WO2014178216A1 - Aqueous solution formulation and method for producing same - Google Patents

Aqueous solution formulation and method for producing same Download PDF

Info

Publication number
WO2014178216A1
WO2014178216A1 PCT/JP2014/054036 JP2014054036W WO2014178216A1 WO 2014178216 A1 WO2014178216 A1 WO 2014178216A1 JP 2014054036 W JP2014054036 W JP 2014054036W WO 2014178216 A1 WO2014178216 A1 WO 2014178216A1
Authority
WO
WIPO (PCT)
Prior art keywords
aqueous solution
etanercept
buffer
concentration
solution
Prior art date
Application number
PCT/JP2014/054036
Other languages
French (fr)
Japanese (ja)
Inventor
健作 松田
里保 中井
恒彦 山根
範彦 山脇
Original Assignee
ニプロ株式会社
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by ニプロ株式会社 filed Critical ニプロ株式会社
Publication of WO2014178216A1 publication Critical patent/WO2014178216A1/en

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/08—Solutions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/177—Receptors; Cell surface antigens; Cell surface determinants
    • A61K38/1793—Receptors; Cell surface antigens; Cell surface determinants for cytokines; for lymphokines; for interferons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/16—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing nitrogen, e.g. nitro-, nitroso-, azo-compounds, nitriles, cyanates
    • A61K47/18—Amines; Amides; Ureas; Quaternary ammonium compounds; Amino acids; Oligopeptides having up to five amino acids
    • A61K47/183—Amino acids, e.g. glycine, EDTA or aspartame
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
    • A61K47/12—Carboxylic acids; Salts or anhydrides thereof

