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WO1986000531A1 - Composition a base de gamma-interferon - Google Patents

Composition a base de gamma-interferon Download PDF

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Publication number
WO1986000531A1
WO1986000531A1 PCT/JP1984/000352 JP8400352W WO8600531A1 WO 1986000531 A1 WO1986000531 A1 WO 1986000531A1 JP 8400352 W JP8400352 W JP 8400352W WO 8600531 A1 WO8600531 A1 WO 8600531A1
Authority
WO
WIPO (PCT)
Prior art keywords
solution
ifn
aqueous solution
freeze
composition
Prior art date
Legal status (The legal status 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 status listed.)
Ceased
Application number
PCT/JP1984/000352
Other languages
English (en)
Japanese (ja)
Inventor
Yasaburo Akagi
Yasumoto Miura
Tetsuo Hoshino
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Takeda Pharmaceutical Co Ltd
Original Assignee
Takeda Chemical Industries Ltd
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 Takeda Chemical Industries Ltd filed Critical Takeda Chemical Industries Ltd
Priority to PCT/JP1984/000352 priority Critical patent/WO1986000531A1/fr
Priority to JP60148093A priority patent/JPS6144826A/ja
Priority to EP85108409A priority patent/EP0168008A3/fr
Priority to ES544991A priority patent/ES8608542A1/es
Priority to KR1019850004888A priority patent/KR860000869A/ko
Publication of WO1986000531A1 publication Critical patent/WO1986000531A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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Classifications

