WO2021249445A1 - 抗il-17a抗体药物组合物及其用途 - Google Patents

抗il-17a抗体药物组合物及其用途 Download PDF

Info

Publication number
WO2021249445A1
WO2021249445A1 PCT/CN2021/099257 CN2021099257W WO2021249445A1 WO 2021249445 A1 WO2021249445 A1 WO 2021249445A1 CN 2021099257 W CN2021099257 W CN 2021099257W WO 2021249445 A1 WO2021249445 A1 WO 2021249445A1
Authority
WO
WIPO (PCT)
Prior art keywords
seq
antibody
polysorbate
amino acid
acid sequence
Prior art date
Application number
PCT/CN2021/099257
Other languages
English (en)
French (fr)
Inventor
刘洪川
刘沛想
张静
王静
刘辉
李园园
冯辉
姚盛
Original Assignee
上海君实生物医药科技股份有限公司
苏州君盟生物医药科技有限公司
上海君实生物工程有限公司
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 上海君实生物医药科技股份有限公司, 苏州君盟生物医药科技有限公司, 上海君实生物工程有限公司 filed Critical 上海君实生物医药科技股份有限公司
Priority to EP21820830.4A priority Critical patent/EP4166159A1/en
Priority to CN202180041325.2A priority patent/CN115768476A/zh
Priority to US18/001,106 priority patent/US20230212278A1/en
Publication of WO2021249445A1 publication Critical patent/WO2021249445A1/zh

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • 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
    • A61K47/02—Inorganic compounds
    • 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
    • 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
    • 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
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • 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
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A61P11/06—Antiasthmatics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/12—Drugs for disorders of the urinary system of the kidneys
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/06—Antipsoriatics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • 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
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/06—Immunosuppressants, e.g. drugs for graft rejection
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/24—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against cytokines, lymphokines or interferons
    • C07K16/244—Interleukins [IL]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/20—Immunoglobulins specific features characterized by taxonomic origin
    • C07K2317/24—Immunoglobulins specific features characterized by taxonomic origin containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
    • C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/90—Immunoglobulins specific features characterized by (pharmaco)kinetic aspects or by stability of the immunoglobulin
    • C07K2317/94—Stability, e.g. half-life, pH, temperature or enzyme-resistance

