigmented villonodular synovitis (PVNS) or tenosynovial giant cell tumor (TGCT).

Claims

1. A pharmaceutical preparation comprising: 40 mg / mL to 200 mg / mL of anti-CSF-1R antibody; 0.01% (w / v) to 0.1% (w / v) surfactant; 5mM to 100mM buffer; 10 mM to 500 mM of at least one stabilizer; The pH value is in the range of 4.5 to 7.

0.

2. The pharmaceutical preparation according to claim 1, wherein the anti-CSF-1R antibody comprises: a heavy chain variable region comprising a heavy chain CDR1 (CDR-H1) of SEQ ID NO: 1, CDR-H2 of SEQ ID NO: 2, and CDR-H3 of SEQ ID NO: 3; and a light chain variable region comprising a light chain CDR1 (CDR-L1) of SEQ ID NO: 4, CDR-L2 of SEQ ID NO: 5, and CDR-L3 of SEQ ID NO:

6.

3. The pharmaceutical preparation according to claim 1 or 2, wherein the anti-CSF-1R antibody comprises: a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 7; and a light chain variable region comprising the amino acid sequence of SEQ ID NO:

8. 4 . The pharmaceutical formulation according to claim 1 , wherein the concentration of the anti-CSF-1R antibody is in the range of 40 mg / mL to 100 mg / mL, in particular 50 mg / mL.

5. The pharmaceutical formulation according to any one of claims 1 to 4, wherein the surfactant is polysorbate.

6. The pharmaceutical formulation according to any one of claims 1 to 5, wherein the polysorbate is present in a concentration ranging from 0.01% (w / v) to 0.1% (w / v), in particular 0.04% (w / v).

7. The pharmaceutical formulation according to any one of claims 1 to 6, wherein the buffer is a histidine buffer, in particular a histidine chloride buffer.

8. The pharmaceutical formulation according to any one of claims 1 to 7, wherein the concentration of the buffer is in the range of 10 mM to 30 mM, in particular 20 mM.

9. The pharmaceutical formulation according to any one of claims 1 to 8, wherein the pH of the formulation is in the range of 5.0 to 6.5, in particular 6.

0.

10. The pharmaceutical formulation according to any one of claims 1 to 9, wherein the at least one stabilizer is selected from the group consisting of salts, sugars and amino acids.

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