WO2019024783A1 - ANTIBODY COMPOSITION FOR TNF-α, AND APPLICATION THEREOF - Google Patents

Abstract

The present invention provides an antibody composition for TNF-α. The solvent of the composition comprises water, and the solute of the composition comprises an anti-TNF-α antibody. The pH value of the composition is 4.0-8.0, and the pH value is adjusted by introducing carbon dioxide to the solvent and solute mixture of the composition. Compared with the prior art, the antibody composition of the present invention contains no common buffer system, and carbon dioxide is introduced to replace a buffer system and keep the pH value of a preparation stable; thus, pains and discomfort of a patient during injection is reduced, and the antibody composition is of great significance to treatment of TNF-α-related diseases.

Description

Antibody composition against TNF-α and application thereof Technical field

The present invention relates to the novel pharmaceutical preparations, and in particular to an antibody composition against TNF-α and uses thereof.

Background technique

Rheumatoid arthritis (RA) is a common arthritis that is an autoimmune disease with an incidence of 0.3-1% in the population. Failure to treat it in time can lead to bone destruction and joint damage. There are a variety of pro-inflammatory cytokines involved in the pathogenesis of RA, such as tumor necrosis factor-α (TNF-α), interleukins such as IL-1, IL-6, IL-8, etc. (Marc Feldmann, Ravinder N. Maini. Discovery of TNF-α as a therapeutic target in rheumatoid arthritis preclinical and clinical studies. Joint Bone Spine, 2009; 69: 12-18.). Therefore, inhibiting the production of pro-inflammatory cytokines or blocking their physiological effects is currently a hot topic in the field of RA research. In recent years, many newly developed biological agents can control the progression of the disease by blocking or down-regulating the activity of pro-inflammatory cytokines, such as TNF-α inhibitors, anti-IL-6R antibodies, etc. The main mechanisms of action of these biological agents are: (1) a monoclonal antibody against a cytokine or a receptor thereof; (2) a soluble receptor antagonist, that is, a cytokine membrane receptor that does not include a transmembrane component and an intracellular functional region. Receptor antagonists bind to free cytokines and inhibit binding of the latter to cell surface receptors. Soluble receptor antagonists have a short half-life and can be prolonged by the addition of certain structures such as Fc receptors of human IgG or polyethylene glycol (PEG); (3) receptor antagonistic proteins, which are biologically inactive. Protein, which competes with cytokines for binding to membrane receptors on the cell surface. Receptor antagonistic proteins must be combined with more than 90% of cell surface receptors to be considered effective.

It is currently believed that TNF-α is one of the most important pro-inflammatory cytokines in many cytokines of RA inflammatory response, and it plays an important role in the development of RA, local inflammation and tissue damage (Fe ldmann M, Brennan FM, Foxwell BMJ, et al. The role of TNF and IL-1 in rheumatoid art hritis. CurrDir Au toimmun, 2001; 3: 188-199.). The levels of TNF-α and TNF-α receptor (TNFR) in RA were significantly elevated in serum, synovium, and synovial fluid, especially in active patients with severe disease. Serum TNF-α levels were positively correlated with joint damage score, erythrocyte sedimentation rate (ESR), and anemia. TNF-α is also associated with weight loss and disease recurrence in RA patients. TNF-α can act on a variety of cells, such as promoting macrophage secretion of inflammatory cytokines and chemokines, and promoting inflammation. The main functions of TNF-α are: (1) induce endothelial cells to express adhesion molecules and vascular endothelial growth factor (VEGF), promote adhesion and penetration of leukocytes and vascular endothelium, leading to local inflammatory reaction; (2) acting on hepatocytes, producing C-reactive protein (CRP); (3) In RA, TNF-α can act on osteoclasts, synoviocytes and chondrocytes, respectively, resulting in the activation of these cells, producing metalloproteinases, collagenases, and basal membrane lytic enzymes ( Stromelysin) and prostaglandin E2 (PGE2) further destroy cartilage causing bone erosion, joint inflammation and cartilage destruction; (4) TNF-α also promotes the production of IL-1 by synoviocytes, macrophages, fibroblasts and chondrocytes IL-8 and TNF-α themselves aggravate tissue damage. Therefore, inhibition of TNF-α is important for controlling the condition of RA and improving prognosis (Daniel Tracey, Lars Klareskog, Eric H. Sssso, etal. Tumor necrosis factor antagon ist mechanisms of action A comprehensive review. Pharmacology and Therapeutics, 2008; 244-279).

There are five TNF-α inhibitors approved by the US FDA: soluble receptor antagonist - Etanercept, human and mouse chimeric antibody - Infliximab, all-human monoclonal antibody Adalimumab, a fully human monoclonal antibody, Golimumab and a polyethylene glycol humanized Fab' fragment, Certolizumab pegol. Etanercept is a recombinant human type II tumor necrosis factor receptor-antibody fusion protein composed of a type II TNF receptor (p75) and a Fc portion of IgG1; Infliximab is composed of a human Ig stable region and a murine Ig variable region. Chimera, specifically binding to TNF; fully human monoclonal antibody can also specifically bind TNF, its stable and variable regions are human; Certolizumab pegol Fab' fragment specifically binds to TNF, C-terminally linked polyethylene glycol Can extend the half-life. The main adverse reactions of TNF inhibitors are injection site reactions, infections, tumors, lymphoproliferative disorders, neurodegenerative lesions, and lupus-like syndrome (DJ Shealy, S. Visvanathan. Anti-TNF Antibodies: Lessons from the Past, Roadmap For the Future. Handbook of Experimental Pharmacology 2008; 181: 101-129. B. Gatto. Biologics targeted at TNF: design, production and challenges. Reumatismo, 2006; 58(2): 94-103).

The above TNF-α inhibitors are intravenous and subcutaneous. Subcutaneous preparations are more convenient for patients to use than intravenous preparations, and the injection time is greatly shortened. It is not required to be injected by a professional physician, and the patient can inject the medicine after receiving the training. However, there are also shortcomings in subcutaneous preparations: factors such as formulation ingredients and pH tend to cause discomfort and even obvious pain when the patient is injected, which may lead to the patient not taking the medicine according to the doctor's advice, especially in the case of non-fatal diseases such as inflammatory diseases. Laursen et al. compared the pain experienced by patients with subcutaneous formulations of different buffer systems and found that citrate buffer formulations caused more pain in patients than the histidine buffer formulation (Laursen T, Hansen B) , Fisker S. Pain Perception after Subcutaneous Injections of Media Containing Different Buffers. Basic & Clinical Pharmacology & Toxicology, 2006; 98(2): 218-221).