Definitions

  • the present invention relates to the field of therapeutic pharmaceutical compositions.
  • the present invention relates to the field of pharmaceutical preparations, and the pharmaceutical composition contains a humanized antibody that specifically binds to IL-17A.
  • Interleukin 17, IL-17 also known as CTLA-8 or IL-17A, plays a key coordination role in the immune system.
  • IL-17A also known as CTLA-8 or IL-17A
  • IL-17A There are six members of the IL-17 family, including IL-17A, IL-17B, IL-17C, IL-17D, IL-17E, and IL-17F, all of which contain four critically conserved cysteine residues.
  • the biological effects of the members differ greatly.
  • IL-17A and IL-17F have the closest homology and biological function, and the current research is also the most in-depth.
  • IL-17A expressed in vivo has an N-terminal signal peptide of 23 amino acids, which is cleaved to produce mature IL-17A.
  • IL-17A is connected by disulfide bonds, usually secreted and present in the form of homodimer, and sometimes also connected with IL-17F to form heterodimer IL-17AF.
  • IL-17A or IL-17 refers to IL-17A homodimer protein, which is mainly produced by helper T cells 17 (T helper 17, Th17), and can also be produced by other immune cells such as ⁇ T cells, LTi (Lymphoid Tissue Inducer cells), ILCs (Innate Lymphoid Cells) and NKT (Natural Killer T) cells are synthesized and secreted.
  • helper T cells 17 T helper 17, Th17
  • LTi Lymphoid Tissue Inducer cells
  • ILCs Innate Lymphoid Cells
  • NKT Natural Killer T
  • Th17 cells are weak and secrete a small amount of IL-17A.
  • the damage effect is small; when IL-23 is present, it promotes the stability of Th17 cells and continues to secrete IL-17A, up-regulates pro-inflammatory factors (IL-22, CSF-2 and IFN- ⁇ ) and down-regulates anti-inflammatory factors (IL- 2. IL-27 and IL-12) expression and other pathways, causing inflammatory outbreaks and tissue damage. Therefore, when IL-23 is abnormally expressed in tissues, the IL-17A pathway plays a key role in tissue damage.
  • IL-17 is usually secreted at a specific site, and it binds to the IL-17 receptor (IL-17R) on the surface of target cells in local tissues to exert an effect.
  • the IL-17R family includes five members: IL-17RA, IL-17RB, IL-17RC, IL-17RD and IL-17RE, which are widely expressed on a variety of cell membranes.
  • IL-17 is mainly combined with IL-17RA/IL-17RC complex on the surface of cells derived from non-hematopoietic cells (e.g.
  • cytokines such as IL-6, G-CSF, GM-CSF, IL-10, TGF- ⁇ , TNF- ⁇
  • chemokines including IL-8, CXCL1 and MCP-1) and prostaglandins (for example, PGE2)
  • ROS reactive oxygen species
  • IL-17 Autoimmune diseases seriously threaten human health, such as psoriasis, rheumatoid arthritis, ankylosing spondylitis, Crohn's disease, multiple sclerosis, etc. Studies have found that the secretion of IL-17 is closely related to the occurrence and development of this type of disease. Antibodies targeting IL-17 can effectively alleviate the symptoms of autoimmune diseases by inhibiting the IL-17-IL-17R signaling pathway. Cosentyx (secukinumab) developed by Novartis is the world's first IL-17 monoclonal antibody.
  • Approved indications include moderate to severe plaque psoriasis (plaque psoriasis), providing an important frontline for the majority of psoriasis groups Biological treatment options.
  • plaque psoriasis plaque psoriasis
  • anti-IL-17 antibodies with different structures, better curative effects, and broader indications for the treatment of autoimmune related diseases such as psoriasis, rheumatoid joints, ankylosing spondylitis, and other IL-17 related diseases Diseases have urgent needs and important significance.
  • the pharmaceutical composition of the present invention is a highly stable pharmaceutical composition containing human antibodies specifically binding to IL-17A.
  • the present invention found that the combination of arginine hydrochloride and sucrose has unexpected characteristics, namely, high stability and low viscosity.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising: (1) a buffer; (2) an anti-IL-17A antibody or an antigen-binding fragment thereof.
  • the HCDR1 of the anti-IL-17A antibody or antigen-binding fragment thereof is selected from SEQ ID NO: 1, 7 and 13; HCDR2 is selected from 2, 8 and 14; HCDR3 is selected from 3, 9 and 15; LCDR1 is selected from From 4, 10, and 16; LCDR2 is selected from 5, 11, and 17; LCDR3 is selected from 6, 12, and 18.
  • the VH of the aforementioned anti-IL-17A antibody is selected from SEQ ID NOs: 19, 21, 23, and 26, and the VL is selected from SEQ ID NOs: 20, 22, 24, 25, and 27.
  • the HC of the anti-IL-17A antibody is selected from SEQ ID NO: 28, 30, 32, and 35; LC is selected from SEQ ID NO: 29, 31, 33, 34, and 36.
  • the above-mentioned anti-IL-17A antibody or antigen-binding fragment thereof has HCDR1, HCDR2, and HCDR3 as shown in SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3, and respectively as shown in SEQ ID. LCDR1, LCDR2, and LCDR3 shown in NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6.
  • the above-mentioned anti-IL-17A antibody or antigen-binding fragment thereof has HCDR1, HCDR2, and HCDR3 as shown in SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 9, respectively, and respectively as shown in SEQ ID LCDR1, LCDR2, and LCDR3 shown in NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12.
  • the above-mentioned anti-IL-17A antibody or antigen-binding fragment thereof has HCDR1, HCDR2, and HCDR3 as shown in SEQ ID NO: 13, SEQ ID NO: 14 and SEQ ID NO: 15, and respectively as shown in SEQ ID. LCDR1, LCDR2, and LCDR3 shown in NO: 16, SEQ ID NO: 17 and SEQ ID NO: 18.
  • the concentration of the anti-IL-17A antibody or antigen-binding fragment thereof in the pharmaceutical composition is about 1-300 mg/mL, preferably about 10-250 mg/mL, preferably about 20-200 mg/mL, preferably It is about 60-180 mg/mL, more preferably about 80-150 mg/mL; more preferably, the concentration of the anti-IL-17A antibody or antigen-binding fragment thereof is about 20 mg/mL, 30 mg/mL, 40 mg/mL, 50 mg/mL mL, 60mg/mL, 70mg/mL, 80mg/mL, 90mg/mL, 100mg/mL, 110mg/mL, 120mg/mL, 130mg/mL, 140mg/mL, 150mg/mL, 160mg/mL, 170mg/mL, 180 mg/mL or 200 mg/mL, preferably about 80 mg/mL or 150 mg/mL.
  • the above-mentioned buffer is selected from one or more of acetate buffer, citrate buffer, succinic acid buffer, and histidine buffer.
  • the above-mentioned buffer is a histidine buffer, preferably, the histidine buffer is selected from histidine-hydrochloride buffer or histidine-acetate buffer, preferably histidine Acid-hydrochloride buffer.
  • the aforementioned histidine-hydrochloride buffer is made of histidine and histidine hydrochloride, preferably L-histidine and L-histidine monohydrochloride.
  • the histidine buffer is made of 1-30 mM L-histidine and 1-30 mM L-histidine monohydrochloride.
  • the histidine buffer is made of histidine and histidine hydrochloride in a molar ratio of 1:1 to 1:4.
  • the histidine buffer is made of histidine and histidine hydrochloride in a molar ratio of 1:1.
  • the histidine buffer is made of histidine and histidine hydrochloride in a molar ratio of 1:3.
  • the histidine preparation is: a histidine buffer with a pH of 5.5 made of 4.5 mM L-histidine and 15.5 mM L-histidine monohydrochloride.
  • the histidine preparation is: a histidine buffer with a pH of 5.5 made of 7.5 mM L-histidine and 22.5 mM L-histidine monohydrochloride.
  • the histidine preparation is: a histidine buffer with a pH of 6.0 made of 15 mM histidine and 15 mM histidine hydrochloride.
  • the above-mentioned histidine buffer is a histidine-acetate buffer, preferably, the molar ratio of the two is 1:1 to 1.5:1, and preferably, the pH of such a buffer is 5.5 ⁇ 0.3, preferably about 5.5.
  • this type of buffer contains 15-20 mM histidine and 12-15 mM acetic acid.
  • the above-mentioned buffer is an acetate buffer.
  • the acetate buffer is an acetic acid-sodium acetate buffer or an acetic acid-potassium acetate buffer, preferably an acetic acid-sodium acetate buffer.
  • the above-mentioned buffer is a citric acid buffer, and preferably, the citric acid buffer is a citric acid-sodium citrate buffer.
  • the above-mentioned buffer is a succinic acid buffer, and preferably, the succinic acid buffer is a succinic acid-sodium succinate buffer.
  • the concentration of the above-mentioned buffer is about 1-200mM, preferably about 5-200mM, preferably about 10-50mM, preferably about 10-30mM; preferably about 10-20mM, the concentration of the above-mentioned buffer is not limited Typical examples are about 5mM, 10mM, 15mM, 20mM, 25mM, 30mM, 40mM, 45mM, 50mM, 60mM, 70mM, 80mM, 90mM, 100mM, 105mM, 110mM, 115mM, 120mM, 130mM, 140mM, 150mM, 160mM, 170mM Or 180 mM or any two values within these ranges as a range formed by endpoints, preferably 10 mM, 15 mM, 20 mM or 30 mM.
  • the pH of the aforementioned buffer is about 5.0-6.5, preferably about 5.0-6.0, preferably about 5.5-6.5, preferably about 5.0-5.5, preferably about 5.5-6.0, preferably about 6.0-6.5 ,
  • a non-limiting example of the pH of the above buffer is about 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, preferably about 5.0, 5.5 or 6.0.
  • the aforementioned pharmaceutical composition further includes a stabilizer selected from the group consisting of arginine hydrochloride, proline, glycine, sodium chloride, mannitol, sorbitol, sucrose, maltose, xylitol, and seaweed One or more types of sugar.
  • a stabilizer selected from the group consisting of arginine hydrochloride, proline, glycine, sodium chloride, mannitol, sorbitol, sucrose, maltose, xylitol, and seaweed One or more types of sugar.
  • the concentration of the aforementioned stabilizer is about 10 mM to 400 mM, preferably 20 mM to 300 mM, more preferably 30 mM to 200 mM.
  • the stabilizer is sodium chloride at a concentration of about 30-200 mM; or the stabilizer is mannitol at a concentration of about 100-300 mM; or the stabilizer is sorbitol at a concentration of about 100-300 mM; or The stabilizer is sucrose with a concentration of about 100-300 mM; or the stabilizer is trehalose with a concentration of about 100-300 mM; or the stabilizer is arginine hydrochloride with a concentration of about 30-200 mM; or the stabilizer Is proline with a concentration of about 100-300 mM; or the stabilizer is glycine with a concentration of about 100-300 mM.
  • the stabilizer is a combination of about 30-200 mM sodium chloride and about 30-200 mM mannitol; or the stabilizer is about 30-200 mM sodium chloride and about 30-200 mM sorbitol Or the stabilizer is a combination of about 30-200mM sodium chloride and about 30-200mM sucrose; or the stabilizer is about 30-200mM arginine hydrochloride and about 30-200mM mannitol Or the stabilizer is a combination of about 30-200mM arginine hydrochloride and about 30-200mM sucrose; or the stabilizer is about 30-200mM arginine hydrochloride and about 30-200mM sorbus A combination of alcohol; or the stabilizer is a combination of about 30-200 mM arginine hydrochloride and about 30-200 mM trehalose.
  • the aforementioned stabilizer is sodium chloride. In some embodiments, the aforementioned stabilizer is sodium chloride at a concentration of about 30-200 mM.
  • the concentration of the aforementioned sodium chloride is preferably about 50-190 mM, preferably about 100-180 mM, preferably about 120-170 mM, preferably about 130. -150mM, non-limiting examples of the above sodium chloride concentration are about 100mM, 110mM, 120mM, 125mM, 130mM, 135mM, 140mM, 145mM, 150mM, 155mM, 160mM, 170mM, 180mM, 190mM, 200mM, preferably 135mM or 140mM .
  • the aforementioned stabilizer is mannitol.
  • the aforementioned stabilizer is mannitol at a concentration of about 100-300 mM, the concentration of the aforementioned mannitol is preferably about 150-300 mM, preferably about 200-280 mM, and a non-limiting example of the aforementioned mannitol concentration is about 200 mM. , 210mM, 220mM, 230mM, 240mM, 250mM, 260mM, 270mM, 280mM, preferably 240mM.
  • the aforementioned stabilizer is sorbitol.
  • the aforementioned stabilizer is sorbitol at a concentration of about 100-300 mM, the concentration of the aforementioned sorbitol is preferably about 150-300 mM, preferably about 200-280 mM, and a non-limiting example of the aforementioned sorbitol concentration is about 200 mM. , 210mM, 220mM, 230mM, 240mM, 250mM, 260mM, 270mM, 280mM, preferably 240mM.
  • the aforementioned stabilizer is sucrose. In some embodiments, the aforementioned stabilizer is sucrose at a concentration of about 100-300 mM. The concentration of the aforementioned sucrose is preferably about 150-300 mM, preferably about 200-280 mM. A non-limiting example of the aforementioned sucrose concentration is about 200 mM, 210 mM, 220mM, 230mM, 240mM, 250mM, 260mM, 270mM, 280mM, preferably 220mM.
  • the aforementioned stabilizer is trehalose.
  • the aforementioned stabilizer is trehalose at a concentration of about 100-300 mM, the concentration of the aforementioned trehalose is preferably about 150-300 mM, preferably about 200-280 mM, and a non-limiting example of the aforementioned trehalose concentration is about 180 mM. , 200mM, 210mM, 220mM, 230mM, 240mM, 250mM, 260mM, 270mM, 280mM, preferably 220mM.
  • the aforementioned stabilizer is arginine hydrochloride.
  • the aforementioned stabilizer is arginine hydrochloride at a concentration of about 30-200 mM, and the concentration of the aforementioned arginine hydrochloride is preferably about 50-190 mM, preferably about 100-180 mM, preferably about 120-170 mM, preferably about It is 130-150mM, the non-limiting example of the above-mentioned arginine hydrochloride concentration is about 100mM, 110mM, 120mM, 125mM, 130mM, 135mM, 140mM, 145mM, 150mM, 155mM, 160mM, 170mM, 180mM, 190mM, 200mM, preferably 135mM or 140mM.
  • the aforementioned stabilizer is proline. In some embodiments, the aforementioned stabilizer is proline at a concentration of about 100-300 mM. The concentration of the aforementioned proline is preferably about 150-300 mM, preferably about 200-280 mM. A non-limiting example of the aforementioned proline concentration It is about 180 mM, 200 mM, 210 mM, 220 mM, 230 mM, 240 mM, 250 mM, 260 mM, 270 mM, 280 mM, preferably 240 mM.
  • the aforementioned stabilizer is glycine. In some embodiments, the aforementioned stabilizer is glycine at a concentration of about 100-300 mM. The concentration of the aforementioned glycine is preferably about 150-300 mM, preferably about 200-280 mM. A non-limiting example of the aforementioned glycine concentration is about 180 mM, 200 mM, 210mM, 220mM, 230mM, 240mM, 250mM, 260mM, 270mM, 280mM, preferably 260mM.
  • the aforementioned stabilizer is a combination of sodium chloride and mannitol. In some aspects, the aforementioned stabilizer is a combination of about 30-200 mM sodium chloride and about 30-200 mM mannitol, preferably a combination of about 40-150 mM sodium chloride and about 40-180 mM mannitol, preferably about A combination of 40-100 mM sodium chloride and about 40-150 mM mannitol, a non-limiting example of the above stabilizer is a combination of about 50 mM sodium chloride and about 120 mM mannitol, and 50 mM sodium chloride and about A combination of 140 mM mannitol, a combination of about 90 mM sodium chloride and about 50 mM mannitol.
  • the aforementioned stabilizer is a combination of sodium chloride and sucrose. In some aspects, the aforementioned stabilizer is a combination of about 30-200 mM sodium chloride and about 30-200 mM sucrose, preferably a combination of about 40-150 mM sodium chloride and about 40-180 mM sucrose, preferably about 40-180 mM sucrose.
  • a combination of 100 mM sodium chloride and about 40-150 mM sucrose, a non-limiting example of the above stabilizer is a combination of about 50 mM sodium chloride and about 120 mM sucrose, about 90 mM sodium chloride and about 50 mM sucrose The combination.
  • the aforementioned stabilizer is a combination of arginine hydrochloride and mannitol. In some embodiments, the aforementioned stabilizer is a combination of about 30-200 mM arginine hydrochloride and about 30-200 mM mannitol, preferably a combination of about 40-150 mM arginine hydrochloride and about 40-100 mM mannitol, A combination of about 60-120 mM arginine hydrochloride and about 40-80 mM mannitol is preferred. A non-limiting example of the above stabilizer is a combination of about 90 mM arginine hydrochloride and about 50 mM mannitol.
  • the aforementioned stabilizer is a combination of arginine hydrochloride and sucrose.
  • the aforementioned stabilizer is a combination of about 30-200 mM arginine hydrochloride and about 30-200 mM sucrose, preferably a combination of about 40-150 mM arginine hydrochloride and about 40-100 mM sucrose, preferably about A combination of 60-120 mM arginine hydrochloride and about 40-80 mM sucrose.
  • a non-limiting example of the above stabilizer is a combination of about 90 mM arginine hydrochloride and about 50 mM sucrose.
  • the aforementioned stabilizer is arginine hydrochloride at a concentration of about 120 mM to about 170 mM; or a combination of arginine hydrochloride at a concentration of about 60 mM to about 120 mM and sucrose at about 40-80 mM; or hydrochloric acid at a concentration of about 60 mM to about 120 mM. Combination of arginine and about 40-80 mM mannitol.
  • the above-mentioned pharmaceutical composition further includes a surfactant, and the surfactant is selected from polysorbate 80, polysorbate 20, or poloxamer 188.
  • the above-mentioned surfactant is selected from polysorbate 80.
  • the above-mentioned surfactant is selected from polysorbate 20.
  • the concentration of the above-mentioned surfactant is about 0.001%-0.1%, preferably about 0.01%-0.1%, preferably about 0.02%-0.08%, more preferably about 0.02%-0.04 %; As a non-limiting example, the concentration of the above-mentioned surfactant is about 0.02%, 0.04% or 0.08%, preferably 0.02%.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof is selected from murine antibodies, chimeric antibodies, and humanized antibodies, preferably humanized antibodies.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain variable region as shown in SEQ ID NO: 19 and a light chain variable region as shown in SEQ ID NO: 20.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain variable region as shown in SEQ ID NO: 21 and a light chain variable region as shown in SEQ ID NO: 20.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain variable region as shown in SEQ ID NO: 21 and a light chain variable region as shown in SEQ ID NO: 22.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain variable region as shown in SEQ ID NO: 23 and a light chain variable region as shown in SEQ ID NO: 24.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain variable region as shown in SEQ ID NO: 23 and a light chain variable region as shown in SEQ ID NO: 25.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain variable region as shown in SEQ ID NO: 26 and a light chain variable region as shown in SEQ ID NO: 27.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain amino acid sequence as shown in SEQ ID NO: 28 and a light chain amino acid sequence as shown in SEQ ID NO: 29.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain amino acid sequence as shown in SEQ ID NO: 30 and a light chain amino acid sequence as shown in SEQ ID NO: 29.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain amino acid sequence as shown in SEQ ID NO: 30 and a light chain amino acid sequence as shown in SEQ ID NO: 31.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain amino acid sequence as shown in SEQ ID NO: 32 and a light chain amino acid sequence as shown in SEQ ID NO: 33.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain amino acid sequence as shown in SEQ ID NO: 32 and a light chain amino acid sequence as shown in SEQ ID NO: 34.
  • the aforementioned anti-IL-17A antibody or antigen-binding fragment thereof has a heavy chain amino acid sequence as shown in SEQ ID NO: 35 and a light chain amino acid sequence as shown in SEQ ID NO: 36.
  • the above-mentioned pharmaceutical composition comprises the following components shown in any one of (1) to (6), wherein the anti-IL-17A antibody or antigen-binding fragment thereof is as described in any one of the embodiments of the present invention :
  • the above-mentioned pharmaceutical composition comprises the following components as shown in any one of (i) to (vi), wherein the anti-IL-17A antibody or antigen-binding fragment thereof is as described in any one of the embodiments of the present invention :
  • the pharmaceutical composition is a liquid formulation or a lyophilized formulation.
  • the pharmaceutical composition is a liquid formulation.
  • the above-mentioned liquid formulation or lyophilized formulation is stable at 2-8°C for at least 3 months, at least 6 months, at least 12 months, at least 18 months, or at least 24 months.
  • the above-mentioned aqueous solution or lyophilized formulation is stable at 40°C for at least 7 days, at least 14 days, or at least 28 days.
  • the present invention also provides the use of the above-mentioned pharmaceutical composition in the preparation of drugs for treating or preventing IL-17A-mediated diseases.
  • the aforementioned diseases include inflammatory diseases and autoimmune diseases, such as arthritis, rheumatoid arthritis, ankylosing spondylitis, chronic obstructive pulmonary disease, systemic lupus erythematosus (SLE), lupus nephritis, Asthma, multiple sclerosis, cystic fibrosis, psoriasis.
  • inflammatory diseases and autoimmune diseases such as arthritis, rheumatoid arthritis, ankylosing spondylitis, chronic obstructive pulmonary disease, systemic lupus erythematosus (SLE), lupus nephritis, Asthma, multiple sclerosis, cystic fibrosis, psoriasis.
  • Figure 1 ELISA to detect the binding effect of humanized antibody and human IL-17A.
  • the antibodies hu31, hu43, hu44, hu59, hu60 and hu250 have high specificity in binding to human IL-17A, and their EC50 is 8.13ng/mL, 8.64ng/mL, 6.764ng/ml, 6.102ng/mL, respectively, 5.776ng/mL and 6.351ng/mL.
  • Figure 2 FACS detection of humanized antibody blocking the binding of human IL-17A to IL-17RA on 293F cells.
  • the antibodies hu31, hu43, hu44, hu59, hu60 and hu250 all have efficient blocking effects, with IC50 of 867.6ng/mL, 780.8ng/mL, 828.5ng/ml, 467.4ng/mL, 482.8ng/mL and 577.8ng/mL.
  • Figure 3 ELISA to detect the effect of humanized antibody blocking IL-17A-mediated epithelial cell secretion of CXCL1.
  • the antibodies hu31, hu43, hu44, hu59, hu60 and hu250 effectively inhibit IL-17A-induced epithelial cells to express CXCL1, and their blocking effect is stronger than that of the control antibody.
  • Figure 4 ELISA detection of humanized antibodies to block IL-17A-mediated biological activity in vivo.
  • the antibodies hu31, hu43, hu44, hu60 and hu250 can effectively inhibit IL-17A-induced expression of CXCL1 in mice, and the blocking effect is stronger than that of the control antibody.
  • Figure 5-1 The effect of humanized antibody administration on the clinical score of imiquimod-induced psoriasis mice.
  • the administration of hu31 and hu44 can significantly inhibit the skin scaly, induration, redness and swelling of the mouse psoriasis model induced by imiquimod, that is, the clinical score is reduced (*P ⁇ 0.05vs KLH).
  • Figure 5-2 The effect of humanized antibody on the degree of ear swelling in mice with psoriasis induced by imiquimod. Administration of hu31 and hu44 can significantly improve ear swelling. (*P ⁇ 0.05vs KLH, **P ⁇ 0.01vs KLH.).
  • Figure 6-1 The effect of humanized antibodies on the body weight of female cynomolgus monkeys induced by type II collagen. Hu31 and hu59 have a certain improvement effect on weight loss caused by arthritis (**P ⁇ 0.01, ****P ⁇ 0.0001, compared with "G2: vehicle group”; One-way ANOVA/Dunnett).
  • Figure 6-2 The effect of humanized antibody on the score of female cynomolgus monkey arthritis induced by type II collagen. hu31 significantly inhibited the increasing trend of clinical scores of cynomolgus arthritis (***P ⁇ 0.001, #P ⁇ 0.05, compared with G2: vehicle group; One-way ANOVA/Dunnett).
  • Figure 7 The effect of humanized antibody on mouse joint swelling induced by NIH3T3-IL17 cells.
  • Figure 8 The effect of humanized antibody on air sac inflammation in mice induced by NIH3T3-IL17 cells.
  • composition means a mixture containing one or more of the antibodies described herein and other components, such as physiologically pharmaceutically acceptable carriers and excipients.
  • the purpose of the pharmaceutical composition is to promote the administration to the organism, facilitate the absorption of the active ingredients and then exert the biological activity.
  • liquid formulation refers to a formulation in a liquid state, and is not intended to refer to a lyophilized formulation that is resuspended.
  • the liquid formulation of the present invention is stable during storage, and its stability does not depend on lyophilization (or other state change methods, such as spray drying).
  • aqueous liquid formulation refers to a liquid formulation using water as a solvent.
  • an aqueous liquid formulation is a formulation that does not require lyophilization, spray drying, and/or freezing to maintain stability (e.g., chemical and/or physical stability and/or biological activity).
  • excipient refers to an agent that can be added to a formulation to provide desired characteristics (e.g., consistency, increased stability) and/or to adjust osmotic pressure.
  • desired characteristics e.g., consistency, increased stability
  • excipients include, but are not limited to, sugars, polyols, amino acids, surfactants, and polymers.
  • the "about” used in this application when referring to a measurable value is intended to cover a variation of ⁇ 20% or ⁇ 10% relative to a specific value, including ⁇ 5%, ⁇ 1%, and ⁇ 0.1 %, because these changes are suitable for carrying out the disclosed method.
  • buffer pH is about 5.0-6.5