Definitions

  • the present invention relates to an aqueous solution preparation and a method for producing the same, and more particularly to an aqueous solution preparation containing etanercept as an active ingredient and a method for producing the same.
  • the etanercept pharmaceutical composition is a glycoprotein comprising an extracellular domain portion of a human p75 tumor necrosis factor (Tumor Necrosis Factor (TNF)) receptor fused to the Fc region of human IgG1 (hereinafter referred to as “etanercept” in the present specification).
  • TNF Tumor Necrosis Factor
  • etanercept As an active ingredient, it captures TNF ⁇ / LT ⁇ produced excessively in the body (receptor binding reaction) and inhibits binding to receptors on the cell surface, thereby exhibiting anti-rheumatic and anti-inflammatory effects
  • Such an etanercept formulation is prepared, for example, as an aqueous solution formulation, and then sealed and stored in a syringe.
  • polypeptides such as etanercept are usually unstable in an aqueous solution and produce impurities composed of aggregates or cleaved bodies in the aqueous solution. These impurities can no longer capture TNF ⁇ / LT ⁇ , and no anti-rheumatic action or anti-inflammatory action is exhibited. Therefore, it is desired to develop a technique for suppressing the generation of impurities in an aqueous solution in an etanercept formulation.
  • Patent Document 1 In order to suppress the production of such impurities, it has been proposed that when L-arginine is contained as an aggregation inhibitor in an aqueous preparation containing etanercept, the production of aggregates which are one of the impurities of etanercept is suppressed.
  • the present inventors can improve the stability of an aqueous preparation containing etanercept by selecting a predetermined stabilizer and a buffer and adjusting the pH to a predetermined value.
  • the present invention was completed.
  • the present invention relates to an aqueous solution preparation containing etanercept, a stabilizer and a buffer, wherein the stabilizer is histidine or a salt thereof, the buffer is an organic acid or a salt thereof, and the pH is from 5.6. It is a formulation that is 6.4.
  • the organic acid or salt thereof is citric acid, citrate, acetic acid, acetate, tartaric acid, tartrate, and tris (hydroxymethyl) aminomethane, and tris (hydroxymethyl) aminomethane hydrochloride.
  • the buffer is contained at a concentration of 1 mM to 1 M with respect to the entire aqueous solution.
  • the stabilizer is contained at a concentration of 1 mM to 1 M with respect to the entire aqueous solution.
  • the aqueous solution formulation of the present invention further contains at least one isotonic agent selected from the group consisting of sodium chloride, sucrose, and glycine.
  • the present invention is also a method for producing an aqueous solution formulation, Preparing a buffer solution having a pH of 5.6 to 6.4 by dissolving histidine or a salt thereof and an organic acid or a salt thereof in water; Adding an etanercept stock solution to the buffer; Concentrating the stock solution to obtain a concentrate of the etanercept; and combining the concentrate with another buffer;
  • a method comprising
  • the organic acid or salt thereof is citric acid, citrate, acetic acid, acetate, tartaric acid, tartrate, and tris (hydroxymethyl) aminomethane, and tris (hydroxymethyl) aminomethane hydrochloride.
  • the isotonic agent is further dissolved in the step of preparing the buffer solution.
  • the tonicity agent is at least one compound selected from the group consisting of sodium chloride, sucrose, and glycine.
  • the present invention it is possible to suppress the generation of impurities composed of aggregates and / or cleaved bodies resulting from etanercept, which is an active ingredient, and improve the stability of the aqueous solution preparation itself. Due to the improved stability, the aqueous solution preparation of the present invention can be stored for a longer period of time.
  • FIG. 4 is a graph showing the ratio of aggregates and cleaved bodies to the total polypeptide at the start of storage when the pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 are stored in an incubator.
  • FIG. 5 is a graph showing the ratio of aggregates and cleaved bodies to the total polypeptide at one week after storage when the pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 are stored in an incubator. is there.
  • FIG. 2 is a graph showing the ratio of aggregates and cleaved bodies to total polypeptides at the time point after 2 weeks of storage when the pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 are stored in an incubator. is there.
  • FIG. 2 is a graph showing the ratio of aggregates and cleaved bodies to total polypeptides at the time point after 3 weeks of storage when the pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 are stored in an incubator. is there.
  • the aqueous solution preparation of the present invention is a pharmaceutical composition containing etanercept.
  • Etanercept constituting the present invention is an active ingredient in the present invention and is useful as an anti-rheumatoid arthritis drug, human IgG1 Fc region and human tumor necrosis factor type II receptor (TNFR-II) having a molecular weight of 75 kDa (p75) It is a glycoprotein (polypeptide) consisting of a dimer of subunits fused with the extracellular domain.
  • Etanercept can be produced by genetic recombination using Chinese hamster ovary cells (CHO). The etanercept subunit consists of 934 amino acids and has a molecular weight of about 150,000 daltons.
  • the content of etanercept in the aqueous solution preparation of the present invention is not particularly limited, but is, for example, 1 mg / mL to 150 mg / mL, preferably 5 mg / mL to 100 mg / mL. If the content of etanercept is less than 1 mg / mL, it may not function sufficiently as a pharmaceutical composition. If the content of etanercept exceeds 150 mg / mL, precipitation may occur in the preparation.
  • the aqueous solution preparation of the present invention contains a stabilizer and a buffer together with the etanercept.
  • the stabilizer in the present invention is histidine or a salt thereof.
  • histidine salts include, but are not limited to, histidine hydrochloride.
  • the stabilizer may be composed of such histidine or a salt thereof alone, or may be composed of a combination of two or more.
  • the concentration of the stabilizer in the aqueous solution preparation of the present invention is not necessarily limited, but is preferably 1 mM to 1 M, more preferably 10 mM to 100 mM, and even more preferably 25 mM to 50 mM, based on the entire aqueous solution. . If the concentration of the stabilizer is less than 1 mM, the generation of impurities of etanercept cannot be sufficiently suppressed, and there is a possibility that many aggregates and cleaved bodies are generated through storage of the aqueous solution preparation. When the concentration of the stabilizer exceeds 1M, the osmotic pressure is no longer too high and may not be suitable as a pharmaceutical composition.
  • the term “aggregate” used in the present specification refers to a high molecular weight polypeptide formed by aggregation of etanercept molecules.
  • the term “cleaved body” used in the present specification refers to a low molecular weight polypeptide consisting of a fragment obtained by cleaving a peptide bond at an arbitrary position in an etanercept molecule.
  • the buffer in the present invention is an organic acid or a salt thereof.
  • buffering agents include, but are not limited to, citric acid and its salts (eg, sodium citrate, sodium dihydrogen citrate, disodium citrate and potassium citrate), acetic acid and its salts (eg, ammonium acetate, Calcium acetate, potassium acetate and sodium acetate), tartaric acid and its salts (eg, sodium tartrate, sodium hydrogen tartrate and potassium sodium tartrate), tris (hydroxymethyl) aminomethane (Tris) and its salts (eg, tris (hydroxymethyl)) Aminomethane hydrochloride).
  • citric acid and its salts are particularly preferable.
  • the concentration of the buffering agent in the aqueous solution preparation of the present invention is not necessarily limited, but is preferably 1 mM to 1 M, more preferably 10 mM to 100 mM, and even more preferably 25 mM to 50 mM, based on the whole aqueous solution.
  • concentration of the buffering agent is less than 1 mM, the generation of impurities of etanercept cannot be sufficiently suppressed, and there is a possibility that many aggregates and cleaved bodies are generated through storage of the aqueous solution preparation. If the concentration of the buffer exceeds 1M, the osmotic pressure is no longer too high and may not be suitable as a pharmaceutical composition.
  • the aqueous solution preparation of the present invention is an aqueous solution in which the above etanercept, stabilizer and buffer are dissolved in water.
  • the aqueous solution preparation of the present invention can also be adjusted to a predetermined pH by adding sodium hydroxide, hydrochloric acid or the like to the aqueous solution of the buffer, stabilizer, tonicity agent and other additives as necessary. You may adjust.
  • the pH of the aqueous solution preparation of the present invention is 5.6 to 6.4, more preferably 5.7 to 6.3, and even more preferably 6.0.
  • the etanercept constituting the present invention greatly changes the production rate of aggregates and cleaved bodies as impurities, depending on the pH of the buffer solution to be dissolved. In particular, when it is tilted to a more basic side above pH 6.4, the formation of aggregates is promoted. On the other hand, when it is tilted to the acidic side from below pH 5.6, the formation of the cut body is promoted. Therefore, in the present invention, such a pH range is maintained.
  • the aqueous solution preparation of the present invention may also contain an isotonic agent as necessary.
  • isotonic agents include, but are not necessarily limited to, sodium chloride, sucrose and glycine, and combinations thereof.
  • the content of the tonicity agent is not necessarily limited, but is set so that the osmotic pressure of the entire aqueous solution is 140 mOsm to 780 mOsm.
  • aqueous solution preparation of the present invention may contain other pharmaceutically acceptable additives or excipients.
  • Additives and / or excipients are not necessarily limited, but include, for example, sugars or polyols (lactose, glycerol, xylitol, sorbitol, mannitol, maltose, inositol, trehalose, glucose); macromolecules (eg, serum albumin (bovine Serum albumin (BSA), human SA or recombinant HA), dextran, PVA, hydroxypropylmethylcellulose (HPMC), polyethyleneimine, gelatin, polyvinylpyrrolidone (PVP), polyoxyethylene (160) polyoxypropylene (30) glycol, Hydroxyethyl cellulose (HEC) lactic acid polymer); non-aqueous solvents (eg polyhydric alcohols such as polyethylene glycol, ethylene glycol, propylene glycol or glycerol) ), Dimethyl
  • the aqueous solution preparation of the present invention can be stored for a long time in the liquid itself or in a frozen state.
  • aqueous solution preparation of the present invention examples include rheumatoid arthritis, juvenile idiopathic arthritis, psoriatic arthritis, ankylosing spondylitis, Wegner's disease, Crohn's disease, chronic obstructive pulmonary disease (COPD), hepatitis C, endometriosis It can be used as a pharmaceutical composition for asthma, cachexia, psoriasis, atopic dermatitis and the like.
  • the dosage of the water-soluble preparation of the present invention depends on the kind and severity of the disease, the patient's age, weight and height, medical history, the degree of response to etanercept, etc. The administration period and the like can be appropriately selected.
  • histidine or a salt thereof and an organic acid or a salt thereof are dissolved in water to prepare a buffer solution having a pH of 5.6 to 6.4.
  • the mode of dissolution of histidine or a salt thereof (the stabilizer) and an organic acid or a salt thereof (the buffer) in water is not particularly limited. Further, the dissolution order of the stabilizer and the buffer is not particularly limited.
  • an isotonic agent in preparing this buffer solution, an isotonic agent, other additives, excipients and the like may be added as necessary in addition to the stabilizer and the buffer.
  • the pH of the buffer solution thus prepared is 5.6 to 6.4, more preferably 5.7 to 6.3, and even more preferably 6.0.
  • the osmotic pressure of the buffer solution is not necessarily limited, but is, for example, 140 mOsm to 780 mOsm, and preferably 250 mOsm to 315 mOsm.
  • the etanercept stock solution is added to the above-obtained buffer solution (one of them) to prepare a stock aqueous solution.
  • the addition mode of the etanercept stock solution to the buffer solution is not particularly limited, and any means commonly used by those skilled in the art can be used. Further, the temperature set at the time of addition is not particularly limited.
  • a concentrated solution in which etanercept is concentrated can be obtained by concentrating the raw aqueous solution.
  • Concentration of the raw aqueous solution is performed using means well known to those skilled in the art.
  • the conditions and temperature applied for the concentration are not particularly limited, and arbitrary conditions and temperatures are selected by those skilled in the art.
  • an ultrafiltration membrane having pores that cannot pass molecules having a certain molecular weight or more is used.
  • the raw aqueous solution is concentrated using, for example, a tangential flow type membrane cassette composed of a molecular filtration membrane having a fractional molecular weight of 30 kDa or less. If the amount is very small, a centrifugal filter unit composed of a tube provided with a desired molecular filtration membrane can be used. The filter unit may be further inserted into a filtrate tube to form a double-structure centrifugal ultrafiltration tube.
  • the concentration of etanercept increases without passing through the molecular filtration membrane, while unnecessary salts and buffer components originally contained in the etanercept stock solution can be efficiently filtered. In this way, a concentrated solution of etanercept in which the solution components constituting the etanercept stock solution are replaced with the buffer solution can be obtained.
  • Such an operation of concentration (substitution) may be repeated until the contents of the stabilizer and the buffering agent are set in advance in the concentrate.
  • the above buffer solution may be newly added to the obtained concentrated solution, and the concentration operation may be performed again.
  • the concentrated solution and the buffer solution may be mixed at a predetermined ratio so that the etanercept, the stabilizer, and the buffering agent have contents preset by those skilled in the art.
  • the aqueous solution obtained by combining may be filtered using a sterile filtration filter or the like as necessary.
  • the obtained aqueous solution is sealed in, for example, a sterilized vial, syringe, or screw cap tube.
  • Example 1 Histidine hydrochloride and glycine in a 50 mM citric acid monohydrate solution so that the concentration of histidine hydrochloride is 25 mM (4.79 g / L) and the concentration of glycine is 120 mM (9.0 g / L) Histidine hydrochloride and glycine were added to the dissolved solution and 50 mM trisodium citrate dihydrate solution, the concentration of histidine hydrochloride was 25 mM (4.79 g / L), and the concentration of glycine was 120 mM (9.0 g). / L) was mixed with an appropriate amount of each solution, and a buffer solution (liquid osmotic pressure 294 mOsm) adjusted to pH 6.0 by adding sodium hydroxide was prepared.
  • a buffer solution liquid osmotic pressure 294 mOsm
  • the buffer solution prepared above was added to the obtained etanercept concentrate, and the combined aqueous solution was frozen once and then melted. Thereafter, the solution is aseptically filtered using a 0.22 ⁇ m filter for aseptic filtration, and sealed in a 1.5 mL screw cap tube that has been sterilized by autoclaving at 121 ° C. for 20 minutes in advance, so that the concentration of etanercept is 47.
  • An aqueous solution of the pharmaceutical composition that was 1 mg / mL was obtained (Table 1).
  • pH of the pharmaceutical composition obtained by the present Example did not change with pH adjusted about the said buffer solution.
  • Example 2 Instead of the buffer prepared in Example 1, histidine hydrochloride and glycine were added to a 25 mM citrate monohydrate solution, the concentration of histidine hydrochloride was 25 mM (4.79 g / L), and the concentration of glycine was Histidine hydrochloride and glycine were added to a solution dissolved to 190 mM (14.25 g / L) and 25 mM trisodium citrate dihydrate solution, and the concentration of histidine hydrochloride was 25 mM (4.79 g / L).
  • a buffer solution liquid osmotic pressure 299 mOsm
  • sodium hydroxide adjusted to pH 6.0 by adding sodium hydroxide to each solution in an appropriate amount so that the glycine concentration is 190 mM (14.25 g / L).
  • a pharmaceutical composition having an etanercept concentration of 49.5 mg / mL was obtained in the same manner as in Example 1 except that was prepared and used (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by the present Example did not change with pH adjusted about the said buffer solution.
  • Example 3 Instead of the buffer solution prepared in Example 1, histidine hydrochloride, sodium chloride and sucrose were added to a 25 mM citric acid monohydrate solution, and the concentration of histidine hydrochloride was 25 mM (4.79 g / L).
  • histidine hydrochloride sodium chloride, and sucrose in a solution dissolved to a concentration of 80 mM (4.68 g / L) and a sucrose concentration of 10 mg / mL and a 25 mM trisodium citrate dihydrate solution was dissolved so that the concentration of histidine hydrochloride was 25 mM (4.79 g / L), the concentration of sodium chloride was 80 mM (4.68 g / L), and the concentration of sucrose was 10 mg / mL.
  • a suitable amount of the solution was mixed, and sodium hydroxide was added to adjust the pH to 6.0 (liquid osmotic pressure 294 mOsm).
  • the concentration of etanercept to obtain a pharmaceutical composition is 47.7 mg / mL (Table 1).
  • pH of the pharmaceutical composition obtained by the present Example did not change with pH adjusted about the said buffer solution.
  • Example 4 instead of the buffer prepared in Example 1, histidine hydrochloride, sodium chloride, and sucrose were added to a 50 mM citric acid monohydrate solution, and the concentration of histidine hydrochloride was 25 mM (4.79 g / L).
  • a solution in which the concentration of sodium is 40 mM (2.34 g / L) and the concentration of sucrose is 10 mg / mL and 50 mM trisodium citrate dihydrate solution are mixed with histidine hydrochloride, sodium chloride, And sucrose so that the concentration of histidine hydrochloride is 25 mM (4.79 g / L), the concentration of sodium chloride is 40 mM (2.34 g / L), and the concentration of sucrose is 10 mg / mL.
  • Buffer solution liquid osmotic pressure 288 mOsm
  • the concentration of etanercept to obtain a pharmaceutical composition is 47.7 mg / mL (Table 1).
  • pH of the pharmaceutical composition obtained by the present Example did not change with pH adjusted about the said buffer solution.
  • Example 1 (Comparative Example 1) Instead of the buffer prepared in Example 1, arginine hydrochloride, sodium chloride and sucrose were added to a 25 mM sodium dihydrogen phosphate solution, and the concentration of arginine hydrochloride was 25 mM (5.27 g / L). Arginine hydrochloride, sodium chloride and sucrose were added to a solution in which the concentration was 100 mM (5.84 g / L) and the sucrose concentration was 10 mg / mL and a 25 mM sodium monohydrogen phosphate dihydrate solution.
  • a buffer solution liquid osmotic pressure of 341 mOsm adjusted to pH 6.5 by adding sodium hydroxide was prepared.
  • the concentration of etanercept to obtain a pharmaceutical composition is 46.7 mg / mL (Table 1).
  • pH of the pharmaceutical composition obtained by this comparative example did not change with pH adjusted about the said buffer solution.
  • the eight types of pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 were stored in an incubator for 3 weeks at a set temperature of 37 ° C. and protected from light.
  • SDS-PAGE Prepare etanercept reduced with 2-mercaptoethanol as a reducing agent and non-reduced (non-reduced) etanercept, and separate polypeptide fragments by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). And stained by silver staining.
  • the band near 75 kDa is the main band, the band with shorter migration distance (above) is cut with the aggregate band, and the band with longer migration distance (below) is cut. It was a body band.
  • the band near 150 kDa is the main band, the band having a shorter migration distance (located above) is the aggregate band, and the band having a longer migration distance (located below). The band was a cut band.
  • a high-intensity band was confirmed in the vicinity of the 75 kDa index, and a high-intensity band was also confirmed in the vicinity of 25 kDa to 37 kDa.
  • a high-intensity band was confirmed in the vicinity of the 150 kDa index, and a high-intensity band was also confirmed in the vicinity of 75 kDa to 100 kDa.
  • Size exclusion chromatography For the eight types of pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4, size exclusion was performed at 4 time points at the start of incubator storage, 1 week after storage, 2 weeks after storage, and 3 weeks after storage. Tested by chromatography. In the test, a sample solution (50 ⁇ L each) prepared by diluting the stored pharmaceutical composition with the buffer used in each Example or Comparative Example so that the etanercept concentration was about 0.2 mg / mL. Used as a test sample.
  • the area of the main peak of the obtained chromatography (A1), the total area of individual peaks eluting before the main peak (A2), and the total area of individual peaks excluding the solvent peak eluting after the main peak ( A3) was measured by an automatic integration method, and the amounts of aggregates and cut bodies, which are impurities of etanercept, were calculated by the following equation.
  • the liquidity of the buffer solution prepared in Comparative Example 3 was on the most acidic side (pH 5.5), and the obtained etanercept was cut most.
  • the buffer solution prepared in Comparative Example 4 was most basic (pH 6.5), and the resulting etanercept aggregate amount was the largest.
  • the storage stability of etanercept in the preparation was effectively improved by adjusting the liquidity of the buffer solution within the range of pH 5.6 to 6.4.
  • the stability of etanercept which is an active ingredient, can be increased.
  • long-term preservation as a pharmaceutical composition can be enhanced.
  • examples of the aqueous solution preparation of the present invention include rheumatoid arthritis, juvenile idiopathic arthritis, psoriatic arthritis, ankylosing spondylitis, Wegner's disease, Crohn's disease, chronic obstructive pulmonary disease (COPD), hepatitis C, endometriosis It is useful as a pharmaceutical composition for asthma, cachexia, psoriasis and atopic dermatitis.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Medicinal Chemistry (AREA)
  • Epidemiology (AREA)
  • Engineering & Computer Science (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Oil, Petroleum & Natural Gas (AREA)
  • Rheumatology (AREA)
  • Organic Chemistry (AREA)
  • Cell Biology (AREA)
  • Zoology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Immunology (AREA)
  • Pain & Pain Management (AREA)
  • Dermatology (AREA)
  • Medicinal Preparation (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

Provided are an aqueous solution formulation which comprises etanercept as an active ingredient, and a method for producing the same. In this aqueous solution formulation, which comprises etanercept, a stabilizer and a buffer, the stabilizer is histidine or a salt thereof, the buffer is an organic acid or a salt thereof, and pH is 5.6 to 6.4. This aqueous solution formulation is stable for a long period of time and is capable of providing excellent shelf life by suppressing generation of both an aggregate and a truncated body caused by the etanercept.