Summary of the invention

Based on this, it is necessary to provide an antibody composition against TNF-α against the problems of discomfort, pain, and the like due to factors such as ingredients, pH, and the like in the conventional subcutaneous injection preparation.

In a first aspect, the present invention provides an antibody composition against TNF-α, wherein the solvent comprises water, the solute comprises an anti-TNF-α antibody; the antibody composition has a pH of 4.0 to 8.0, and the pH is The liquid component of the antibody composition is pressurized with carbon dioxide.

In some of these embodiments, the solute further comprises a stabilizer, a surfactant; the anti-TNF-α antibody comprises a recombinant human anti-TNF-α monoclonal antibody BAT1406.

In some of these embodiments, the antibody composition for TNF-[alpha], including the type and amount of solute, comprises:

The concentration of the anti-TNF-α antibody is 20.0-130.0 mg/ml;

The stabilizer comprises one or two of 20.0-60.0 mg/ml polyol and 70.0-100.0 mg/ml sugar;

The surfactant comprises 0.5-2.0 mg / ml polysorbate 80;

The antibody composition has a pH of from 4.0 to 8.0, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 10-300 mmHg.

In some of these embodiments, the antibody composition for TNF-[alpha], including the type and amount of solute, satisfies one or more of the following preferred conditions:

The concentration of the anti-TNF-α antibody is 20.0-110.0 mg/ml;

The type of the polyol includes mannitol or sorbitol, the concentration of the polyol is 30.0-50.0 mg/ml; the kind of the saccharide includes sucrose or trehalose, and the concentration of the saccharide is 80.0-90.0 mg. /ml;

The concentration of the polysorbate 80 is 0.8-1.5 mg / ml;

The antibody composition has a pH of from 4.9 to 5.7, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 100-300 mmHg.

In some of these embodiments, the antibody composition for TNF-[alpha], including the type and amount of solute, satisfies one or more of the following preferred conditions:

The concentration of the anti-TNF-α antibody is 50 mg/ml or 100 mg/ml;

The type of the polyol includes mannitol or sorbitol, the concentration of the polyol is 42 mg/ml, and the type of the sugar includes sucrose or trehalose, and the concentration of the saccharide is 84 mg/ml;

The concentration of the polysorbate 80 is 1.0 mg / ml;

The antibody composition has a pH of 5.0 to 5.4, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 220 mmHg.

In some of these embodiments, the antibody composition for TNF-[alpha], the species of the solute and its content comprises:

The concentration of the anti-TNF-α antibody is 100.0 mg/ml,

The stabilizer is mannitol at a concentration of 42.0 mg/ml.

The surfactant is polysorbate 80, and the concentration is 1.0 mg/ml.

The antibody composition has a pH of 5.0 to 5.4, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 220 mmHg.

In some of these embodiments, the antibody composition for TNF-[alpha], including the type and amount of solute, comprises:

The concentration of the anti-TNF-α antibody is 70.0-130.0 mg/ml;

The stabilizer includes one or two of 3.0-15.0 mg/ml polyol and 5.0-8.0 mg/ml sodium chloride.

The surfactant comprises 3.0-10 mg / ml polysorbate 80;

The antibody composition has a pH of from 4.0 to 6.0, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 20-80 mmHg.

In some of these embodiments, the antibody composition for TNF-[alpha], including the type and amount of solute, satisfies one or more of the following preferred conditions:

The concentration of the anti-TNF-α antibody is 90.0-110.0 mg/ml;

The type of the stabilizer includes mannitol, sodium chloride, the concentration of the mannitol is 8.0-12.0 mg/ml, and the concentration of the sodium chloride is 6.0-7.0 mg/ml;

The surfactant comprises 4.0-8.0 mg/ml polysorbate 80;

The antibody composition has a pH of from 4.8 to 5.5, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 40-60 mmHg.

In some of these embodiments, the antibody composition for TNF-[alpha], including the type and amount of solute, satisfies one or more of the following preferred conditions:

The concentration of the anti-TNF-α antibody is 100 mg/ml;

The concentration of mannitol is 10.0 mg/ml, and the concentration of the sodium chloride is 6.5 mg/ml;

The concentration of the polysorbate 80 is 6.0 mg / ml;

The antibody composition has a pH of 5.0 to 5.4, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 50 mmHg.

In some of these embodiments, the antibody composition for TNF-[alpha], including the type and amount of solute, is as follows:

The concentration of the anti-TNF-α antibody is 100.0 mg/ml,

The stabilizer comprises 10.0 mg/ml of mannitol and 6.5 mg/ml of sodium chloride.

The surfactant is polysorbate 80, and the concentration is 6 mg/ml.

The antibody composition has a pH of 5.0 to 5.4, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 50 mmHg.

In a second aspect, the invention provides an antibody preparation for TNF-[alpha] comprising a closed container, and said antibody composition for TNF-[alpha] in a closed container, said composition having a pH of from 4.0 to 8.0.

In some embodiments, the closed container comprises a pressure can, a vial, an injection needle, and the like.

In a third aspect, the present invention provides a use according to the antibody composition for TNF-α in the preparation of a therapeutic agent for a TNF-α-causing disorder.

In some of these embodiments, the therapeutic agent comprises a subcutaneous injection formulation.

Compared with the prior art, the present invention has the following beneficial effects:

At present, the type and concentration of the buffer contained in the subcutaneous injection are closely related to the pain and discomfort generated during the injection of the patient, and have been confirmed by a large number of experiments. The antibody composition of the present invention does not contain a common buffer system, and replaces the buffer system by filling carbon dioxide and maintains a stable pH of the preparation, thereby reducing the pain and discomfort of the patient during injection, and is important for treating TNF-α-related diseases.