  • the buffer used in the formulation of the present invention may have a pH in the range of about 5.0 to about 6.5, or a pH in the range of about 5.5 to about 6.5, or a pH in the range of about 5.0 to about 6.0.
  • examples of the "buffer” for controlling the pH within this range include acetate (such as sodium acetate), succinate (such as sodium succinate), gluconic acid, histidine, histidine hydrochloride , Methionine, citrate, phosphate, citrate/phosphate, imidazole, acetic acid, acetate, citrate, combinations thereof and other organic acid buffers.
  • Hetidine buffer is a buffer containing histidine ions.
  • histidine buffers include histidine and histidine salts, such as histidine hydrochloride, histidine acetate, histidine phosphate and histidine sulfate, etc., such as containing histidine
  • the histidine buffer of acid and histidine hydrochloride; the histidine buffer of the present invention also includes histidine buffer containing histidine and acetate (such as sodium salt or potassium salt).
  • citrate buffer is a buffer that includes citrate ions.
  • citrate buffers include citric acid-sodium citrate, citric acid-potassium citrate, citric acid-calcium citrate, citric acid-magnesium citrate, and the like.
  • the preferred citrate buffer is citric acid-sodium citrate buffer.
  • Acetate buffer is a buffer that includes acetate ions.
  • acetate buffers include acetic acid-sodium acetate, acetic acid-potassium acetate, acetic acid-calcium acetate, acetic acid-magnesium acetate, and the like.
  • the preferred acetate buffer is acetic acid-sodium acetate buffer.
  • succinate buffer is a buffer that includes succinate ions.
  • succinate buffer include succinate-sodium succinate, succinate-potassium succinate, succinate-calcium succinate, succinate-magnesium succinate, and the like.
  • the preferred succinate buffer is succinate-sodium succinate buffer.
  • stabilizer refers to a pharmaceutically acceptable excipient that protects the active pharmaceutical ingredient and/or formulation from chemical and/or physical degradation during manufacture, storage, and application.
  • Stabilizers include but are not limited to sugars, amino acids, salts, polyols and their metabolites as defined below, such as sodium chloride, calcium chloride, magnesium chloride, mannitol, sorbitol, sucrose, trehalose, arginine or its Salts (such as arginine hydrochloride), glycine, alanine ( ⁇ -alanine, ⁇ -alanine), betaine, leucine, lysine, glutamic acid, aspartic acid, proline Acid, 4-hydroxyproline, sarcosine, ⁇ -aminobutyric acid (GABA), opines, alanine, octopine, glycinine (strombine) and trimethylamine N- Oxide (TMAO), human serum albumin (hsa), bovine serum albumin
  • Some stabilizers such as sodium chloride, calcium chloride, magnesium chloride, mannitol, sorbitol, sucrose, etc. can also play a role in controlling osmotic pressure.
  • the stabilizer specifically used in the present invention is selected from one or more of polyols, amino acids, salts, and sugars.
  • the preferred salt is sodium chloride
  • the preferred sugars are sucrose and trehalose
  • the preferred polyols are sorbitol and mannitol.
  • Preferred amino acids are arginine or its salts (such as arginine hydrochloride), glycine, and proline.
  • Preferred stabilizers are sodium chloride, mannitol, sorbitol, sucrose, trehalose, arginine hydrochloride, glycine, proline, sodium chloride-sorbitol, sodium chloride-mannitol, sodium chloride-sucrose , Sodium chloride-trehalose, arginine hydrochloride-mannitol, arginine hydrochloride-sucrose, more preferably arginine hydrochloride, sodium chloride-sucrose, arginine hydrochloride-mannitol, arginine hydrochloride- Sucrose is more preferably arginine hydrochloride-sucrose.
  • viscosity as used herein may be “kinematic viscosity” or “absolute viscosity”.
  • Kininematic viscosity is a measure of the resistive flow of a fluid under the influence of gravity.
  • the dimension of kinematic viscosity is L2/T, where L is length and T is time. Generally, kinematic viscosity is expressed in centistokes (cSt).
  • the SI unit of kinematic viscosity is mm 2 /s, which is lcSt.
  • the absolute viscosity is expressed in centipoise (cP) units.
  • the term "low-level viscosity" as used herein means an absolute viscosity of less than about 15 centipoise (cP).
  • cP centipoise
  • the formulation exhibits an absolute viscosity of about 15cP, about 14cP, about 13cP, about 12cP, about 11cP, about 10cP, about 9cP, about 8cP, about 7cP, about 6cP, about 5cP, about 4cP, about 3cP, about 2cP, about 1cP, or lower, the pharmaceutical composition of the present invention will be considered to have "low viscosity".
  • the term "medium level viscosity" as used herein will mean an absolute viscosity between about 35 cP and about 15 cP.
  • the formulation exhibits an absolute viscosity of about 34cP, about 33cP, about 32cP, about 31cP, about 30cP, about 29cP, about 28cP, about 27cP, about 26cP, about 25cP, about 24cP, about 23cP, about 22cP, about 21cP, about 20cP, about 19cP, 18cP, about 17cP, about 16cP, or about 15.lcP
  • the pharmaceutical composition of the present invention will be considered to have a "medium viscosity”.
  • the pharmaceutical composition of the present invention may exhibit a low level of viscosity in certain embodiments.
  • comparing the viscosity of different excipients it is found that arginine or its salt can obtain a viscosity level significantly lower than that of other excipients.
  • it is found from the comparison of the buffer system that the viscosity level of the histidine buffer system is lower than other buffer systems.
  • surfactant generally includes agents that protect proteins, such as antibodies, from stress induced by the air/solution interface, solution/surface-induced stress to reduce antibody aggregation or minimize the formation of particulates in the formulation.
  • exemplary surfactants include, but are not limited to, nonionic surfactants such as polyoxyethylene sorbitan fatty acid esters (such as polysorbate 20 and polysorbate 80), polyethylene-polypropylene copolymers, poly Ethylene-polypropylene glycol, polyoxyethylene-stearate, polyoxyethylene alkyl ether, such as polyoxyethylene monolauryl ether, alkylphenyl polyoxyethylene ether (Triton-X), polyoxyethylene- Polyoxypropylene copolymer (Pluronic), sodium dodecyl sulfate (SDS).
  • nonionic surfactants such as polyoxyethylene sorbitan fatty acid esters (such as polysorbate 20 and polysorbate 80), polyethylene-polypropylene copolymers,
  • isotonic means that the preparation has substantially the same osmotic pressure as human blood. Isotonic formulations generally have an osmotic pressure of about 250 to 350 mOsm. The isotonicity can be measured using a vapor pressure or freezing point drop osmometer.
  • stable formulation is a formulation in which the antibody substantially maintains its physical and/or chemical stability and/or biological activity during the manufacturing process and/or during storage. Even if the contained antibody fails to maintain 100% of its chemical structure or biological function after a certain period of storage, the pharmaceutical preparation can be stable. In some cases, after a certain period of storage, it can maintain about 90%, about 95%, about 96%, about 97%, about 98%, or about 99% of the structure or function of the antibody, which can also be considered as " stable”.
  • Various analytical techniques for measuring protein stability are available in this technical field, and are reviewed in "Peptide and Protein Drug Delivery” 247-301, Vincent Lee, Editor-in-Chief, Marcel Dekker, Inc. ., New York, NY, Pubs. (1991)), and Jones, A. (1993) Adv. Drug Delivery Rev. 10: 29-90 (both are incorporated by reference).
  • the stability of the protein is determined in terms of the percentage of monomeric protein in a solution that has a low percentage of degradation (e.g., fragmentation) and/or aggregated protein.
  • the formulation can be stored stably at room temperature, about 25-30°C, or 40°C for at least 2 weeks, at least 28 days, at least 1 month, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, at least 12 months, at least 18 months, at least 24 months, or Longer, no more than about 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1% antibody in aggregate form at most.
  • the percentage (and other methods) of antibody (and other methods) that migrate in the more acidic fraction of this antibody main fraction ("mainly charged form") during ion exchange stability can be measured, where stability is the same as The percentage of acidic form of antibody is inversely proportional.
  • the percentage of "acidified" antibodies can be measured by ion exchange chromatography, such as cation exchange high performance liquid chromatography [CEX-HPLC].
  • an acceptable degree of stability means that when the formulation is stored at a certain temperature for a certain period of time, the acidic form of the antibody that can be detected therein does not exceed about 49%, 45%, 40%, 35 %, 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2%, 1%, 0.5% or 0.1%.
  • the certain period of storage before measuring stability can be at least 2 weeks, at least 28 days, at least 1 month, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, at least 12 months, at least 18 months, at least 24 months, or longer.
  • a certain temperature that allows the storage of pharmaceutical preparations can be any temperature in the range of about -80°C to about 45°C, for example, storage at about -80°C, about -30°C, about -20°C, or about 0°C , About 2-8°C, about 5°C, about 25°C, or about 40°C.
  • the antibody does not substantially show signs of aggregation, precipitation, and/or denaturation during visual inspection of color and/or clarity or by UV light scattering or by pore exclusion chromatography, then the antibody is in the drug combination
  • the substance "maintains its physical stability". Aggregation is the process by which single molecules or complexes associate covalently or non-covalently to form aggregates. Aggregation can proceed to the point where visible precipitates are formed.
  • the stability of the formulation can be evaluated by methods known in the art, including measuring the apparent extinction (absorbance or optical density) of the sample. Such extinction measurement is related to the turbidity of the formulation.
  • the turbidity of a formulation is partly an inherent property of the protein dissolved in the solution, and is usually measured by turbidimetry and measured in turbidity units (NTU).
  • the level of turbidity that varies with, for example, the concentration of one or more components in the solution (eg, protein and/or salt concentration) is also referred to as the "opacity" or "opaque appearance" of the formulation.
  • the turbidity level can be calculated with reference to a standard curve generated using a suspension of known turbidity.
  • the reference standard used to determine the turbidity level of the pharmaceutical composition can be based on the standards of the European Pharmacopoeia (European Pharmacopoeia, fourth edition, Directorate for the Quality of Medicine). of the Council of Europe (EDQM), France).
  • a clear solution is defined as a solution with a turbidity lower than or equal to the turbidity of a reference suspension of about 3 according to the "European Pharmacopoeia” standard.
  • Turbidity measurement by turbidimetry can detect Rayleigh scattering in the absence of association or non-ideal effects, which usually varies linearly with concentration. Other methods for assessing physical stability are well known in the art.
  • the antibody "retains its chemical stability" in the pharmaceutical composition.
  • the chemical stability can be assessed by, for example, detecting or quantifying the form of chemical changes of the antibody.
  • Chemical changes can include size changes (e.g., cropping), which can be assessed using, for example, size exclusion chromatography, SDS-PAGE, and/or matrix-assisted laser desorption/time-of-flight mass spectrometry (MALDI/TOF MS).
  • Other types of chemical changes include charge changes (for example, occurring as a result of deamidation or oxidation), which can be assessed by, for example, ion exchange chromatography.
  • the antibody in the pharmaceutical composition is biologically active for its intended purpose, the antibody "retains its biological activity" in the pharmaceutical composition.
  • the preparation is stored at temperatures such as 5°C, 25°C, and 45°C for a certain period of time (for example, 1 to 12 months)
  • the binding affinity of the anti-IL-17A antibody contained in the preparation to IL-17A is the storage
  • the binding affinity of the antibody is at least 90%, 95% or more, the formulation of the present invention can be considered stable.
  • the binding affinity can also be determined using techniques such as ELISA or plasmon resonance.
  • the "therapeutically effective amount” or “effective amount” of the antibody refers to an amount effective in preventing or treating or alleviating the symptoms of a disorder that the antibody can effectively treat.
  • the "therapeutically effective dose” or “therapeutically effective dose” of the drug is any amount of drug that protects the subject from the onset of disease or promotes the regression of the disease when used alone or in combination with another therapeutic agent. The regression of the disease is evidenced by a reduction in the severity of the symptoms of the disease, an increase in the frequency and duration of the asymptomatic period of the disease, or the prevention of injury or disability caused by the pain of the disease.
  • a therapeutically effective amount of a drug includes a "prophylactically effective amount", that is, any amount that inhibits the development or recurrence of the disease when administered to a subject at risk or a subject who has a recurrence of the disease, alone or in combination with other therapeutic drugs Drug.
  • subject or "patient” is intended to include mammalian organisms.
  • subjects/patients include humans and non-human mammals, such as non-human primates, dogs, cows, horses, pigs, sheep, goats, cats, mice, rabbits, rats, and transgenic non-human animals.
  • the subject is a human.
  • Anti-PD-1 antibody administration routes include intravenous, intramuscular, subcutaneous, peritoneal, spinal, or other parenteral administration routes, such as injection or infusion.
  • Parenter administration refers to administration methods usually by injection other than enteral or local administration, including but not limited to intravenous, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, and intrasaccular , Intraframe, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subepidermal, intraarticular, subcapsular, subarachnoid, intraspine, intradural and intrasternal injection and infusion, and intracorporeal electroporation.
  • antibody as used herein should be understood to include intact antibody molecules and antigen-binding fragments thereof.
  • antigen-binding portion or “antigen-binding fragment” (or simply “antibody portion” or “antibody fragment”) of an antibody as used herein refers to an antibody that retains human IL-17A (interleukin 17A)) or One or more fragments of its epitope-specific binding ability.
  • full-length antibody or “complete antibody molecule” as used herein refers to an immunoglobulin molecule comprising four peptide chains, two heavy (H) chains (about 50-70kDa at full length) and two light (L) chains (Approximately 25kDa at full length) are connected to each other by disulfide bonds.
  • Each heavy chain is composed of a heavy chain variable region (abbreviated as VH herein) and a heavy chain constant region (abbreviated as CH herein).
  • the heavy chain constant region is composed of three structural domains CH1, CH2 and CH3.
  • Each light chain is composed of a light chain variable region (abbreviated as VL herein) and a light chain constant region.
  • the constant region of the light chain consists of a domain CL.
  • VH and VL regions can be further subdivided into complementarity determining regions (CDR) with high variability and more conservatively spaced regions called framework regions (FR).
  • CDR complementarity determining regions
  • FR framework regions
  • Each VH or VL region consists of 3 CDRs and 4 FRs arranged in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4 from the amino terminus to the carboxy terminus.
  • the variable regions of the heavy and light chains contain binding domains that interact with antigens.
  • the constant region of the antibody can mediate the binding of immunoglobulins to host tissues or factors (including various cells of the immune system (for example, effector cells) and the first component (Clq) of the classical complement system).
  • CDR refers to the complementarity determining region within the variable sequence of an antibody.
  • the precise boundaries of these CDRs have different definitions according to different systems.
  • the system described by Kabat (supra) not only provides a clear residue numbering system applicable to any variable region of an antibody, but also provides precise residue boundaries that define the 3 CDRs.
  • These CDRs can be called Kabat CDRs. Chothia et al.
  • CDR boundary definitions may not strictly follow one of the systems described in this article, but still overlap with Kabat CDR, although they may be shortened or lengthened due to specific residues or residue groups or even discovered based on predictions or experiments The entire CDR does not significantly affect antigen binding.
  • the methods used herein can utilize CDRs defined in accordance with any of these systems, although certain embodiments use CDRs defined by Kabat or Chothia.
  • antigen-binding fragment includes antibody fragments or derivatives thereof, and generally includes at least one fragment of the antigen-binding region or variable region (eg, one or more CDRs) of the parent antibody, which retains at least some of the parent antibody Binding specificity.
  • antigen-binding fragments include, but are not limited to, Fab, Fab', F(ab')2 and Fv fragments; diabodies; linear antibodies; single-chain antibody molecules, such as sc-Fv; nanobodies formed from antibody fragments (nanobody) And multispecific antibodies.
  • the binding fragment or derivative thereof When the binding activity of an antibody is expressed on a molar concentration basis, the binding fragment or derivative thereof generally retains at least 10% of the antigen binding activity of the parent antibody.
  • the binding fragment or derivative thereof retains at least 20%, 50%, 70%, 80%, 90%, 95% or 100% or more of the antigen binding affinity of the parent antibody. It is also expected that the antigen-binding fragment of an antibody may include conservative or non-conservative amino acid substitutions that do not significantly alter its biological activity (referred to as “conservative variants” or “functionally conservative variants” of the antibody).
  • the anti-IL-17A antibody or antigen-binding fragment thereof of the present invention includes any one of the anti-IL-17A antibodies described in the application number PCT/CN2019/124486, and the entire content disclosed herein is incorporated herein by way of introduction.
  • the CDR sequences of the antibodies used in the methods and compositions of the present invention include the CDR sequences from the antibody hu31 described in PCT/CN2019/124486.
  • the non-limiting and exemplary antibodies used in the examples herein are selected from the humanized antibodies hu31, hu43, hu44, hu59, hu60 and hu250 described in CN201811515045.7.
  • the variable regions and their amino acid CDR sequences are shown in Table 1.
  • the full-length amino acid sequence of its light/heavy chain is shown in Table 2.
  • HC Humanized antibody Heavy chain
  • LC Light chain
  • the pharmaceutical composition of the present invention is a highly stable pharmaceutical composition containing a humanized antibody specifically binding to IL-17A.
  • the present invention found that the combination of arginine salt (such as arginine hydrochloride) and sucrose can significantly improve the stability of the pharmaceutical composition.
  • the pharmaceutical composition of the present invention is a liquid preparation that contains a high concentration of active antibodies and has high stability and low viscosity.
  • the present invention found that the viscosity of the formulation containing arginine salt was significantly lower than that of the formulation containing only other auxiliary materials.
  • the viscosity level of the histidine buffer system is significantly lower than other buffer systems.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising: (1) a buffer; (2) an anti-IL-17A antibody or an antigen-binding fragment thereof.
  • the antibodies in the pharmaceutical composition of the present invention may be murine antibodies, chimeric antibodies, humanized antibodies, preferably humanized antibodies, which may have SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3, respectively.
  • the antibody in the pharmaceutical composition of the present invention has a heavy chain variable region as shown in SEQ ID NO: 19 and a light chain variable region as shown in SEQ ID NO: 20, or has a variable region as shown in SEQ ID NO: 21
  • the antibody in the pharmaceutical composition of the present invention has a heavy chain amino acid sequence as shown in SEQ ID NO: 28 and a light chain amino acid sequence as shown in SEQ ID NO: 29, or the present invention
  • the antibody in the pharmaceutical composition of the invention has a heavy chain amino acid sequence as shown in SEQ ID NO: 30 and a light chain amino acid sequence as shown in SEQ ID NO: 29, or the antibody in the pharmaceutical composition of the invention has a heavy chain amino acid sequence as shown in SEQ ID NO: 29, respectively.
  • the heavy chain amino acid sequence shown in ID NO: 30 and the light chain amino acid sequence shown in SEQ ID NO: 31; more preferably, the antibody in the pharmaceutical composition of the present invention has the heavy chain shown in SEQ ID NO: 28, respectively.
  • the humanized antibody antibody in the pharmaceutical composition of the present invention may preferably have HCDR1, HCDR2, and HCDR3 as shown in SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 9, and respectively as shown in SEQ ID NO: 10. LCDR1, LCDR2, and LCDR3 shown in SEQ ID NO: 11 and SEQ ID NO: 12.
  • the antibody in the pharmaceutical composition of the present invention has a heavy chain variable region as shown in SEQ ID NO: 23 and a light chain variable region as shown in SEQ ID NO: 24, or has a variable region as shown in SEQ ID NO: 23
  • the heavy chain variable region shown in SEQ ID NO: 25 and the light chain variable region shown in SEQ ID NO: 25; more preferably, the antibody in the pharmaceutical composition of the present invention has heavy chain amino acids shown in SEQ ID NO: 32, respectively The sequence and the light chain amino acid sequence shown in SEQ ID NO: 33, or the heavy chain amino acid sequence shown in SEQ ID NO: 32 and the light chain amino acid sequence shown in SEQ ID NO: 34, respectively.
  • the humanized antibody antibody in the pharmaceutical composition of the present invention may preferably have HCDR1, HCDR2, and HCDR3 as shown in SEQ ID NO: 13, SEQ ID NO: 14 and SEQ ID NO: 15, and respectively as shown in SEQ ID NO: 16. LCDR1, LCDR2, and LCDR3 shown in SEQ ID NO: 17 and SEQ ID NO: 18.
  • the antibody in the pharmaceutical composition of the present invention has a heavy chain variable region as shown in SEQ ID NO: 26 and a light chain variable region as shown in SEQ ID NO: 27; more preferably, the pharmaceutical composition of the present invention
  • the antibody in the product has a heavy chain amino acid sequence as shown in SEQ ID NO: 35 and a light chain amino acid sequence as shown in SEQ ID NO: 36, respectively.
  • the concentration of the anti-IL-17A antibody or antigen-binding fragment thereof is about 1-300 mg/mL, preferably about 10-250 mg/mL, preferably about 20-200 mg/mL, more preferably 60-180 mg /mL, more preferably about 80-150 mg/mL.
  • the concentration of anti-IL-17A antibody or its antigen-binding fragment is about 20 mg/mL, 30 mg/mL, 40 mg/mL, 50 mg/mL, 60 mg/mL, 70 mg/mL, 80 mg/mL, 90 mg/mL mL, 100mg/mL, 110mg/mL, 120mg/mL, 130mg/mL, 140mg/mL, 150mg/mL, 160mg/mL, 170mg/mL, 180mg/mL or 200mg/mL, preferably about 80mg/mL or 150mg /mL.
  • the buffer in the pharmaceutical composition of the present invention can be selected from acetate buffer, citrate buffer, succinic acid buffer and histidine buffer to provide 5.0 to 6.5, preferably 5.0 to 6.0, for the pharmaceutical composition of the present invention, It is more preferably 5.5 ⁇ 0.3, and more preferably a pH of about 5.5.
  • the pH of the buffer used in the pharmaceutical composition of the present invention may be 5.0-6.5, preferably 5.0-6.0, more preferably 5.5 ⁇ 0.3, more preferably about 5.5.
  • the particularly preferred buffer in the pharmaceutical composition of the present invention is histidine buffer, including histidine-hydrochloride buffer or histidine-acetate buffer, preferably histidine-hydrochloride buffer. More preferably, the histidine-hydrochloride buffer is made of histidine and histidine hydrochloride, preferably L-histidine and L-histidine monohydrochloride. In some protocols, the histidine buffer is made of 1-20 mM L-histidine and 1-20 mM L-histidine monohydrochloride. In some schemes, the histidine buffer is made of histidine and histidine hydrochloride in a molar ratio of 1:1 to 1:4.
  • the histidine buffer is made of histidine and histidine hydrochloride in a molar ratio of 1:1. In some schemes, the histidine buffer is made of histidine and histidine hydrochloride in a molar ratio of 1:3. In some schemes, the histidine preparation is: a histidine buffer with a pH of 5.5 ⁇ 0.3, preferably about 5.5, made of 4.5 mM L-histidine and 15.5 mM L-histidine monohydrochloride Agent.