    • 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
    • A61K38/19—Cytokines; Lymphokines; Interferons
    • A61K38/21—Interferons [IFN]
    • A61K38/217—IFN-gamma
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/52—Cytokines; Lymphokines; Interferons
    • C07K14/555—Interferons [IFN]
    • C07K14/57—IFN-gamma
    • 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
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1629—Organic macromolecular compounds
    • A61K9/1652—Polysaccharides, e.g. alginate, cellulose derivatives; Cyclodextrin
    • 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
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/19—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles lyophilised, i.e. freeze-dried, solutions or dispersions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates to a human r-type interferon composition.
  • ⁇ Type 9 and type 9 interferon are relatively stable, are provided to the clinic mainly in the form of parenteral administration, and are undergoing systematic clinical research.
  • r-type interferon FN-sometimes abbreviated as r
  • FN-sometimes abbreviated as r is extremely unstable, and its activity is easily reduced during storage, freezing or freeze-drying operations of aqueous solutions.
  • turbidity is observed in a solution obtained by re-dissolving the lyophilized product, and it has been extremely difficult to obtain a stable composition that can be used clinically.
  • inorganic ions used for adjusting pH and ionic strength are also present in the purified aqueous solution of interfering solution.
  • Running it has been disclosed that the addition of an inorganic salt stabilizes interferon having no ⁇ chain] (Japanese Patent Application Laid-Open No. 59-25364). .
  • the present inventors surprisingly formulate using an aqueous IFN- r solution with a reduced inorganic salt concentration, and surprisingly, in the freezing and freeze-drying ft crop, a conventional IFN-r aqueous solution containing an inorganic salt was used.
  • a stable IFN-r composition can be obtained in which the decrease in the IFN-r activity of the ⁇ layer is less and no turbidity is observed in the solution obtained by re-dissolving the obtained composition.
  • the present inventors have further studied and completed the present invention.
  • An object of the present invention is to provide a human r-type interfering mouth composition which has been frozen or freeze-dried under conditions in which substantially no inorganic salt is present and amino acids coexist.
  • IFN-r used in the present invention may be any of IFN-r that is derived from human or natural or obtained by genetic recombination technology.
  • an aqueous solution having a high concentration of human IFN- r (rIFN-) obtained by genetic recombination technology is advantageously used.
  • the specific activity of human IFN-r is preferably 1 ⁇ 10 5 to 1 ⁇ 10 7 international units / ⁇ (I u ⁇ ), and 1 ⁇ 10 2 to 1 1 0 7 I UZ, which has a 1 X 1 0 4 ⁇ 1 X 1 0 7 IU ⁇ activity divided and are preferred.
  • the concentration of the inorganic salt may be 0.1 M or less, and is 0.05 5 or less. Is preferred.
  • the inorganic ion strength may be such that the ion strength calculated from the inorganic ions is 0.1 or less. 0.05 or less, and! The reason is preferably 0.01 or less.
  • the inorganic salt may be 30% or less, preferably 15% or less, and preferably 0.3% or less based on the total weight.
  • the FN- ⁇ 7_ ⁇ solution that substantially contains inorganic salts can be prepared by using a buffer solution that does not contain inorganic salts as the buffer used in the purification step of IF-r and in the final chromatographic operation. ], Or by removing inorganic salts from the purified aqueous solution of IFN-r]).
  • Preferred amino acids are glycine, na-alanin,
  • Mono-amino aliphatic amino acids such as glycine are preferred, and glycine is particularly preferred.
  • a physiologically acceptable salt or derivative thereof may be used.
  • One or more of these amino acids can be used.
  • the amino acids to be used can be commercially available, but in order to apply this composition to clinical applications, it should be of a quality sufficient for parenteral administration. Are preferred.
  • the total amount of amino acids is 1 or more, preferably 5 to 50, equivalent to an IFN-r aqueous solution.
  • composition of the present invention may contain a reducing sulfur compound.
  • reducing sulfur compound examples include gnoletathione (reduced form), thioctic acid, thiocyanate diglycolone, thiocyanate ethanoamine, monothiocyanate glycerone, thiocyanate slate and carbon number ⁇ . And thioalkanoic acids of from 7 to 7].
  • the IFN- r aqueous solution 1D reducing sulfur compound is preferably 0.1 ⁇ or more, and 0.5 to 10 is preferred.
  • Still other stabilizers such as sugars, such as dextran, hydroxyshetti
  • Substitute Sir G- One or more substances selected from polysaccharides such as starch, disaccharides such as sucrose, maltose and lactose, and monosaccharides such as glucose, fructose, mannose and galactose can also be added.
  • polysaccharides such as starch
  • disaccharides such as sucrose, maltose and lactose
  • monosaccharides such as glucose, fructose, mannose and galactose
  • dextran and hydroxyxechinole starch commercially available ones can be used, but in order to clinically apply the present composition, those having a quality such that it can be used as a plasma substitute for parenteral administration are preferable.
  • Dextran has an average molecular weight of 10,000 to 100,000, and more than 40,000 to 70,000, while hydroxyethyl starch has an average molecular weight of 10,000 to 200,000, and] 9 more than 20,000 to 60,000. 200,000 are advantageously used.
  • the aqueous solution be contained in an IFN-r aqueous solution in an amount of 1 ⁇ or more, preferably 3 to 50 ⁇ .
  • the composition may further contain the above-mentioned saccharide or (and) amino acids as an osmotic pressure adjusting agent, but the addition of an inorganic salt such as sodium chloride may adversely affect the quality of the composition. Is inferior, so it is preferable.
  • the above-mentioned preferable osmotic pressure adjusting agent may be added to the composition in advance, or may be added to a solvent for re-dissolving the freeze-dried product.
  • the frozen and freeze-dried human FN- r composition of the present invention can be produced, for example, by the following method.