Further, the present invention adds a stabilizer, a surfactant to the antibody composition, and further prepares the formulation of the antibody preparation as follows (1) or (2): (1) solvent water containing anti-TNF-α antibody The concentration is 20.0-130.0 mg/ml; the stabilizer contained includes one or two of 20.0-60.0 mg/ml polyol and 80.0-90.0 mg/ml sugar; the surfactant contained includes 0.5-2.0 mg/ Ml polysorbate 80; the formulation composition contained has a pH of 4.0 to 8.0, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 10-300 mmHg. (2) a solvent water having a concentration of the anti-TNF-α antibody of 70.0 to 130.0 mg/ml; and the stabilizer comprises one of 3.0-15.0 mg/ml of a polyol and 5.0-8.0 mg/ml of sodium chloride. Or both, the surfactant comprises 3.0-10 mg/ml polysorbate 80; the antibody composition has a pH of 4.0-6.0, and the pH is obtained by into the liquid component of the antibody composition 20-80mmHg pressure is pressed into the carbon dioxide to adjust. The technical effect of the shelf life of the contained antibody component in the liquid state is 30 months, and the advantageous effect of the antibody composition being stored at room temperature for 2 weeks and the shelf life of 2 months at 2-8 ° C is 30 months; The antibody composition exhibits excellent effects in terms of high temperature, light, antibacterial, and the like.

In addition, on the basis of the above (1) or (2) optimized formula, the inventor has further optimized the carbon dioxide pressure range through multiple experiments, and optimized the solute type and content of the antibody composition to realize the antibody combination. The material is further improved in terms of shelf life, high temperature, light, and antibacterial.

DRAWINGS

Fig. 1 (Fig. A, Fig. B, Fig. C) Results of room temperature stability test of the preparation of recombinant human anti-TNF-α monoclonal antibody BAT1406; wherein: Figure A is the size exclusion chromatography test result of each sample of Example 1. Figure B is a graph showing the results of capillary electrophoresis test for each sample of Example 1; Figure C is a graph showing the results of biological activity test of each sample of Example 1.

Figure 2 is a graph showing the results of a 15-day stability test under high temperature (40 ° C) conditions of two recombinant human anti-TNF-α monoclonal antibodies BAT1406;

Figure 3 is a graph showing the results of stability experiments of two recombinant preparations of recombinant human anti-TNF-α monoclonal antibody BAT1406 under room temperature illumination (4000 lx);

Figure 4 is a diagram showing the results of a screening test for the mannitol concentration of the recombinant A anti-TNF-α monoclonal antibody BAT1406;

Figure 5A Effect of recombinant human anti-TNF-α monoclonal antibody BAT1406 on experimental mouse tg197 in vivo arthritis score; Figure 5B Effect of recombinant human anti-TNF-α monoclonal antibody BAT1406 on experimental histopathological score of experimental mouse tg197;

Figure 6 is a graph showing the blood concentration of the recombinant human anti-TNF-α monoclonal antibody BAT1406A preparation when administered at low, medium and high doses;

Figure 7 is a graph showing the plasma concentration of the recombinant human anti-TNF-α monoclonal antibody BAT1406A preparation and the buffer system-containing preparation.

Detailed ways

The antibody preparation composition of the present invention and its use will be further described in detail below in conjunction with specific examples. In order to more clearly understand the technical content of the present invention, the following embodiments are specifically described. It is to be understood that the examples are not intended to limit the scope of the invention. The experimental methods in the following examples which do not specify the specific conditions are usually carried out according to the conditions described in the conventional conditions, for example, Sambrook et al., Molecular Cloning: Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or according to the manufacturer. The suggested conditions. The various common chemical reagents used in the examples are commercially available products.

Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning meaning The terms used in the description of the present invention are for the purpose of describing the specific embodiments and are not intended to limit the invention. The term "and/or" used in the present invention includes any and all combinations of one or more of the associated listed items.

The present invention provides an antibody composition against TNF-α, wherein the solvent comprises water, the solute comprises an anti-TNF-α antibody; the antibody composition has a pH of 4.0 to 8.0, and the pH is obtained by combining the antibodies The liquid component of the substance is pressurized with carbon dioxide; further, the above composition further includes a surfactant and a stabilizer.

Further, the antibody composition against TNF-α is preferably of two formulations, as follows:

Formula A: solvent water, the concentration of the anti-TNF-α antibody contained is 20.0-130.0 mg/ml; the stabilizer contained includes one of 20.0-60.0 mg/ml polyol, 80.0-90.0 mg/ml sugar or Two kinds; the surfactant contained includes 0.5-2.0 mg/ml polysorbate 80; the antibody composition contained has a pH of 4.0-8.0, and the pH is obtained by into the liquid component of the antibody composition 10-300mmHg pressure is pressed into the carbon dioxide-regulated; the antibody composition prepared in the A formula is referred to as the A preparation, such as Table 1A below;

Formula B: solvent water, the concentration of the anti-TNF-α antibody contained is 70.0-130.0 mg/ml; the stabilizer includes one of 3.0-15.0 mg/ml polyol, 5.0-8.0 mg/ml sodium chloride. Or both, the surfactant comprises 3.0-10 mg/ml polysorbate 80; the antibody composition has a pH of 4.0-6.0, and the pH is obtained by into the liquid component of the antibody composition 20-80mmHg pressure is pressed into the carbon dioxide to adjust. An antibody composition prepared in Formula B is referred to as a B preparation, such as Table 1B below;

Table 1A. List of A Formulations for Recombinant Human Anti-TNF-α Monoclonal Antibody BAT1406

Features	Ingredient name	Dosage	concentration
anti-TNF-α antibody	Antibody BAT1406	40mg	100mg/ml
stabilizer	Mannitol	16.8mg	42mg/ml
Surfactant	Polysorbate 80	0.4mg	1mg/ml
pH regulator	carbon dioxide	220mmHg	pH5.0-5.4
Solvent	Water for Injection	Added to 0.4ml	-

Table 1B. List of B Formulations for Recombinant Human Anti-TNF-α Monoclonal Antibody BAT1406

Features	Ingredient name	Dosage	concentration
anti-TNF-α antibody	Antibody BAT1406	40mg	100mg/ml
stabilizer	Mannitol	4mg	10mg/ml
Surfactant	Polysorbate 80	2.4mg	6mg/ml
stabilizer	Sodium chloride	2.6mg	6.5mg/ml
pH regulator	carbon dioxide	50mmHg	pH5.0-5.4
Solvent	Water for Injection	Added to 0.4ml	-

Example 1: Preparation of antibody composition

This example illustrates the preparation of a 20 L A formulation:

(1) The following weight components were weighed: 840 g of mannitol (stabilizer), 20 g of polysorbate 80 (surfactant) and 20 L of water for injection.