Description

水溶液製剤およびその製造方法Aqueous preparation and method for producing the same
　本発明は、水溶液製剤およびその製造方法に関し、より詳細には有効成分としてエタネルセプトを含有する水溶液製剤およびその製造方法に関する。 The present invention relates to an aqueous solution preparation and a method for producing the same, and more particularly to an aqueous solution preparation containing etanercept as an active ingredient and a method for producing the same.
　エタネルセプト医薬組成物は、ヒトＩｇＧ１のＦｃ領域に融合したヒトｐ７５腫瘍壊死因子［Ｔｕｍｏｒ　Ｎｅｃｒｏｓｉｓ　Ｆａｃｔｏｒ（ＴＮＦ）］受容体の細胞外ドメイン部分からなる糖蛋白質（以下、本願明細書において「エタネルセプト」という）を有効成分として含有し、体内で過剰に産生されたＴＮＦα／ＬＴαを捕捉（レセプター結合反応）して、細胞表面のレセプターとの結合を阻害することにより、抗リウマチ作用や抗炎症作用を発揮する、医薬組成物である（非特許文献１）。 The etanercept pharmaceutical composition is a glycoprotein comprising an extracellular domain portion of a human p75 tumor necrosis factor (Tumor Necrosis Factor (TNF)) receptor fused to the Fc region of human IgG1 (hereinafter referred to as “etanercept” in the present specification). As an active ingredient, it captures TNFα / LTα produced excessively in the body (receptor binding reaction) and inhibits binding to receptors on the cell surface, thereby exhibiting anti-rheumatic and anti-inflammatory effects It is a pharmaceutical composition (Non-Patent Document 1).
　このようなエタネルセプト製剤は、例えば、水溶液製剤として調製された後、シリンジに封入して保管される。 Such an etanercept formulation is prepared, for example, as an aqueous solution formulation, and then sealed and stored in a syringe.
　しかし、エタネルセプトのような多くのポリペプチドは、通常、水溶液中では不安定であり、水溶液中で凝集体あるいは切断体からなる不純物を生成することが知られている。そして、これらの不純物はもはやＴＮＦα／ＬＴαを捕捉し得ず、抗リウマチ作用や抗炎症作用は発揮されない。そのため、エタネルセプト製剤における水溶液中での不純物の生成を抑制する技術開発が所望されている。 However, it is known that many polypeptides such as etanercept are usually unstable in an aqueous solution and produce impurities composed of aggregates or cleaved bodies in the aqueous solution. These impurities can no longer capture TNFα / LTα, and no anti-rheumatic action or anti-inflammatory action is exhibited. Therefore, it is desired to develop a technique for suppressing the generation of impurities in an aqueous solution in an etanercept formulation.
　当該不純物の生成を抑制するために、エタネルセプトを含有する水性製剤中に凝集抑制剤としてＬ－アルギニンを含有させると、エタネルセプトの不純物の一つである凝集体の生成が抑制されることが提案されている（特許文献１）。 In order to suppress the production of such impurities, it has been proposed that when L-arginine is contained as an aggregation inhibitor in an aqueous preparation containing etanercept, the production of aggregates which are one of the impurities of etanercept is suppressed. (Patent Document 1).
　しかし、前述の通り、エタネルセプトは水溶液中では凝集体だけではなく、切断体をも生成する。特許文献１の方法では切断体の生成が抑制されないため、不純物の生成を抑制する技術としては不十分である。 However, as described above, etanercept produces not only aggregates but also cut bodies in an aqueous solution. Since the method of Patent Document 1 does not suppress the generation of the cut body, it is insufficient as a technique for suppressing the generation of impurities.
　そこで、エタネルセプトの不純物である凝集体および切断体のいずれの生成をも抑制する医薬組成物の提供が所望されている。 Therefore, it is desired to provide a pharmaceutical composition that suppresses the formation of both aggregates and cleaved bodies, which are impurities of etanercept.
特表２００５－５２７５０３号公報JP 2005-527503 Gazette
　本発明は、上記問題の解決を課題とするものであり、その目的とするところは、有効成分としてエタネルセプトを含有する水溶液製剤において、エタネルセプトの不純物である凝集体および切断体の生成が抑制された、安定性に優れた水溶液製剤およびその製造方法を提供することにある。 An object of the present invention is to solve the above problems, and the object of the present invention is to suppress the formation of aggregates and cut bodies that are impurities of etanercept in an aqueous solution preparation containing etanercept as an active ingredient. Another object of the present invention is to provide an aqueous solution preparation excellent in stability and a method for producing the same.
　本発明者らは、上記課題を解決するために、所定の安定化剤および緩衝剤を選択し、かつ所定のｐＨに調整することにより、エタネルセプトを含有する水溶液製剤の安定性を向上させ得ることを見出し、本発明を完成させた。 In order to solve the above-mentioned problems, the present inventors can improve the stability of an aqueous preparation containing etanercept by selecting a predetermined stabilizer and a buffer and adjusting the pH to a predetermined value. The present invention was completed.
　本発明は、エタネルセプト、安定化剤および緩衝剤を含有する水溶液製剤であって、安定化剤がヒスチジンまたはその塩であり、緩衝剤が有機酸またはその塩であり、そしてｐＨが５．６から６．４である、製剤である。 The present invention relates to an aqueous solution preparation containing etanercept, a stabilizer and a buffer, wherein the stabilizer is histidine or a salt thereof, the buffer is an organic acid or a salt thereof, and the pH is from 5.6. It is a formulation that is 6.4.
　１つの実施形態では、上記有機酸またはその塩は、クエン酸、クエン酸塩、酢酸、酢酸塩、酒石酸、酒石酸塩、およびトリス（ヒドロキシメチル）アミノメタン、およびトリス（ヒドロキシメチル）アミノメタン塩酸塩からなる群から選択される少なくとも１種の化合物である。 In one embodiment, the organic acid or salt thereof is citric acid, citrate, acetic acid, acetate, tartaric acid, tartrate, and tris (hydroxymethyl) aminomethane, and tris (hydroxymethyl) aminomethane hydrochloride. At least one compound selected from the group consisting of:
　１つの実施形態では、上記緩衝剤を、水溶液全体に対し１ｍＭから１Ｍの濃度で含有する。 In one embodiment, the buffer is contained at a concentration of 1 mM to 1 M with respect to the entire aqueous solution.
　１つの実施形態では、上記安定化剤を、水溶液全体に対し１ｍＭから１Ｍの濃度で含有する。 In one embodiment, the stabilizer is contained at a concentration of 1 mM to 1 M with respect to the entire aqueous solution.
　１つの実施形態では、本発明の水溶液製剤は、さらに、塩化ナトリウム、スクロース、およびグリシンからなる群から選択される少なくとも１種の等張化剤を含有する。 In one embodiment, the aqueous solution formulation of the present invention further contains at least one isotonic agent selected from the group consisting of sodium chloride, sucrose, and glycine.
　本発明はまた、水溶液製剤の製造方法であって、
　ヒスチジンまたはその塩と、有機酸またはその塩とを水に溶解させて、ｐＨが５．６から６．４である緩衝液を調製する工程；
　該緩衝液にエタネルセプト原液を添加する工程；
　該原液を濃縮することにより、該エタネルセプトの濃縮液を得る工程；および
　該濃縮液を別の該緩衝液と合わせる工程；
を包含する、方法である。 The present invention is also a method for producing an aqueous solution formulation,
Preparing a buffer solution having a pH of 5.6 to 6.4 by dissolving histidine or a salt thereof and an organic acid or a salt thereof in water;
Adding an etanercept stock solution to the buffer;
Concentrating the stock solution to obtain a concentrate of the etanercept; and combining the concentrate with another buffer;
A method comprising
　１つの実施形態では、上記有機酸またはその塩は、クエン酸、クエン酸塩、酢酸、酢酸塩、酒石酸、酒石酸塩、およびトリス（ヒドロキシメチル）アミノメタン、およびトリス（ヒドロキシメチル）アミノメタン塩酸塩からなる群から選択される少なくとも１種の化合物である。 In one embodiment, the organic acid or salt thereof is citric acid, citrate, acetic acid, acetate, tartaric acid, tartrate, and tris (hydroxymethyl) aminomethane, and tris (hydroxymethyl) aminomethane hydrochloride. At least one compound selected from the group consisting of:
　１つの実施形態では、上記緩衝液を調製する工程において、さらに等張化剤が溶解される。 In one embodiment, the isotonic agent is further dissolved in the step of preparing the buffer solution.
　さらなる実施形態では、上記等張化剤は、塩化ナトリウム、スクロース、およびグリシンからなる群から選択される少なくとも１種の化合物である。 In a further embodiment, the tonicity agent is at least one compound selected from the group consisting of sodium chloride, sucrose, and glycine.
　本発明によれば、有効成分であるエタネルセプトに起因する凝集体および／または切断体から構成される不純物の生成を抑制して、水溶液製剤自体の安定性を向上させることができる。当該安定性の向上により、本発明の水溶液製剤はより長期間の保管が可能である。 According to the present invention, it is possible to suppress the generation of impurities composed of aggregates and / or cleaved bodies resulting from etanercept, which is an active ingredient, and improve the stability of the aqueous solution preparation itself. Due to the improved stability, the aqueous solution preparation of the present invention can be stored for a longer period of time.
実施例１～４および比較例１～４で得られた医薬組成物をインキュベータで保存した際の、保存開始時の時点での凝集体および切断体の全ポリペプチドに占める割合を表すグラフである。FIG. 4 is a graph showing the ratio of aggregates and cleaved bodies to the total polypeptide at the start of storage when the pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 are stored in an incubator. . 実施例１～４および比較例１～４で得られた医薬組成物をインキュベータで保存した際の、保存１週間後の時点での凝集体および切断体の全ポリペプチドに占める割合を表すグラフである。FIG. 5 is a graph showing the ratio of aggregates and cleaved bodies to the total polypeptide at one week after storage when the pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 are stored in an incubator. is there. 実施例１～４および比較例１～４で得られた医薬組成物をインキュベータで保存した際の、保存２週間後の時点での凝集体および切断体の全ポリペプチドに占める割合を表すグラフである。FIG. 2 is a graph showing the ratio of aggregates and cleaved bodies to total polypeptides at the time point after 2 weeks of storage when the pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 are stored in an incubator. is there. 実施例１～４および比較例１～４で得られた医薬組成物をインキュベータで保存した際の、保存３週間後の時点での凝集体および切断体の全ポリペプチドに占める割合を表すグラフである。FIG. 2 is a graph showing the ratio of aggregates and cleaved bodies to total polypeptides at the time point after 3 weeks of storage when the pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 are stored in an incubator. is there.
（水溶液製剤）
　まず、本発明の水溶液製剤について説明する。 (Aqueous solution formulation)
First, the aqueous solution preparation of the present invention will be described.
　本発明の水溶液製剤は、エタネルセプトを含有する医薬組成物である。 The aqueous solution preparation of the present invention is a pharmaceutical composition containing etanercept.
　本発明を構成するエタネルセプトは、本発明における有効成分であって、抗関節リウマチ薬として有用な、ヒトＩｇＧ１のＦｃ領域と分子量７５ｋＤａ（ｐ７５）のヒト腫瘍壊死因子ＩＩ型受容体（ＴＮＦＲ－ＩＩ）の細胞外ドメインが融合したサブユニットの二量体からなる糖蛋白質（ポリペプチド）である。エタネルセプトは、チャイニーズハムスター卵巣細胞（ＣＨＯ）を利用した遺伝子組換えにより産生され得る。エタネルセプトサブユニットは、９３４アミノ酸からなり、約１５０，０００ダルトンの分子量を有する。 Etanercept constituting the present invention is an active ingredient in the present invention and is useful as an anti-rheumatoid arthritis drug, human IgG1 Fc region and human tumor necrosis factor type II receptor (TNFR-II) having a molecular weight of 75 kDa (p75) It is a glycoprotein (polypeptide) consisting of a dimer of subunits fused with the extracellular domain. Etanercept can be produced by genetic recombination using Chinese hamster ovary cells (CHO). The etanercept subunit consists of 934 amino acids and has a molecular weight of about 150,000 daltons.
　本発明の水溶液製剤におけるエタネルセプトの含有量は、特に限定されないが、例えば、１ｍｇ／ｍＬ～１５０ｍｇ／ｍＬであり、好ましくは５ｍｇ／ｍＬ～１００ｍｇ／ｍＬである。エタネルセプトの含有量が、１ｍｇ／ｍＬ未満であると、医薬組成物として充分に機能しないおそれがある。エタネルセプトの含有量が、１５０ｍｇ／ｍＬを上回ると、製剤中で沈殿を生じるおそれがある。 The content of etanercept in the aqueous solution preparation of the present invention is not particularly limited, but is, for example, 1 mg / mL to 150 mg / mL, preferably 5 mg / mL to 100 mg / mL. If the content of etanercept is less than 1 mg / mL, it may not function sufficiently as a pharmaceutical composition. If the content of etanercept exceeds 150 mg / mL, precipitation may occur in the preparation.
　本発明の水溶液製剤は、当該エタネルセプトとともに安定化剤および緩衝剤を含有する。 The aqueous solution preparation of the present invention contains a stabilizer and a buffer together with the etanercept.
　本発明における安定化剤はヒスチジンまたはその塩である。ヒスチジンの塩の例としては、特に限定されないが、ヒスチジンの塩酸塩が挙げられる。本発明において、安定化剤は、このようなヒスチジンまたはその塩の単独から構成されていてもよく、２種以上を組合せて構成されていてもよい。 The stabilizer in the present invention is histidine or a salt thereof. Examples of histidine salts include, but are not limited to, histidine hydrochloride. In the present invention, the stabilizer may be composed of such histidine or a salt thereof alone, or may be composed of a combination of two or more.
　本発明の水溶液製剤における安定化剤の濃度は、必ずしも限定されないが、水溶液全体を基準として、好ましくは１ｍＭ～１Ｍであり、より好ましくは１０ｍＭ～１００ｍＭであり、さらにより好ましくは２５ｍＭ～５０ｍＭである。安定化剤の濃度が１ｍＭ未満であると、エタネルセプトの不純物の生成を充分に抑制することができず、水溶液製剤の保管を通じて凝集体や切断体が多く生成するおそれがある。安定化剤の濃度が１Ｍを超えると、もはや浸透圧が過度に高く、医薬組成物として適切でなくなるおそれがある。 The concentration of the stabilizer in the aqueous solution preparation of the present invention is not necessarily limited, but is preferably 1 mM to 1 M, more preferably 10 mM to 100 mM, and even more preferably 25 mM to 50 mM, based on the entire aqueous solution. . If the concentration of the stabilizer is less than 1 mM, the generation of impurities of etanercept cannot be sufficiently suppressed, and there is a possibility that many aggregates and cleaved bodies are generated through storage of the aqueous solution preparation. When the concentration of the stabilizer exceeds 1M, the osmotic pressure is no longer too high and may not be suitable as a pharmaceutical composition.