  • the histidine preparation is: a histidine buffer with a pH of 5.5 ⁇ 0.3, preferably about 5.5, made of 7.5 mM L-histidine and 22.5 mM L-histidine monohydrochloride Agent.
  • the histidine preparation is: a histidine buffer with a pH of 6.0 ⁇ 0.3, preferably about 6.0, made of 15 mM histidine and 15 mM histidine hydrochloride.
  • the histidine buffer in the pharmaceutical composition of the present invention can be a histidine-acetate buffer, preferably, the molar ratio of the two is 1:1 to 1.5:1, preferably, the pH of such a buffer It is 5.5 ⁇ 0.3, preferably about 5.5.
  • this type of buffer contains 15-20 mM histidine and 12-15 mM acetic acid.
  • the pharmaceutical composition of the present invention may contain: a histidine-histidine hydrochloride buffer with a pH of 5.0-6.0, the concentration of which in the pharmaceutical composition is 10-30 mM; and 60-180 mg/mL
  • a histidine-histidine hydrochloride buffer with a pH of 5.0-6.0 the concentration of which in the pharmaceutical composition is 10-30 mM
  • 60-180 mg/mL The anti-IL-17A antibody or antigen-binding fragment thereof according to any one of the preceding embodiments, especially hu31, hu43, hu44, hu59, hu60 and hu250 or antigen-binding fragments thereof described herein.
  • the pharmaceutical composition of the present invention also contains a stabilizer.
  • the stabilizer is selected from one or more of arginine hydrochloride, proline, glycine, sodium chloride, mannitol, sorbitol, sucrose, maltose, xylitol and trehalose.
  • the stabilizer in the pharmaceutical composition contains at least arginine hydrochloride, and optionally contains one or more of mannitol, sorbitol, sucrose and trehalose.
  • the pharmaceutical composition may contain arginine hydrochloride and mannitol, arginine hydrochloride and sorbitol, arginine hydrochloride and sucrose, or arginine hydrochloride and trehalose.
  • concentration of the stabilizer in the pharmaceutical composition of the present invention is about 10 mM to 400 mM, preferably 20 mM to 300 mM, more preferably 30 mM to 200 mM.
  • the stabilizer is sodium chloride at a concentration of about 30-200 mM; or the stabilizer is mannitol at a concentration of about 100-300 mM, preferably 200-300 mM; or the stabilizer is a concentration of about 100-300 mM, Preferably 200-300mM sorbitol; or said stabilizer is sucrose with a concentration of about 100-300mM, preferably 50-250mM; or said stabilizer is trehalose with a concentration of about 100-300mM, preferably 200-300mM; or The stabilizer is arginine hydrochloride with a concentration of about 30-200 mM, preferably about 60-150 mM; or the stabilizer is proline with a concentration of about 100-300 mM; or the stabilizer is glycine with a concentration of about 100-300 mM.
  • the stabilizer is a combination of about 30-200 mM sodium chloride and about 30-200 mM mannitol; or the stabilizer is about 30-200 mM sodium chloride and about 30-200 mM sucrose Or the stabilizer is a combination of about 30-200mM arginine hydrochloride and about 30-200mM mannitol; or the stabilizer is about 30-200mM, preferably about 60-120mM arginine hydrochloride In combination with about 30-200 mM, preferably about 40-80 mM sucrose.
  • the pharmaceutical composition of the present invention contains: a histidine-histidine hydrochloride buffer with a pH of 5.0-6.0, the concentration of which in the pharmaceutical composition is 10-30 mM; 60- 180 mg/mL of the anti-IL-17A antibody or antigen-binding fragment thereof described in any one of the preceding embodiments, especially hu31, hu43, hu44, hu59, hu60 and hu250 or antigen-binding fragments thereof described herein; and 20mM-300mM
  • the stabilizer includes at least arginine hydrochloride, optionally including one of mannitol, sorbitol, sucrose and trehalose, preferably 30-200mM arginine hydrochloride, or 30- 200mM arginine hydrochloride and 30-200mM mannitol, or 30-200mM arginine hydrochloride and 30-200mM sucrose.
  • the stabilizer is 30-200 mM arginine hydrochloride and 30-200 mM sucrose. In some embodiments, the stabilizer is 60-120 mM arginine hydrochloride and 40-80 mM sucrose.
  • the pharmaceutical composition of the present invention also includes a surfactant.
  • Preferred surfactants are selected from polysorbate 80, polysorbate 20 and poloxamer 188.
  • the most preferred surfactant is polysorbate 20.
  • the concentration of the surfactant in the pharmaceutical composition of the present invention is about 0.001%-0.1%, preferably about 0.02%-0.08%, preferably about 0.02%-0.04%.
  • the concentration of the surfactant in the pharmaceutical composition of the present invention is about 0.02%, 0.04% or 0.08%, preferably 0.02%.
  • the pharmaceutical composition of the present invention contains: a histidine-histidine hydrochloride buffer with a pH of 5.0-6.0, the concentration of which in the pharmaceutical composition is 10-30 mM; 60- 180 mg/mL of the anti-IL-17A antibody or antigen-binding fragment thereof according to any one of the preceding embodiments, especially hu31, hu43, hu44, hu59, hu60 and hu250 or antigen-binding fragments thereof as described herein; 20mM-300mM
  • the stabilizer preferably, preferably, the stabilizer includes at least arginine hydrochloride, optionally including one of mannitol, sorbitol, sucrose and trehalose, preferably 100-180mM arginine hydrochloride, or 60-120 mM arginine hydrochloride and 40-80 mM mannitol, or 60-120 mM arginine hydrochloride and 40-80 m
  • the pharmaceutical composition of the present invention may be a liquid preparation or a lyophilized preparation.
  • the present invention also provides a pharmaceutical composition according to any embodiment of the present invention for the treatment or prevention of IL-17A-mediated diseases.
  • the pharmaceutical composition according to any embodiment of the present invention is used to treat or prevent IL-17A.
  • IL-17A-mediated diseases refer to diseases in which IL-17A participates in the occurrence and development of diseases, including but not limited to inflammatory diseases and autoimmune diseases.
  • diseases suitable for the treatment and prevention of the pharmaceutical composition of the present invention include but are not limited to arthritis, rheumatoid arthritis, ankylosing spondylitis, chronic obstructive pulmonary disease, systemic lupus erythematosus (SLE), lupus Nephritis, asthma, multiple sclerosis, cystic fibrosis, psoriasis.
  • the buffer system and pH closely affect the stability of the antibody, and each antibody with unique physical and chemical properties has the most suitable buffer type and pH.
  • This example aims to screen an optimal buffer system and pH so that the anti-IL-17A antibody disclosed in the present invention has the best stability for clinical application.
  • This example was performed with the antibody hu31 at a concentration of about 80 mg/mL and 150 mg/mL.
  • the sample was concentrated by ultrafiltration with Millipore Pellicon3 0.11m 2 membrane and the solution was exchanged. After the solution was changed, the sample was placed in the corresponding prescription, and the sample was placed in a closed centrifuge tube for buffer screening. Screened acetate buffer, succinate buffer and histidine buffer, pH from 5.0 to 6.5 (as shown in Table 3). Place the samples in an environment of 40 ⁇ 2°C, and take them out at the 0th week, the 2nd week, and the 4th week for analysis and testing.
  • the main pathway of protein degradation is the formation of aggregates, lysate products and charged variants.
  • SEC-HPLC Size exclusion chromatography
  • CEX-HPLC cation exchange chromatography
  • the preparation of the excipient combination of arginine hydrochloride and sucrose has the strongest stability and the lowest viscosity of the antibody. Specifically, after being placed at a high temperature of 40 ⁇ 2°C for 4 weeks, the preparation group containing a combination of arginine hydrochloride and sucrose: (1) For the stability of the antibody structure, the rate of decrease in the purity of the antibody monomer was significantly the lowest, as low as 0.2%/ Week, about 40% of the highest mannitol group, the purity of antibody monomer is as high as 99.2%; (2) For the stability of antibody charge, the rate of decrease of the main charge of the antibody is low, as low as 6.00%/week, the main charge As high as 86.0%; (3) Regarding the viscosity of the preparation, arginine hydrochloride can significantly reduce the viscosity of the preparation, and the viscosity of the preparation containing only arginine hydrochloride is lower than 6cP (Table 7), which is significantly lower than other
  • a high-concentration antibody solution is generally more likely to cause antibody aggregation, precipitation, etc., which reduces the stability of the antibody, and the increase in solution viscosity makes injection (especially subcutaneous or intramuscular injection) difficult to administer.
  • arginine salt can not only ensure the stability of the antibody, but also significantly reduce the viscosity of the liquid.
  • the buffer is a histidine buffer with a pH of 5.5 and a stabilizer containing arginine hydrochloride and sucrose, the effect of antibody stability and solution viscosity is optimal.
  • Surfactants added to liquid formulations are often used to protect proteins such as antibodies from stress induced by the air/solution interface and solution/surface-induced stress during storage to reduce antibody aggregation or minimize the formation of particulate matter in the formulation.
  • Reagent which is conducive to the stability of the physical and chemical properties of the antibody.
  • the polysorbate 20 or polysorbate 80 was placed at 40 ⁇ 2°C for 2 weeks and 4 weeks, and then analyzed and tested. The straight line was fitted and the falling slope (%/week) was calculated. The results are shown in Table 10.
  • Example 4 The formulation dilution stability and prescription confirmation
  • the antibody hu31 selects histidine buffer to adjust pH, arginine hydrochloride and sucrose adjust the osmotic pressure and viscosity of the formulation, and adds polysorbate 20 to increase the solubility of the formulation.
  • Liquid pharmaceutical products containing therapeutic antibodies usually need to be stored at 2-8°C, so it is very important that the formulations maintain high stability during long-term storage. Based on the above screening results, we selected 4 batches of antibody stock solutions to prepare the No. 31 prescription preparation shown in Table 13 for long-term stability studies of the preparation.
  • CE-SDS sodium dodecyl sulfate capillary electrophoresis
  • SEC-HPLC SEC-HPLC to measure the antibody monomer (quality standard: ⁇ 97.0%), aggregate (Quality standard: ⁇ 3.0%) or the content of fragments (Quality standard: ⁇ 1.0%)
  • CEX-HPLC measurement of antibody main charge quality standard: ⁇ 70.0%
  • acid charge quality standard: ⁇ 30.0%
  • alkaline charge content quality standard: ⁇ 15.0%
  • biological activity quality standard: 70%-130% of the reference product.
  • NA in the table means that it is a non-critical inspection item at this point in time, and no test will be done at this sampling point;
  • NR means that it has not arrived at this point in time, the same below;
  • ND means that it has not been detected.
  • CEX-HPLC Stored under accelerated stability conditions (25 ⁇ 2°C, 60 ⁇ 5%RH) for 6 months, the main peak of CEX-HPLC decreased, and the acidic peak increased, which exceeded the quality standard for 6 months; SEC-HPLC monomer content and R- The purity of CE-SDS has a downward trend, but it does not exceed the quality standard; there is no significant change in biological activity.
  • NA in the table means that it is a non-key inspection item at this time point, and no test is done at this sampling point;
  • NR means that the time point has not arrived;
  • ND means that it has not been detected.
  • Example 7 Binding specificity of humanized antibody to human IL-17A
  • the conventional ELISA method was used to detect the binding specificity of different humanized antibodies (prescription configuration according to prescription 31) and human IL-17A. That is, 0.5 ⁇ g/ml of human IL-17A-mFc was coated on a 96-well microtiter plate, and incubated at 37 degrees Celsius for 60-90 minutes. Then the solution in the well was discarded, washed 3 times with washing buffer, and blocked with PBS solution containing 2% BSA for 60 minutes.
  • washing buffer After washing 3 times with washing buffer, add different concentrations of humanized antibody diluents, incubate at 37°C for 60 minutes, rinse with washing buffer 3 times, then add 1:10000 times diluted biotin-anti-IgG4, and incubate at 37°C After washing three times with washing buffer for 1 hour, add HPR-Strep diluted 1:10000 with washing buffer, incubate at room temperature for 1 hour, after washing with washing buffer 3 times, add 100 ⁇ L of TMB substrate solution for color development. After reacting at room temperature for 30 minutes, stop the reaction with 100 ⁇ L of 2M hydrochloric acid solution and read the absorbance at 450 nm.
  • the humanized antibodies hu31, hu43, hu44, hu59, hu60 and hu250 all specifically bind to IL-17A. Its EC50 were 8.13ng/mL, 8.64ng/mL, 6.76ng/ml, 6.10ng/mL, 5.78ng/mL and 6.35ng/mL, respectively.
  • Example 8 Humanized antibody blocks the binding effect of human IL-17A and IL-17RA
  • the mixture was incubated with the cell suspension (293F IL-17RA stably transfected cell line obtained in Example 2, 1.5 ⁇ 10 5 cells/well) at 37 degrees Celsius for 15 minutes, eluted with PBS 3 times, and then added 5 ⁇ g/ml Anti-mouse IgG and incubate at room temperature for 30 minutes. After eluting with PBS three times, the inhibitory effect of humanized antibody on the binding of IL-17A to IL-17RA on the surface of 293F cells was detected by flow cytometry. As shown in Figure 2, the humanized antibodies hu31, hu43, hu44, hu59, hu60 and hu250 can significantly inhibit the specific binding of IL-17A to IL-17RA on cells. Its IC50 are 867.6ng/mL, 780.8ng/mL, 828.5ng/ml, 467.4ng/mL, 482.8ng/mL and 577.8ng/mL, respectively.
  • Example 9 Humanized antibody antagonizes IL-17A to induce epithelial cells to express CXCL1
  • IL-17A can stimulate a variety of epithelial cells and other cells to secrete the expression and release of cytokine CXCL1.
  • the changes in CXCL1 expression in the cell supernatant can be quantitatively detected by ELISA to determine the effects of different humanized antibodies (prescription 31) on IL
  • the effect of -17A on cell-mediated biological activity can be quantitatively detected by ELISA to determine the effects of different humanized antibodies (prescription 31) on IL
  • -17A on cell-mediated biological activity.
  • HT-29 cells human colorectal adenocarcinoma epithelial cells, ATCC
  • ATCC human colorectal adenocarcinoma epithelial cells
  • humanized antibodies hu31, hu43, hu44, hu59, hu60, and hu250 have stronger antagonistic effects on IL-17A-stimulated epithelial cells to release CXCL1 than the reference antibody Secukinumab.
  • Example 10 Humanized antibody antagonizes IL-17A and induces expression of CXCL1 in mice
  • mice By detecting changes in serum CXCL1 levels in mice, the effects of different humanized antibodies (configured according to prescription 31) on IL-17A-mediated biological activity in vivo are judged.
  • 40 female Balb/c mice aged 10 weeks were selected and divided into 8 groups with 5 mice in each group. 4 days before the administration, the serum was collected, and the expression of CXCL1 was detected as the basic value.
  • the candidate antibodies humanized antibodies hu31, hu43, hu44, hu60 and hu250
  • IgG4 isotype control hIgG
  • the dose was administered by subcutaneous injection of human IL-17A; 2 hours after the injection of human IL-17A, the serum was collected to detect the concentration of CXCL1 in the blood, and compared with the basic value to calculate the multiple of the change in the concentration of CXCL1 before and after the administration in each group (average Value ⁇ standard error).
  • the comparative analysis of the candidate antibody and the IgG4 isotype control used the Student’s-t test test. P ⁇ 0.05 was considered as a significant difference, *P ⁇ 0.05, **P ⁇ 0.01, ***P ⁇ 0.001.
  • the candidate antibodies hu31, hu43, hu44, hu60 and hu250 have stronger antagonistic effects on IL-17A-stimulated release of CXCL1 in mice.
  • Example 11 Study on the effect of humanized antibody on improving the efficacy of imiquimod-induced mouse psoriasis model
  • Applying imiquimod to the skin of the back of the ears of mice can induce psoriasis-like pathological features, that is, hyperproliferation of keratinocytes, aggregation of inflammatory cells, and vascular proliferation in the dermal papillae to construct a mouse model of psoriasis.
  • the clinical score and ear swelling degree were used as indicators to judge the therapeutic effect of drugs on mice with psoriasis.
  • group I was sham operation group
  • group II was PBS group
  • group III was KLH control group (identical IgG), and KLH was given
  • group IV was hu31 group
  • V The group is the hu43 administration group
  • the VI group is the hu44 administration group, and the dosage of each group is 50mg/kg
  • the above groups are injected intraperitoneally on the 0th day and the 3rd day of the grouping.
  • day 1 mice in group II-VI applied about 62.5 mg of imiquimod cream (Aidale, 5%, 3M Health Care Limited) to the right ear and back skin for 4 consecutive days.
  • the humanized antibody is configured according to prescription 31.
  • the thickness of the mouse's right ear was measured with a spiral micrometer every day, and the thickness of the mouse's ear swelling was calculated with the thickness of the right ear on the first day as a control.
  • the mice were weighed daily to observe the skin scaly, induration, and erythema, and scored using a 4-level scoring method: 0 points, no disease; 1 point, mild; 2 points, moderate; 3 points, severe; 4 points ,very serious. The results are expressed as mean ⁇ standard error.
  • ANOVA one-way analysis of variance
  • administration of the humanized antibody of the present invention can significantly inhibit imiquimod-induced skin scaly, induration, redness and swelling in the mouse psoriasis model, that is, the score value is small.
  • the humanized antibody of the present invention can obviously resist the onset of mice, and the phenotype is the reduction of the mouse clinical score and the degree of ear swelling.
  • Example 12 Study on the effect of humanized antibody on improving type II collagen-induced female cynomolgus arthritis
  • Type II collagen-induced arthritis is an animal model widely used in rheumatoid arthritis (RA) research. It has the same histopathological characteristics as human RA, and is characterized by inflammation of facet joints and progressive erosion of cartilage and bone . Human/humanized biological macromolecules including antibodies often have better cross-reactivity with antigens in cynomolgus monkeys. Therefore, the cynomolgus monkey arthritis model is one of the anti-rheumatic effects of the humanized antibody IL-17A in the present invention. Effective system. This experiment is to evaluate the efficacy of candidate antibodies on the cynomolgus rheumatoid arthritis model.
  • Dissolve type II bovine collagen (CII, Sichuan University) in acetic acid (Cat. No. 10000218; Sinopharm; Shanghai; China), put it in a 4°C refrigerator and stir overnight, and then use an equal volume of complete Freund’s adjuvant (Cat. No.: F5881, Sigma- Aldrich, USA) emulsified collagen, the final concentration of emulsion collagen was 2mg/ml.
  • the animals were anesthetized with Shutai (1.5-5 mg/kg, im), and multiple collagen emulsion injections were performed on the back and the base of the tail. If necessary, 1.5%-5% isoflurane was used to maintain anesthesia during immunization. .
  • G1 is the normal animal group ("naive"), without arthritis induction
  • G2 is the vehicle control group ("vehicle”
  • G3 is the antibody hu31 administration group
  • G4 is the antibody hu59 administration Group.
  • the clinical score of an animal reaches 5% of the maximum value of the clinical score (192 ⁇ 5% ⁇ 10), it is assigned to each experimental group in order, and the group that reaches the score first enters the group, and so on until all eligible animals They are all sorted into each group in order.
  • the drug was administered, once a week, 7.5 mg/kg each time, and the infusion pump (infusion bump) was continuously pumped for 30 minutes for 5 weeks.
  • the humanized antibody is configured according to prescription 31.
  • Weight measurement The animal's weight was measured the day before immunization, and then once a week until the end of the experiment.
  • Arthritis score score the inflammation degree of arthritis in the limbs of the monkeys on day 0 and day 21, and score once a week after 21 days until the end of the experiment (if there is an early onset, the corresponding arthritis score once a week in advance) .
  • Scoring criteria for arthritis 0 points, normal; 1 point, mild arthritis, mild but clearly distinguishable; 2 points, moderate swelling; 3 points, severe arthritis, severe swelling or obvious joint deformation.
  • MCP metacarpophalangeal joints
  • PIP proximal finger joints
  • DIP distal finger joints
  • the weight of the normal cynomolgus monkey (G1) is stable; the cynomolgus monkey after arthritis induction, the average weight of the cynomolgus monkey in the vehicle treatment group (G2) continues to decrease.
  • Both antibodies hu31 and hu59 controlled this downward trend. Therefore, under the experimental conditions, hu31 and hu59 have a certain improvement effect on weight loss caused by arthritis (**P ⁇ 0.01, ****P ⁇ 0.0001, compared with "G2: vehicle group”; One-way ANOVA/Dunnett).
  • Example 13 Effect of humanized antibody on mouse joint swelling induced by NIH3T3-IL17 cells
  • Cells NIH3T3 cells, NIH3T3 cells expressing human IL-17.
  • NIH3T3-IL-17+control IgG antibody group (30mg/kg);
  • NIH3T3-IL-17+ test antibody high-dose administration group (antibody hu31, 3mg/kg);
  • NIH3T3-IL-17+ test antibody mid-dose administration group (antibody hu31, 10mg/kg);
  • NIH3T3-IL-17+ test antibody low-dose administration group (antibody hu31, 30mg/kg);
  • NIH3T3-IL-17+ positive drug administration group (cosentyx, 10mg/kg).
  • the humanized antibody is configured according to prescription 31.
  • NIH3T3-IL-17 cells and NIH3T3 control cells were injected into the articular cavity of the right ankle joint of each group of mice.
  • the antibody hu31 (3, 10, 30 mg/kg) and cosentyx (10 mg/kg) were administered by intraperitoneal injection 1 day before the model selection, once every 3 days, to investigate the effect of antibody hu31 injection on the mouse arthritis model .
  • the thickness of the mouse ankle joint was measured with a vernier caliper, and the degree of swelling was calculated.
  • NIH3T3 cells stably transformed with hIL-17 were injected into the joint cavity of the mouse metaphyseal joint, and severe swelling of the mouse metaphyseal joint was observed on the second day.
  • the swelling inhibition rate is calculated based on the thickness of the mouse ankle joint.
  • the swelling inhibition rate of cosentyx (10mg/kg) was 67.1%. Calculating the average inhibition rate of each group for different days, the results showed that the average inhibition rate of each group (3, 10, and 30 mg/kg) of the antibody hu31 was 45.2%, 57.0%, and 73.9%, respectively.
  • the swelling inhibition rate of cosentyx (10mg/kg) was 60.4%.
  • the experimental results indicate that the antibody hu31 can inhibit IL-17-induced joint ankle swelling in mice in a dose-dependent manner, and its effect at 10 mg/kg is equivalent to the positive control drug cosentyx (10 mg/kg). The effect of 30mg/kg is better than the positive control drug cosentyx (10mg/kg).
  • Example 14 Effect of humanized antibody on mouse air sac inflammation induced by NIH3T3-IL17 cells
  • Cells NIH3T3 cells, NIH3T3 cells expressing human IL-17.
  • Reagents Gr1-FITC antibody, Biolegend company.
  • NIH3T3-IL-17 cell group + test antibody high-dose administration group (antibody hu31, 30mg/kg);
  • NIH3T3-IL-17 cell group + test antibody mid-dose administration group (antibody hu31, 10mg/kg);
  • NIH3T3-IL-17 cell group + test antibody low-dose administration group (antibody hu31, 3mg/kg);
  • Administration route intraperitoneal injection, humanized antibody is configured according to prescription 31.
  • Air sac 2.5ml of air was injected into the back of mice on day 0 and 3 respectively. On the 5th day, the cells were injected into the air sac. The number of cells injected per mouse was 2 ⁇ 10 5 cells/500 microliters of PBS. Each group has 8 mice.
  • NIH-3T3 cells stably transformed into hIL-17A were injected into the air sac of the mouse back. From the total number of infiltrated cells, compared with the NIH3T3 cell group, the NIH3T3-IL-17 cell group was in the air sac. The number of infiltrated white blood cells increased significantly, the proportion and number of Grl+ cells also increased significantly, and the selection of the model was successful.
  • the antibody hu31 dose of 3, 10, and 30 mg/kg
  • the inhibition rate is calculated based on the total number of cells infiltrated and the number of Grl+ cells.
  • inhibition rate (%) (number of cells in NIH3T3-ILI7-lgG group-number of cells in administration group)/(NIH3T3-ILl7-IgG group Cell number-NIH3T3 cell number) ⁇ 100.
  • the results showed that the inhibition rates of the total number of infiltrating cells in each group (3, 10, 30 mg/kg) of antibody hu31 were 50.0%, 56.7% and 78.3%, respectively.
  • Calculating the inhibition rate of the number of Grl+ cells the results showed that the average Grl+ cell inhibition rates of each group (3, 10, 30 mg/kg) of antibody hu31 were 59.1%, 54.5% and 81.8%, respectively.
  • the antibody hu31 can inhibit IL- in a dose-dependent manner. 17 induced infiltration of mouse total cells and inflammatory cells.