The carbon dioxide is charged after the preparation of the preparation.

(2) A solvent system is prepared by dissolving the said mannitol (stabilizer) and polysorbate 80 (surfactant) components in about 90% by weight of the water for injection.

It has been shown that the order of adding mannitol (stabilizer) and polysorbate 80 (surfactant) does not affect the quality of the preparation, and can be selected at will.

(3) The BAT1406 antibody concentrate containing 2.0 kg of total protein was thawed in a water bath, and then the solvent system was added to the antibody concentrate under stirring until the final weight of the total solution was reached.

(4) The mixed solution obtained in the step (3) is filled with carbon dioxide to adjust the pH, a final volume of water for injection is added, and then the preparation is sterilized by a filter membrane of a hydrophilic polyvinylidene fluoride having a pore size of 0.22 μm. The filter membrane can filter the formulation into a sterile container. The filter medium used is filter sterilized ammonia.

The formulation containing all of its ingredients was then filtered by filtration as described above, except that the formulation was filtered through two layers of sterile 0.22 μm membrane filters. After disinfection, the reagents are packaged for use in vials or pre-filled syringes, all of which are verified by microbial intrusion tests and meet the requirements of the Good Manufacturing Practice (2010 Revision).

The skilled person will understand that the weight and/or weight to volume ratios referred to herein may be converted to molar and/or molar concentrations using well known molecular weights of the ingredients. The weights listed herein are for the volume. The skilled person will understand that the weight can be adjusted proportionally when different formulation volumes are required. For example, the 16L, 14L, 12L, 10L, 5L formulations each comprise 80%, 70%, 60%, 50%, 25% of the listed weight.

The preparation of the B preparation is the same as the preparation of the above A.

Example 2: Stability study at room temperature

Prepare BAT1406 antibody in PBS sample A1 (20 mg/mL) and A formulation low concentration sample A2 (4 mg/mL), medium concentration sample A3 (20 mg/mL), high concentration sample A4 (100 mg/mL), and place at room temperature Save for 2 weeks. The formulation of the A1-A4 numbered samples is shown in Table 2A:

Table 2A

The samples were tested by capillary electrophoresis (CE), size-exclusion chromatography (SEC) and biological activity (TNF-α-induced L929 cytotoxicity neutralization test). The results of the assay are shown in Table 2B and 1.

The results showed that the BAT1406 antibody A preparations were low (A2 sample), medium (A3 sample), high (A4 sample) three samples at room temperature for 2 weeks, the main peak degradation trend was consistent, the range was 98.95-95.17%, obviously Better than PBS sample A1 (98.66-87.27%); CE-SDS (NR) results show that BAT1406 antibody A preparation is low (A2 sample), medium (A3 sample), high (A4 sample) three concentrations of sample at room temperature After 10 days of storage, the degradation trend of the main peak was consistent, ranging from 93.20 to 89.05%, which was significantly better than PBS sample A1 (93.79-82.36%). The results of biological activity showed that the BAT1406 antibody A preparation was low, medium and high. After 2 weeks at room temperature, the activity remained essentially unchanged, ranging from 98.52 to 97.49%, which was significantly better than PBS sample A1 (98.49-90.27%). The overall comparison shows that the preparation prepared by the invention is significantly better than the preparation prepared by using the PBS buffer; for the preparation of the present application, the two preparations of the low, medium and high concentrations of the A preparation have two concentrations of medium and high. The sample is more stable.

Table 2B. Results of room temperature stability test for A1-A4 numbered samples

Example 3: Study on factors affecting the pH of the preparation

Prepare BAT1406 Antibody A preparation (see Table 1A for specific formulation) 10L sample at a concentration of 100mg/mL, store in a pressure tank, charge different amounts of carbon dioxide, place at 4 ° C, test the pH of the preparation in the pressure tank when different carbon dioxide pressure is measured . The results of the measurements are shown in Table 3A.

Table 3A. Experimental Results of Carbon Dioxide Pressure on pH of BAT1406A Formulation

Carbon dioxide pressure (mmHg)	Formulation pH
300	4.76
270	4.98
240	5.06
210	5.15
180	5.30
150	5.51
100	5.84

Prepare BAT1406 Antibody A preparation (see Table 1A for specific formulation) 10L sample at 100mg/mL, store in vial and injection needle (see Table 3B for material information), fill the vial and injection needle with carbon dioxide, and the pressure is 220mmHg. It was placed at room temperature to investigate the effect of sealability of the vial and the injection needle and changes in carbon dioxide pressure on the pH of the preparation. The results of the measurements are shown in Table 3C.

Table 3B. Packaging Information

name	Vendor
Vial bottle	SCHOTT Glass Technology (Suzhou) Co., Ltd.
Rubber stopper	French STEMI
Injection needle	BD Medical Devices (Shanghai) Co., Ltd.

Table 3C. The pH of the BAT1406A preparation is affected by the preservation time.

save time	Formulation pH
0d	5.16
3d	5.18
6d	5.21
12d	5.22
15d	5.25

The inventors have proved through many experiments that the preparation of the present invention can achieve better comfort when the carbon dioxide is pressurized at a pressure of 10-300 mmHg to adjust the pH to 4.0-8.0.

Further, the results of the above examples show that when the carbon dioxide gas pressure ranges from 100 to 300 mmHg, the preparation pH is 4.76-5.84; the seal bottle of the preservation preparation A and the injection needle have good sealing property, and the charged carbon dioxide gas pressure is 220 mmHg. After storage for 15 days at room temperature, the pH of the preparation varied from 5.16 to 5.25, which basically met the experimental requirements.

Similarly, the B formulation of BAT1406 (see Table 1B for specific formulation) has a gas pressure range of 40-60 mmHg, preferably 50 mmHg.