　ここで、本明細書中に用いられる用語「凝集体」とは、エタネルセプト分子が凝集してできた高分子量のポリペプチドを指して言う。また、本明細書中に用いられる用語「切断体」とは、エタネルセプト分子内の任意の箇所におけるペプチド結合が切断して得られた断片からなる低分子量のポリペプチドを指して言う。 Here, the term “aggregate” used in the present specification refers to a high molecular weight polypeptide formed by aggregation of etanercept molecules. In addition, the term “cleaved body” used in the present specification refers to a low molecular weight polypeptide consisting of a fragment obtained by cleaving a peptide bond at an arbitrary position in an etanercept molecule.
　本発明における緩衝剤は有機酸またはその塩である。緩衝剤の例としては、特に限定されないが、クエン酸およびその塩（例えば、クエン酸ナトリウム、クエン酸二水素ナトリウム、クエン酸二ナトリウムおよびクエン酸カリウム）、酢酸およびその塩（例えば、酢酸アンモニウム、酢酸カルシウム、酢酸カリウムおよび酢酸ナトリウム）、酒石酸およびその塩（例えば、酒石酸ナトリウム、酒石酸水素ナトリウムおよび酒石酸ナトリウムカリウム）、トリス（ヒドロキシメチル）アミノメタン（Ｔｒｉｓ）およびその塩（例えば、トリス（ヒドロキシメチル）アミノメタン塩酸塩）が挙げられる。緩衝剤としては、特にクエン酸およびその塩が好ましい。 The buffer in the present invention is an organic acid or a salt thereof. Examples of buffering agents include, but are not limited to, citric acid and its salts (eg, sodium citrate, sodium dihydrogen citrate, disodium citrate and potassium citrate), acetic acid and its salts (eg, ammonium acetate, Calcium acetate, potassium acetate and sodium acetate), tartaric acid and its salts (eg, sodium tartrate, sodium hydrogen tartrate and potassium sodium tartrate), tris (hydroxymethyl) aminomethane (Tris) and its salts (eg, tris (hydroxymethyl)) Aminomethane hydrochloride). As the buffer, citric acid and its salts are particularly preferable.
　本発明の水溶液製剤における緩衝剤の濃度は、必ずしも限定されないが、水溶液全体を基準として、好ましくは１ｍＭ～１Ｍであり、より好ましくは１０ｍＭ～１００ｍＭであり、さらにより好ましくは２５ｍＭ～５０ｍＭである。緩衝剤の濃度が１ｍＭ未満であると、エタネルセプトの不純物の生成を充分に抑制することができず、水溶液製剤の保管を通じて凝集体や切断体が多く生成するおそれがある。緩衝剤の濃度が１Ｍを超えると、もはや浸透圧が過度に高く、医薬組成物として適切でなくなるおそれがある。 The concentration of the buffering agent in the aqueous solution preparation of the present invention is not necessarily limited, but is preferably 1 mM to 1 M, more preferably 10 mM to 100 mM, and even more preferably 25 mM to 50 mM, based on the whole aqueous solution. When the concentration of the buffering agent is less than 1 mM, the generation of impurities of etanercept cannot be sufficiently suppressed, and there is a possibility that many aggregates and cleaved bodies are generated through storage of the aqueous solution preparation. If the concentration of the buffer exceeds 1M, the osmotic pressure is no longer too high and may not be suitable as a pharmaceutical composition.
　本発明の水溶液製剤は、上記エタネルセプト、安定化剤および緩衝剤を水に溶解させてなる水溶液である。 The aqueous solution preparation of the present invention is an aqueous solution in which the above etanercept, stabilizer and buffer are dissolved in water.
　本発明の水溶液製剤はまた、上記緩衝剤、安定化剤、等張化剤およびその他の添加剤の水溶液に対し、必要に応じて水酸化ナトリウム、塩酸などを添加することにより、所定のｐＨに調整してもよい。本発明の水溶液製剤のｐＨは、５．６～６．４であり、より好ましくは５．７～６．３であり、さらにより好ましくは６．０である。本発明を構成するエタネルセプトは、溶解される緩衝液のｐＨによっても、その不純物である凝集体および切断体の生成率を大きく変化させる。特に、ｐＨ６．４を上回るより塩基性側に傾けると、凝集体の生成が促進される。一方、ｐＨ５．６を下回るより酸性側に傾けると、切断体の生成が促進される。したがって、本発明においてはこのようなｐＨ範囲が保持されている。 The aqueous solution preparation of the present invention can also be adjusted to a predetermined pH by adding sodium hydroxide, hydrochloric acid or the like to the aqueous solution of the buffer, stabilizer, tonicity agent and other additives as necessary. You may adjust. The pH of the aqueous solution preparation of the present invention is 5.6 to 6.4, more preferably 5.7 to 6.3, and even more preferably 6.0. The etanercept constituting the present invention greatly changes the production rate of aggregates and cleaved bodies as impurities, depending on the pH of the buffer solution to be dissolved. In particular, when it is tilted to a more basic side above pH 6.4, the formation of aggregates is promoted. On the other hand, when it is tilted to the acidic side from below pH 5.6, the formation of the cut body is promoted. Therefore, in the present invention, such a pH range is maintained.
　本発明の水溶液製剤はまた、必要に応じて等張化剤を含有していてもよい。等張化剤の例としては、必ずしも限定されないが、塩化ナトリウム、スクロースおよびグリシン、ならびにこれらの組合せが挙げられる。等張化剤の含有量もまた、必ずしも限定されないが、水溶液全体の浸透圧が、１４０ｍＯｓｍ～７８０ｍＯｓｍとなるよう設定される。 The aqueous solution preparation of the present invention may also contain an isotonic agent as necessary. Examples of isotonic agents include, but are not necessarily limited to, sodium chloride, sucrose and glycine, and combinations thereof. The content of the tonicity agent is not necessarily limited, but is set so that the osmotic pressure of the entire aqueous solution is 140 mOsm to 780 mOsm.
　本発明の水溶液製剤は、薬学的に許容され得る他の添加剤または賦形剤を含有していてもよい。添加剤および／または賦形剤は、必ずしも限定されないが、例えば、糖またはポリオール類（ラクトース、グリセロール、キシリトール、ソルビトール、マンニトール、マルトース、イノシトール、トレハロース、グルコース）；高分子（例えば、血清アルブミン（ウシ血清アルブミン（ＢＳＡ）、ヒトＳＡまたは組換えＨＡ）、デキストラン、ＰＶＡ、ヒドロキシプロピルメチルセルロース（ＨＰＭＣ）、ポリエチレンイミン、ゼラチン、ポリビニルピロリドン（ＰＶＰ）、ポリオキシエチレン（１６０）ポリオキシプロピレン（３０）グリコール、ヒドロキシエチルセルロース（ＨＥＣ）乳酸重合体）；非水性溶媒（例えば、ポリエチレングリコール、エチレングリコール、プロピレングリコールまたはグリセロールのような多価アルコール）、ジメチルスルホキシド（ＤＭＳＯ）およびジメチルホルムアミド（ＤＭＦ））；アミノ酸（例えばアスパラギン酸、メチオニン、プロリン、Ｌ－セリン、グルタミン酸またはその塩、アラニン、フェニルアラニン、リシン塩酸塩、サルコシンおよびγ－アミノ酪酸）；界面活性剤（例えばＴｗｅｅｎ８０、Ｔｗｅｅｎ２０、ＳＤＳ、ポリソルベート、およびポリオキシエチレンコポリマー）；その他の添加剤（例えば、リン酸カリウム、酢酸ナトリウム、硫酸アンモニウム、硫酸マグネシウム、硫酸ナトリウム、トリメチルアミンＮ－オキシド、ベタイン、金属イオン（例えば、亜鉛、銅、カルシウム、マンガン及びマグネシウム）、ＣＨＡＰＳ、モノラウレート、および２－Ｏ－β－マンノグリセレート）；またはこれらの組合せが挙げられる。当該添加剤および／または賦形剤の含有量は、特に限定されず、上記エタネルセプト、安定化剤および緩衝剤による効果を阻害しない程度において、当業者によって適切な量が設定され得る。 The aqueous solution preparation of the present invention may contain other pharmaceutically acceptable additives or excipients. Additives and / or excipients are not necessarily limited, but include, for example, sugars or polyols (lactose, glycerol, xylitol, sorbitol, mannitol, maltose, inositol, trehalose, glucose); macromolecules (eg, serum albumin (bovine Serum albumin (BSA), human SA or recombinant HA), dextran, PVA, hydroxypropylmethylcellulose (HPMC), polyethyleneimine, gelatin, polyvinylpyrrolidone (PVP), polyoxyethylene (160) polyoxypropylene (30) glycol, Hydroxyethyl cellulose (HEC) lactic acid polymer); non-aqueous solvents (eg polyhydric alcohols such as polyethylene glycol, ethylene glycol, propylene glycol or glycerol) ), Dimethyl sulfoxide (DMSO) and dimethylformamide (DMF)); amino acids (eg aspartic acid, methionine, proline, L-serine, glutamic acid or salts thereof, alanine, phenylalanine, lysine hydrochloride, sarcosine and γ-aminobutyric acid) Surfactants (eg Tween 80, Tween 20, SDS, polysorbate and polyoxyethylene copolymers); other additives (eg potassium phosphate, sodium acetate, ammonium sulfate, magnesium sulfate, sodium sulfate, trimethylamine N-oxide, betaine, Metal ions (eg, zinc, copper, calcium, manganese and magnesium), CHAPS, monolaurate, and 2-O-β-mannoglycerate); or combinations thereof It is below. The content of the additive and / or excipient is not particularly limited, and an appropriate amount can be set by those skilled in the art to the extent that the effects of the etanercept, stabilizer and buffer are not inhibited.
　本発明の水溶液製剤は、液体自体の状態でまたは凍結した状態で長期間保存することができる。 The aqueous solution preparation of the present invention can be stored for a long time in the liquid itself or in a frozen state.
　本発明の水溶液製剤は、例えば、関節リウマチ、若年性特発性関節炎、乾癬性関節炎、強直性脊椎炎、ヴェーグナー病、クローン病、慢性閉塞性肺疾患（ＣＯＰＤ）、Ｃ型肝炎、子宮内膜症、喘息、悪液質、乾癬及びアトピー性皮膚炎などのための医薬組成物として使用され得る。本発明の水溶性製剤の用量は、疾患の種類および重篤度、患者の年齢、体重、および身長、病歴、ならびにエタネルセプトに対する応答の程度等に依存して、医師により適切な量、投与方法、投与期間等が適宜選択され得る。 Examples of the aqueous solution preparation of the present invention include rheumatoid arthritis, juvenile idiopathic arthritis, psoriatic arthritis, ankylosing spondylitis, Wegner's disease, Crohn's disease, chronic obstructive pulmonary disease (COPD), hepatitis C, endometriosis It can be used as a pharmaceutical composition for asthma, cachexia, psoriasis, atopic dermatitis and the like. The dosage of the water-soluble preparation of the present invention depends on the kind and severity of the disease, the patient's age, weight and height, medical history, the degree of response to etanercept, etc. The administration period and the like can be appropriately selected.
　（水溶液製剤の製造方法）
　次に、本発明の水溶液製剤の製造方法の一例について説明する。 (Method for producing aqueous solution preparation)
Next, an example of the manufacturing method of the aqueous solution formulation of this invention is demonstrated.
　本発明の製造方法においては、まず、ヒスチジンまたはその塩と、有機酸またはその塩とが水に溶解され、ｐＨが５．６から６．４である緩衝液が調製される。 In the production method of the present invention, first, histidine or a salt thereof and an organic acid or a salt thereof are dissolved in water to prepare a buffer solution having a pH of 5.6 to 6.4.
　水（例えば、純水、蒸留水、またはイオン交換水）への、ヒスチジンまたはその塩（上記安定化剤）および有機酸またはその塩（上記緩衝剤）の溶解の様式は特に限定されない。また、当該安定化剤および緩衝剤の溶解順序も特に限定されない。 The mode of dissolution of histidine or a salt thereof (the stabilizer) and an organic acid or a salt thereof (the buffer) in water (for example, pure water, distilled water, or ion exchange water) is not particularly limited. Further, the dissolution order of the stabilizer and the buffer is not particularly limited.
　この緩衝液の調製にあたっては、上記安定化剤および緩衝剤以外に、必要に応じて等張化剤、他の添加剤、賦形剤等が添加されてもよい。 In preparing this buffer solution, an isotonic agent, other additives, excipients and the like may be added as necessary in addition to the stabilizer and the buffer.
　このようにして調製された緩衝液のｐＨは、５．６～６．４であり、より好ましくは５．７～６．３であり、さらにより好ましくは６．０である。さらに当該緩衝液の浸透圧は、必ずしも限定されないが、例えば、１４０ｍＯｓｍ～７８０ｍＯｓｍであり、好ましくは２５０ｍＯｓｍ～３１５ｍＯｓｍである。なお、この調製された緩衝液は、その後の工程に使用するために、予め少なくとも２つに分けておくか、あるいは予め少なくとも２種類を別々に調製しておくことが好ましい。 The pH of the buffer solution thus prepared is 5.6 to 6.4, more preferably 5.7 to 6.3, and even more preferably 6.0. Further, the osmotic pressure of the buffer solution is not necessarily limited, but is, for example, 140 mOsm to 780 mOsm, and preferably 250 mOsm to 315 mOsm. In addition, it is preferable to divide this prepared buffer solution into at least two beforehand, or to prepare at least two types separately beforehand, in order to use it for a subsequent process.
　次いで、上記得られた緩衝液（の１つ）にエタネルセプト原液が添加され、原水溶液が調製される。 Next, the etanercept stock solution is added to the above-obtained buffer solution (one of them) to prepare a stock aqueous solution.
　緩衝液へのエタネルセプト原液の添加様式は特に限定されず、当業者が通常用いる任意の手段を使用することができる。さらに添加の際に設定される温度もまた特に限定されない。 The addition mode of the etanercept stock solution to the buffer solution is not particularly limited, and any means commonly used by those skilled in the art can be used. Further, the temperature set at the time of addition is not particularly limited.
　その後、上記原水溶液を濃縮することにより、エタネルセプトが濃縮された濃縮液を得ることができる。 Thereafter, a concentrated solution in which etanercept is concentrated can be obtained by concentrating the raw aqueous solution.
　原水溶液の濃縮は、当業者に周知の手段を用いて行われる。濃縮にあたって付与される条件および温度もまた特に限定されず、当業者によって任意の条件および温度が選択される。 Concentration of the raw aqueous solution is performed using means well known to those skilled in the art. The conditions and temperature applied for the concentration are not particularly limited, and arbitrary conditions and temperatures are selected by those skilled in the art.
　なお、本発明で用いられる濃縮においては、一定の分子量以上の分子は通過し得ない程度の細孔を有する限外ろ過膜が使用される。１つの実施形態では、濃縮にあたり、原水溶液は、例えば、分画分子量３０ｋＤａ以下の分子濾過膜からなるタンジェンシャルフロータイプの膜カセットを用いて濃縮が施される。ごく少量であれば、所望の分子ろ過膜の施されたチューブで構成される遠心式フィルターユニットが使用され得る。当該フィルターユニットは、さらに濾液チューブに挿入されて２重構造の遠心式限外ろ過チューブを形成していてもよい。このような濃縮を通じて、エタネルセプトは分子濾過膜を通過することなく、濃度が高くなる一方で、当初エタネルセプト原液に含まれていた不要な塩類,緩衝液成分を効率的に濾過することができる。このようにして、エタネルセプト原液を構成する溶液成分が緩衝液に置換されたエタネルセプトの濃縮液を得ることができる。 In addition, in the concentration used in the present invention, an ultrafiltration membrane having pores that cannot pass molecules having a certain molecular weight or more is used. In one embodiment, for concentration, the raw aqueous solution is concentrated using, for example, a tangential flow type membrane cassette composed of a molecular filtration membrane having a fractional molecular weight of 30 kDa or less. If the amount is very small, a centrifugal filter unit composed of a tube provided with a desired molecular filtration membrane can be used. The filter unit may be further inserted into a filtrate tube to form a double-structure centrifugal ultrafiltration tube. Through such concentration, the concentration of etanercept increases without passing through the molecular filtration membrane, while unnecessary salts and buffer components originally contained in the etanercept stock solution can be efficiently filtered. In this way, a concentrated solution of etanercept in which the solution components constituting the etanercept stock solution are replaced with the buffer solution can be obtained.