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Immunology (AREA)
  • Epidemiology (AREA)
  • Oil, Petroleum & Natural Gas (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biochemistry (AREA)
  • Physical Education & Sports Medicine (AREA)
  • Rheumatology (AREA)
  • Pulmonology (AREA)
  • Orthopedic Medicine & Surgery (AREA)
  • Genetics & Genomics (AREA)
  • Biophysics (AREA)
  • Dermatology (AREA)
  • Neurology (AREA)
  • Biomedical Technology (AREA)
  • Neurosurgery (AREA)
  • Urology & Nephrology (AREA)
  • Pain & Pain Management (AREA)
  • Microbiology (AREA)
  • Mycology (AREA)
  • Transplantation (AREA)
  • Inorganic Chemistry (AREA)
  • Medicinal Preparation (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Abstract

本发明提供一种稳定的抗IL-17A抗体的药物组合物及其在医药上的应用。该药物组合物含有抗IL-17A抗体或其抗原结合片段、缓冲液，还可以含有至少一种稳定剂，任选的还可以含有表面活性剂。

Description

抗IL-17A抗体药物组合物及其用途 技术领域
本发明涉及治疗性药物组合物领域。尤其是，本发明涉及医药制剂领域，该药物组合物含有一种与IL-17A特异性结合的人源化抗体。
背景技术
白细胞介素17(Interleukin 17,IL-17)，也称为CTLA-8或IL-17A，在免疫系统中扮演着关键的协调作用。IL-17家族共六个成员，包括IL-17A、IL-17B、IL-17C、IL-17D、IL-17E和IL-17F，都含有至关重要的4个高度保守的半胱氨酸残基，但成员之间生物学效应差异较大，其中，IL-17A和IL-17F同源性和生物学功能最接近，目前研究也最深入。体内表达的IL-17A具有23个氨基酸的N-末端信号肽，其经切割产生成熟的IL-17A。成熟的IL-17A通过二硫键连接，一般以同源二聚体形式分泌与存在，有时也与IL-17F连接形成异源二聚体IL-17AF。一般IL-17A或IL-17指IL-17A同源二聚体蛋白，其主要由辅助性T细胞17(T helper 17,Thl7)产生，也可以由其它免疫细胞如γδT细胞、LTi(Lymphoid Tissue inducer cells)、ILCs(Innate Lymphoid Cells)和NKT(Natural Killer T)细胞合成分泌。IL-17A的表达调控非常复杂，研究发现细胞因子IL-6、IL-1β和TGFβ等诱导初始CD4+T细胞分化为Th17，但此时Th17细胞稳定性弱，分泌少量IL-17A，其组织损伤作用小；当IL-23存在时，其通过促进Th17细胞稳定性并持续分泌IL-17A、上调促炎因子(IL-22、CSF-2和IFN-γ)和下调抑炎因子(IL-2、IL-27和IL-12)表达等途径，引起炎性爆发和组织损伤。所以，当组织中IL-23异常表达时，IL-17A途径在组织损伤中发挥着关键性的作用。
IL-17一般定点分泌，在局部组织中与靶细胞表面的IL-17受体(IL-17R)结合发挥效应。IL-17R家族包括IL-17RA、IL-17RB、IL-17RC、IL-17RD及IL-17RE五个成员，广泛表达于多种细胞膜上。IL-17主要与非造血细胞来源的细胞(如上皮细胞、间质细胞)表面的IL-17RA/IL-17RC复合体结合发挥效应，促进细胞分泌 细胞因子(诸如IL-6、G-CSF、GM-CSF、IL-10、TGF-β、TNF-α)、趋化因子(包括IL-8、CXCL1和MCP-1)和前列腺素(例如，PGE2)，诱导中性粒细胞和巨噬细胞聚集，释放活性氧类物质(ROS)而损伤组织。
自身免疫性疾病严重威胁着人类的健康，如银屑病、类风湿性关节炎、强直性脊柱炎、克罗恩病、多发性硬化病等。研究发现，IL-17的分泌失调与该类疾病发生发展密切相关。靶向IL-17的抗体通过抑制IL-17-IL-17R信号通路，能有效地缓解自身免疫性疾病的症状。诺华公司开发的Cosentyx(secukinumab)是全球首个IL-17单抗，获批的适应症包括中度至重度斑块型银屑病(plaque psoriasis)，为广大银屑病群体提供一个重要的一线生物治疗选择。但是，开发结构不同、疗效更优、适应症更广等不同特性的抗IL-17抗体，治疗银屑病、类风湿性关节、强直性脊柱炎等自身免疫相关病症以及IL-17相关的其它疾病具有迫切的需求和重要的意义。
发明内容
本发明所述的药物组合物是一种含有与IL-17A特异性结合的人源抗体的高稳定性药物组合物。特别地，本发明发现盐酸精氨酸和蔗糖的组合具有出人意料的特征，即具有高稳定性和低粘度。
本发明提供了一种药物组合物，包含：(1)缓冲液；(2)抗IL-17A抗体或其抗原结合片段。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段的HCDR1选自SEQ ID NO:1、7和13；HCDR2选自2、8和14；HCDR3选自3、9和15；LCDR1选自4、10和16；LCDR2选自5、11和17；LCDR3选自6、12和18。
在一些方案中，上述抗IL-17A抗体的VH选自SEQ ID NO:19、21、23和26，且VL选自SEQ ID NO:20、22、24、25和27。
在一些方案中，上述抗IL-17A抗体的HC选自SEQ ID NO:28、30、32和35；LC选自SEQ ID NO:29、31、33、34和36。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有分别如SEQ ID NO:1、SEQ ID NO:2和SEQ ID NO:3所示的HCDR1、HCDR2和HCDR3，和分别如SEQ ID NO:4、SEQ ID NO:5和SEQ ID NO:6所示的LCDR1、LCDR2和LCDR3。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有分别如SEQ ID NO:7、 SEQ ID NO:8和SEQ ID NO:9所示的HCDR1、HCDR2和HCDR3，和分别如SEQ ID NO:10、SEQ ID NO:11和SEQ ID NO:12所示的LCDR1、LCDR2和LCDR3。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有分别如SEQ ID NO:13、SEQ ID NO:14和SEQ ID NO:15所示的HCDR1、HCDR2和HCDR3，和分别如SEQ ID NO:16、SEQ ID NO:17和SEQ ID NO:18所示的LCDR1、LCDR2和LCDR3。
在一些方案中，所述药物组合物中抗IL-17A抗体或其抗原结合片段的浓度约为1-300mg/mL，优选约为10-250mg/mL，优选约为20-200mg/mL，优选约为60-180mg/mL，更优选约为80-150mg/mL；更优选地，上述抗IL-17A抗体或其抗原结合片段浓度约为20mg/mL，30mg/mL，40mg/mL，50mg/mL，60mg/mL，70mg/mL，80mg/mL，90mg/mL，100mg/mL，110mg/mL，120mg/mL，130mg/mL，140mg/mL，150mg/mL，160mg/mL，170mg/mL，180mg/mL或200mg/mL，优选约为80mg/mL或150mg/mL。
在一些方案中，上述缓冲液选自醋酸缓冲液、柠檬酸缓冲液、琥珀酸缓冲液和组氨酸缓冲液中一种或一种以上。
在一些方案中，上述缓冲液为组氨酸缓冲液，优选地，所述组氨酸缓冲液选自组氨酸-盐酸盐缓冲液或组氨酸-醋酸盐缓冲液，优选组氨酸-盐酸盐缓冲液。
在一些方案中，上述组氨酸-盐酸盐缓冲液由组氨酸和组氨酸盐酸盐制成，优选L-组氨酸和L-组氨酸单盐酸盐。在一些方案中，组氨酸缓冲液由1-30mM的L-组氨酸和1-30mM的L-组氨酸单盐酸盐制成。在一些方案中，组氨酸缓冲液由摩尔比为1:1到1:4的组氨酸和组氨酸盐酸盐制成。在一些方案中，组氨酸缓冲液由摩尔比为1:1组氨酸和组氨酸盐酸盐制成。在一些方案中，组氨酸缓冲液由摩尔比为1:3的组氨酸和组氨酸盐酸盐制成。在一些方案中，组氨酸制剂为：由4.5mM的L-组氨酸和15.5mM的L-组氨酸单盐酸盐制成的pH为5.5的组氨酸缓冲剂。在一些方案中，组氨酸制剂为：由7.5mM的L-组氨酸和22.5mM的L-组氨酸单盐酸盐制成的pH为5.5的组氨酸缓冲剂。在一些方案中，组氨酸制剂为：由15mM的组氨酸和15mM的组氨酸盐酸盐制成的pH为6.0的组氨酸缓冲液。
在一些方案中，上述组氨酸缓冲液为组氨酸-醋酸盐缓冲液，优选地，两者的摩尔比为1:1到1.5:1，优选地，此类缓冲液的pH为5.5±0.3，优选约为5.5，优选 地，这类缓冲液含有15-20mM的组氨酸和12-15mM的醋酸。
在一些方案中，上述缓冲液为醋酸缓冲液，优选地，所述醋酸缓冲液为醋酸-醋酸钠缓冲液或醋酸-醋酸钾缓冲液，优选醋酸-醋酸钠缓冲液。
在一些方案中，上述缓冲液为柠檬酸缓冲液，优选地，所述柠檬酸缓冲液为柠檬酸-柠檬酸钠缓冲液。
在一些方案中，上述缓冲液为琥珀酸缓冲液，优选地，所述琥珀酸缓冲液为琥珀酸-琥珀酸钠缓冲液。
在一些方案中，上述缓冲液的浓度约为1-200mM，优选约为5-200mM，优选约为10-50mM，优选约为10-30mM；优选约为10-20mM，上述缓冲液浓度非限制性实施例约为5mM，10mM，15mM，20mM，25mM，30mM、40mM、45mM、50mM、60mM、70mM、80mM、90mM、100mM、105mM、110mM、115mM、120mM、130mM、140mM、150mM、160mM、170mM或180mM或这些范围内任意两个数值作为端点形成的范围，优选为10mM、15mM、20mM或30mM。
在一些方案中，上述缓冲液的pH约为5.0-6.5，优选约为5.0-6.0，优选约为5.5-6.5，优选约为5.0-5.5，优选约为5.5-6.0，优选约为6.0-6.5，上述缓冲液的pH非限制性实施例约为5.0，5.1，5.2，5.3，5.4，5.5，5.6，5.7，5.8，5.9，6.0，6.1，6.2，6.3，6.4，6.5，优选约为5.0，5.5或6.0。
在一些方案中，上述的药物组合物还包括稳定剂，所述稳定剂选自盐酸精氨酸、脯氨酸、甘氨酸、氯化钠、甘露醇、山梨醇、蔗糖、麦芽糖、木糖醇和海藻糖中的一种或一种以上。
在一些方案中，上述稳定剂的浓度为约10mM～400mM，优选20mM～300mM，更优选30mM～200mM。
在一些方案中，上述稳定剂为浓度约30-200mM的氯化钠；或所述稳定剂为浓度约100-300mM的甘露醇；或所述稳定剂为浓度约100-300mM的山梨醇；或所述稳定剂为浓度约100-300mM的蔗糖；或所述稳定剂为浓度约100-300mM的海藻糖；或所述稳定剂为浓度约30-200mM的盐酸精氨酸；或所述稳定剂为浓度约100-300mM的脯氨酸；或所述稳定剂为浓度约100-300mM的甘氨酸。
在一些方案中，上述稳定剂为约30-200mM的氯化钠与约30-200mM的甘露醇的组合；或所述稳定剂为约30-200mM的氯化钠与约30-200mM的山梨醇的组合； 或所述稳定剂为约30-200mM的氯化钠与约30-200mM的蔗糖的组合；或所述稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的甘露醇的组合；或所述稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的蔗糖的组合；或所述稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的山梨醇的组合；或所述稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的海藻糖的组合。
在一些方案中，上述稳定剂为氯化钠。在一些方案中，上述稳定剂为浓度约30-200mM的氯化钠，上述氯化钠的浓度优选约为50-190mM，优选约为100-180mM，优选约为120-170mM，优选约为130-150mM，上述氯化钠浓度的非限制性实施例为约100mM，110mM，120mM，125mM，130mM，135mM，140mM，145mM，150mM，155mM，160mM，170mM，180mM，190mM，200mM，优选135mM或140mM。
在一些方案中，上述稳定剂为甘露醇。在一些方案中，上述稳定剂为浓度约100-300mM的甘露醇，上述甘露醇的浓度优选约为150-300mM，优选约为200-280mM，上述甘露醇浓度的非限制性实施例为约200mM，210mM，220mM，230mM，240mM，250mM，260mM，270mM，280mM，优选为240mM。
在一些方案中，上述稳定剂为山梨醇。在一些方案中，上述稳定剂为浓度约100-300mM的山梨醇，上述山梨醇的浓度优选约为150-300mM，优选约为200-280mM，上述山梨醇浓度的非限制性实施例为约200mM，210mM，220mM，230mM，240mM，250mM，260mM，270mM，280mM，优选为240mM。
在一些方案中，上述稳定剂为蔗糖。在一些方案中，上述稳定剂为浓度约100-300mM的蔗糖，上述蔗糖的浓度优选约为150-300mM，优选约为200-280mM，上述蔗糖浓度的非限制性实施例为约200mM，210mM，220mM，230mM，240mM，250mM，260mM，270mM，280mM，优选为220mM。
在一些方案中，上述稳定剂为海藻糖。在一些方案中，上述稳定剂为浓度约100-300mM的海藻糖，上述海藻糖的浓度优选约为150-300mM，优选约为200-280mM，上述海藻糖浓度的非限制性实施例为约180mM，200mM，210mM，220mM，230mM，240mM，250mM，260mM，270mM，280mM，优选为220mM。
在一些方案中，上述稳定剂为盐酸精氨酸。在一些方案中，上述稳定剂为浓度约30-200mM的盐酸精氨酸，上述盐酸精氨酸的浓度优选约为50-190mM，优选约 为100-180mM，优选约为120-170mM，优选约为130-150mM，上述盐酸精氨酸浓度的非限制性实施例为约100mM，110mM，120mM，125mM，130mM，135mM，140mM，145mM，150mM，155mM，160mM，170mM，180mM，190mM，200mM，优选135mM或140mM。
在一些方案中，上述稳定剂为脯氨酸。在一些方案中，上述稳定剂为浓度约100-300mM的脯氨酸，上述脯氨酸的浓度优选约为150-300mM，优选约为200-280mM，上述脯氨酸浓度的非限制性实施例为约180mM，200mM，210mM，220mM，230mM，240mM，250mM，260mM，270mM，280mM，优选为240mM。
在一些方案中，上述稳定剂为甘氨酸。在一些方案中，上述稳定剂为浓度约100-300mM的甘氨酸，上述甘氨酸的浓度优选约为150-300mM，优选约为200-280mM，上述甘氨酸浓度的非限制性实施例为约180mM，200mM，210mM，220mM，230mM，240mM，250mM，260mM，270mM，280mM，优选为260mM。
在一些方案中，上述稳定剂为氯化钠与甘露醇的组合。在一些方案中，上述稳定剂为约30-200mM的氯化钠与约30-200mM的甘露醇的组合，优选约40-150mM的氯化钠与约40-180mM的甘露醇的组合，优选约40-100mM的氯化钠与约40-150mM的甘露醇的组合，上述稳定剂的非限制性实施例约为50mM的氯化钠与约120mM的甘露醇的组合，50mM的氯化钠与约140mM的甘露醇的组合，约90mM的氯化钠与约50mM的甘露醇的组合。
在一些方案中，上述稳定剂为氯化钠与蔗糖的组合。在一些方案中，上述稳定剂为约30-200mM的氯化钠与约30-200mM的蔗糖的组合，优选约40-150mM的氯化钠与约40-180mM的蔗糖的组合，优选约40-100mM的氯化钠与约40-150mM的蔗糖的组合，上述稳定剂的非限制性实施例约为50mM的氯化钠与约120mM的蔗糖的组合，约90mM的氯化钠与约50mM的蔗糖的组合。
在一些方案中，上述稳定剂为盐酸精氨酸与甘露醇的组合。在一些方案中，上述稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的甘露醇的组合，优选约40-150mM的盐酸精氨酸与约40-100mM的甘露醇的组合，优选约60-120mM的盐酸精氨酸与约40-80mM的甘露醇的组合，上述稳定剂的非限制性实施例约为90mM的盐酸精氨酸与约50mM的甘露醇的组合。
在一些方案中，上述稳定剂为盐酸精氨酸与蔗糖的组合。在一些方案中，上述 稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的蔗糖的组合，优选约40-150mM的盐酸精氨酸与约40-100mM的蔗糖的组合，优选约60-120mM的盐酸精氨酸与约40-80mM的蔗糖的组合，上述稳定剂的非限制性实施例约为90mM的盐酸精氨酸与约50mM的蔗糖的组合。
在一些方案中，上述稳定剂为浓度约120mM至约170mM的盐酸精氨酸；或约60mM至约120mM的盐酸精氨酸与约40-80mM的蔗糖的组合；或约60mM至约120mM的盐酸精氨酸与约40-80mM的甘露醇的组合。
在一些方案中，上述药物组合物还包括表面活性剂，所述表面活性剂选自聚山梨醇酯80、聚山梨醇酯20或泊洛沙姆188。
在一些方案中，上述表面活性剂选自聚山梨醇酯80。
在一些方案中，上述表面活性剂选自聚山梨醇酯20。
在一些方案中，以w/v计算，上述表面活性剂浓度约为0.001％-0.1％，优选约为0.01％-0.1％，优选约为0.02％-0.08％，更优选约为0.02％-0.04％；作为非限制性实施例，上述表面活性剂的浓度约为0.02％，0.04％或0.08％，优选0.02％。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段选自鼠源抗体、嵌合抗体、人源化抗体，优选人源化抗体。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:19所示的重链可变区，和如SEQ ID NO:20所示的轻链可变区。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:21所示的重链可变区，和如SEQ ID NO:20所示的轻链可变区。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:21所示的重链可变区，和如SEQ ID NO:22所示的轻链可变区。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:23所示的重链可变区，和如SEQ ID NO:24所示的轻链可变区。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:23所示的重链可变区，和如SEQ ID NO:25所示的轻链可变区。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:26所示的重链可变区，和如SEQ ID NO:27所示的轻链可变区。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:28 所示的重链氨基酸序列，和如SEQ ID NO:29所示的轻链氨基酸序列。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:30所示的重链氨基酸序列，和如SEQ ID NO:29所示的轻链氨基酸序列。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:30所示的重链氨基酸序列，和如SEQ ID NO:31所示的轻链氨基酸序列。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:32所示的重链氨基酸序列，和如SEQ ID NO:33所示的轻链氨基酸序列。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:32所示的重链氨基酸序列，和如SEQ ID NO:34所示的轻链氨基酸序列。
在一些方案中，上述抗IL-17A抗体或其抗原结合片段具有如SEQ ID NO:35所示的重链氨基酸序列，和如SEQ ID NO:36所示的轻链氨基酸序列。
在一些方案中，上述药物组合物，其包含如下(1)-(6)任一项所示的组分，其中，抗IL-17A抗体或其抗原结合片段如本发明任一实施方案所述：
(1)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约100-300mM的山梨醇；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
(2)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的盐酸精氨酸；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
(3)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的氯化钠与约30-200mM的甘露醇的组合；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
(4)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的氯化钠与约30-200mM的蔗糖的组合；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
(5)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的盐酸 精氨酸与约30-200mM的甘露醇的组合；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或(6)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的盐酸精氨酸与约30-200mM的蔗糖的组合；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80。在一些方案中，上述的药物组合物包含如下(7)-(12)任一项所示的组分：
(7)(a)约80mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约20mM组氨酸缓冲液，pH约为5.5；(c)约50mM的氯化钠与约140mM的甘露醇的组合；(d)以及约0.02％的聚山梨醇酯20；或
(8)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约30mM组氨酸缓冲液，pH约为5.5；(c)约135mM的盐酸精氨酸；(d)以及约0.02％的聚山梨醇酯20；或
(9)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约30mM组氨酸缓冲液，pH约为5.5；(c)约50mM的氯化钠与约120mM的蔗糖的组合；(d)以及约0.02％的聚山梨醇酯20；
(10)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约30mM组氨酸缓冲液，pH约为5.5；(c)约90mM的盐酸精氨酸与约50mM的甘露醇的组合；(d)以及约0.02％的聚山梨醇酯20；
(11)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约30mM组氨酸缓冲液，pH约为5.5；(c)约90mM的盐酸精氨酸与约50mM的蔗糖的组合；(d)以及约0.02％的聚山梨醇酯20；
(12)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约15mM组氨酸缓冲液，pH约为5.5；(c)约90mM的盐酸精氨酸与约50mM的蔗糖的组合；(d)以及约0.02％的聚山梨醇酯20。
在一些方案中，上述药物组合物，其包含如下(i)-(vi)任一项所示的组分，其中，抗IL-17A抗体或其抗原结合片段如本发明任一实施方案所述：
(i)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约200-280mM的山梨醇；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
(ii)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约120mM至约170mM的盐酸精氨酸；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
(iii)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约40mM至约100mM的氯化钠与约40-150mM的甘露醇的组合；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
(iv)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约40mM至约100mM的氯化钠与约40-150mM的蔗糖的组合；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
(v)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约60mM至约120mM的盐酸精氨酸与约40-80mM的甘露醇的组合；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
(vi)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约60mM至约120mM的盐酸精氨酸与约40-80mM的蔗糖的组合；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80。
在一些方案中，所述药物组合物为液体制剂或冻干制剂。
在一些方案中，所述药物组合物为液体制剂。
在一些方案中，上述液体制剂或冻干制剂于2-8℃稳定至少3个月，至少6个月，至少12个月，至少18个月或至少24个月。
在一些方案中，上述水溶液或冻干制剂于40℃稳定至少7天，至少14天或至少28天。
本发明还提供了上述药物组合物在制备治疗或预防IL-17A介导的疾病的药物中的用途。
在一些方案中，上述疾病包括炎性疾病和自身免疫性疾病，例如关节炎、类风湿性关节炎、强直性脊柱炎、慢性阻塞性肺疾病、系统性红斑狼疮(SLE)、狼疮 性肾炎、哮喘、多发性硬化、囊性纤维化、银屑病。
附图说明
图1：ELISA检测人源化抗体与人IL-17A的结合作用。其中抗体hu31，hu43，hu44，hu59，hu60和hu250与人IL-17A结合具有较高的特异性，其EC50分别为8.13ng/mL，8.64ng/mL，6.764ng/ml，6.102ng/mL，5.776ng/mL和6.351ng/mL。
图2：FACS检测人源化抗体阻断人IL-17A与293F细胞上的IL-17RA结合作用。其中抗体hu31，hu43，hu44，hu59，hu60和hu250均具有高效的阻断作用，其IC50分别为867.6ng/mL，780.8ng/mL，828.5ng/ml，467.4ng/mL，482.8ng/mL和577.8ng/mL。
图3：ELISA检测人源化抗体阻断IL-17A介导的上皮细胞分泌CXCL1的效应。其中抗体hu31，hu43，hu44，hu59，hu60和hu250均高效抑制IL-17A诱导上皮细胞表达CXCL1，并且比对照抗体的阻断作用更强。
图4：ELISA检测人源化抗体阻断IL-17A介导的体内生物活性作用。其中抗体hu31，hu43，hu44，hu60和hu250具有高效抑制IL-17A诱导小鼠表达CXCL1，并且比对照抗体的阻断作用更强。