Example 4: Stability study under high temperature (40 ° C) conditions

Prepare B preparation of BAT1406 (see Table 1A for specific formulation) and preparation B (see Table 1B for specific formulation), and store at high temperature (40 °C) for 15 days. The samples were separately subjected to SEC and ion exchange chromatography. , IEC) test, the results of the test are shown in Table 4 and Figure 2.

Table 4. Results of 15 days stability test of A and B preparations under high temperature (40 ° C) conditions

The results showed that the samples of A and B were placed at high temperature (40 ° C) for 15 days. The main peak of B preparation was lower than that of A preparation, and the SEC aggregates and fragments were also more. The downward trend of IEC main peak of A preparation was consistent with that of B preparation. However, the performance on the IEC acid zone is better than that of the B formulation. It is indicated that the stability of the antibody A preparation at high temperature is better than that of the B preparation.

Example 5: Stability study under room temperature illumination (4000 lx)

A preparation of BAT1406 antibody (see Table 1A for specific formulation) and preparation B (see Table 1B for specific formulation) were prepared in a vial with a light-shielding material and stored under room temperature (4000 lx) for 2 weeks. The samples were tested by IEC and the results of the measurements are shown in Figure 3.

The results showed that: BAT1406 antibody A preparation and B preparation sample were placed under room temperature illumination (4000lx) for 2 weeks, the main peak of IEC of A preparation remained basically unchanged, and the main peak of B preparation decreased faster than that of A preparation, indicating that although both preparation A and preparation B were It has better photostability, while the antibody A preparation is better than the B preparation under light (4000lx).

Example 6: Mannitol concentration screening test

In order to optimize the concentration of mannitol in the preparation, the concentration of mannitol in the A preparation is from 60 mg/ml to 3 mg/ml, up to 18 mg/ml, and 15 (numbers 1-15, decreasing in concentration) are different. The BAT1406 antibody A preparation (100 mg/ml) of mannitol concentration was different from Table 1A except that the A preparation mannitol concentration was partially the same as Table 1A. The storage temperature was 40 ° C and the time period was 4 weeks. The samples were tested by SEC and IEC respectively, and the results are shown in Figure 4.

The results showed that the combined SEC main peak and IEC main peak data can be seen that mannitol can protect BAT1406 antibody at 20-60mg/ml, while mannitol concentration is 42mg/ml (ie No. 7 preparation). The best protection for antibodies.

Similarly, the mannitol concentration of the B formula was optimized, and it was found that mannitol can protect the BAT1406 antibody at 3-15 mg/ml, while the mannitol concentration is 10 mg/ml for the antibody. The protection is best.

Example 7: Microbial research

By performing microbiological studies on the A preparation and the B preparation, it is determined whether the preparation can support microbial growth.

Specifically, inoculating Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Bacillus cereus to the A preparation (see Table 1A for specific formulation) and the B preparation (see Table 1B for specific formulation), respectively. The amount was 100 cfu/ml, and the whole microbial growth in the inoculated preparation was measured by room temperature for 15 days after inoculation. The evaluation indexes mainly included the number of microorganisms under the microscope and the change of the turbidity of the preparation, wherein the lack of turbidity indicates that there was no overall microbial growth. . Among them, the microorganisms used in the examples include Escherichia coli (CICC-10003), Staphylococcus aureus (CICC-10306), Pseudomonas aeruginosa (WDCM-00024), and Bacillus cereus (CICC-10352).

The test results are shown in Table 5: Storage at room temperature 20-25 ° C for 15 days, the A preparation, B preparation does not support microbial growth.

Table 5. Microbiological test results of A and B preparations

Example 8: Insoluble particulate study

The insoluble particles in the preparation have a particle size of 1-50 μm, are invisible to the naked eye, can not be metabolized with blood flow, and may cause hard-to-find and potentially serious harm to the human body. Therefore, insoluble particle control is included in the injection quality standard.

Prepare low, medium and high concentration samples of BAT1406A preparation, wherein: A1 sample antibody partial concentration is 4mg/mL, the rest are the same as Table 1A; A2 sample antibody partial concentration is 20mg/mL, the rest are the same as Table 1A; A3 The sample composition is the same as Table 1A;

Prepare low, medium and high concentration samples of BAT1406B preparation, wherein: B1 sample antibody partial concentration is 4mg/mL, the rest is the same as Table 1B; B2 sample antibody partial concentration is 20mg/mL, the rest are the same as Table 1B; A3 The sample composition is the same as Table 1B;

The insoluble particles in the formulation were counted using a Beckman Laboratories Liquid Particle Counter (model HIAC 9703+). The test results are shown in Table 6.

Table 6. Insoluble particle test results for both types of formulations

preparation	Preparation concentration	≥10μm particles	≥25μm particles
A1	4mg/mL	8.9	1.4
A2	20mg/mL	11.2	2.3
A3	100mg/mL	15.8	2.5
B1	4mg/mL	9.5	1.8
B2	20mg/mL	13.2	2.5
B3	100mg/mL	17.1	2.9

It can be seen from Table 6 that the insoluble particles of the A preparation and the B preparation are in compliance with the Chinese Pharmacopoeia (2015 edition) insoluble particulate inspection standard.

Example 9: Comparative study on biological activity of two preparations

100 mg/ml BAT1406 antibody preparation composition (Table 1A preparation) was stored in a 2-8 ° C refrigerator, and an in vitro TNF-α neutralization test (L-929 biological activity test) was performed every 3 months, and each sample was repeated 2 The time period is 36 months, and the activity of the A preparation (Table 1A) of the month is taken as 100% relative activity, and the normal relative activity range of the preparation is set to be 80-125%;

The B preparation (Table 1B) was tested with reference to the detection method of the above A preparation;

The test results of the above A preparation and B preparation are shown in Table 7.

Table 7. Long-term storage activity test results of the two formulations

The results show that the A preparation of the present invention has a shelf life of at least 30 months, and the B preparation has a shelf life of at least 24 months.

Example 10: Recombinant human anti-TNF-α monoclonal antibody BAT1406 protein sequence

The recombinant human anti-TNF-α monoclonal antibody BAT1406 was expressed in CHO cells by genetic engineering techniques and purified by a series of standard chromatographic steps. BAT1406 is an IgG1 antibody with a molecular weight of approximately 148 kDa and consists of two IgG1 ^{z, a} heavy chains and two kappa light chains.