　このような濃縮（置換）の操作は、濃縮液において安定化剤、および緩衝剤について予め設定した含有量となるまで繰り返し行われてもよい。例えば、得られた濃縮液に新たに上記緩衝液を添加し、再度濃縮の操作が行われてもよい。 Such an operation of concentration (substitution) may be repeated until the contents of the stabilizer and the buffering agent are set in advance in the concentrate. For example, the above buffer solution may be newly added to the obtained concentrated solution, and the concentration operation may be performed again.
　次いで、得られた濃縮液が別の上記緩衝液と合わされる。 Next, the obtained concentrated solution is combined with another buffer solution.
　上記濃縮液と緩衝液とは、上記エタネルセプト、安定化剤、および緩衝剤について当業者が予め設定した含有量となるように、所定の割合で混合され得る。なお、合わせて得られた水溶液は必要に応じて無菌濾過フィルター等を用いて濾過されてもよい。 The concentrated solution and the buffer solution may be mixed at a predetermined ratio so that the etanercept, the stabilizer, and the buffering agent have contents preset by those skilled in the art. In addition, the aqueous solution obtained by combining may be filtered using a sterile filtration filter or the like as necessary.
　得られた水溶液は、例えば、滅菌済みのバイアル、シリンジやスクリューキャップチューブに封入される。 The obtained aqueous solution is sealed in, for example, a sterilized vial, syringe, or screw cap tube.
　このようにして、本発明の水溶液製剤を製造することができる。 In this way, the aqueous solution preparation of the present invention can be produced.
　以下、実施例により本発明をより具体的に説明するが、本発明はこれらの実施例により限定されるものではない。 Hereinafter, the present invention will be described more specifically with reference to examples, but the present invention is not limited to these examples.
（実施例１）
　５０ｍＭクエン酸一水和物溶液にヒスチジン塩酸塩およびグリシンを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、かつグリシンの濃度が１２０ｍＭ（９．０ｇ／Ｌ）となるように溶解させた液と５０ｍＭクエン酸三ナトリウム二水和物溶液にヒスチジン塩酸塩およびグリシンを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、かつグリシンの濃度が１２０ｍＭ（９．０ｇ／Ｌ）となるように溶解させた液とをそれぞれ適量混合し、水酸化ナトリウムを加えてｐＨ６．０に調整した緩衝液（液浸透圧２９４ｍＯｓｍ）を調製した。 (Example 1)
Histidine hydrochloride and glycine in a 50 mM citric acid monohydrate solution so that the concentration of histidine hydrochloride is 25 mM (4.79 g / L) and the concentration of glycine is 120 mM (9.0 g / L) Histidine hydrochloride and glycine were added to the dissolved solution and 50 mM trisodium citrate dihydrate solution, the concentration of histidine hydrochloride was 25 mM (4.79 g / L), and the concentration of glycine was 120 mM (9.0 g). / L) was mixed with an appropriate amount of each solution, and a buffer solution (liquid osmotic pressure 294 mOsm) adjusted to pH 6.0 by adding sodium hydroxide was prepared.
　エタネルセプトを原薬として１００ｍｇを含む水溶液（エタネルセプト原液）４ｍＬに、上記で調製したクエン酸緩衝液約１１ｍＬを混合して原水溶液を調製し、当該原水溶液を遠心式フィルターユニット（分画分子量：３０ｋＤａ，メルク株式会社製）に注入し、チューブにセットして限外ろ過デバイスを得た。このデバイスを遠心機（型番：ＣＲ－２１／日立工機株式会社製）にセットして、設定温度２０℃にて回転数３０００ｒｐｍで遠心し、フィルターユニット内の液量が１ｍＬ以下となるまで原水溶液を濃縮した。遠心後、限外濾過膜を通過した廃液を捨てた後、該フィルターユニット内に残存する濃縮液にさらに上記緩衝液１４ｍＬを注入して、遠心濾過を上記と同様にして行った。なお、各遠心濾過の操作において、得られた濃縮液の吸光度を測定し、吸光度から算出されるエタネルセプトの濃度が約５０ｍｇ／ｍＬとなるまで当該遠心濃縮の操作を（２回以上）繰り返し、エタネルセプト濃縮液を得た。 About 11 mL of the citrate buffer solution prepared above was mixed with 4 mL of an aqueous solution containing 100 mg of etanercept as the drug substance (etanercept stock solution) to prepare a raw aqueous solution, and the raw aqueous solution was subjected to a centrifugal filter unit (fractionated molecular weight: 30 kDa). , Manufactured by Merck Ltd.) and set in a tube to obtain an ultrafiltration device. This device is set in a centrifuge (model number: CR-21 / manufactured by Hitachi Koki Co., Ltd.), and centrifuged at a set temperature of 20 ° C. at a rotation speed of 3000 rpm. The aqueous solution was concentrated. After centrifugation, the waste solution that passed through the ultrafiltration membrane was discarded, and then 14 mL of the buffer solution was further injected into the concentrated solution remaining in the filter unit, and centrifugal filtration was performed in the same manner as described above. In each centrifugal filtration operation, the absorbance of the obtained concentrated solution was measured, and the centrifugal concentration operation was repeated (twice or more) until the concentration of etanercept calculated from the absorbance reached about 50 mg / mL. A concentrated solution was obtained.
　次いで、得られたエタネルセプト濃縮液に上記で調製した緩衝液を添加して、合わせた水溶液を１回凍結させた後、融解処理した。その後、０．２２μｍの無菌濾過用フィルターを用いて無菌濾過し、予め１２１℃にて２０分間オートクレーブ処理することにより滅菌した１．５ｍＬのスクリューキャップチューブに封入することにより、エタネルセプトの濃度が４７．１ｍｇ／ｍＬである医薬組成物の水溶液を得た（表１）。なお、本実施例で得られた医薬組成物のｐＨは上記緩衝液について調整したｐＨと変化がないことを確認した。 Next, the buffer solution prepared above was added to the obtained etanercept concentrate, and the combined aqueous solution was frozen once and then melted. Thereafter, the solution is aseptically filtered using a 0.22 μm filter for aseptic filtration, and sealed in a 1.5 mL screw cap tube that has been sterilized by autoclaving at 121 ° C. for 20 minutes in advance, so that the concentration of etanercept is 47. An aqueous solution of the pharmaceutical composition that was 1 mg / mL was obtained (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by the present Example did not change with pH adjusted about the said buffer solution.
（実施例２）
　実施例１で調製した緩衝液の代わりに、２５ｍＭクエン酸一水和物溶液にヒスチジン塩酸塩およびグリシンを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、かつグリシンの濃度が１９０ｍＭ（１４．２５ｇ／Ｌ）となるように溶解させた液と２５ｍＭクエン酸三ナトリウム二水和物溶液にヒスチジン塩酸塩およびグリシンを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、かつグリシンの濃度が１９０ｍＭ（１４．２５ｇ／Ｌ）となるように溶解させた液とをそれぞれ適量混合し、水酸化ナトリウムを加えてｐＨ６．０に調整した緩衝液（液浸透圧２９９ｍＯｓｍ）を調製して用いたこと以外は、実施例１と同様にして、エタネルセプトの濃度が４９．５ｍｇ／ｍＬである医薬組成物を得た（表１）。なお、本実施例で得られた医薬組成物のｐＨは上記緩衝液について調整したｐＨと変化がないことを確認した。 (Example 2)
Instead of the buffer prepared in Example 1, histidine hydrochloride and glycine were added to a 25 mM citrate monohydrate solution, the concentration of histidine hydrochloride was 25 mM (4.79 g / L), and the concentration of glycine was Histidine hydrochloride and glycine were added to a solution dissolved to 190 mM (14.25 g / L) and 25 mM trisodium citrate dihydrate solution, and the concentration of histidine hydrochloride was 25 mM (4.79 g / L). And a buffer solution (liquid osmotic pressure 299 mOsm) adjusted to pH 6.0 by adding sodium hydroxide to each solution in an appropriate amount so that the glycine concentration is 190 mM (14.25 g / L). A pharmaceutical composition having an etanercept concentration of 49.5 mg / mL was obtained in the same manner as in Example 1 except that was prepared and used (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by the present Example did not change with pH adjusted about the said buffer solution.
（実施例３）
　実施例１で調製した緩衝液の代わりに、２５ｍＭクエン酸一水和物溶液にヒスチジン塩酸塩、塩化ナトリウムおよびスクロースを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、塩化ナトリウムの濃度が８０ｍＭ（４．６８ｇ／Ｌ）であり、かつスクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液と２５ｍＭクエン酸三ナトリウム二水和物溶液にヒスチジン塩酸塩、塩化ナトリウムおよびスクロースを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、塩化ナトリウムの濃度が８０ｍＭ（４．６８ｇ／Ｌ）であり、かつスクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液とをそれぞれ適量混合し、水酸化ナトリウムを加えてｐＨ６．０に調整した緩衝液（液浸透圧２９４ｍＯｓｍ）を調製して用いたこと以外は、実施例１と同様にして、エタネルセプトの濃度が４７．７ｍｇ／ｍＬである医薬組成物を得た（表１）。なお、本実施例で得られた医薬組成物のｐＨは上記緩衝液について調整したｐＨと変化がないことを確認した。 (Example 3)
Instead of the buffer solution prepared in Example 1, histidine hydrochloride, sodium chloride and sucrose were added to a 25 mM citric acid monohydrate solution, and the concentration of histidine hydrochloride was 25 mM (4.79 g / L). Of histidine hydrochloride, sodium chloride, and sucrose in a solution dissolved to a concentration of 80 mM (4.68 g / L) and a sucrose concentration of 10 mg / mL and a 25 mM trisodium citrate dihydrate solution Was dissolved so that the concentration of histidine hydrochloride was 25 mM (4.79 g / L), the concentration of sodium chloride was 80 mM (4.68 g / L), and the concentration of sucrose was 10 mg / mL. A suitable amount of the solution was mixed, and sodium hydroxide was added to adjust the pH to 6.0 (liquid osmotic pressure 294 mOsm). And except for the use, in the same manner as in Example 1, the concentration of etanercept to obtain a pharmaceutical composition is 47.7 mg / mL (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by the present Example did not change with pH adjusted about the said buffer solution.
（実施例４）
　実施例１で調製した緩衝液の代わりに、５０ｍＭクエン酸一水和物溶液にヒスチジン塩酸塩、塩化ナトリウム、およびスクロースを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、塩化ナトリウムの濃度が４０ｍＭ（２．３４ｇ／Ｌ）であり、かつスクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液と５０ｍＭクエン酸三ナトリウム二水和物溶液にヒスチジン塩酸塩、塩化ナトリウム、およびスクロースを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、塩化ナトリウムの濃度が４０ｍＭ（２．３４ｇ／Ｌ）であり、かつスクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液とをそれぞれ適量混合し、水酸化ナトリウムを加えてｐＨ６．０に調整した緩衝液（液浸透圧２８８ｍＯｓｍ）を調製して用いたこと以外は、実施例１と同様にして、エタネルセプトの濃度が４７．７ｍｇ／ｍＬである医薬組成物を得た（表１）。なお、本実施例で得られた医薬組成物のｐＨは上記緩衝液について調整したｐＨと変化がないことを確認した。 Example 4
Instead of the buffer prepared in Example 1, histidine hydrochloride, sodium chloride, and sucrose were added to a 50 mM citric acid monohydrate solution, and the concentration of histidine hydrochloride was 25 mM (4.79 g / L). A solution in which the concentration of sodium is 40 mM (2.34 g / L) and the concentration of sucrose is 10 mg / mL and 50 mM trisodium citrate dihydrate solution are mixed with histidine hydrochloride, sodium chloride, And sucrose so that the concentration of histidine hydrochloride is 25 mM (4.79 g / L), the concentration of sodium chloride is 40 mM (2.34 g / L), and the concentration of sucrose is 10 mg / mL. Buffer solution (liquid osmotic pressure 288 mOsm) adjusted to pH 6.0 by adding sodium hydroxide. Except for the use and preparation, in the same manner as in Example 1, the concentration of etanercept to obtain a pharmaceutical composition is 47.7 mg / mL (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by the present Example did not change with pH adjusted about the said buffer solution.
（比較例１）
　実施例１で調製した緩衝液の代わりに、２５ｍＭリン酸二水素ナトリウム溶液にアルギニン塩酸塩、塩化ナトリウムおよびスクロースを、アルギニン塩酸塩の濃度が２５ｍＭ（５．２７ｇ／Ｌ）であり、塩化ナトリウムの濃度が１００ｍＭ（５．８４ｇ／Ｌ）であり、かつスクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液と２５ｍＭリン酸一水素ナトリウム二水和物溶液にアルギニン塩酸塩、塩化ナトリウムおよびスクロースを、アルギニン塩酸塩の濃度が２５ｍＭ（５．２７ｇ／Ｌ）であり、塩化ナトリウムの濃度が１００ｍＭ（５．８４ｇ／Ｌ）であり、かつスクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液とをそれぞれ適量混合した、ｐＨ６．３の緩衝液（液浸透圧３０６ｍＯｓｍ）を調製して用いたこと以外は、実施例１と同様にして、エタネルセプトの濃度が４７．２ｍｇ／ｍＬである医薬組成物を得た（表１）。なお、本比較例で得られた医薬組成物のｐＨは上記緩衝液について調整したｐＨと変化がないことを確認した。 (Comparative Example 1)
Instead of the buffer prepared in Example 1, arginine hydrochloride, sodium chloride and sucrose were added to a 25 mM sodium dihydrogen phosphate solution, and the concentration of arginine hydrochloride was 25 mM (5.27 g / L). Arginine hydrochloride, sodium chloride and sucrose were added to a solution in which the concentration was 100 mM (5.84 g / L) and the sucrose concentration was 10 mg / mL and a 25 mM sodium monohydrogen phosphate dihydrate solution. Was dissolved so that the concentration of arginine hydrochloride was 25 mM (5.27 g / L), the concentration of sodium chloride was 100 mM (5.84 g / L), and the concentration of sucrose was 10 mg / mL. A buffer solution of pH 6.3 (liquid osmotic pressure 306 mOsm) prepared by mixing appropriate amounts of each solution was used. , The same procedure as in Example 1, the concentration of etanercept to obtain a pharmaceutical composition is 47.2 mg / mL (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by this comparative example did not change with pH adjusted about the said buffer solution.