  • the above-mentioned amino acid can be added to the aqueous solution containing IFN- r during the production process. When an aqueous solution of IFN-r also containing this amino acid is used, it is necessary or necessary. Amino acids can be added up to the concentration of the amino acids and subjected to the following step. Change Further, the above-mentioned saccharides and the like can be added together.
  • the aqueous IF 1 N-r solution may contain a reducing sulfur compound of at least 0.1 ⁇ , preferably 0.5 to 1) ⁇ and a trace amount of a surfactant, and Like agents, these can be added fresh.
  • the frozen human IFN-r composition of the present invention can be obtained, for example, by freezing the above aqueous solution at a temperature of usually 180 ° to ⁇ 30! ) Can be manufactured.
  • the frozen composition is stored at 180 ° C to 110 ° C.
  • the freeze-dried human IFN-r composition of the present invention can be obtained, for example, by drying the above-mentioned frozen composition under reduced pressure or by thawing the above-mentioned aqueous solution or the above-mentioned frozen composition!
  • the resulting aqueous solution can be subdivided as desired], frozen in the same manner as described above, and dried under reduced pressure by a conventional method.
  • the aqueous solution of the composition or its component is purified by sterilization and filtration before subdivision, and sterilized. It is preferable to dispense into a vial or the like by an operation and then subject to the freeze-drying treatment.
  • the frozen or freeze-dried human IFN-r composition of the present invention has very little decrease in F-r activity and quality during the freeze-freezing or freeze-drying operation and storage, and becomes turbid when re-dissolved! ) Is useful because it does not occur. Also, the lyophilized composition was obtained as a stabilized human IFN-r powder! In particular, it can be advantageously used as a parenteral preparation.
  • the freeze-dried human IFN-r composition of the present invention When used as a preparation for injection, the lyophilized composition is usually used in a vial, i.e., distilled water for injection or an injection solution of sugar, etc. Dissolve and use the solution within the physiologically acceptable range of osmotic pressure. It can be used as a dosage form for ophthalmic, otic and nasal administration using appropriate carriers, excipients and diluents.
  • the frozen or freeze-dried human IFN-II composition of the present invention has low toxicity, and can be used for the same purpose as known human IFN-r and in the same manner.
  • I U the international unit (I U) was determined as the activity of IFN-r (antiviral activity) as follows.
  • the international standard IFN- r whose unit (unit) was determined and the target data were measured using the cytopathic effect inhibition test of Sindbis Winores (S indbi Virus) against FL cell line derived from human amniotic membrane, and the ratio was used to determine the power. The value was calculated.
  • FIG. 1 shows the amino acid sequence of IFN-r contained in the aqueous solution of rIFN-r disclosed in Reference Example.
  • Example 1 was carried out in the same manner as in Example 1 except that hydroxyzinole starch (average molecular weight: 200,000) 37.5 ⁇ was added together with glycine 15. The results are shown in Table 1.
  • Example 1 was carried out in the same manner as in Example 1 except that glycine was increased to 30 ⁇ and sodium glutamate (7.5 W) was further added. The results are shown in Table 1.
  • the titer of the human IFN-r solution before freeze-drying was 94%.
  • the residual ratio to the titer at the start of storage after storage at 40 ° for one month was 100%, which was stable.
  • the titer of the human IFN-r solution before freeze-drying was 102%.
  • the residual ratio to the titer at the start of storage after storage at 40 ° for one month was stable at 106%.
  • a strain RRl (pR248cIts, 25 pRC23I / IF1-900) having a working human IFN-r gene was prepared according to the description in Example 8 of EPC 0 08 9 6 7 9 Cultured frozen cells were replaced with 100 M by 7 M Add 300 mM 100 mM Tris-HCl buffer ( ⁇ 7.0) containing guanidine hydrochloride and 2 mM phenylmethinolesulfonyl phenololeide, stir at 4 ° C for 1 hour, and centrifuge (17,000 rp 0 min) to obtain a clear supernatant. This supernatant was diluted with 1337 mM sodium chloride, .27.
  • the mixture was diluted 70-fold with a buffer solution containing potassium (hereinafter abbreviated as PBS), and the resulting precipitate was removed by a Sharples centrifuge (10, OO rpm). Then, the obtained supernatant solution 220 was subjected to Pellicon (Millipore Corporation).
  • PBS buffer solution containing potassium
  • Reference Example 1 3 of an aqueous solution (protein content; 0.952 / ml) containing human IFN-r in a high concentration obtained by the method of ( ⁇ ) was previously added to 10 mM reduced gnoretathion and 40 mM 25 mM acetate buffer containing sodium chloride
  • the specific activity of this IFN-r was 4.6 ⁇ 10 6 IU ⁇ ⁇ protein.
  • the FN- ⁇ concentration of this solution was 0.047 / d.
  • the solution was then aged for 2 days at 4 and then concentrated to 14 by the ultrafiltration method of Diaflo PM-10 membrane (Amicon's ultra furnace membrane), with a protein content of 0.711 ⁇ Substitute in real A clear rIFN-r solution that does not contain inorganic salts (less than 10 ppm) was obtained.
  • the thus obtained high concentration rIFN-r converged on the monomer by SDS-PAGE.
  • the specific activity of r I ⁇ 1 N—r was 3.6 10 6 KW-protein.
  • Substantially absent inorganic salts of the present invention human I 5 1 N of amino acid was sintered or lyophilized freeze under conditions that coexist - 7 "compositions, the frozen or freeze-drying operation and during storage In this case, the decrease in FN-r activity is extremely small, and no turbidity 9 is produced upon re-dissolving, so that it can be advantageously used as a pharmaceutical or the like.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Medicinal Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Animal Behavior & Ethology (AREA)
  • Epidemiology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Immunology (AREA)
  • Biophysics (AREA)
  • Biochemistry (AREA)
  • Genetics & Genomics (AREA)
  • Molecular Biology (AREA)
  • Toxicology (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Peptides Or Proteins (AREA)
  • Medicinal Preparation (AREA)