Each heavy chain contains 451 amino acids, a molecular weight of 49 kDa, and its heavy chain amino acid sequence is shown in SEQ ID No. 1.

Each light chain contains 214 amino acids with a molecular weight of 24 kDa and the light chain amino acid sequence is set forth in SEQ ID No. 2.

Example 11: Expression and purification of recombinant human anti-TNF-α monoclonal antibody BAT1406

A monoclonal antibody that specifically binds TNF-α is expressed in CHO cells according to the method of Wood et al., J Immunol. 145: 3011 (1990). The expression vector containing the antibody gene was constructed by a conventional molecular biological method (Molecular Cloning) and expressed as a host cell of a CHO cell (ATCC CCL61). Yield build process of stable cell lines described as follows: a host cell grown in suspension in CDCHO AGT ^TM media (Gibco, CA), taken in the logarithmic growth phase by centrifugation host cells, resuspended in fresh medium of CD CHO AGT ^TM, The cell density was counted and adjusted to 1.24×10 ⁷ cells/ml, 800 ul of the above cell suspension was added to the electric shock cup, and then 40 ug of the linearized plasmid was added, and the cells were mixed with the plasmid by pipetting. It was transformed with a Bio-rad electrorotator, and the instrument parameters were set to capacitance: 960 uFD, voltage: 300V. Usually the electric shock time is 15-20 milliseconds is normal. The electroporated cells were immediately resuspended in 37 ° C pre-warmed CD CHO AGT ^TM medium, 100 μl per well was added to a 96-well plate, and 2-3 days later, an equal amount of screening medium (CDCHO AGT media + 50 uM methionine imino generation) was added. Sulfone (MSX)). The expression level of the antibody in the 96-well plate cell culture supernatant was measured. The clones with higher expression levels were transferred from 96-well plates to 24-well plates. After the cells were grown to a certain amount, the cells were transferred to 6-well plates, and 2×10 ⁵ cells were cultured in 5 ml of each well to measure the antibody production of the cells. And yield. Usually 20-30 clones are transferred to shake flasks for further evaluation. The last 5-8 clones with the highest expression were subcloned and further screened for expression. The culture solution was harvested, and the cells and the medium were separated by low speed centrifugation, and the supernatant was further clarified by high speed centrifugation. The preliminary purification using cation exchange chromatography (POROS XS medium) can achieve the intended purpose, effectively removing most of the heteroproteins while capturing the target protein, and the purity is over 90%. After preliminary purification, anion (Giga Cap Q-650M) and cation (CM Ceramic HyperD F) exchange chromatography were used for further separation and purification through detailed process research to remove endotoxin, HCP, DNA, and acidic isomers. Etc.

Example 12: Pharmacodynamic study of recombinant human anti-TNF-α monoclonal antibody BAT1406

Pharmacodynamic studies mainly carried out in vitro pharmacodynamic tests of recombinant human anti-TNF-α monoclonal antibodies and pharmacodynamic studies in in vivo animal models. In vitro tests mainly carry out the following pharmacodynamic tests: binding ability to TNF, antibody specificity, competition for binding of TNF to receptor, inhibition of biological activity of TNF-α, and cytotoxicity experiments. . The binding ability of recombinant human anti-TNF-α monoclonal antibody to TNF-α was compared by taking antibody specific binding as an example.

The specific experimental method is as follows: firstly dilute rhTNF-α, rhTNF-β or rmTNF-α into 50 ng/50 μl with PBS, and add diluted rhTNF-α, rhTNF-β or rmTNF-α to a micro 96-well microtiter plate. , 50 μl/well, overnight at 4 °C. The next day, it was blocked with PBS containing 3% BSA, 100 μl/well, and placed at 37 ° C for 2 hours. After discarding the supernatant, different concentrations of recombinant human anti-TNFα monoclonal antibody BAT1406 and human IgG were added to the corresponding wells (final antibody concentration of 60.75 ug/ml, 3-fold dilution, 10 gradients, duplicate wells), at 37 ° C The effect is 2 hours. After discarding the supernatant, PBST washed the plate 5 times and patted dry. Then, goat anti-human AP secondary antibody was added at 1:10000, 50 μl/well, and 37 ° C for 2 hours. After discarding the supernatant, the plate was washed 8 times with PBST, and PNPP coloring solution was added, 50 μl/well. OD405 reading as a standard curve. The results showed that different concentrations of recombinant human anti-TNFα monoclonal antibody, BAT1406 specific binding to rhTNF-α and concentration gradient results, the results are consistent; but different concentrations of recombinant human anti-TNFα monoclonal antibody BAT1406 to rhTNF-β or rmTNF-α There is no specific combination. Negative control human IgG did not specifically bind to rhTNF-α, rhTNF-β or rmTNF-α, demonstrating that BAT1406 binds only to rhTNF-α (IC50=3×10 ^-9 M), but not to rhTNF-β and rmTNF- Alpha binding.

The pharmacodynamic evaluation of animals mainly carried out acute animal models and chronic animal models. Among them, the acute animal model is D-galactose-sensitive mouse and rhTNF-α-induced rabbit fever model. A mouse model of intraperitoneal injection of 1μg rhTNF-α+20mg D-galactosamine was used as a model to study the toxic effects of recombinant human anti-TNF-α monoclonal antibody BAT1406 against rhTNF-α and D-galactosamine. TNF-α monoclonal antibody injection has the effect of antagonizing the toxic effect of 1 μg rhTNF-α+20 mg D-galactosamine and improving the survival rate of mice. According to the model of rhTNF-α-induced changes in the temperature of rabbit basal body, the experiment showed that recombinant human anti-TNF-α monoclonal antibody BAT1406 antagonized the effect of rhTNF-α induced rabbit temperature.