（比較例２）
　実施例１で調製した緩衝液の代わりに、２５ｍＭリン酸二水素ナトリウム溶液にヒスチジン塩酸塩およびグリシンを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、かつグリシンの濃度が２６６ｍＭ（１９．９５ｇ／Ｌ）となるように溶解させた液と２５ｍＭリン酸一水素ナトリウム二水和物溶液にヒスチジン塩酸塩およびグリシンを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、かつグリシンの濃度が２６６ｍＭ（１９．９５ｇ／Ｌ）となるように溶解させた液とをそれぞれ適量混合した、ｐＨ６．０の緩衝液（液浸透圧３４９ｍＯｓｍ）を調製して用いたこと以外は、実施例１と同様にして、エタネルセプトの濃度が４７．２ｍｇ／ｍＬである医薬組成物を得た（表１）。なお、本比較例で得られた医薬組成物のｐＨは上記緩衝液について調整したｐＨと変化がないことを確認した。 (Comparative Example 2)
Instead of the buffer prepared in Example 1, histidine hydrochloride and glycine were added to a 25 mM sodium dihydrogen phosphate solution, the concentration of histidine hydrochloride was 25 mM (4.79 g / L), and the concentration of glycine was 266 mM. (19.95 g / L) and 25 mM sodium monohydrogen phosphate dihydrate solution were mixed with histidine hydrochloride and glycine, and the concentration of histidine hydrochloride was 25 mM (4.79 g / L). Except for the preparation and use of a pH 6.0 buffer solution (liquid osmotic pressure 349 mOsm), which is mixed with an appropriate amount of each solution so that the concentration of glycine is 266 mM (19.95 g / L). Obtained a pharmaceutical composition in which the concentration of etanercept was 47.2 mg / mL in the same manner as in Example 1 (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by this comparative example did not change with pH adjusted about the said buffer solution.
（比較例３）
　実施例１で調製した緩衝液の代わりに、２５ｍＭクエン酸一水和物溶液にヒスチジン塩酸塩、塩化ナトリウム、およびスクロースを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、塩化ナトリウムの濃度が１００ｍＭ（５．８５ｇ／Ｌ）であり、かつスクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液と２５ｍＭクエン酸三ナトリウム二水和物溶液にヒスチジン塩酸塩、塩化ナトリウム、およびスクロースを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、塩化ナトリウムの濃度が１００ｍＭ（５．８５ｇ／Ｌ）であり、かつスクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液とをそれぞれ適量混合した、ｐＨ５．５の緩衝液（液浸透圧３３２ｍＯｓｍ）を調製して用いたこと以外は、実施例１と同様にして、エタネルセプトの濃度が４８．９ｍｇ／ｍＬである医薬組成物を得た（表１）。なお、本比較例で得られた医薬組成物のｐＨは上記緩衝液について調整したｐＨと変化がないことを確認した。 (Comparative Example 3)
Instead of the buffer prepared in Example 1, histidine hydrochloride, sodium chloride, and sucrose were added to a 25 mM citric acid monohydrate solution, the concentration of histidine hydrochloride was 25 mM (4.79 g / L), A solution in which the concentration of sodium is 100 mM (5.85 g / L) and the concentration of sucrose is 10 mg / mL and 25 mM trisodium citrate dihydrate solution are mixed with histidine hydrochloride, sodium chloride, And sucrose so that the concentration of histidine hydrochloride is 25 mM (4.79 g / L), the concentration of sodium chloride is 100 mM (5.85 g / L), and the concentration of sucrose is 10 mg / mL. Except for preparing and using a pH 5.5 buffer solution (liquid osmotic pressure of 332 mOsm), each of which was mixed with an appropriate amount. In the same manner as in Example 1, the concentration of etanercept to obtain a pharmaceutical composition is 48.9 mg / mL (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by this comparative example did not change with pH adjusted about the said buffer solution.
（比較例４）
　実施例１で得られた緩衝液の代わりに、２５ｍＭクエン酸一水和物溶液にヒスチジン塩酸塩、塩化ナトリウム、およびスクロースを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、塩化ナトリウムの濃度が１００ｍＭ（５．８５ｇ／Ｌ）であり、スクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液と２５ｍＭクエン酸三ナトリウム二水和物溶液にヒスチジン塩酸塩、塩化ナトリウム、およびスクロースを、ヒスチジン塩酸塩の濃度が２５ｍＭ（４．７９ｇ／Ｌ）であり、塩化ナトリウムの濃度が１００ｍＭ（５．８５ｇ／Ｌ）であり、スクロースの濃度が１０ｍｇ／ｍＬとなるように溶解させた液とをそれぞれ適量混合し、水酸化ナトリウムを加えてｐＨ６．５に調製した緩衝液（液浸透圧３４１ｍＯｓｍ）を調製して用いたこと以外は、実施例１と同様にして、エタネルセプトの濃度が４６．７ｍｇ／ｍＬである医薬組成物を得た（表１）。なお、本比較例で得られた医薬組成物のｐＨは上記緩衝液について調整したｐＨと変化がないことを確認した。 (Comparative Example 4)
In place of the buffer obtained in Example 1, histidine hydrochloride, sodium chloride, and sucrose were added to a 25 mM citrate monohydrate solution, and the concentration of histidine hydrochloride was 25 mM (4.79 g / L). A solution in which the concentration of sodium chloride is 100 mM (5.85 g / L) and the concentration of sucrose is 10 mg / mL and a 25 mM trisodium citrate dihydrate solution is mixed with histidine hydrochloride, sodium chloride, And sucrose were dissolved so that the concentration of histidine hydrochloride was 25 mM (4.79 g / L), the concentration of sodium chloride was 100 mM (5.85 g / L), and the concentration of sucrose was 10 mg / mL. A buffer solution (liquid osmotic pressure of 341 mOsm) adjusted to pH 6.5 by adding sodium hydroxide was prepared. And except for the use, in the same manner as in Example 1, the concentration of etanercept to obtain a pharmaceutical composition is 46.7 mg / mL (Table 1). In addition, it confirmed that pH of the pharmaceutical composition obtained by this comparative example did not change with pH adjusted about the said buffer solution.
Figure JPOXMLDOC01-appb-T000001
Figure JPOXMLDOC01-appb-T000001
　実施例１～４および比較例１～４で得られた８種類の医薬組成物を、設定温度３７℃にて遮光下でインキュベータに３週間保存した。 The eight types of pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4 were stored in an incubator for 3 weeks at a set temperature of 37 ° C. and protected from light.
　各保存サンプルをドデシル硫酸ナトリウム－ポリアクリルアミドゲル電気泳動（ＳＤＳ－ＰＡＧＥ）およびサイズ排除クロマトグラフィーでそれぞれ以下のようにして分析した。 Each preserved sample was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and size exclusion chromatography as follows.
（ＳＤＳ－ＰＡＧＥ）
　還元剤として２－メルカプトエタノールを用いて還元したエタネルセプトと、還元しない（非還元の）エタネルセプトをそれぞれ用意して、ドデシル硫酸ナトリウム－ポリアクリルアミドゲル電気泳動（ＳＤＳ－ＰＡＧＥ）によりポリペプチド断片を分離し、銀染色法により染色した。 (SDS-PAGE)
Prepare etanercept reduced with 2-mercaptoethanol as a reducing agent and non-reduced (non-reduced) etanercept, and separate polypeptide fragments by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). And stained by silver staining.
　還元したエタネルセプトのＳＤＳ－ＰＡＧＥでは、７５ｋＤａ付近のバンドを主バンドとし、これより泳動距離の短い（上方に位置する）バンドを凝集体のバンド、泳動距離の長い（下方に位置する）バンドを切断体のバンドとした。一方、非還元のエタネルセプトのＳＤＳ－ＰＡＧＥでは、１５０ｋＤａ付近のバンドを主バンドとし、これより泳動距離の短い（上方に位置する）バンドを凝集体のバンド、泳動距離の長い（下方に位置する）バンドを切断体のバンドとした。 In SDS-PAGE of reduced etanercept, the band near 75 kDa is the main band, the band with shorter migration distance (above) is cut with the aggregate band, and the band with longer migration distance (below) is cut. It was a body band. On the other hand, in the SDS-PAGE of non-reducing etanercept, the band near 150 kDa is the main band, the band having a shorter migration distance (located above) is the aggregate band, and the band having a longer migration distance (located below). The band was a cut band.
　測定条件は、表２の通りであった。 The measurement conditions were as shown in Table 2.
Figure JPOXMLDOC01-appb-T000002
Figure JPOXMLDOC01-appb-T000002
　還元したエタネルセプトのＳＤＳ－ＰＡＧＥでは、７５ｋＤａの指標付近に高い強度のバンドを確認することができ、２５ｋＤａ～３７ｋＤａ付近にも高い強度のバンドを確認することができた。一方、非還元のエタネルセプトのＳＤＳ－ＰＡＧＥでは、１５０ｋＤａの指標付近に高い強度のバンドを確認することができ、７５ｋＤａ～１００ｋＤａ付近にも高い強度のバンドを確認することができた。 In the reduced etanercept SDS-PAGE, a high-intensity band was confirmed in the vicinity of the 75 kDa index, and a high-intensity band was also confirmed in the vicinity of 25 kDa to 37 kDa. On the other hand, in non-reducing etanercept SDS-PAGE, a high-intensity band was confirmed in the vicinity of the 150 kDa index, and a high-intensity band was also confirmed in the vicinity of 75 kDa to 100 kDa.
（サイズ排除クロマトグラフィー）
　実施例１～４および比較例１～４で得られた８種類の医薬組成物について、インキュベータ保存開始時、保存１週間後、保存２週間後、および保存３週間後の４時点において、サイズ排除クロマトグラフィーにより試験した。試験には、保存された医薬組成物に、エタネルセプト濃度として約０．２ｍｇ／ｍＬとなるようにそれぞれの実施例または比較例で使用した緩衝液で希釈して調製した試料溶液（各５０μＬ）を試験サンプルとして使用した。 (Size exclusion chromatography)
For the eight types of pharmaceutical compositions obtained in Examples 1 to 4 and Comparative Examples 1 to 4, size exclusion was performed at 4 time points at the start of incubator storage, 1 week after storage, 2 weeks after storage, and 3 weeks after storage. Tested by chromatography. In the test, a sample solution (50 μL each) prepared by diluting the stored pharmaceutical composition with the buffer used in each Example or Comparative Example so that the etanercept concentration was about 0.2 mg / mL. Used as a test sample.
　試験条件は、表３の通りであった。 The test conditions were as shown in Table 3.
Figure JPOXMLDOC01-appb-T000003
Figure JPOXMLDOC01-appb-T000003
　得られたクロマトグラフィーの主ピークの面積（Ａ１）、主ピークより前に溶出する個々のピークの合計面積（Ａ２）、および主ピークより後に溶出する溶媒ピークを除いた個々のピークの合計面積（Ａ３）を自動積分法により測定し、次式によりエタネルセプトの不純物である凝集体および切断体の量を算出した。 The area of the main peak of the obtained chromatography (A1), the total area of individual peaks eluting before the main peak (A2), and the total area of individual peaks excluding the solvent peak eluting after the main peak ( A3) was measured by an automatic integration method, and the amounts of aggregates and cut bodies, which are impurities of etanercept, were calculated by the following equation.
Figure JPOXMLDOC01-appb-M000004
Figure JPOXMLDOC01-appb-M000004
　得られた結果を表４および図１～４に示す。 The obtained results are shown in Table 4 and FIGS.
Figure JPOXMLDOC01-appb-T000005
Figure JPOXMLDOC01-appb-T000005
　表４および図１～４に示すように、エタネルセプトの不純物の合計量について、実施例１～４と比較例１～４とを比較すると、実施例１～４それぞれの不純物の合計量は、特に、保存１週間後、および保存２週間後の時点において比較例１～４の不純物の合計量を下回っていた。 As shown in Table 4 and FIGS. 1 to 4, when the total amount of impurities in etanercept is compared between Examples 1 to 4 and Comparative Examples 1 to 4, the total amount of impurities in Examples 1 to 4 is The total amount of impurities in Comparative Examples 1 to 4 was less than 1 week after storage and 2 weeks after storage.
　したがって、実施例１～４で得られた医薬組成物は、製剤中のエタネルセプトの保存安定性を向上させていたことがわかる。特に、比較例１は安定化剤および緩衝剤のいずれもが実施例で使用したものと異なるが、比較例１が最も不純物量が高い値を示していた。このことから考えて、本実施例で使用した安定化剤および緩衝剤の組合せは、製剤中のエタネルセプトの保存安定性の向上に寄与していたことがわかる。 Therefore, it can be seen that the pharmaceutical compositions obtained in Examples 1 to 4 improved the storage stability of etanercept in the preparation. In particular, Comparative Example 1 was different from that used in the Examples for both the stabilizer and the buffer, but Comparative Example 1 showed the highest amount of impurities. In view of this, it can be seen that the combination of the stabilizer and the buffer used in this example contributed to the improvement of the storage stability of etanercept in the preparation.
　また、比較例３で調製した緩衝液の液性が最も酸性側（ｐＨ５．５）にあり、得られたエタネルセプトの切断体量が最も多い結果となった。一方、比較例４で調製した緩衝液の液性が最も塩基性側（ｐＨ６．５）にあり、得られたエタネルセプトの凝集体量が最も多い結果となった。これに対し、緩衝液の液性をｐＨ５．６～６．４の範囲内に調整することにより、製剤中のエタネルセプトの保存安定性が効果的に向上していたことがわかる。 Moreover, the liquidity of the buffer solution prepared in Comparative Example 3 was on the most acidic side (pH 5.5), and the obtained etanercept was cut most. On the other hand, the buffer solution prepared in Comparative Example 4 was most basic (pH 6.5), and the resulting etanercept aggregate amount was the largest. On the other hand, it can be seen that the storage stability of etanercept in the preparation was effectively improved by adjusting the liquidity of the buffer solution within the range of pH 5.6 to 6.4.
　本発明によれば、有効成分であるエタネルセプトの安定性を高めることができる。このことにより、医薬組成物としての長期間の保存性を高めることができる。本発明の水溶液製剤は、例えば、関節リウマチ、若年性特発性関節炎、乾癬性関節炎、強直性脊椎炎、ヴェーグナー病、クローン病、慢性閉塞性肺疾患（ＣＯＰＤ）、Ｃ型肝炎、子宮内膜症、喘息、悪液質、乾癬及びアトピー性皮膚炎などのための医薬組成物として有用である。 According to the present invention, the stability of etanercept, which is an active ingredient, can be increased. Thereby, long-term preservation as a pharmaceutical composition can be enhanced. Examples of the aqueous solution preparation of the present invention include rheumatoid arthritis, juvenile idiopathic arthritis, psoriatic arthritis, ankylosing spondylitis, Wegner's disease, Crohn's disease, chronic obstructive pulmonary disease (COPD), hepatitis C, endometriosis It is useful as a pharmaceutical composition for asthma, cachexia, psoriasis and atopic dermatitis.