图5-1：人源化抗体给药对咪喹莫特诱导的银屑病小鼠临床评分影响。而给予hu31和hu44可明显抑制咪喹莫特诱导的小鼠银屑病模型皮肤鳞屑、硬结、红肿等情况,即临床评分降低(*P<0.05vs KLH)。
图5-2：人源化抗体对咪喹莫特诱导的银屑病小鼠耳肿胀程度的影响。给予hu31和hu44可明显改善耳肿胀程度。(*P<0.05vs KLH,**P<0.01vs KLH.)。
图6-1：人源化抗体对II型胶原蛋白诱导的雌性食蟹猴体重的影响。hu31和hu59对关节炎引起的体重下降具有一定改善作用(**P<0.01，****P<0.0001，和“G2:溶媒组”相比较；One-way ANOVA/Dunnett)。
图6-2：人源化抗体对II型胶原蛋白诱导的雌性食蟹猴关节炎评分的影响。hu31显著抑制了食蟹猴关节炎临床评分的增加趋势(***P<0.001，#P<0.05，和G2:溶媒组相比较；One-way ANOVA/Dunnett)。
图7：人源化抗体对NIH3T3-IL17细胞诱导的小鼠关节肿胀的作用。
图8：人源化抗体对NIH3T3-IL17细胞诱导的小鼠空气囊炎症的作用。
具体实施方式
定义和说明
为了更容易理解本发明，以下具体定义了某些技术和科学术语。除非在本文中另有明确定义，本文使用的所有其它技术和科学术语都具有本发明所属领域的一般技术人员通常理解的含义。应理解本发明不限于具体的方法、试剂、化合物、组合物或生物系统，当然可以对以上进行变化。还应理解本申请所用术语仅为了描述具体的实施方式，并不旨在进行限制。
除非该内容被另外明确说明，否则本说明书以及所附权利要求中所用的单数形式"一个"、"一种"和"该"包括复数指代。因此，例如，提及"一种多肽"包括了两种或更多种多肽等的组合。
术语"药物组合物"或“制剂”表示含有一种或多种本文所述抗体与其他组分的混合物，所述其他组分例如生理学可药用的载体和赋形剂。药物组合物的目的是促进对生物体的给药，利于活性成分的吸收进而发挥生物活性。
术语“液体制剂”是指处于液体状态下的制剂，且不意图指称重悬浮的冻干制剂。本发明的液体制剂在储存时稳定，并且其稳定性不依赖于冻干(或其他状态改变方法，例如喷雾干燥)。
术语“水性液体制剂”是指使用水作为溶剂的液体制剂。在一些方案中，水性液体制剂是不需冻干、喷雾干燥和/或冷冻来维持稳定性(例如化学和/或物理稳定性和/或生物活性)的制剂。
术语“赋形剂”是指可以向制剂添加以提供所需特性(例如稠度、提高的稳定性)和/或调节渗透压的试剂。常用赋形剂的实例包括但不限于糖类、多元醇、氨基酸、表面活性剂和聚合物。
本申请所用的"约"在指代可测量数值(如量、持续时间等)时意在涵盖相对于具体数值±20％或±10％的变化，包括±5％、±1％和±0.1％，因为这些变化适于进行所公开的方法。
术语"缓冲液pH约为5.0-6.5"是指这样的试剂，通过其酸/碱共轭组分的作用使得包含该试剂的溶液能抵抗pH变化。本发明的制剂中使用的缓冲液可具有约5.0至约6.5范围内的pH、或约5.5至约6.5范围内的pH、或约5.0至约6.0范围内的 pH。
在本文中，将pH控制在该范围内的“缓冲液”实例包括乙酸盐(例如乙酸钠)、琥珀酸盐(例如琥珀酸钠)、葡萄糖酸、组氨酸、组氨酸盐酸盐、甲硫氨酸、柠檬酸盐、磷酸盐、柠檬酸盐/磷酸盐、咪唑、醋酸、醋酸盐、枸橼酸盐、其组合和其他有机酸缓冲剂。
"组氨酸缓冲液"为包含组氨酸离子的缓冲液。组氨酸缓冲液的实例包括组氨酸和组氨酸的盐，如组氨酸盐酸盐、组氨酸乙酸盐、组氨酸磷酸盐和组氨酸硫酸盐等，如含有组氨酸与组氨酸盐酸盐的组氨酸缓冲液；本发明的组氨酸缓冲液也包括含有组氨酸和醋酸盐(如钠盐或钾盐)的组氨酸缓冲液。
“柠檬酸盐缓冲液”是包括柠檬酸根离子的缓冲液。柠檬酸盐缓冲液的实例包括柠檬酸-柠檬酸纳、柠檬酸-柠檬酸钾、柠檬酸-柠檬酸钙、柠檬酸-柠檬酸镁等。优选的柠檬酸盐缓冲液为柠檬酸-柠檬酸纳缓冲液。
“醋酸盐缓冲液”是包括醋酸根离子的缓冲液。醋酸盐缓冲液的实例包括醋酸-醋酸纳、醋酸-醋酸钾、醋酸-醋酸钙、醋酸-醋酸镁等。优选的醋酸盐缓冲液为醋酸-醋酸纳缓冲液。
“琥珀酸盐缓冲液”是包括琥珀酸根离子的缓冲液。琥珀盐缓冲液的实例包括琥珀酸-琥珀酸纳、琥珀-琥珀酸钾、琥珀酸-琥珀酸钙、琥珀酸-琥珀酸镁等。优选的琥珀酸盐缓冲液为琥珀-琥珀酸纳缓冲液。
术语“稳定剂”表示药学上可接受的赋形剂，其在制造，储存和应用过程中保护活性药物成分和/或制剂免受化学和/或物理降解。稳定剂包括但不限于如以下定义的糖，氨基酸，盐，多元醇和他们的代谢产物，例如氯化钠、氯化钙、氯化镁、甘露醇、山梨醇、蔗糖、海藻糖、精氨酸或其盐(如盐酸精氨酸)、甘氨酸、丙氨酸(α-丙氨酸、β-丙氨酸)、甜菜碱、亮氨酸、赖氨酸、谷氨酸、天冬氨酸、脯氨酸、4-羟基脯氨酸、肌氨酸、γ-氨基丁酸(GABA)、奥品类(opines)、丙氨奥品、章鱼碱、甘氨奥品(strombine))和三甲胺的N-氧化物(TMAO)、人血清白蛋白(hsa)、牛血清白蛋白(bsa)、α-酪蛋白、球蛋白、α-乳白蛋白、LDH、溶菌酶、肌红蛋白、卵清蛋白和RNAaseA。部分稳定剂，如氯化钠、氯化钙、氯化镁、甘露醇、山梨醇、蔗糖等也可起到控制渗透压的作用。在本发明中具体地使用的稳定剂选自多元醇、氨基酸、盐、糖中的一种或一种以上。优选的盐为氯化钠，优选的糖为蔗糖和海藻糖， 优选的多元醇为山梨醇和甘露醇。优选的氨基酸为精氨酸或其盐(如盐酸精氨酸)、甘氨酸、脯氨酸。优选的稳定剂为氯化钠、甘露醇、山梨醇、蔗糖、海藻糖、盐酸精氨酸、甘氨酸、脯氨酸、氯化钠-山梨醇、氯化钠-甘露醇、氯化钠-蔗糖、氯化钠-海藻糖、盐酸精氨酸-甘露醇、盐酸精氨酸-蔗糖，更优选为盐酸精氨酸、氯化钠-蔗糖、盐酸精氨酸-甘露醇、盐酸精氨酸-蔗糖，更优选为盐酸精氨酸-蔗糖。
本文所用的术语“粘度”可以是“运动粘度”或“绝对粘度”。“运动粘度”是对流体在重力影响下所产生的抵抗性流动的一种测量指标。“绝对粘度”，有时称为动态粘度或简单粘度，是运动粘度与流体密度的乘积(绝对粘度＝运动粘度X密度)。运动粘度的量纲是L2/T，其中L是长度，T是时间。通常，运动粘度以厘沲(cSt)表示。运动粘度的国际单位制单位是mm 2/s，即lcSt。绝对粘度以厘泊(cP)单位表示。绝对粘度的国际单位制单位是毫帕斯卡·秒(mPa-s)，其中1cP＝lmPa·s。
对于本发明的药物组合物，本文所用的术语“低水平粘度”表示低于约15厘泊(cP)的绝对粘度。例如，如果当使用标准粘度测量技术测量时，该制剂展示的绝对粘度为约15cP、约14cP、约13cP、约12cP、约11cP、约10cP、约9cP、约8cP、约7cP、约6cP、约5cP、约4cP、约3cP、约2cP、约1cP，或更低，则本发明药物组合物将被认为是具有“低粘度”。对于本发明药物组合物，本文所用的术语“中等水平粘度”将表示介于约35cP和约15cP之间的绝对粘度。例如，如果当使用标准粘度测量技术测量时，该制剂展示的绝对粘度为约34cP、约33cP、约32cP、约31cP、约30cP、约29cP、约28cP、约27cP、约26cP、约25cP、约24cP、约23cP、约22cP、约21cP、约20cP、约19cP、18cP、约17cP、约16cP，或约15.lcP，则本发明药物组合物将被认为是具有“中等粘度”。本发明药物组合物在某些实施方式中可展示低水平的粘度。在一些实施方式中，不同辅料粘度对比发现，精氨酸或其盐可以获得粘度水平显著低于其它辅料。在一些实施方式中，从缓冲体系方面对比发现，组氨酸缓冲体系的粘度水平低于其它缓冲体系。
术语“表面活性剂”一般包括保护蛋白质例如抗体免受空气/溶液界面诱导的应力、溶液/表面诱导的应力的影响以减少抗体的聚集或使制剂中颗粒物的形成最小化的试剂。示例性的表面活性剂包括但不限于非离子型表面活性剂例如聚氧乙烯脱水山梨醇脂肪酸酯(如聚山梨醇酯20和聚山梨醇酯80)、聚乙烯-聚丙烯共聚物、聚 乙烯-聚丙烯二醇、聚氧乙烯-硬脂酸酯、聚氧乙烯烷基醚、例如聚氧乙烯单月桂基醚、烷基苯基聚氧乙烯醚(Triton-X)、聚氧乙烯-聚氧丙烯共聚物(泊洛沙姆，Pluronic)、十二烷基硫酸钠(SDS)。
术语"等渗"是指该制剂具有与人血液基本相同的渗透压。等渗制剂一般具有约250至350mOsm的渗透压。可使用蒸汽压或冰点下降式的渗透压计测量等渗性。
术语“稳定的”制剂是其中的抗体在制造过程期间和/或储存时基本上保持其物理稳定性和/或化学稳定性和/或生物活性的制剂。即使所含的抗体在经过一定时间储存之后未能保持其100％的化学结构或生物功能，医药制剂也可以是稳定的。在某些情况下，在经过一定时间储存之后，能维持约90％、约95％、约96％、约97％、约98％或约99％的抗体结构或功能，也可被认为是“稳定的”。用于测量蛋白质稳定性的各种分析技术在本技术领域中是可得的，并综述在《肽和蛋白质药物递送》(Peptide and Protein Drug Delivery)247-301，Vincent Lee主编，Marcel Dekker，Inc.，New York，N.Y.，Pubs.(1991))，和Jones，A.(1993)Adv.Drug Delivery Rev.10：29-90中(二者引入作为参考)。
制剂在一定温度下经过一定时间的储存之后，通过测定其中剩余的天然抗体的百分比(及其它方法)，可以测量其稳定性。除其它方法外，天然抗体的百分比可以通过尺寸排阻色谱法(例如尺寸排阻高效液相色谱法[SEC-HPLC])来测量，“天然的”指未聚集的和未降解的。在一些方案中，蛋白质的稳定性按照具有低百分比的降解(例如片段化)和/或聚集蛋白质的溶液中单体蛋白质的百分数来确定。在一些方案中，制剂可以在室温、约25-30℃或40℃下稳定储存至少2周、至少28天、至少1个月、至少2个月、至少3个月、至少4个月、至少5个月、至少6个月、至少7个月、至少8个月、至少9个月、至少10个月、至少11个月、至少12个月、至少18个月、至少24个月，或更长，最多不超过约6％、5％、4％、3％、2％、1％、0.5％，或0.1％聚集形式的抗体。
通过测定在离子交换期间在此抗体主馏分(“主要荷电形式”)较为酸性的馏分中迁移的抗体(“酸性形式”)的百分比(及其它方法)，可以测量稳定性，其中稳定性与酸性形式抗体的百分比成反比。除其它方法外，“酸化”抗体的百分比可以通过离子交换色谱法(例如阳离子交换高效液相色谱法[CEX-HPLC])来测量。在一些实施方式中，可接受程度的稳定性意为当制剂在一定温度下经过一定时间的储存之 后，其中可检测出的酸性形式的抗体最多不超过约49％、45％、40％、35％、30％、25％、20％、15％、10％、5％、4％、3％、2％、1％、0.5％或0.1％。在测量稳定性之前储存的一定时间可以是至少2周、至少28天、至少1个月、至少2个月、至少3个月、至少4个月、至少5个月、至少6个月、至少7个月、至少8个月、至少9个月、至少10个月、至少11个月、至少12个月、至少18个月、至少24个月，或更长。当评估稳定性时，容许储存医药制剂的一定温度可以是约-80℃至约45℃范围内的任何温度，例如储存于约-80℃、约-30℃、约-20℃、约0℃、约2-8℃、约5℃、约25℃，或约40℃。
如果抗体在颜色和/或澄清度目测检查时或通过UV光散射或通过孔径排阻层析测量时基本上不显示出例如聚集、沉淀和/或变性的迹象，则所述抗体在该药物组合物中“保持其物理稳定性”。聚集是单个分子或复合物共价或非共价缔合以形成聚集体的过程。聚集可以进行到形成可见沉淀物的程度。
制剂的稳定性例如物理稳定性可以通过本技术领域中公知的方法来评估，包括测量样品的表观消光度(吸光度或光密度)。这样的消光测量与制剂的浊度相关。制剂的浊度部分地是溶解在溶液中的蛋白质的固有性质，并且通常通过比浊法来测量，并用比浊法浊度单位(NTU)来量度。
随着例如溶液中一种或多种组分的浓度(例如蛋白质和/或盐浓度)而变化的浊度水平也被称为制剂的“乳浊”或“乳浊外观”。浊度水平可以参照使用已知浊度的悬液产生的标准曲线来计算。用于测定药物组合物的浊度水平的参比标准品可以基于《欧洲药典》标准(《欧洲药典》(European Pharmacopoeia)，第四版，“欧洲药品质量委员会指令”(Directorate for the Quality of Medicine of the Council of Europe)(EDQM)，Strasbourg，France)。根据《欧洲药典》标准，澄清溶液被定义为浊度低于或等于按照《欧洲药典》标准具有约3的参比悬液的浊度的溶液。比浊法的浊度测量可以检测在不存在缔合或非理想效应的情况下的瑞利散射，其通常随浓度线性变化。用于评估物理稳定性的其他方法在本技术领域中是公知的。
如果抗体在给定时间点的化学稳定性使得抗体被认为仍保持如下文中所定义的其生物活性，则所述抗体在药物组合物中“保持其化学稳定性”。可以通过例如检测或定量抗体的化学改变的形式来评估化学稳定性。化学改变可以包括尺寸改变(例如剪短)，其可以使用例如孔径排阻层析、SDS-PAGE和/或基质辅助的激光解吸 电离/飞行时间质谱(MALDI/TOF MS)来评估。其他类型的化学改变包括电荷改变(例如作为脱酰胺或氧化的结果而发生)，其可以通过例如离子交换层析来评估。
如果药物组合物中的抗体对于其预期目的来说是生物活性的，则所述抗体在药物组合物中“保持其生物活性”。例如，如果制剂于例如5℃、25℃、45℃等温度下储存一定时间(例如1至12个月)之后，该制剂所含抗IL-17A抗体与IL-17A结合的亲和力为所述储存之前抗体结合亲和力的至少90％、95％或以上，则可认为本发明之制剂是稳定的。结合亲和力也可用例如ELISA或等离子共振技术测定。
在本发明的情形中，在药理学意义上，抗体的“治疗有效量”或“有效量”是指在抗体可以有效治疗的障碍的症状的预防或治疗或减轻方面有效的量。本发明中，药物的“治疗有效量”或“治疗有效剂量”是当单独使用或与另一种治疗剂组合使用时保护受试者免于疾病发作或促进疾病消退的任何量的药物，所述疾病消退通过疾病症状的严重性的降低，疾病无症状期的频率和持续时间的增加，或由疾病痛苦引起的损伤或失能的预防来证明。药物促进疾病消退的能力可以使用本领域技术人员已知的多种方法来评价，比如在临床试验期间的人受试者中，在预测人类功效的动物模型系统中，或通过在体外测定法中测定所述药剂的活性。药物治疗有效量包括“预防有效量”，即当单独或如与其它治疗药物组合给与处于患病风险的受试者或患病复发的受试者时，抑制疾病的发展或复发的任何量的药物。
术语“受试者”或“患者”意图包括哺乳动物生物体。受试者/患者的实例包括人类和非人类哺乳动物，例如非人类灵长动物、狗、奶牛、马、猪、绵羊、山羊、猫、小鼠、兔、大鼠和转基因非人类动物。在本发明的特定实施方式中，受试者是人类。
术语“施用”、“给与”及“处理”是指采用本领域技术人员已知的各种方法或递送系统中的任意一种将包含治疗剂的组合物引入受试者。抗PD-1抗体的给药途径包括静脉内、肌内、皮下、腹膜、脊髓或其他胃肠外给药途径，比如注射或输注。“胃肠外给药”是指除了肠内或局部给药以外的通常通过注射的给药方式，包括但不限于静脉内、肌内、动脉内、鞘内、淋巴内、损伤内、囊内、框内、心内、皮内、腹膜内、经气管、皮下、表皮下、关节内、囊下、蛛网膜下、脊柱内、硬膜内和胸骨内注射和输注以及经体内电穿孔。
抗IL-17A抗体
本文所用的术语“抗体”应被理解为包括完整抗体分子及其抗原结合片段。本文所用的术语抗体的“抗原结合部分”或“抗原结合片段”(或简称为“抗体部分”或“抗体片段”)是指抗体中保持了与人IL-17A(白细胞介素17A))或其表位特异性结合能力的一个或多个片段。
本文所用的术语“全长抗体”或“完整抗体分子”指包含四条肽链的免疫球蛋白分子，两条重(H)链(全长时约50-70kDa)和两条轻(L)链(全长时约25kDa)通过二硫键互相连接。每一条重链由重链可变区(在本文中缩写为VH)和重链恒定区(在本文中缩写为CH)组成。重链恒定区由3个结构域CH1、CH2和CH3组成。每一条轻链由轻链可变区(在本文中缩写为VL)和轻链恒定区组成。轻链恒定区由一个结构域CL组成。VH和VL区可被进一步细分为具有高可变性的互补决定区(CDR)和其间隔以更保守的称为框架区(FR)的区域。每一个VH或VL区由按下列顺序：FR1、CDR1、FR2、CDR2、FR3、CDR3、FR4从氨基末端至羧基末端排列的3个CDR和4个FR组成。重链和轻链的可变区含有与抗原相互作用的结合结构域。抗体的恒定区可介导免疫球蛋白对宿主组织或因子(包括免疫系统的各种细胞(例如，效应细胞)和经典补体系统的第一组分(Clq))的结合。
当在本文中使用时，术语“CDR”是指抗体可变序列内的互补决定区。在重链和轻链的各个可变区中存在3个CDR，其对于各个重链和轻链可变区被命名为HCDR1、HCDR2和HCDR3或LCDR1、LCDR2和LCDR3。这些CDR的准确边界按照不同的系统有不同的定义。由Kabat(同上)描述的系统不仅提供了可适用于抗体的任何可变区的明确的残基编号系统，而且提供了定义3个CDR的准确残基边界。这些CDR可以被称为Kabat CDR。Chothia等发现，Kabat CDR内的某些子部分采取几乎一致的肽骨架构型，尽管在氨基酸序列水平上具有大的多样性(Chothia等，(1987)Mol.Biol.196:901-917；Chothia等，(1989)Nature 342:877-883)。定义与Kabat CDR交叠的CDR的其他边界已由Padlan(1995)FASEB J.9:133-139和MacCalIum(1996)J.Mol.Biol.262(5):732-45描述。其他的CDR边界定义可能不严格遵从本文中描述的系统之一，但仍然与Kabat CDR交叠，尽管它们可能被缩短或加长，这是由于根据预测或实验发现特定残基或残基组或甚至整个CDR不显著影响抗原结合。本文中使用的方法可以利用按照任何这些系统所定义的CDR，尽管某些实施方式使用了Kabat或Chothia定义的CDR。
本文所使用的“抗原结合片段”包括抗体的片段或其衍生物，通常包括亲代抗体的抗原结合区或可变区(例如一个或多个CDR)的至少一个片段，其保持亲代抗体的至少一些结合特异性。抗原结合片段的实例包括但不限于Fab，Fab'，F(ab')2和Fv片段；双抗体；线性抗体；单链抗体分子，例如sc-Fv；由抗体片段形成的纳米抗体(nanobody)和多特异性抗体。当抗体的结合活性在摩尔浓度基础上表示时，结合片段或其衍生物通常保持亲代抗体抗原结合活性的至少10％。优选结合片段或其衍生物保持亲代抗体的抗原结合亲和力的至少20％、50％、70％、80％、90％、95％或100％或更高。还预期抗体的抗原结合片段可包括不明显改变其生物活性的保守或非保守氨基酸取代(称为抗体的“保守变体”或“功能保守变体”)。
本发明所述的抗IL-17A抗体或其抗原结合片段包括申请号为PCT/CN2019/124486中描述的任意一个抗IL-17A抗体，本文将其所公开的全部内容以引入的方式纳入本文。在一些方案中，在本发明的方法和组合物中使用的抗体的CDR序列包括来自于PCT/CN2019/124486中描述的抗体hu31的CDR序列。
在本文实施例中所用的非限制性、示范性抗体选自CN201811515045.7中描述的人源化抗体hu31，hu43，hu44，hu59，hu60和hu250，其可变区及其氨基酸CDR序列见表1，其轻/重链氨基酸全长序列见表2。
表1：人源化抗体可变区和CDR序列
抗体 hu31 hu43 hu44 hu59 hu60 hu250
HCDR1 SEQ ID NO:1 SEQ ID NO:1 SEQ ID NO:1 SEQ ID NO:7 SEQ ID NO:7 SEQ ID NO:13
HCDR2 SEQ ID NO:2 SEQ ID NO:2 SEQ ID NO:2 SEQ ID NO:8 SEQ ID NO:8 SEQ ID NO:14
HCDR3 SEQ ID NO:3 SEQ ID NO:3 SEQ ID NO:3 SEQ ID NO:9 SEQ ID NO:9 SEQ ID NO:15
LCDR1 SEQ ID NO:4 SEQ ID NO:4 SEQ ID NO:4 SEQ ID NO:10 SEQ ID NO:10 SEQ ID NO:16
LCDR2 SEQ ID NO:5 SEQ ID NO:5 SEQ ID NO:5 SEQ ID NO:11 SEQ ID NO:11 SEQ ID NO:17
LCDR3 SEQ ID NO:6 SEQ ID NO:6 SEQ ID NO:6 SEQ ID NO:12 SEQ ID NO:12 SEQ ID NO:18
VH SEQ ID NO:19 SEQ ID NO:21 SEQ ID NO:21 SEQ ID NO:23 SEQ ID NO:23 SEQ ID NO:26
VL SEQ ID NO:20 SEQ ID NO:20 SEQ ID NO:22 SEQ ID NO:24 SEQ ID NO:25 SEQ ID NO:27
表2：人源化抗体氨基酸全长序列
人源化抗体 重链(HC) 轻链(LC)
hu31 SEQ ID NO:28 SEQ ID NO:29
hu43 SEQ ID NO:30 SEQ ID NO:29
hu44 SEQ ID NO:30 SEQ ID NO:31
hu59 SEQ ID NO:32 SEQ ID NO:33
hu60 SEQ ID NO:32 SEQ ID NO:34
hu250 SEQ ID NO:35 SEQ ID NO:36
医药制剂
本发明所述的药物组合物是一种含有与IL-17A特异性结合的人源化抗体的高稳定性药物组合物。本发明发现精氨酸盐(如盐酸精氨酸)与蔗糖的组合能明显提高药物组合物的稳定性。
本发明所述的药物组合物是一种包含高浓度活性抗体且具有高稳定性、低粘度的液体性制剂。特别地，本发明发现含有精氨酸盐的制剂粘度明显低于仅含有其它辅料的配制液。另外，组氨酸缓冲体系的粘度水平明显低于其它缓冲体系。
本发明提供了一种药物组合物，包含：(1)缓冲液；(2)抗IL-17A抗体或其抗原结合片段。
本发明药物组合物中的抗体可以是鼠源抗体、嵌合抗体、人源化抗体，优选人源化抗体，可具有分别如SEQ ID NO:1、SEQ ID NO:2和SEQ ID NO:3所示的HCDR1、HCDR2和HCDR3，和分别如SEQ ID NO:4、SEQ ID NO:5和SEQ ID NO:6所示的LCDR1、LCDR2和LCDR3。优选地，本发明药物组合物中的抗体具有如SEQ ID NO:19所示的重链可变区和如SEQ ID NO:20所示的轻链可变区，或具有如SEQ ID NO:21所示的重链可变区和如SEQ ID NO:20所示的轻链可变区，或具有如SEQ ID NO:21所示的重链可变区和如SEQ ID NO:22所示的轻链可变区；更优选地，本发明药物组合物中的抗体具有分别如SEQ ID NO:28所示的重链氨基酸序列和如SEQ ID NO:29所示的轻链氨基酸序列，或本发明药物组合物中的抗体具有分别如SEQ ID NO:30所示的重链氨基酸序列和如SEQ ID NO:29所示的轻链氨基酸序列，或本发明药物组合物中的抗体具有分别如SEQ ID NO:30所示的重链氨基酸序列和如SEQ ID NO:31所示的轻链氨基酸序列；更优选地，本发明药物组合 物中的抗体具有分别如SEQ ID NO:28所示的重链氨基酸序列和如SEQ ID NO:29所示的轻链氨基酸序列。
本发明药物组合物中的人源化抗体抗体优选可具有分别如SEQ ID NO:7、SEQ ID NO:8和SEQ ID NO:9所示的HCDR1、HCDR2和HCDR3，和分别如SEQ ID NO:10、SEQ ID NO:11和SEQ ID NO:12所示的LCDR1、LCDR2和LCDR3。优选地，本发明药物组合物中的抗体具有如SEQ ID NO:23所示的重链可变区和如SEQ ID NO:24所示的轻链可变区，或具有如SEQ ID NO:23所示的重链可变区和如SEQ ID NO:25所示的轻链可变区；更优选地，本发明药物组合物中的抗体具有分别如SEQ ID NO:32所示的重链氨基酸序列和如SEQ ID NO:33所示的轻链氨基酸序列，或具有分别如SEQ ID NO:32所示的重链氨基酸序列和如SEQ ID NO:34所示的轻链氨基酸序列。
本发明药物组合物中的人源化抗体抗体优选可具有分别如SEQ ID NO:13、SEQ ID NO:14和SEQ ID NO:15所示的HCDR1、HCDR2和HCDR3，和分别如SEQ ID NO:16、SEQ ID NO:17和SEQ ID NO:18所示的LCDR1、LCDR2和LCDR3。优选地，本发明药物组合物中的抗体具有如SEQ ID NO:26所示的重链可变区和如SEQ ID NO:27所示的轻链可变区；更优选地，本发明药物组合物中的抗体具有分别如SEQ ID NO:35所示的重链氨基酸序列和如SEQ ID NO:36所示的轻链氨基酸序列。
本发明的药物组合物中，抗IL-17A抗体或其抗原结合片段浓度约为1-300mg/mL，优选约为10-250mg/mL，优选约为20-200mg/mL，更优选60-180mg/mL，更优选约为80-150mg/mL。作为非限制性实施例；抗IL-17A抗体或其抗原结合片段浓度约为20mg/mL，30mg/mL，40mg/mL，50mg/mL，60mg/mL，70mg/mL，80mg/mL，90mg/mL，100mg/mL，110mg/mL，120mg/mL，130mg/mL，140mg/mL，150mg/mL，160mg/mL，170mg/mL，180mg/mL或200mg/mL，优选约为80mg/mL或150mg/mL。
本发明药物组合物中的缓冲液可选自醋酸缓冲液、柠檬酸缓冲液、琥珀酸缓冲液和组氨酸缓冲液，用以为本发明的药物组合物提供5.0到6.5、优选5.0到6.0，更优选5.5±0.3，更优选为约5.5的pH。另一方面，用于本发明药物组合物中的缓冲液的pH可为5.0-6.5，优选为5.0-6.0，更优选5.5±0.3，更优选为约5.5。
本发明药物组合物中特别优选的缓冲液是组氨酸缓冲液，包括组氨酸-盐酸盐缓冲液或组氨酸-醋酸盐缓冲液，优选组氨酸-盐酸盐缓冲液。更优选地，所述组氨酸-盐酸盐缓冲液由组氨酸和组氨酸盐酸盐制成，优选L-组氨酸和L-组氨酸单盐酸盐。在一些方案中，组氨酸缓冲液由1-20mM的L-组氨酸和1-20mM的L-组氨酸单盐酸盐制成。在一些方案中，组氨酸缓冲液由摩尔比为1:1到1:4的组氨酸和组氨酸盐酸盐制成。在一些方案中，组氨酸缓冲液由摩尔比为1:1组氨酸和组氨酸盐酸盐制成。在一些方案中，组氨酸缓冲液由摩尔比为1:3的组氨酸和组氨酸盐酸盐制成。在一些方案中，组氨酸制剂为：由4.5mM的L-组氨酸和15.5mM的L-组氨酸单盐酸盐制成的pH为5.5±0.3、优选约5.5的组氨酸缓冲剂。在一些方案中，组氨酸制剂为：由7.5mM的L-组氨酸和22.5mM的L-组氨酸单盐酸盐制成的pH为5.5±0.3、优选约5.5的组氨酸缓冲剂。在一些方案中，组氨酸制剂为：由15mM的组氨酸和15mM的组氨酸盐酸盐制成的pH为6.0±0.3、优选约6.0的组氨酸缓冲液。
本发明药物组合物中的组氨酸缓冲液可为组氨酸-醋酸盐缓冲液，优选地，两者的摩尔比为1:1到1.5:1，优选地，此类缓冲液的pH为5.5±0.3，优选约为5.5，优选地，这类缓冲液含有15-20mM的组氨酸和12-15mM的醋酸。
因此，本发明的药物组合物可含有：pH为5.0-6.0的组氨酸-组氨酸盐酸盐缓冲液，其在药物组合物中的浓度为10-30mM；和60-180mg/mL的前文任一实施方案所述的抗IL-17A抗体或其抗原结合片段，尤其是本文所述的hu31，hu43，hu44，hu59，hu60和hu250或其抗原结合片段。
在一些方案中，本发明的药物组合物还含有稳定剂。优选地，所述稳定剂选自盐酸精氨酸、脯氨酸、甘氨酸、氯化钠、甘露醇、山梨醇、蔗糖、麦芽糖、木糖醇和海藻糖中的一种或一种以上。优选地，药物组合物中的稳定剂中至少含有盐酸精氨酸，任选含有甘露醇、山梨醇、蔗糖和海藻糖中的一种或多种。例如，药物组合物中可含有盐酸精氨酸与甘露醇、盐酸精氨酸与山梨醇、盐酸精氨酸与蔗糖、或者盐酸精氨酸与海藻糖。本发明的药物组合物中稳定剂的浓度为约10mM-400mM，优选20mM-300mM，更优选30mM-200mM。在一些方案中，稳定剂为浓度约30-200mM的氯化钠；或所述稳定剂为浓度约100-300mM、优选200-300mM的甘露醇；或所述稳定剂为浓度约100-300mM、优选200-300mM的山梨醇；或所述稳定剂为浓度约100-300mM、优选50-250mM的蔗糖；或所述稳定剂为浓度约 100-300mM、优选200-300mM的海藻糖；或所述稳定剂为浓度约30-200mM、优选约60-150mM的盐酸精氨酸；或所述稳定剂为浓度约100-300mM的脯氨酸；或所述稳定剂为浓度约100-300mM的甘氨酸。在一些方案中，所述稳定剂为约30-200mM的氯化钠与约30-200mM的甘露醇的组合；或所述稳定剂为约30-200mM的氯化钠与约30-200mM的蔗糖的组合；或所述稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的甘露醇的组合；或所述稳定剂为约30-200mM、优选约60-120mM的盐酸精氨酸与约30-200mM、优选约40-80mM的蔗糖的组合。