The chronic animal model is a model of spontaneous polyarthritis in human soluble TNF transgenic mice (Tg197). In vivo efficacy evaluation of recombinant human anti-TNF-α monoclonal antibody BAT1406 was performed using a Tg197 (stable expression of sTNF) transgenic mouse model. The injection method used in this study was intraperitoneal injection. The determination of the intraperitoneal injection protocol was based on the historical data of other anti-TNF-α monoclonal antibodies in the tg197 mouse test. The transgenic mice were divided into 4 groups (G1). -G4): Each group contained 8 mice of different ages and genders. The G1 group was injected with normal saline, and the G2-G4 group was injected with 3 mg/kg, 10 mg/kg, 30 mg/kg of BAT1406 antibody. In order to prevent and treat arthritis, the experimental product was injected twice a week from the third week, starting from the third week, to the tenth week, before the formation of arthritis. To control the histopathological state at the onset of the lesion, a control group of transgenic mice (4) was added, and the group was sacrificed before the first dose. All animals were sacrificed in the tenth week and serum and ankle joints were collected. Serum was stored at -80 °C for further analysis and the ankle joint was used for pathological evaluation. The degree of arthritic lesions was assessed using a scoring system, and histopathological evaluation was performed using a pathology scoring system. The experimental results show that the experimental product BAT1406 has a typical drug response in accordance with the pathological and in vivo inhibitory effects of Tg197 arthritis in the dose range of 3 mg/kg, 10 mg/kg, and 30 mg/kg. The 10 mg/kg and 30 mg/kg doses of BAT1406 have a good inhibitory effect on the histopathological pathology and histopathology of the histological pathology. There is no statistical difference, and the scores obtained are lower than before treatment. The BAT1406 antibody was shown to have a therapeutic effect on the tg197 arthritis model (see Figures 5A, 5B).

Example 13: Pharmacokinetic Study of Recombinant Human Anti-TNF-α Monoclonal Antibody BAT1406

The purpose of this example is to evaluate the safety of the BAT1406 antibody preparation composition (A preparation) containing no buffer system in the BAT1406 preparation containing the buffer system as a control, and provide a reference for future clinical rational drug use.

The plasma concentration of the BAT1406 antibody preparation composition in a single dose of SD rats was determined by the BAT1406 antibody ELISA assay in SD rat plasma, and the high, medium and low doses of the BAT1406 antibody preparation composition were studied. , the relationship between exposure and dose of BAT1406 antibody in SD rats, and its pharmacokinetic characteristics; by comparing the kinetic characteristics of the same antibody dose BAT1406A preparation and BAT1406 buffer system preparation (pre-development) in SD rats, The bioequivalence of the two formulations of the buffer-free formulation of the present invention and the BAT1406 buffer-containing formulation (pre-developed) were evaluated.

Taking the A preparation as an example, the research scheme is as follows:

The low, medium and high doses of BAT1406A preparation were 10mg/kg, 30mg/kg and 100mg/kg respectively (the dose here refers specifically to the dose according to the weight of SD rats), and the dose of BAT1406 containing buffer system is 30mg. /kg, 8 in each group, half male and half female. The mode of administration is intraperitoneal injection. The rats are cut at 1h, 4h, 24h, 48h, 72h, 96h, 7day, 9day, 11day, 13day, 15day, 22day, 29day after administration. The blood sample was taken from the tail, and the collected blood sample was placed in an ice bath heparin anticoagulation centrifuge tube, centrifuged within 5 minutes, and the separated plasma was stored in a -70 ° C refrigerator for later use.

Determination of BAT1406 antibody content in plasma samples: firstly coated with the recombinant rhTNF-α protein which specifically binds to the Fab segment of the BAT1406 antibody to capture the BAT1406 antibody in the biological sample, and then add the enzyme-labeled goat anti-human The antibody (IgG-HRP) was bound to the captured BAT1406 antibody, and finally the substrate was added for color development. After the reaction was terminated with a 2 mol/L sulfuric acid solution, the OD value of each well was read at 450 nm. The absorbance of the BAT1406 antibody standard sample and the concentration of the standard sample were fitted to a standard curve by four parameters, and the concentration of the test sample 1406 antibody measured on the same plate was quantified using the obtained standard curve. The mean of the pharmacokinetic parameters obtained from 8 rats represented the pharmacokinetic parameters of each dose group; the blood concentration of BAT1406 antibody at different doses and different preparation groups was statistically judged by independent sample t-test. The results are shown in Table 10 and Table 11.

Table 10. Comparison of mean plasma concentrations after administration of different doses of BAT1406A preparation

Blood collection time (h)	Low dose group (μg/mL)	Medium dose group (μg/mL)	High dose group (μg/mL)
0	0.2	0.5	1.9
1	415.3	1013.5	4502.5
4	193.4	553.6	2241.3
twenty four	180.9	421.8	1831.4
48	124.2	337.5	1733.8
96	145.6	277.6	1114.4
168	93.7	226.2	956.3
216	102.5	180.5	754.7
264	80.3	141.6	402.3
312	51.4	87.0	278.4
360	23.3	49.1	106.2
528	8.1	22.7	71.6
696	4.3	6.8	13.8

Table 11 Comparison of blood concentration of subcutaneous injections containing buffer system preparation (30 mg/kg) and A preparation (30 mg/kg)

The results (Tables 10 and 11) showed that the blood concentration of each group reached the highest value 1 h after administration, and then gradually decreased. Comparing the blood concentration of the A preparation with the buffered system preparation at the corresponding time point, it can be seen from the p-value of the t test listed in Table 11, the two preparations adopt the same administration method, and there is no significant difference in the blood concentration of the corresponding point. (P>0.05).

Example 14

The present embodiment provides a sample of the antibody composition, the raw material composition is shown in the Z1 sample and the Z2 sample in Table 12, and at the same time, a control (D-Z1 sample, D-Z2 sample) is also set for the Z1 sample and the Z2 sample, and The comfort performance of each sample was tested. For pain test, refer to Birtha Hansen et al. Pain Perception after Subcutaneous Injections of Media Containing Different Buffers. Basic & Clinical Pharmacology & Toxicology. 2006, 98, 218-221. The test results are shown in Table 13.

Table 12

Table 13

	Z1 sample	Z2 sample	D-Z1 sample	D-Z2 sample
Pain score		Pain score 2	Pain score 1	Pain score 4	Pain score 3

According to the test results of Table 13, the pain score of the sample of the present invention was significantly lower than that of the control sample, indicating that the present invention significantly lowered the pain level.