Claims (9)

  1. 　エタネルセプト、安定化剤および緩衝剤を含有する水溶液製剤であって、
    　安定化剤がヒスチジンまたはその塩であり、
    　緩衝剤が有機酸またはその塩であり、そして
    　ｐＨが５．６から６．４である、製剤。     An aqueous solution formulation containing etanercept, stabilizer and buffer,
    The stabilizer is histidine or a salt thereof,
    A formulation wherein the buffer is an organic acid or salt thereof and the pH is 5.6 to 6.4.
  2. 　前記有機酸またはその塩が、クエン酸、クエン酸塩、酢酸、酢酸塩、酒石酸、酒石酸塩、およびトリス（ヒドロキシメチル）アミノメタン、およびトリス（ヒドロキシメチル）アミノメタン塩酸塩からなる群から選択される少なくとも１種の化合物である、請求項１に記載の水溶液製剤。 The organic acid or salt thereof is selected from the group consisting of citric acid, citrate, acetic acid, acetate, tartaric acid, tartrate, and tris (hydroxymethyl) aminomethane, and tris (hydroxymethyl) aminomethane hydrochloride The aqueous solution preparation according to claim 1, which is at least one compound.
  3. 　前記緩衝剤を、水溶液全体に対し１ｍＭから１Ｍの濃度で含有する、請求項１または２に記載の水溶液製剤。 The aqueous solution preparation according to claim 1 or 2, comprising the buffer at a concentration of 1 mM to 1 M with respect to the entire aqueous solution.
  4. 　前記安定化剤を、水溶液全体に対し１ｍＭから１Ｍの濃度で含有する、請求項１から３のいずれかに記載の水溶液製剤。 The aqueous solution preparation according to any one of claims 1 to 3, wherein the stabilizer is contained at a concentration of 1 mM to 1 M with respect to the entire aqueous solution.
  5. 　さらに、塩化ナトリウム、スクロースおよびグリシンからなる群から選択される少なくとも１種の等張化剤を含有する、請求項１から４のいずれかに記載の水溶液製剤。 The aqueous solution preparation according to any one of claims 1 to 4, further comprising at least one isotonic agent selected from the group consisting of sodium chloride, sucrose and glycine.
  6. 　水溶液製剤の製造方法であって、
    　ヒスチジンまたはその塩と、有機酸またはその塩とを水に溶解させて、ｐＨが５．６から６．４である緩衝液を調製する工程；
    　該緩衝液にエタネルセプト原液を添加する工程；
    　該原液を濃縮することにより、該エタネルセプトの濃縮液を得る工程；および
    　該濃縮液を別の該緩衝液と合わせる工程；
    を包含する、方法。     A method for producing an aqueous solution formulation comprising:
    Preparing a buffer solution having a pH of 5.6 to 6.4 by dissolving histidine or a salt thereof and an organic acid or a salt thereof in water;
    Adding an etanercept stock solution to the buffer;
    Concentrating the stock solution to obtain a concentrate of the etanercept; and combining the concentrate with another buffer;
    Including the method.
  7. 　前記有機酸またはその塩が、クエン酸、クエン酸塩、酢酸、酢酸塩、酒石酸、酒石酸塩、およびトリス（ヒドロキシメチル）アミノメタン、およびトリス（ヒドロキシメチル）アミノメタン塩酸塩からなる群から選択される少なくとも１種の化合物である、請求項６に記載の方法。 The organic acid or salt thereof is selected from the group consisting of citric acid, citrate, acetic acid, acetate, tartaric acid, tartrate, and tris (hydroxymethyl) aminomethane, and tris (hydroxymethyl) aminomethane hydrochloride The method of claim 6, wherein the method is at least one compound.
  8. 　前記緩衝液を調製する工程において、さらに等張化剤が溶解される、請求項６または７に記載の方法。 The method according to claim 6 or 7, wherein an isotonic agent is further dissolved in the step of preparing the buffer solution.
  9. 　前記等張化剤が、塩化ナトリウム、スクロースおよびグリシンからなる群から選択される少なくとも１種の化合物である、請求項８に記載の方法。 The method according to claim 8, wherein the tonicity agent is at least one compound selected from the group consisting of sodium chloride, sucrose and glycine.
PCT/JP2014/054036 2013-04-30 2014-02-20 Aqueous solution formulation and method for producing same WO2014178216A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2013095979A JP2014214153A (en) 2013-04-30 2013-04-30 Pharmaceutical preparation of aqueous solution and method for producing the same
JP2013-095979 2013-04-30