Abstract

Composition à base de gamma-interféron humain ne contenant sensiblement pas de sels inorganiques, et qui est congelée ou lyophilisée en présence d'acides aminés. Une solution aqueuse de gamma-interféron humain ne contenant sensiblement pas de sels inorganiques est préparée en utilisant un tampon exempt de sels inorganiques comme tampon destiné à être utilisé lors du processus chromatographique final ou en éliminant les sels inorganiques d'une solution aqueuse purifiée. Cette solution aqueuse peut être congelée ou lyophilisée en présence d'un acide aminé avec une réduction extrêmement faible de l'activité de l'interféron; la composition résultante possède une bonne stabilité au stockage et, lorsqu'elle est redissoute, ne forme pas de turbidité, ce qui permet de l'utiliser avantageusement comme agent antiviral, agent antinéoplastique, etc.
PCT/JP1984/000352 1984-07-10 1984-07-10 Composition a base de gamma-interferon Ceased WO1986000531A1 (fr)

Priority Applications (5)

Application Number Priority Date Filing Date Title
PCT/JP1984/000352 WO1986000531A1 (fr) 1984-07-10 1984-07-10 Composition a base de gamma-interferon
JP60148093A JPS6144826A (ja) 1984-07-10 1985-07-04 γ型インタ−フエロン組成物
EP85108409A EP0168008A3 (fr) 1984-07-10 1985-07-06 Composition stable d'interféron-gamma
ES544991A ES8608542A1 (es) 1984-07-10 1985-07-09 Un metodo para producir una composicion de gamma-interferon de ser humano.
KR1019850004888A KR860000869A (ko) 1984-07-10 1985-07-09 γ-인터페론 안정화 조성물의 제조방법

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
PCT/JP1984/000352 WO1986000531A1 (fr) 1984-07-10 1984-07-10 Composition a base de gamma-interferon

Publications (1)

Publication Number Publication Date
WO1986000531A1 true WO1986000531A1 (fr) 1986-01-30

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Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP1984/000352 Ceased WO1986000531A1 (fr) 1984-07-10 1984-07-10 Composition a base de gamma-interferon

Country Status (3)

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JP (1) JPS6144826A (fr)
KR (1) KR860000869A (fr)
WO (1) WO1986000531A1 (fr)

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS6156195A (ja) * 1985-03-01 1986-03-20 Asahi Chem Ind Co Ltd 新規生理活性ペプチド
JP2577744B2 (ja) * 1986-07-18 1997-02-05 中外製薬株式会社 安定な顆粒球コロニ−刺激因子含有製剤

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS55102519A (en) * 1979-01-31 1980-08-05 Green Cross Corp:The Stabilization of interferon
JPS5821691A (ja) * 1981-07-29 1983-02-08 Mochida Pharmaceut Co Ltd α−及びβ−インタ−フェロンの精製方法
JPS5892622A (ja) * 1981-11-28 1983-06-02 Sunstar Inc インタ−フエロン安定配合製剤

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS55102519A (en) * 1979-01-31 1980-08-05 Green Cross Corp:The Stabilization of interferon
JPS5821691A (ja) * 1981-07-29 1983-02-08 Mochida Pharmaceut Co Ltd α−及びβ−インタ−フェロンの精製方法
JPS5892622A (ja) * 1981-11-28 1983-06-02 Sunstar Inc インタ−フエロン安定配合製剤

Also Published As

Publication number Publication date
JPS6144826A (ja) 1986-03-04
KR860000869A (ko) 1986-02-20

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