因此，在一些实施方案中，本发明的药物组合物含有：pH为5.0-6.0的组氨酸-组氨酸盐酸盐缓冲液，其在药物组合物中的浓度为10-30mM；60-180mg/mL的前文任一实施方案所述的抗IL-17A抗体或其抗原结合片段，尤其是本文所述的hu31，hu43，hu44，hu59，hu60和hu250或其抗原结合片段；以及20mM-300mM的稳定剂，优选地，所述稳定剂至少包括盐酸精氨酸，任选包括甘露醇、山梨醇、蔗糖和海藻糖中的一种，优选为30-200mM的盐酸精氨酸，或30-200mM的盐酸精氨酸与30-200mM的甘露醇，或30-200mM的盐酸精氨酸与30-200mM的蔗糖。在一些实施方案中，所述稳定剂为30-200mM的盐酸精氨酸与30-200mM的蔗糖。在一些实施方案中，所述稳定剂为60-120mM的盐酸精氨酸与40-80mM的蔗糖。
在一些方案中，本发明的药物组合物还包括表面活性剂。优选的表面活性剂选自聚山梨醇酯80、聚山梨醇酯20和泊洛沙姆188。最优选的表面活性剂是聚山梨醇酯20。以w/v计，本发明药物组合物中表面活性剂的浓度约为0.001％-0.1％，优选约为0.02％-0.08％，优选约为0.02％-0.04％。作为非限制性实施例，本发明药物组合物中表面活性剂的浓度约为0.02％，0.04％或0.08％，优选0.02％。
因此，在一些实施方案中，本发明的药物组合物含有：pH为5.0-6.0的组氨酸-组氨酸盐酸盐缓冲液，其在药物组合物中的浓度为10-30mM；60-180mg/mL的前文任一实施方案所述的抗IL-17A抗体或其抗原结合片段，尤其是本文所述的hu31，hu43，hu44，hu59，hu60和hu250或其抗原结合片段；20mM-300mM的稳定剂，优选地，优选地，所述稳定剂至少包括盐酸精氨酸，任选包括甘露醇、山梨醇、蔗糖和海藻糖中的一种，优选为100-180mM的盐酸精氨酸，或60-120mM的盐酸精氨酸与40-80mM的甘露醇，或60-120mM的盐酸精氨酸与40-80mM的蔗糖；以及以w/v计0.02％-0.04％的聚山梨醇酯20。
本发明的药物组合物可以是液体制剂，或者是冻干制剂。
医药用途和方法
本发明还提供了用于治疗或预防IL-17A介导的疾病的本发明任一实施方案所述的药物组合物，本发明任一实施方案所述的药物组合物在制备治疗或预防IL-17A介导的疾病的药物中的用途，以及给予需要的个体或患者治疗有效量的本发明任一实施方案所述的药物组合物以治疗或预防IL-17A介导的疾病的方法。
本发明中，IL-17A介导的疾病指在IL-17A参与了疾病的发生和发展的疾病，包括但不限于炎性疾病和自身免疫性疾病。本发明中，适用于本发明药物组合物进行治疗和预防的疾病包括但不限于关节炎、类风湿性关节炎、强直性脊柱炎、慢性阻塞性肺疾病、系统性红斑狼疮(SLE)、狼疮性肾炎、哮喘、多发性硬化、囊性纤维化、银屑病。
下文将以具体实施例的方式阐述本发明。应理解，这些实施例仅仅是阐述性的，并非意图限制本发明的范围。实施例中所用到的方法和材料，除非另有说明，否则为本领域的常规方法和材料。
实施例1：缓冲液体系和pH筛选实验
液体型药物组合物中，缓冲液体系和pH密切影响抗体的稳定性，每种具有独特理化性质的抗体都具有最适宜的缓冲液的种类和pH。本实施例旨在筛选一种最佳缓冲液体系和pH，使本发明公开的抗IL-17A抗体具有最佳的稳定性以适宜临床应用。
本实施例以约80mg/mL和150mg/mL浓度的抗体hu31进行。样品使用Millipore Pellicon3 0.11m 2膜进行超滤浓缩换液，换液后样品处于相应的处方中，样品放置在密闭的离心管中进行缓冲液筛选。筛选了醋酸缓冲液、琥珀酸缓冲液和组氨酸缓冲液，pH从5.0到6.5(如表3所示)。将样品放置在40±2℃环境下，分别在第0周、第2周、第4周取出进行分析检测。蛋白降解的主要途径是聚集物、裂解产品和带电变体的形成。采用尺寸排阻色谱法(SEC-HPLC)测定天然形式(蛋白单体)与聚集形式所占的百分比，采用阳离子交换色谱法(CEX-HPLC)测定酸性与碱性形 式抗体所占的百分比。以放置四周(4W)的SEC-HPLC单体含量和CEX-HPLC主峰含量，拟合直线并计算下降斜率(％/周)考察不同缓冲液体系和pH对抗体hu31抗体稳定性的影响，结果汇总见表4和表5所示。
表3：缓冲液体系和pH筛选实验中的处方信息
Figure PCTCN2021099257-appb-000001
表4：缓冲液体系和pH筛选实验中SEC-HPLC单体含量降解速率
Figure PCTCN2021099257-appb-000002
Figure PCTCN2021099257-appb-000003
表5：缓冲液体系和pH筛选实验中CEX-HPLC电荷异质体含量降解速率
Figure PCTCN2021099257-appb-000004
由表4和表5所示，SEC-HPLC实验检测中，高温40±2℃放置4周后，单体含量随蛋白浓度的增高下降速率增快，但是在pH 5.5的组氨酸缓冲体系下不同蛋白浓度的单体含量下降速率均较低，平均下降速率0.24％/周；CEX-HPLC实验检测中，可以看出CEX-HPLC主峰含量降解速率与pH有一定的相关性，pH越高主峰含量降解速率越快，在pH 5.0～5.5的缓冲体系中，主峰含量降解速度相对较低。
根据上述筛选结果，综合考虑该产品的临床使用剂量、产品规格以及产品的目标质量属性(聚体含量和电荷变异含量水平)，选择蛋白浓度150mg/ml和pH5.5组氨酸缓冲体系进行后续的处方筛选。
实施例2：稳定剂(辅料)筛选实验
2.1辅料对粘度影响考察
为了进一步探究不同辅料对抗体稳定性和粘度的影响，我们选取氯化钠、蔗糖、盐酸精氨酸、脯氨酸、甘氨酸、海藻糖、山梨醇或甘露醇之一的制剂进行了比较测试。考察20mM组氨酸缓冲体系和其他缓冲体系组合在相同渗透压和pH5.5，单抗hu31浓度80mg/ml和150mg/ml条件下，上述不同辅料对粘度的影响，具体处方信息如表6所示。各处方制剂分装后放置于40±2℃条件下，在第4周取出，按标准方法进行分析检测制剂粘度，结果见表7。
表6：不同辅料粘度考察处方信息
Figure PCTCN2021099257-appb-000005
注：“-”表示未添加。
表7：不同辅料粘度测定数据
Figure PCTCN2021099257-appb-000006
Figure PCTCN2021099257-appb-000007
根据表7可以看出，粘度和蛋白浓度成正比关系，随着蛋白浓度增高粘度变大，从含不同辅料粘度对比(处方7～处方14)，盐酸精氨酸的粘度明显低于其他辅料，其粘度数据仅相当于甘露醇的约65％。
从缓冲体系方面对比(处方15～处方18)，组氨酸缓冲体系的粘度低于其他缓冲体系。在此基础上我们进行后续的处方设计及筛选。
2.2辅料对稳定性影响考察
为了进一步探究不同辅料对抗体稳定性的影响，我们选取氯化钠、蔗糖、盐酸精氨酸、海藻糖、山梨醇或甘露醇之一或其组合的制剂进行了比较测试。即将上述不同的辅料或其组合分别加入含约150mg/mL抗体hu31的30mM组氨酸缓冲液中，具体处方信息如表8所示。各处方制剂分装后放置于40±2℃条件下，分别在第0周、第2周、第4周取出进行分析检测。通过分子排阻高效液相色谱法(SEC-HPLC)检测抗体hu31单体含量变化、弱阳离子高效液相色谱法(CEX-HPLC)检测抗体hu31电荷主峰含量。结果如表9所示。
根据稳定性考察结果，不同辅料的制剂处方样品在40±2℃高温条件下放置4周后，抗体均有较强的热稳定性。
综合各项数据分析，对比处方19～处方24发现，单一辅料对比可以明显看出含盐酸精氨酸的单一辅料明显优于氯化钠、甘露醇、山梨醇、蔗糖和海藻糖的单一辅料，表现在SEC-HPLC的单体含量与CEX-HPLC的主峰含量的降解速率更低。对比处方25～处方30发现，含辅料盐酸精氨酸和蔗糖组合最为稳定，在SEC-HPLC的单体含量下降速率低至0.2％/周，明显优于含单一辅料的，CEX-HPLC的主峰含 量的降解速率没有明显差异。
因此，选用盐酸精氨酸和蔗糖的辅料组合更有利于产品的稳定性。
表8：辅料筛选实验中的处方信息
Figure PCTCN2021099257-appb-000008
注：“-”表示未添加。
表9：辅料筛选实验结果汇总
Figure PCTCN2021099257-appb-000009
Figure PCTCN2021099257-appb-000010
综合各项数据分析，选用盐酸精氨酸和蔗糖的辅料组合的制剂，抗体的稳定性最强且粘度最低。具体地，高温40±2℃放置4周后，含盐酸精氨酸和蔗糖组合的制剂组：(1)对于抗体结构的稳定性，抗体单体纯度的下降速率明显最低，低至0.2％/周，约为最高的甘露醇组的40％，抗体的单体纯度高达99.2％；(2)对于抗体电荷的稳定性，抗体主要电荷的下降速率较低，低至6.00％/周，主要电荷高达86.0％；(3)对于制剂的粘度，盐酸精氨酸能显著降低制剂的粘度，仅含盐酸精氨酸的制剂粘度低于6cP(表7)，明显低于其它各组处方，特别是仅为含甘露醇制剂组。
高浓度的抗体溶液一般更容易引起抗体聚集、沉淀等，使抗体稳定性降低，并且溶液粘度增加导致注射(特别是皮下或肌内注射)施用难度。综合以上实验，我们发现，液体制剂中，精氨酸盐不仅能保证抗体的稳定性，还能显著降低液体粘度。特别是，当缓冲液为pH为5.5的组氨酸缓冲液，含盐酸精氨酸和蔗糖的稳定剂时，抗体稳定性和溶液的粘度效果最优。
实施例3：表面活性剂筛选实验
液体制剂中添加表面活性剂常用于保护蛋白质例如抗体在储存过程中免受空气/溶液界面诱导的应力、溶液/表面诱导的应力的影响以减少抗体的聚集或使制剂中颗粒物的形成最小化的试剂，其利于抗体理化性质的稳定。在含有30mM组氨酸缓冲液(组氨酸-组氨酸盐酸盐，pH为5.5)和150mg/ml的抗体hu31的制剂(辅料1和辅料2均按照处方30添加)中分别加入不同浓度的聚山梨酯20或聚山梨酯 80，40±2℃放置2周、4周后分析检测，拟合直线并计算下降斜率(％/周)，结果如表10所示。
综合分析显示，表面活性剂筛选试验(处方31～36)结果显示添加不同浓度的聚山梨酯80或聚山梨酯20对SEC-HPLC的单体含量影响不明显。
表10：表面活性剂筛选结果
Figure PCTCN2021099257-appb-000011
实施例4：制剂的处方稀释稳定性和处方确认
选用抗体原液，配制如表8所示的30号处方制剂，用生理盐水、葡萄糖注射液和不同处方稀释样品至不同浓度考察样品稳定性，同时考察反复冻融条件下的样品稳定性，进行处方条件确认(表11)。筛选结果见表12。
表11：考察信息(处方30)
反复冻融 -40±5℃冻融5次
生理盐水稀释 稀释浓度至10mg/ml
葡萄糖注射液 稀释浓度至10mg/ml
处方30缓冲液 稀释浓度至2mg/ml、10mg/ml
表12：处方稳定性筛选结果
Figure PCTCN2021099257-appb-000012
注：“/”表示未测定或未添加。
通过对样品在此处方条件下，进行25±2℃反复冻融、葡萄糖注射液、生理盐水以及处方30缓冲液(即30mM组氨酸缓冲液)稀释样品至不同浓度对单体含量没有大的影响，表现了处方良好的稳定性。
综上所述，我们通过对不同缓冲体系，不同pH条件、不同抗体浓度以及不同辅料和表面活性剂组成进行考察，探索研究重组人源化抗IL-17A单克隆抗体hu31的稳定性，并确定最佳液体制剂配方。抗体hu31选择组氨酸缓冲液来调节pH，盐酸精氨酸和蔗糖调节制剂渗透压和粘度，添加聚山梨酯20来增加制剂溶解性。
实施例5：制剂长期稳定性研究
含有治疗性抗体的液体药物制品通常需要储存在2～8℃条件下，所以制剂能在长期储存中保持高稳定性非常重要。根据上述的筛选结果，我们选用4批次抗体原液，配制如表13所示的31号处方制剂进行制剂的长期稳定性研究。
放置2～8℃条件下12个月，对各样品进行分析测试。通过以下参数评估稳定性：(a)CE-SDS(十二烷基硫酸钠毛细管电泳)法检测抗体的分子量；(b)SEC-HPLC测量抗体单体(质量标准：≥97.0％)、聚体(质量标准：≤3.0％)或片段的含量(质量标准：≤1.0％)；(c)CEX-HPLC测量抗体主要电荷(质量标准：≥70.0％)、酸性电荷(质量标准：≤30.0％)或碱性电荷含量(质量标准：≤15.0％)；生物学活性(质量标准：参比品的70％-130％)。
结果显示，4批次原液在31号制剂处方的纯度(SEC-HPLC(分子排阻高效液相色谱法)、CEX-HPLC(弱阳离子交换高效液相色谱法)、R-CE-SDS(还原电泳法)、NR-CE-SDS(非还原电泳法)和生物学活性均无显著变化，具体见表14。结果表明，上述4批次原液在31号制剂处方保存在2-8℃条件下，0-12月储存过程具有非常好的稳定性。
表13：制剂长期稳定性研究的制剂处方
Figure PCTCN2021099257-appb-000013
表14：制剂处方的长期稳定性数据
Figure PCTCN2021099257-appb-000014
Figure PCTCN2021099257-appb-000015
Figure PCTCN2021099257-appb-000016
注：表格中NA表示在此时间点为非重点检项，在该取样点不做检测；NR表示未到该时间点，下同；ND表示未检出。
实施例6：制剂的加速稳定性研究
选用4批次原液，配制如表13所示的31号处方制剂分别储存于透明小瓶中，放置25±2℃，60±5％相对湿度(RH)条件下，0-6个月后，对各样品进行分析测试。通过以下参数评估稳定性：(a)CE-SDS(十二烷基硫酸钠毛细管电泳)法检测抗体的分子量；(b)SEC-HPLC测量抗体单体(质量标准：≥97.0％)、聚体(质量标准：≤3.0％)或片段的含量(质量标准：≤1.0％)；(c)CEX-HPLC测量抗体主要电荷(质量标准：≥70.0％)、酸性电荷(质量标准：≤30.0％)或碱性电荷含量(质量标准：≤15.0％)；(d)生物学活性(质量标准：参比品的70％-130％)具体见表15。
在加速稳定性条件(25±2℃，60±5％RH)下储存6个月，CEX-HPLC主峰下降，酸性峰升高，6个月超出质量标准；SEC-HPLC单体含量和R-CE-SDS的纯 度有下降趋势，但未超出质量标准；生物学活性无显著变化。
表15：制剂处方的加速稳定性数据
Figure PCTCN2021099257-appb-000017
Figure PCTCN2021099257-appb-000018
注：表格中NA表示在此时间点为非重点检项，在该取样点不做检测；NR表示未到该时间点；ND表示未检出。
实施例7：人源化抗体与人IL-17A的结合特异性
利用常规ELISA检测方法检测不同人源化抗体(按处方31的配方配置)与人IL-17A结合特异性。即将0.5μg/ml的人IL-17A-mFc包被于96孔酶标板，37摄氏度恒温孵育60-90分钟。然后弃去孔内溶液，用洗涤缓冲液洗3次，加入含有2％BSA的PBS溶液封闭60分钟。用洗涤缓冲液洗3次后加入不同浓度的人源化抗体稀释液，37摄氏度孵育60分钟后用洗涤缓冲液冲洗3次，然后加入1：10000倍稀释的生物素-抗IgG4，37摄氏度孵育1小时，经洗涤缓冲液冲洗三次后，加入 用洗涤缓冲液以1：10000倍稀释的HPR-Strep，室温孵育1小时，经洗涤缓冲液冲洗3次后，加入100μL TMB底物溶液显色，室温反应30分钟后，以100μL 2M的盐酸溶液终止反应并在450nm处读出吸光度。如图1所示，人源化抗体hu31，hu43，hu44，hu59，hu60和hu250均与IL-17A特异性结合。其EC50分别为8.13ng/mL，8.64ng/mL，6.76ng/ml，6.10ng/mL，5.78ng/mL和6.35ng/mL。
实施例8：人源化抗体阻断人IL-17A与IL-17RA的结合作用
采用竞争性基于细胞的流式细胞测定(FACS)，检测不同人源化抗体(按处方31的配方配置)阻断IL-17A与细胞上IL-17RA的结合作用。简而言之，将不同浓度的人源化抗体稀释液(起始10ug/ml，3倍滴定)与预先生物素标记的实施例1获得的人IL-17A-mFC(3ug/ml)混合，室温孵育30分钟。然后将混合物与细胞悬液(实施例2获得的293F IL-17RA稳转细胞株，1.5×10 5个细胞/孔)在37摄氏度孵育15分钟，以PBS洗脱3次后，加入5μg/ml的抗小鼠IgG并室温孵育30分钟。以PBS洗脱3次后，通过流式细胞仪检测人源化抗体对IL-17A与293F细胞表面的IL-17RA结合的抑制作用。如图2所示，人源化抗体hu31，hu43，hu44，hu59，hu60和hu250均能显著地抑制IL-17A与细胞上的IL-17RA特异性结合。其IC50分别为867.6ng/mL，780.8ng/mL，828.5ng/ml，467.4ng/mL，482.8ng/mL和577.8ng/mL。
实施例9：人源化抗体拮抗IL-17A诱导上皮细胞表达CXCL1
IL-17A可以刺激多种上皮细胞和其它细胞分泌细胞因子CXCL1表达与释放，可通过ELISA定量检测细胞上清中CXCL1表达量变化，判断不同人源化抗体(按处方31的配方配置)对IL-17A在细胞中介导的生物活性的影响。
使用标准技术在组织培养物处理的烧瓶中的培养/测定培养基中维持HT-29细胞(人结肠直肠腺癌上皮细胞，ATCC)。HT-29在组织培养瓶中生长直至它们在测定当天达到50-80％汇合。在测定当天，用PBS冲洗细胞并用胰蛋白酶+EDTA从培养瓶中脱离细胞，并制成细胞悬液。取人源化抗体hu31、hu43、hu44、hu59、hu60、hu250或参照抗体(Secukinumab，诺华)稀释液(起始浓度为55ug/ml，3倍浓度梯度稀释)与人IL-17A(1ug/ml)混合，铺入96孔板中，孵育1h。向每孔 中加入100ul(2×104个)HT-29细胞(ATCC，人结肠直肠腺癌上皮细胞)悬液，37℃，7％CO 2培养48h。离心，将培养上清转入新的96孔板中，利用ELISA试剂盒检测CXCL1的表达。
如图3所示，相比参照抗体Secukinumab，人源化抗体hu31、hu43、hu44、hu59、hu60和hu250对IL-17A刺激上皮细胞释放CXCL1具有更强的拮抗作用。
实施例10：人源化抗体拮抗IL-17A诱导小鼠表达CXCL1
通过检测小鼠血清CXCL1水平变化，判断不同人源化抗体(按处方31的配方配置)对IL-17A在体内介导生物活性的影响。简言之，选择40只10周龄的雌性Balb/c小鼠，分为8组，每组5只。给药前4天，搜集血清，并检测CXCL1表达量作为基础值。给药当天，按1mg/kg，分别心内注射候选抗体(人源化抗体hu31，hu43，hu44，hu60和hu250)和IgG4同种型对照(hIgG)；给药后1小时，按150ug/kg给药量通过皮下注射人IL-17A；人IL-17A注射后2小时，搜集血清检测血液中CXCL1的浓度，并与基础值进行对比，计算各组在给药前后CXCL1的浓度变化倍数(平均值±标准误差)。候选抗体与IgG4同种型对照的比较分析使用Student’s-t test检验，P<0.05认为有显著差异，*P<0.05，**P<0.01，***P<0.001。
如图4所示，相较于IgG4同种型对照，候选人源化抗体hu31，hu43，hu44，hu60和hu250对IL-17A刺激小鼠体内释放CXCL1具有更强的拮抗作用。
实施例11：人源化抗体改善咪喹莫特诱导的小鼠银屑病模型的疗效研究
在小鼠耳背皮肤涂抹咪喹莫特可诱导银屑病样病理学特征，即角质形成细胞过度增生、炎症细胞聚集和真皮乳头部血管增生等，构建银屑病小鼠模型。以临床评分、耳部肿胀度等为指标判断药物对银屑病小鼠的治疗作用。
1、实验方法
取6-8周C57BL/6雌性小鼠(南京大学模式动物研究所购买，动物合格证号为201605578)48只，背部脱毛，除假手术组(“sham”)外，二天后致敏。致敏前两天随机分成6组(每组8只)：I组为假手术组；II组为PBS组，III组为KLH对照组(同型IgG)，给予KLH；IV组为hu31组；V组为hu43给药组；VI组为hu44给药组，各组给药剂量均为50mg/kg；上述各组于分组第0天及第3天 腹腔注射药物1次。致敏当天(第1天)，II-VI组小鼠于右耳及背部皮肤上涂抹约62.5mg咪喹莫特乳膏(艾达乐，5％，3M Health Care Limited)，连续4天。人源化抗体按处方31配置。
2、评价方法
致敏当天起每天用螺旋测微仪测量小鼠右耳厚度，以第1天右耳厚度为对照，计算小鼠耳肿胀厚度数值。同时给小鼠每天称重，观察皮肤鳞屑、硬结、红斑情况，进行评分，采用4级评分法：0分，不发病；1分，轻微；2分，中度；3分，严重；4分，非常严重。结果以平均值±标准误差表示，先使用单因素方差分析(ANOVA)，有差异后两组之间的比较使用Student’s-t test检验，P<0.05认为有显著差异。
如图5-1所示，给予本发明人源化抗体可明显抑制咪喹莫特诱导的小鼠银屑病模型皮肤鳞屑、硬结、红肿等情况，即评分值较小。
如图5-2所示，自致敏当天起于小鼠右耳涂抹咪喹莫特乳膏，其右耳会发生严重肿胀，耳厚度增加，而本发明人源化抗体均可明显改善耳肿胀程度。
在咪喹莫特诱导的小鼠银屑病模型中，本发明人源化抗体可明显抵抗小鼠的发病，表型为小鼠临床评分及耳部肿胀程度的降低。
实施例12：人源化抗体改善II型胶原蛋白诱导的雌性食蟹猴关节炎的疗效研究
II型胶原蛋白诱导的关节炎是广泛用于类风湿关节炎(RA)研究的动物模型，具有与人RA相同的组织病理学特征，其特征在于小关节的炎症和软骨及骨的进行性侵蚀。人/人源化的生物大分子包括抗体常常与食蟹猴体内抗原具有更优的交叉反应，所以食蟹猴关节炎模型是检测本发明中人源化抗体IL-17A抗风湿作用的一种有效体系。本实验即在食蟹猴类风关节炎模型上评价候选抗体药效。
1、实验方法
将II型牛胶原(CII，四川大学)溶解在醋酸(货号10000218；国药；上海；中国)中放入4℃冰箱搅拌过夜，然后用等体积的完全弗氏佐剂(货号：F5881，Sigma-Aldrich，美国)乳化胶原，乳剂胶原终浓度2mg/ml。第0天，使用舒泰(1.5-5mg/kg，i.m)麻醉动物，于背部及尾根部进行多点的胶原乳剂免疫注射，根据需要， 免疫时用1.5％-5％的异氟烷维持麻醉。3个星期后(第21天)重新注射一次胶原，方法同第一次。本实验分为4组，G1为正常动物组(“naive”)，不进行关节炎的诱导；G2为溶媒对照组(“vehicle”)；G3为抗体hu31给药组；G4为抗体hu59给药组。当某只动物的临床评分达到临床评分最大值的5％(192×5％≈10)时，按顺序分别分到各实验组，先达分值的先入组，如此循环直到所有符合条件的动物都依序分入各组中。入组后开始给药，每周一次，每次7.5mg/kg，由输液泵(infusion bump)30分钟持续泵入，持续5周。人源化抗体按处方31配置。
2、评价方法
体重测量：免疫前一天测量动物体重，以后每周测量一次体重直到实验结束。
关节炎评分：在第0天和第21天对猴子四肢关节炎发炎程度进行评分，并在21天后每周评分一次直至实验结束(如果出现提早发病，则相应提前每周一评的关节炎评分)。关节炎的评分标准：0分，正常；1分，轻微关节炎，发病轻微但可以明确分辨出；2分，中度肿胀；3分，严重关节炎，严重肿胀或有明显的关节变形。对每个爪掌的下述15个关节进行评分：5个掌指关节(MCP)，4个近端指关节(PIP)，5个远端指关节(DIP)，1个腕关节或踝关节。同时还需要评估四肢膝盖/肘关节的发病程度。各个关节评分的总和即为这只动物的关节炎评分，最大分值为192(16×3×4)。
实验数据表示成平均值±标准误(mean±S.E.M)。统计分析溶媒对照、参照药和测试药组各项参数的组间差异，p<0.05被认为具有统计学差异(One-way ANOVA/Dunnett)。
如图6-1所示，正常的食蟹猴(G1)体重稳定；关节炎诱导后的食蟹猴，溶媒处理组(G2)的食蟹猴平均体重持续下降，与之相比，受试抗体hu31和hu59均使这种下降趋势得到控制。故在本实验条件下，hu31和hu59对关节炎引起的体重下降具有一定的改善作用(**P<0.01，****P<0.0001，和“G2:溶媒组”相比较；One-way ANOVA/Dunnett)。
如图6-2所示，动物入组后，正常对照动物(G1)关节炎临床评分保持为0；溶媒对照组(G2)动物关节炎评分呈渐进性增加，而受试抗体hu31和hu59显著抑制了动物的关节炎临床评分的增加趋势。故受试抗体hu31和hu59有抑制关节炎病情渐进性发展的作用，受试抗体hu31显著抑制了食蟹猴关节炎临床评分的增加 趋势(***P<0.001，#P<0.05，和溶媒对照组相比较；One-way ANOVA/Dunnett)。
实施例13：人源化抗体对NIH3T3-IL17细胞诱导的小鼠关节肿胀的作用
1.动物：C57BL/6，雌性，6-8周，北京维通利华实验动物技术有限公司。
2.细胞：NIH3T3细胞，表达人IL-17的NIH3T3细胞。
3.分组及给药方案：
NIH3T3组；
NIH3T3-IL-17+对照IgG抗体组(30mg/kg)；
NIH3T3-IL-17+受试抗体高剂量给药组(抗体hu31,3mg/kg)；
NIH3T3-IL-17+受试抗体中剂量给药组(抗体hu31,10mg/kg)；
NIH3T3-IL-17+受试抗体低剂量给药组(抗体hu31,30mg/kg)；
NIH3T3-IL-17+阳性药给药组(cosentyx,10mg/kg)。
人源化抗体按处方31配置。
4.造模及给药：
将NIH3T3-IL-17细胞及NIH3T3对照细胞(2.5×10 5个/只，每只注射体积为25uL)分别注射入各组小鼠的右踝关节的关节腔内。
选模前1天开始腹腔注射给予抗体hu31(3、10、30mg/kg)和cosentyx(10mg/kg)干预给药，每3天1次，考察抗体hu31注射液对于小鼠关节炎模型的影响。
5.检测：
游标卡尺测量小鼠踝关节厚度，计算肿胀度。
6.实验结果：
如图7所示，在小鼠跺关节腔内注射稳转hIL-17的NIH3T3细胞，第二天可观察到小鼠跺关节处严重肿胀。
依据小鼠踝关节的厚度进行肿胀抑制率的计算，计算公式为：抑制率(％)＝(NIH3T3-ILI7组踝关节厚度-给药组踝关节厚度)/(NIH3-ILl7细踝关节厚度-NIH3T3组踝关节厚度)×100。结果显示，给药后第2天，观察到各剂量药物组别对小鼠踝关节肿胀的抑制作用，直至试验结束，在给药后第6天达到最大抑制效率。第10天时，抗体hu31(3、10、30mg/kg)各组抑制率分别为49.4％、65.9％和74.1％。cosentyx(10mg/kg)肿胀抑制率为67.1％。计算各组不同天数的平均抑 制率，结果显示，抗体hu31各组(3、10和、30mg/kg)的平均抑制率分别为45.2％、57.0％和73.9％。cosentyx(10mg/kg)肿胀抑制率为60.4％。实验结果提示：抗体hu31能剂量依赖性地抑制IL-17诱导的小鼠关节踝肿胀，其10mg/kg的效果与阳性对照药物cosentyx(10mg/kg)相当。30mg/kg的效果优于阳性对照药物cosentyx(10mg/kg)。
实施例14：人源化抗体对NIH3T3-IL17细胞诱导的小鼠空气囊炎症的作用
1.动物：C57BL/6，雄性，6-8周，由北京维通利华实验动物技术有限公司购买)。
2.细胞：NIH3T3细胞，表达人IL-17的NIH3T3细胞。
3.试剂：Gr1-FITC抗体，Biolegend公司。
4.分组及给药方案：
NIH3T3细胞组
NIH3T3-IL-17细胞组+对照IgG抗体组(30mg/kg)；
NIH3T3-IL-17细胞组+受试抗体高剂量给药组(抗体hu31,30mg/kg)；
NIH3T3-IL-17细胞组+受试抗体中剂量给药组(抗体hu31,10mg/kg)；
NIH3T3-IL-17细胞组+受试抗体低剂量给药组(抗体hu31,3mg/kg)；
给药途径：腹腔注射，人源化抗体按处方31配置。
5.造模：
空气囊：分别在第0和3天在小鼠背部注射2.5ml空气。第5天开始将细胞注入空气囊内。每只小鼠注射的细胞数量为2×10 5个/500微升PBS。每组8只小鼠。
6.检测：
白细胞向空气囊的迁移，计算灌洗液的细胞总数，流式测定Gr1+细胞比例，计算中性粒细胞数量；其中，中性粒细胞数量＝细胞总数×Gr1+细胞比例。
7.实验结果
如图8所示，在小鼠背部空气囊内注射稳转hlL-17A的NIH-3T3细胞，从浸润的细胞总数来看，相比于NIH3T3细胞组，NIH3T3-IL-17细胞组空气囊中浸润的白细胞数显著增加，Grl+细胞比例和数量也显著增加，选模成功。造模当天腹腔 注射给予抗体hu31(剂量分别为3、l0、30mg/kg)干预给药。依据浸润的总细胞数和Grl+细胞数进行抑制率的计算，计算公式为：抑制率(％)＝(NIH3T3-ILI7-lgG组细胞数-给药组细胞数)/(NIH3T3-ILl7-IgG组细胞数-NIH3T3细胞数)×100。结果显示，抗体hu31各组(3、10、30mg/kg)浸润细胞总数抑制率分别为50.0％、56.7％和78.3％。计算Grl+细胞数的抑制率，结果显示：抗体hu31各组(3、10、30mg/kg)平均Grl+细胞抑制率分别为59.1％、54.5％和81.8％，抗体hu31能剂量依赖性的抑制IL-17诱导的小鼠总细胞及炎症细胞浸润。