Publications (1)

Publication Number Publication Date
WO2014178216A1 true WO2014178216A1 (en) 2014-11-06

Family

ID=51843354

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2014/054036 WO2014178216A1 (en) 2013-04-30 2014-02-20 Aqueous solution formulation and method for producing same

Country Status (2)

Country Link
JP (1) JP2014214153A (en)
WO (1) WO2014178216A1 (en)

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2016088897A (en) * 2014-11-06 2016-05-23 持田製薬株式会社 Lyophilized products of etanercept
WO2016102328A1 (en) * 2014-12-22 2016-06-30 Ares Trading S.A. Liquid pharmaceutical composition
CN108367094A (en) * 2015-12-10 2018-08-03 株式会社目立康 Peptide combinations
CN112313727A (en) * 2018-06-12 2021-02-02 瑞乐文特生物有限公司 Method for preparing buffer solution for drug solubility in vitro test, package for preparing buffer solution, and kit for clinical state test

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
SG11202103137WA (en) * 2018-10-26 2021-04-29 Novo Nordisk As Stable semaglutide compositions and uses thereof

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2008520551A (en) * 2004-10-20 2008-06-19 ジェネンテック・インコーポレーテッド Antibody preparation
JP2009525986A (en) * 2006-02-03 2009-07-16 メディミューン，エルエルシー Protein preparation
JP2011519848A (en) * 2008-05-01 2011-07-14 アレコー リミテッド Protein preparation
WO2012143418A1 (en) * 2011-04-20 2012-10-26 Sandoz Ag STABLE PHARMACEUTICAL LIQUID FORMULATIONS OF THE FUSION PROTEIN TNFR:Fc
WO2012165917A1 (en) * 2011-06-03 2012-12-06 Lg Life Sciences Ltd. Stable liquid formulation of etanercept

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2008520551A (en) * 2004-10-20 2008-06-19 ジェネンテック・インコーポレーテッド Antibody preparation
JP2009525986A (en) * 2006-02-03 2009-07-16 メディミューン，エルエルシー Protein preparation
JP2011519848A (en) * 2008-05-01 2011-07-14 アレコー リミテッド Protein preparation
WO2012143418A1 (en) * 2011-04-20 2012-10-26 Sandoz Ag STABLE PHARMACEUTICAL LIQUID FORMULATIONS OF THE FUSION PROTEIN TNFR:Fc
WO2012165917A1 (en) * 2011-06-03 2012-12-06 Lg Life Sciences Ltd. Stable liquid formulation of etanercept

Cited By (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2016088897A (en) * 2014-11-06 2016-05-23 持田製薬株式会社 Lyophilized products of etanercept
WO2016102328A1 (en) * 2014-12-22 2016-06-30 Ares Trading S.A. Liquid pharmaceutical composition
CN107205925A (en) * 2014-12-22 2017-09-26 阿雷斯贸易股份有限公司 Composition of liquid medicine
CN107205925B (en) * 2014-12-22 2020-12-11 阿雷斯贸易股份有限公司 Liquid pharmaceutical composition
US10959939B2 (en) 2014-12-22 2021-03-30 Ares Trading S.A. Liquid pharmaceutical composition of etanercept with lysine and proline
CN108367094A (en) * 2015-12-10 2018-08-03 株式会社目立康 Peptide combinations
US10702606B2 (en) 2015-12-10 2020-07-07 Menicon Co., Ltd. Peptide composition
RU2740185C2 (en) * 2015-12-10 2021-01-12 Меникон Ко., Лтд. Peptide composition
CN108367094B (en) * 2015-12-10 2021-09-14 株式会社目立康 Peptide composition
CN112313727A (en) * 2018-06-12 2021-02-02 瑞乐文特生物有限公司 Method for preparing buffer solution for drug solubility in vitro test, package for preparing buffer solution, and kit for clinical state test
CN112313727B (en) * 2018-06-12 2023-12-22 瑞乐文特生物有限公司 Method and package for preparing buffer solution and kit for clinical test

Also Published As

Publication number Publication date
JP2014214153A (en) 2014-11-17

Similar Documents

Publication Publication Date Title
US20210007991A1 (en) Etanercept formulations exhibiting marked reduction in sub-visible particles
US11129876B2 (en) Etanercept formulations stabilized with amino acids
WO2014178216A1 (en) Aqueous solution formulation and method for producing same
JP7536640B2 (en) VEGFR-Fc fusion protein preparation
US20220288167A1 (en) A stable lyophilized formulation for hybrid fc fused g-csf
WO2015056613A1 (en) Stabilized polypeptide aqueous preparation

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 14791739

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 14791739

Country of ref document: EP

Kind code of ref document: A1