Example 15

This example provides a sample of an antibody preparation obtained by formula A. The composition of the raw materials is shown in Table 14, and the performance of each sample was tested. The test results are shown in Table 15.

Table 14

Table 15

Based on the above Examples 1-13, according to Tables 14 and 15 above, the effects of the A1 sample and the A2 sample are slightly lower than those of the A3 sample and the A4 sample, and the effects of the A3 sample and the A4 sample are relatively less than the table. Formulation formulation as described in 1A, which illustrates that the present invention has a preferred embodiment.

Example 16

This example provides a sample of the antibody preparation sample composition obtained by the B formula. The composition of the raw materials is shown in Table 16, and the performance of each sample was tested. The test results are shown in Table 17.

Table 16

Table 17

Based on the above Examples 1-13, according to Tables 16 and 17 above, the effects of the B1 sample and the B2 sample are slightly lower than those of the B3 sample and the B4 sample, and the effects of the B3 sample and the B4 sample are relatively less than the table. Formulation formulation as described in 1B, which illustrates that the present invention has a preferred embodiment.

Comparative example

This comparative example provides the following four comparative samples, wherein: D1 sample and D2 sample are comparative samples of the A1 sample in Example 15, and D3 sample and D4 sample are comparative samples of the B1 sample in Example 16, the comparative sample is composed as follows Table 18 shows the results of the detection of each sample as shown in Table 19 below.

Table 18

Table 19

From the effects of Table 19, it is understood that for the composition of the present application, the adjustment of the pH and the selection of the range of pressure play a crucial role in the realization of the effect.

The technical features of the above-described embodiments may be arbitrarily combined. For the sake of brevity of description, all possible combinations of the technical features in the above embodiments are not described. However, as long as there is no contradiction between the combinations of these technical features, All should be considered as the scope of this manual.

The above-described embodiments are merely illustrative of several embodiments of the present invention, and the description thereof is more specific and detailed, but is not to be construed as limiting the scope of the invention. It should be noted that a number of variations and modifications may be made by those skilled in the art without departing from the spirit and scope of the invention. Therefore, the scope of the invention should be determined by the appended claims.

Claims (10)

1. An antibody composition against TNF-α, characterized in that the solvent comprises water, the solute comprises an anti-TNF-α antibody; the composition has a pH of 4.0 to 8.0, and the pH is obtained by the composition The solvent solute mixture is pressurized with carbon dioxide.
2. The antibody composition for TNF-α according to claim 1, wherein the solute further comprises a stabilizer, a surfactant; the anti-TNF-α antibody comprises a recombinant human anti-TNF-α monoclonal antibody BAT1406 .
3. The antibody composition for TNF-α according to claim 2, wherein the type and content of the solute comprises:

   The concentration of the anti-TNF-α antibody is 20.0-130.0 mg/ml;

   The stabilizer comprises one or two of 20.0-60.0 mg/ml polyol and 70.0-100.0 mg/ml sugar;

   The surfactant comprises 0.5-2.0 mg / ml polysorbate 80;

   The composition has a pH of from 4.0 to 8.0, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 10-300 mmHg.
4. The antibody composition for TNF-α according to claim 3, wherein the type and content of the solute satisfy one or more of the following preferred conditions:

   The concentration of the anti-TNF-α antibody is 20.0-110.0 mg/ml;

   The type of the polyol includes mannitol or sorbitol, the concentration of the polyol is 30.0-50.0 mg/ml; the kind of the saccharide includes sucrose or trehalose, and the concentration of the saccharide is 80.0-90.0 mg. /ml;

   The concentration of the polysorbate 80 is 0.8-1.5 mg / ml;

   The formulation composition has a pH of from 4.9 to 5.7, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 100-300 mmHg.
5. The antibody composition for TNF-α according to claim 4, wherein the type and content of the solute satisfy one or more of the following preferred conditions:

   The concentration of the anti-TNF-α antibody is 50.0 mg/ml or 100.0 mg/ml;

   The type of the polyol includes mannitol or sorbitol, the concentration of the polyol is 42.0 mg/ml, and the type of the sugar includes sucrose or trehalose, and the concentration of the saccharide is 84.0 mg/ml;

   The concentration of the polysorbate 80 is 1.0 mg / ml;

   The composition has a pH of from 5.0 to 5.4, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 220 mmHg.
6. The antibody composition for TNF-α according to claim 2, wherein the species and content including the solute comprises:

   The concentration of the anti-TNF-α antibody is 70.0-130.0 mg/ml;

   The stabilizer comprises one or two of 3.0-15.0 mg/ml polyol, 5.0-8.0 mg/ml sodium chloride;

   The surfactant comprises 3.0-10.0 mg / ml polysorbate 80;

   The composition has a pH of from 4.0 to 6.0, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 20-80 mmHg.
7. The antibody composition for TNF-α according to claim 6, wherein the species and content including the solute satisfy one or more of the following preferred conditions:

   The concentration of the anti-TNF-α antibody is 90.0-110.0 mg/ml,

   The type of the stabilizer includes mannitol, sodium chloride, the concentration of the mannitol is 8.0-12.0 mg/ml, and the concentration of the sodium chloride is 6.0-7.0 mg/ml;

   The surfactant comprises 4.0-8.0 mg/ml polysorbate 80;

   The antibody composition has a pH of from 4.8 to 5.5, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 40-60 mmHg.
8. The antibody composition for TNF-α according to claim 7, wherein the species and content including the solute satisfy one or more of the following preferred conditions:

   The concentration of the anti-TNF-α antibody is 100 mg/ml;

   The concentration of mannitol is 10.0 mg/ml, and the concentration of the sodium chloride is 6.5 mg/ml;

   The concentration of the polysorbate 80 is 6.0 mg / ml;

   The antibody composition has a pH of 5.0 to 5.4, and the pH is adjusted by injecting carbon dioxide into the liquid component of the antibody composition at a pressure of 50 mmHg.
9. An antibody preparation for TNF-α, comprising: a closed container, and an antibody composition for TNF-α according to any one of claims 1 to 8 stored in a closed container, said composition It has a pH of 4.0-8.0.
10. Use of an antibody composition against TNF-α according to any one of claims 1 to 8 for the preparation of a medicament for the treatment of a TNF-α causing disorder.

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