Claims (15)

  1. 一种药物组合物，包含：
    (1)缓冲液；和
    (2)抗IL-17A抗体或其抗原结合片段；
    其中所述抗IL-17A抗体或其抗原结合片段包含选自以下(Ⅰ)至(Ⅲ)中的任一项：
    (Ⅰ)氨基酸序列分别如SEQ ID NO:1、SEQ ID NO:2和SEQ ID NO:3所示的HCDR1、HCDR2和HCDR3，和氨基酸序列分别如SEQ ID NO:4、SEQ ID NO:5和SEQ ID NO:6所示的LCDR1、LCDR2和LCDR3；或
    (Ⅱ)氨基酸序列分别如SEQ ID NO:7、SEQ ID NO:8和SEQ ID NO:9所示的HCDR1、HCDR2和HCDR3，和氨基酸序列分别如SEQ ID NO:10、SEQ ID NO:11和SEQ ID NO:12所示的LCDR1、LCDR2和LCDR3；或
    (Ⅲ)氨基酸序列分别如SEQ ID NO:13、SEQ ID NO:14和SEQ ID NO:15所示的HCDR1、HCDR2和HCDR3，和分别如SEQ ID NO:16、SEQ ID NO:17和SEQ ID NO:18所示的LCDR1、LCDR2和LCDR3。
  2. 如权利要求1所述的药物组合物，其中所述缓冲液选自醋酸缓冲液、柠檬酸缓冲液、琥珀酸缓冲液和组氨酸缓冲液；优选地，所述缓冲液为组氨酸缓冲液；优选地，所述组氨酸缓冲液为组氨酸-盐酸盐缓冲液。
  3. 如权利要求1或2所述的药物组合物，其中所述缓冲液的浓度约为10～30mM。
  4. 如权利要求1-3中任一项所述的药物组合物，其中所述缓冲液的pH约为5.0-6.5，优选约为5.0-6.0，更优选约为5.5。
  5. 如权利要求1-4中任一项所述的药物组合物，其中所述药物组合物还包括稳定剂，所述稳定剂选自盐酸精氨酸、脯氨酸、甘氨酸、氯化钠、甘露醇、山梨醇、蔗糖、麦芽糖、木糖醇和海藻糖中的一种或多种；优选地，所述稳定剂选自盐酸精氨酸、盐酸精氨酸和甘露醇的组合、或盐酸精氨酸和蔗糖的组合。
  6. 如权利要求5所述的药物组合物，其中所述稳定剂为浓度约30-200mM的氯化钠；或所述稳定剂为浓度约100-300mM的甘露醇；或所述稳定剂为浓度约100-300mM的山梨醇；或所述稳定剂为浓度约100-300mM的蔗糖；或所述稳定剂为浓度约100-300mM的海藻糖；或所述稳定剂为浓度约30-200mM的盐酸精氨酸； 或所述稳定剂为浓度约100-300mM的脯氨酸；或所述稳定剂为浓度约100-300mM的甘氨酸；或所述稳定剂为约30-200mM的氯化钠与约30-200mM的甘露醇的组合；或所述稳定剂为约30-200mM的氯化钠与约30-200mM的蔗糖的组合；或所述稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的甘露醇的组合；或所述稳定剂为约30-200mM的盐酸精氨酸与约30-200mM的蔗糖的组合。
  7. 如权利要求6所述的药物组合物，其中所述稳定剂为浓度约120mM至约170mM的盐酸精氨酸；或约60-120mM的盐酸精氨酸与约40-80mM的蔗糖的组合；或约60-120mM的盐酸精氨酸与约40-80mM的甘露醇的组合。
  8. 如权利要求1-7中任一项所述的药物组合物，其中所述药物组合物还包括表面活性剂，所述表面活性剂选自聚山梨醇酯80、聚山梨醇酯20或泊洛沙姆188。
  9. 如权利要求8所述的药物组合物，其中所述表面活性剂浓度约为0.01％-0.1％，优选约0.02％-0.08％，更优选约为0.02％-0.04％；优选地，所述表面活性剂为约0.01％-0.1％、优选约0.02％-0.08％、更优选约为0.02％-0.04％的聚山梨醇酯20。
  10. 如权利要求1-9中任一项所述的药物组合物，其中，所述抗IL-17A抗体包含选自以下(Ⅰ)至(Ⅵ)中的任一项：
    (Ⅰ)氨基酸序列如SEQ ID NO:19所示的重链可变区和氨基酸序列如SEQ ID NO:20所示的轻链可变区；或
    (Ⅱ)氨基酸序列如SEQ ID NO:21所示的重链可变区和氨基酸序列如SEQ ID NO:20所示的轻链可变区；或
    (Ⅲ)氨基酸序列如SEQ ID NO:21所示的重链可变区和如氨基酸序列SEQ ID NO:22所示的轻链可变区；或
    (Ⅳ)氨基酸序列如SEQ ID NO:23所示的重链可变区和氨基酸序列如SEQ ID NO:24所示的轻链可变区；或
    (Ⅴ)氨基酸序列如SEQ ID NO:23所示的重链可变区和氨基酸序列如SEQ ID NO:25所示的轻链可变区；或
    (Ⅵ)氨基酸序列如SEQ ID NO:26所示的重链可变区和氨基酸序列如SEQ ID NO:27所示的轻链可变区。
  11. 如权利要求1-10中任一项所述的药物组合物，其中所述抗IL-17A抗体包含选自以下(Ⅰ)至(Ⅵ)中的任一项：
    (Ⅰ)如SEQ ID NO:28所示的重链氨基酸序列和如SEQ ID NO:29所示的轻链氨基酸序列；或
    (Ⅱ)如SEQ ID NO:30所示的重链氨基酸序列和如SEQ ID NO:29所示的轻链氨基酸序列；或
    (Ⅲ)如SEQ ID NO:30所示的重链氨基酸序列和如SEQ ID NO:31所示的轻链氨基酸序列；或
    (Ⅳ)如SEQ ID NO:32所示的重链氨基酸序列和如SEQ ID NO:33所示的轻链氨基酸序列；或
    (Ⅴ)如SEQ ID NO:32所示的重链氨基酸序列和如SEQ ID NO:34所示的轻链氨基酸序列；或
    (Ⅵ)如SEQ ID NO:35所示的重链氨基酸序列和如SEQ ID NO:36所示的轻链氨基酸序列。
  12. 如权利要求1-11中任一项所述的药物组合物，其中所述抗IL-17A抗体或其抗原结合片段浓度约为20-200mg/mL，优选约为60-180mg/mL，更优选约为80-150mg/mL。
  13. 如权利要求1-12中任一项所述的药物组合物，其包含如下(1)-(12)中任一项所示的组分：
    (1)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约100-300mM的山梨醇；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
    (2)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的盐酸精氨酸；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
    (3)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的氯化钠与约30-200mM的甘露醇的组合；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
    (4)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的氯化 钠与约30-200mM的蔗糖的组合；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
    (5)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的盐酸精氨酸与约30-200mM的甘露醇的组合；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
    (6)(a)约60mg/mL-180mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.0-6.0；(c)约30mM至约200mM的盐酸精氨酸与约30-200mM的蔗糖的组合；(d)以及约0.01％-0.1％的聚山梨醇酯20或聚山梨醇酯80；或
    (7)(a)约80mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约20mM组氨酸缓冲液，pH约为5.5；(c)约50mM的氯化钠与约140mM的甘露醇的组合；(d)以及约0.02％的聚山梨醇酯20；或
    (8)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约30mM组氨酸缓冲液，pH约为5.5；(c)约135mM的盐酸精氨酸；(d)以及约0.02％的聚山梨醇酯20；或
    (9)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约30mM组氨酸缓冲液，pH约为5.5；(c)约50mM的氯化钠与约120mM的蔗糖的组合；(d)以及约0.02％的聚山梨醇酯20；
    (10)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约30mM组氨酸缓冲液，pH约为5.5；(c)约90mM的盐酸精氨酸与约50mM的甘露醇的组合；(d)以及约0.02％的聚山梨醇酯20；
    (11)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约30mM组氨酸缓冲液，pH约为5.5；(c)约90mM的盐酸精氨酸与约50mM的蔗糖的组合；(d)以及约0.02％的聚山梨醇酯20；
    (12)(a)约150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约15mM组氨酸缓冲液，pH约为5.5；(c)约90mM的盐酸精氨酸与约50mM的蔗糖的组合；(d)以及约0.02％的聚山梨醇酯20。
  14. 如权利要求1-12中任一项所述的药物组合物，其包含如下(i)-(vi)任 一项所示的组分：
    (i)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约200-280mM的山梨醇；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
    (ii)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约120mM至约170mM的盐酸精氨酸；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
    (iii)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约40mM至约100mM的氯化钠与约40-150mM的甘露醇的组合；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
    (iv)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约40mM至约100mM的氯化钠与约40-150mM的蔗糖的组合；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
    (v)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约60mM至约120mM的盐酸精氨酸与约40-80mM的甘露醇的组合；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80；或
    (vi)(a)约80mg/mL-150mg/mL的抗IL-17A抗体或其抗原结合片段；(b)约10-30mM组氨酸缓冲液，pH约为5.5-6.0；(c)约60mM至约120mM的盐酸精氨酸与约40-80mM的蔗糖的组合；(d)以及约0.02％-0.04％的聚山梨醇酯20或聚山梨醇酯80。
  15. 如权利要求1-14中任一项所述的药物组合物在制备治疗或预防IL-17A介导的疾病的药物中的用途；优选地，所述疾病包括炎性疾病和自身免疫性疾病，优选选自：关节炎、类风湿性关节炎、强直性脊柱炎、慢性阻塞性肺疾病、系统性红斑狼疮(SLE)、狼疮性肾炎、哮喘、多发性硬化、囊性纤维化和银屑病。
PCT/CN2021/099257 2020-06-10 2021-06-09 抗il-17a抗体药物组合物及其用途 WO2021249445A1 (zh)

Priority Applications (3)

Application Number Priority Date Filing Date Title
EP21820830.4A EP4166159A1 (en) 2020-06-10 2021-06-09 Pharmaceutical composition of anti-il-17a antibodies and use thereof
CN202180041325.2A CN115768476A (zh) 2020-06-10 2021-06-09 抗il-17a抗体药物组合物及其用途
US18/001,106 US20230212278A1 (en) 2020-06-10 2021-06-09 Pharmaceutical composition of anti-il-17a antibodies and use thereof

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN202010524523.1A CN113769082A (zh) 2020-06-10 2020-06-10 抗il-17a抗体药物组合物及其用途
CN202010524523.1 2020-06-10

Publications (1)

Publication Number Publication Date
WO2021249445A1 true WO2021249445A1 (zh) 2021-12-16

Family

ID=78834685

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2021/099257 WO2021249445A1 (zh) 2020-06-10 2021-06-09 抗il-17a抗体药物组合物及其用途

Country Status (4)

Country Link
US (1) US20230212278A1 (zh)
EP (1) EP4166159A1 (zh)
CN (2) CN113769082A (zh)
WO (1) WO2021249445A1 (zh)

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2024120518A1 (zh) * 2022-12-09 2024-06-13 上海华奥泰生物药业股份有限公司 Il-17a抗体制剂及其用途
CN118078988A (zh) * 2024-04-19 2024-05-28 正大天晴药业集团南京顺欣制药有限公司 包含靶向il-17a的抗体药物组合物

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009082624A2 (en) * 2007-12-10 2009-07-02 Zymogenetics, Inc. Antagonists of il-17a, il-17f, and il-23 and methods of using the same
CN106474470A (zh) * 2015-08-28 2017-03-08 江苏恒瑞医药股份有限公司 一种抗il‑17a抗体的组合物
WO2019101582A1 (en) * 2017-11-23 2019-05-31 Ucb Biopharma Sprl Formulation
CN110585430A (zh) * 2019-09-29 2019-12-20 华博生物医药技术（上海）有限公司 一种人源化抗人il-17a单克隆抗体的药物组合物
CN111303283A (zh) * 2018-12-12 2020-06-19 上海君实生物医药科技股份有限公司 抗il-17a抗体及其应用

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009082624A2 (en) * 2007-12-10 2009-07-02 Zymogenetics, Inc. Antagonists of il-17a, il-17f, and il-23 and methods of using the same
CN106474470A (zh) * 2015-08-28 2017-03-08 江苏恒瑞医药股份有限公司 一种抗il‑17a抗体的组合物
WO2019101582A1 (en) * 2017-11-23 2019-05-31 Ucb Biopharma Sprl Formulation
CN111303283A (zh) * 2018-12-12 2020-06-19 上海君实生物医药科技股份有限公司 抗il-17a抗体及其应用
CN110585430A (zh) * 2019-09-29 2019-12-20 华博生物医药技术（上海）有限公司 一种人源化抗人il-17a单克隆抗体的药物组合物

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
CHOTHIA ET AL., MOL. BIOL., vol. 196, 1987, pages 901 - 917
CHOTHIA ET AL., NATURE, vol. 342, 1989, pages 877 - 883
MACCALLUM, J. MOL. BIOL., vol. 262, no. 5, 1996, pages 732 - 45
PADLAN, FASEB J, vol. 9, 1995, pages 133 - 139

Also Published As

Publication number Publication date
US20230212278A1 (en) 2023-07-06
CN115768476A (zh) 2023-03-07
CN113769082A (zh) 2021-12-10
EP4166159A1 (en) 2023-04-19

Similar Documents

Publication Publication Date Title
CN114616249B (zh) 含有抗pd-l1抗体的稳定制剂
CN107257692B (zh) Il-17抗体的药物产品和稳定液体组合物
WO2021249445A1 (zh) 抗il-17a抗体药物组合物及其用途
WO2020259605A1 (zh) 包含抗cd47/pd-l1双特异性抗体的制剂及其制备方法和用途
CN113797333A (zh) 一种新型冠状病毒抗体的药物组合物及其用途
US20230174638A1 (en) Formulations of anti-il-33 antibodies
TW202011995A (zh) 高濃度液體抗體配製物
WO2022152245A1 (zh) 抗tigit抗体药物组合物及其用途
KR20210078514A (ko) 항-rsv 항체의 제제 및 그의 사용 방법
WO2020200099A1 (zh) 包含抗il-5抗体的药物组合物及其用途
EP4151233A1 (en) Preparation comprising anti-il-23p19 antibody, preparation method therefor and use thereof
JP7502286B2 (ja) 抗ｐｃｓｋ９抗体を含む安定製剤
WO2023226617A1 (zh) 一种稳定的抗体制剂
CN116725961A (zh) 抗cd39抗体药物组合物及其用途
CN116803420A (zh) 靶向PD-1和TGFβ的双功能蛋白药物组合物及其用途
CN116725960A (zh) 新型冠状病毒抗体药物组合物及其用途
WO2022223028A1 (zh) 抗BLyS抗体、其药物组合物及其用途
CN118078988A (zh) 包含靶向il-17a的抗体药物组合物
KR20240021800A (ko) 항체 조성물
CN117999085A (zh) 用于治疗或预防冠状病毒感染的干扰素相关的抗原结合蛋白
CN116019907A (zh) 抗pd-1抗体药物组合物及其用途

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: 21820830

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

ENP Entry into the national phase

Ref document number: 2021820830

Country of ref document: EP

Effective date: 20230110